Dopamine D1-Like, D2 and D3 Receptor Subtypes in Catalepsy Sensitization and Conditioning in Rats: Implications for Motor Function, Motivation and Learning

DOPAMINE D1-LIKE, D2 AND D3 RECEPTOR SUBTYPES IN CATALEPSY SENSITIZATION AND CONDITIONING IN RATS: IMPLICATIONS FOR MOTOR FUNCTION, MOTIVATION AND LEARNING

Tomek Jan Banasikowski

A thesis submitted to the Centre for Neuroscience Studies

In conformity with the requirements for

the degree of Doctor of Philosophy

Queen’s University

Kingston, Ontario, Canada

July, 2012

The behavioral effects of drugs that act on the brain’s dopamine (DA) system change with repeated exposure to the drug. Antipsychotic drugs, that block DA receptors, produce progressively greater effects on behavior with repeated testing. For example, rats repeatedly treated with a low dose of the D2 receptor-preferring antagonist haloperidol do not initially exhibit catalepsy, a response quantified by time spent on a horizontal bar without active movement. However, with repeated drug-environment pairings animals show a reduction in exploration and increases in catalepsy. The current thesis examined the drug-environment relationship in catalepsy sensitization, and how different DA receptor subtypes control this phenomenon.

Treatment with a D2, but not D3 or D1-like receptor-preferring antagonist produced catalepsy sensitization. Catalepsy sensitization developed in one test environment did not transfer to another environment. Similarly, rats with a history of haloperidol treatments outside of the test environment (unpaired group) did not exhibit significant catalepsy when given haloperidol for the first time prior to catalepsy testing. Previous exposure to the catalepsy test environment led to a more rapid development of catalepsy sensitization. Thus, drug-environment interaction is critical for the development and expression of catalepsy sensitization.

Rats previously given haloperidol and tested with saline in the drug paired environment exhibited conditioned catalepsy. The acquisition of conditioned catalepsy is dependent on D1- like receptors, while its expression is dependent on D3 receptors. Conditioned catalepsy showed gradual day-to-day extinction with repeated saline treatment in the previously haloperidol-paired environment. Following extinction, the response to haloperidol in previously sensitized rats shifted from environment-specific to environment-independent suggesting that a putative ii haloperidol drug cue alone can elicit conditioned catalepsy.

In summary, treatment with a D2, but not D1-like or D3 receptor-preferring antagonist in a particular test environment produces catalepsy sensitization, while acquisition of conditioned catalepsy is dependent on D1-like receptors, and its expression is dependent on D3 receptors.

Importantly, the acquisition and expression of sensitization to haloperidol is conditional on the presence of drug-associated environmental stimuli. Our findings provide further insight into the current understanding of learning processes involved in the action of antipsychotic drugs and the dissociable effects of D1-like, D2 and D3 receptors controlling this phenomenon.

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Co-Authorship

In all cases, I (Tomek J. Banasikowski) designed the experiments, collected and analyzed data and wrote the manuscripts. Richard J. Beninger contributed to data analysis and manuscript preparation.

Chapter 2 has been published in its entirety online, and can be cited as:

Banasikowski TJ, Beninger RJ (2012). Haloperidol conditioned catalepsy in rats: a possible role for D1-like receptors. The International Journal of Neuropsychopharmacology, 1-10, DOI: http://dx.doi.org/10.1017/S1461145711001696 (published online: 18 November 2011).

Chapter 3 has been published in its entirety online, and can be cited as:

Banasikowski TJ, Beninger RJ (2012). Reduced expression of haloperidol conditioned catalepsy in rats by the dopamine D3 receptor antagonists nafadotride and NGB 2904. European Neuropsychopharmacology, 1-8, DOI: http://dx.doi.org/10.1016/j.euroneuro.2012.02.004, (published online: 10 March 2012).

Chapter 4 is to be submitted for publication:

Banasikowski TJ, Beninger RJ (2012). Drug-environment interaction controls the acquisition and expression of haloperidol catalepsy sensitization and conditioning. (To be submitted July 2012).

Know rather that we must turn to nature itself, to the observation of the body in health and

disease, to learn the truth.

Hippocrates

Table of Contents

Abstract ...... ii Co-Authorship...... iv List of Figures ...... ix List of Abbreviations……………………………………………………………………………....x Chapter 1 General Introduction...... 1 1.1 Dopamine Neuron Anatomy ...... 2 1.2 Dopamine Neuron Physiology ...... 4 1.3 Dopamine Receptor Subtypes ...... 5 1.4 Dopamine and Motor Function ...... 7 1.5 Dopamine and Motivation ...... 9 1.6 Dopamine and Stimulus-Reward Learning ...... 14 1.7 Dopamine and Behavioral Sensitization ...... 18 1.8 Hypothesis...... 21 1.9 References ...... 23 Chapter 2 Haloperidol Conditioned Catalepsy in Rats: A Possible Role for D1-like Receptors ... 37 2.1 Abstract ...... 38 2.2 Introduction ...... 39 2.3 Materials and Methods ...... 41 2.3.1 Subjects ...... 41 2.3.2 Drugs ...... 41 2.3.3 Behavioral Testing ...... 41 2.3.4 Experimental Design ...... 42 2.3.5 Statistics ...... 44 2.4 Results ...... 44 2.4.1 Experiment 1: Haloperidol Catalepsy ...... 44 2.4.2 Experiment 2: SCH 23390 Catalepsy ...... 47 2.4.3 Experiment 3: Haloperidol + SCH 23390 Catalepsy ...... 47 2.5 Discussion ...... 48 2.6 References ...... 59 Chapter 3 Reduced Expression of Haloperidol Conditioned Catalepsy in Rats by the Dopamine D3 Receptor Antagonists Nafadotride and NGB 2904 ...... 67 3.1 Abstract ...... 68

3.2 Introduction ...... 69 3.3 Materials and Methods ...... 71 3.3.1 Subjects ...... 71 3.3.2 Drugs ...... 71 3.3.3 Behavioral Testing ...... 72 3.3.4 Experimental Design ...... 72 3.3.5 Statistics ...... 75 3.4 Results ...... 75 3.4.1 Experiment 1: Nafadotride Effects on Acquisition and Expression of Catalepsy ...... 75 3.4.2 Experiment 2: NGB 2904 Effects on Acquisition and Expression of Catalepsy ...... 78 3.5 Discussion ...... 81 3.6 References ...... 88 Chapter 4 Drug-Environment Interaction Controls the Acquisition and Expression of Haloperidol Catalepsy Sensitization and Conditioning ...... 93 4.1 Abstract ...... 94 4.2 Introduction ...... 95 4.3 Materials and Methods ...... 98 4.3.1 Subjects ...... 98 4.3.2 Drugs ...... 98 4.3.3 Behavioral Testing ...... 99 4.3.4 Experimental Design ...... 99 4.3.5 Statistics ...... 103 4.4 Results ...... 104 4.4.1 Experiment 1: Environment-Specific Expression of Haloperidol Catalepsy Sensitization and Conditioning...... 104 4.4.2 Experiment 2: Associative or Non-Associative Processes Involved in Expression of Haloperidol Catalepsy Sensitization ...... 107 4.4.3 Experiment 3: Environment Pre-Exposure and Acquisition of Future Haloperidol Catalepsy Sensitization ...... 108 4.5 Discussion ...... 111 4.6 References ...... 118 Chapter 5 General Discussion ...... 125 5.1 Experimental Summary ...... 126 5.2 Dopamine Receptor Subtypes and Behavior ...... 128 vii

5.2.1 Behavioral Sensitization ...... 128 5.2.2 Incentive (Stimulus – Outcome) Learning and Motivation ...... 132 5.3 Dopamine and Striatal Plasticity ...... 135 5.4 Dopamine, Schizophrenia and Antipsychotic Drug Effects ...... 138 5.5 References ...... 141

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List of Figures

Figure 1 Haloperidol Catalepsy and Conditioning...... 46 Figure 2 SCH 23390 Catalepsy and Conditioning ...... 49 Figure 3 Haloperidol + SCH 23390 Catalepsy and Conditioning ...... 50 Figure 4 Nafadotride Catalepsy and Conditioning...... 77 Figure 5 NGB 2904 Catalepsy and Conditioning ...... 80 Figure 6 Environment-Specific Expression of Catalepsy Sensitization and Conditioning ...... 105 Figure 7 Environment-Specific Acquisition of Catalepsy Sensitization ...... 109 Figure 8 Effect of Test Environment Pre-Exposure on Catalepsy Sensitization ...... 110

List of Abbreviations

6-OHDA 6-hydroxydopamine

AMPH amphetamine

ANOVA analysis of variance

CPP conditioned place preference

Cyclic AMP cyclic adenosine monophosphate

DA dopamine

GABA γ-Aminobutyric acid

Glu glutamate

GPe external globus pallidus

GPi internal globus pallidus

G-protein guanine nucleotide-binding protein

HPC hippocampus

IEG immediate early gene

MFB medial forebrain bundle

MSN medium spiny neuron mRNA messenger ribonucleic acid

NAc nucleus accumbens

NMDA N-Methyl-D-aspartate acid

LTD long-term depression

LTP long-term potentiation

PD Parkinson’s disease

PET positron emission tomography

PFC prefrontal cortex

SNpc substantia nigra pars compacta

SNr substantia nigra pars reticulata

STN subthalamic nucleus

VTA ventral tegmental area

Chapter 1

General Introduction

Pathological disturbances of the brain dopamine (DA) system have been implicated in a number of neurological and neuropsychiatric disorders, including

Parkinson’s disease (PD), schizophrenia, depression and drug addiction (Schmidt and

Beninger, 2006). The behavioral effects of drugs that act on the brain’s DA system change with repeated exposure to the drug. This may be particularly important in understanding the response to treatment of psychosis in schizophrenia or motor symptoms in PD. Despite efforts invested in understanding the role of DA in normal and abnormal behaviors, its involvement in motor, motivation and learning processes remains elusive and continues to be a focus of debate.

1.1 Dopamine Neuron Anatomy

Catecholamine histochemical techniques revealed that major groups of DA neurons in the midbrain provide dense ascending axonal projections via the medial forebrain bundle (MFB) to sub-cortical and cortical brain regions (Bjorklund and

Dunnett, 2007; Wise, 2009). For the most part, there are no clear boundaries between these groups as they form a single continuous system sending axons along the long- distance trajectories to influence neuronal function widely throughout the brain, particularly the ventral/dorsal striatum and prefrontal cortex (PFC). Two of these groups

(A8 and A10) originate in the retrorubral area and ventral tegmental area (VTA) and the third group (A9) in the substantia nigra pars compacta (SNpc). The A8 and A10 axons make contact with the limbic regions, the ventral striatum and PFC and are collectively referred to as the mesocorticolimbic DA system. The A9 axons project vastly to the

caudate-putamen (dorsal striatum) and this pathway is referred to as nigrostriatal DA system. Parts of the SNpc also project to some limbic areas as well as the PFC (Wise,

2009).

Projections along the mesocorticolimbic pathway have been the major focus of studies on motivation, cognition and reward-related processes. On the other hand, studies of the significantly larger projections along the nigrostriatal pathway have focused on execution of motor habits because of its association with neurodegeneration of SNpc and

PD. Nevertheless, there is considerable intermixing of the three cell groups and their target projections to the striatum, the main input of basal ganglia (Tepper and Bolam,

2004; Wise, 2009) and thus the two DA systems are often collectively referred to as the mesotelencephalic dopamine system (Banasikowski and Beninger, 2010).

At the level of the striatum, the principal role of the dense DA innervations is to modulate the flow of cortical and thalamic information throughout the basal ganglia.

Most of the DA inputs terminate on spines of GABA releasing medium spiny neurons

(MSN), which comprise 95% of neurons in the ventral and dorsal striatum. The striatal neurons, in turn, project back onto DA cells, facilitating a bidirectional communication pathway. The MSNs are among the most densely spined neurons in the brain, making these cells well designed to integrate information from different sources (Tepper and

Bolam, 2004). As well as dopaminergic afferents, the dorsal striatum receives glutamate

(Glu) inputs from virtually all sensory and motor regions of the cortex and also from specific regions of the thalamus (Fuxe et al., 1977; McGeer et al., 1977). The ventral

striatum, on the other hand, receives Glu inputs largely from the limbic regions such as the amygdala, PFC, hippocampus (HPC) and anterior cingulate cortex. Studies using various lesions, inactivation or neuropharmacological manipulations show that the striatum can be segregated into heterogeneous functional behavioral domains which act in parallel and collectively contribute to optimal performance (Belin and Everitt, 2008;

Belin et al., 2009). Close anatomical relationships between the cortex and the striatal regions exist in the form of cortico-striato-thalamo-cortical loops that allow for parallel processes to converge and integrate emotional, motivational, and cognitive inputs with sensorimotor processes (Alexander et al., 1986; Haber et al., 2000; Sesack and Grace,

2010). DA signals play an important role linking these processes.

1.2 Dopamine Neuron Physiology

DA neurons are characterized by their large cell bodies and long and complicated axonal arbors that include terminals specialized to release its transmitter (DA) into the extracellular space. This allows DA to achieve a broad distribution of its signal with a minimal number of axons. For example, it is estimated that an individual DA neuron gives rise to approximately 400,000 synapses in the striatum (Arbuthnott et al., 1998;

Tepper and Bolam, 2004; Wickens et al., 2007). Furthermore, DA neurons generate atypically long-duration action potentials, as long as 2-3 ms, which limits the maximal firing rates that these neurons can produce. Thus, it is clear that DA neurons are designed to distribute the same ‘promiscuous’ signal to a large number of targets in the striatum, thalamus and the cortex (Arbuthnott et al., 1998; Glimcher, 2011; Sesack and Grace,

2010).

DA neurons exhibit two distinct activity states (Grace and Bunney, 1984). Firstly,

DA neurons exhibit spontaneous baseline activity (3-5 Hz) driven primarily via a pacemaker-like mechanism that controls their resting membrane potential. This activity state has been termed the tonic firing rate. Secondly, when a spontaneously firing DA neuron is activated via glutamatergic inputs, it will exhibit burst firing (20-30 Hz). Burst firing is associated with a rapid change of state that signifies a behaviorally relevant event

(Glimcher, 2011). Burst and population firing activity leads to phasic and tonic DA release, respectively (Grace et al., 2007). Burst firing produces fast and large phasic DA release that activates both intra-synaptic and extra-synaptic D1-like and D2-like receptors. On the other hand, population-firing produces a slower tonic DA release that mainly activates the higher affinity extra-synaptic D2-like receptors (Floresco et al.,

2003; Hong and Hikosaka, 2011; Richfield et al., 1989a; Richfield et al., 1989b).

1.3 Dopamine Receptor Subtypes

Although DA receptors are similar in structure, receptor subtypes differ by their affinity for DA and coupling to downstream effectors, including guanine nucleotide- binding proteins (G proteins). To date, five subtypes of dopamine receptors have been cloned and differentiated into two families based on their pharmacological profile and function: the D1-like receptor family, which includes the D1 and D5 receptors, and the

D2-like receptor family, which includes the D2, D3, and D4 receptors. The D1 receptor binds DA with lower affinity than the D5 receptor, or any of the D2-like receptor family

subtypes (Richfield et al., 1989a). Among the D2-like receptors, the D2 receptor binds to

DA with lower affinity than D3 or D4 receptors (Levant et al., 1995; Roth et al., 1995;

Van Tol et al., 1991). D1-like receptors are post-synaptic receptors and are coupled to stimulatory G proteins that activate adenylate cyclase, and consequently, cyclic adenosine monophosphate (cyclic AMP) (Stoof and Kebabian, 1981). Conversely, dopamine D2,

D3, and D4 receptors are coupled to inhibitory G proteins, which inhibit adenylate cyclase and in turn the production of intracellular cyclic AMP. Unlike D1-like receptors that act post-synaptically, D2 and D3 (but not D4) receptors are thought to function both pre- and post-synaptically. Thus, D2 and D3 receptors may serve as autoreceptors that regulate the release and synthesis of DA and the activity of DA neurons (Jaber et al.,

1996; Missale et al., 1998; Rankin et al., 2010).

Dopamine receptors are widely distributed in the central and peripheral nervous systems, with largest densities found in the striatum, the limbic system and cortex

(Rankin et al., 2010; Sesack and Grace, 2010). Both D1 and D2 receptors are present in the striatum, NAc, olfactory tubercle, HPC, amygdala, septum, hypothalamus, VTA and

SNpc and PFC (Missale et al., 1998; Rankin et al., 2010). The D3 receptor is expressed at roughly 10-fold lower abundance than the D2 receptor, and its distribution is more restricted to the limbic regions with high concentrations in the islands of Calleja, NAc, amygdala and olfactory tubercle (Levant, 1997; Vallone et al., 2000). Smaller levels of

D3 receptor densities have been reported in the striatum, HPC, VTA and SNpc. DA D4 receptors are predominantly expressed in the prefrontal, sensory, motor and cingulate

cortical regions, as well as in the amygdala and HPC. Finally, D5 receptors are mainly expressed within limbic regions including the HPC, NAc, olfactory tubercle, but also in the cortex, hypothalamus and SNpc (Rankin et al., 2010; Vallone et al., 2000).

1.4 Dopamine and Motor Function

Lesions of the DA system with the neurotoxin 6-hydroxydopamine (6-OHDA) lead to severe motor impairments, making the animal akinetic and generally unresponsive to external stimuli (Marshall et al., 1976; Ungerstedt, 1971). Similar effect has been observed in DA-deficient animals (Zhou and Palmiter, 1995). Movement deficits associated with PD have been linked to a loss DA neurons resulting in striatal, and to a lesser extent limbic, DA hypofunction (Braak et al., 2002; Hornykiewicz, 1973; Lim et al., 2009; Papeschi, 1972; Remy et al., 2005). Similarly to lesion studies, DA receptor antagonists dose-dependently suppress spontaneous, and psychomotor stimulant drug- induced locomotor activity (Beninger, 1983; Pijnenburg et al., 1976; Pijnenburg et al.,

1975) while high doses induce catalepsy, a state of akinesia and rigidity where animals maintain externally imposed postures for a prolonged period of time (Costall and Olley,

1971; Fog et al., 1970; Fuxe et al., 1977; Hornykiewicz, 1975; Ossowska, 2002; Sanberg,

1980; Sanberg et al., 1988).

The loss of motor function associated with pharmacological or lesion-induced DA loss can be reversed under certain stimulus conditions (Lynch and Carey, 1987). For example, people with PD who are severely akinetic can “wake up” and show normal gait in response to salient stimuli such as the loud sound of a fire alarm (Schwab and Zieper,

1965; Souques, 1921). This phenomenon is termed paradoxical kinesis (Martin, 1967;

Whishaw et al., 1989). Rats made akinetic with extensive DA depleting lesions also respond appropriately to salient stimuli such as cold baths, tail pinch or the forced swim test (Antelman et al., 1976; Keefe et al., 1989; Marshall et al., 1976). Thus, certain motor impairments associated with DA hypofunction appear to be less motor specific, and more sensory-motor in nature, where motivationally salient stimuli are able to elicit motor responses.

Both DA D1-like and D2 receptors are involved in locomotor activity where they function synergistically (Bateup et al., 2009; Tran et al., 2002; Tran et al., 2005; Tran et al., 2008). In contrast, D3 and D4 receptors do not appear to be significantly involved in motor processes (Beninger and Banasikowski, 2008; Bristow et al., 1997; Levant, 1997).

The classical model of basal ganglia circuitry provides a basic overview of the relationship between D1 and D2 receptors in locomotion (Gerfen, 1988; Mink, 1996).

Depending on their output, the MSNs in the ventral and dorsal striatum express either D1 or D2 receptors but not both (Berendse et al., 1992a; Berendse et al., 1992b; Gerfen and

Young, 1988). MSNs that co-express substance P and dynorphin with GABA, project to the internal capusule of globus pallidus (GPi) and the adjacent substantia nigra pars reticulata (SNr) and belong to the striatonigral or “direct” output pathway of the basal ganglia. On the other hand, neurons that co-express enkephalin and GABA and project to external globus pallidus (GPe) belong to the striatopallidal or “indirect” pathway

(Matamales et al., 2009). D1 receptors are predominantly found in the “direct”, and D2

receptors in the “indirect” pathway respectively (Berendse et al., 1992b; Gerfen and

Young, 1988).

According to the classical model, activation of the direct pathway via D1 receptors is thought to release desired movements due to excitatory action of DA, resulting in increased GABA-ergic input to the GPi/SNr. The inhibition of GPi/SNr in turn relieves inhibition of thalamic neurons thereby facilitating excitation of cortical motor neurons (Bateup et al., 2009). In agreement, pharmacological stimulation of D1 receptors, as well as optogenetic activation of the striatonigral pathway via D1 receptors increases locomotor behavior (Desai et al., 2005; Kravitz et al., 2010). Similarly, activation of D2 receptors can increase locomotion by inhibition of striatal neuron activity in the indirect pathway. This results in a decreased GABA-ergic input into the

GPe, in turn causing strong inhibitory influence on the subthalamic nucleus (STN).

Inhibition of STN results in attenuated excitation of the GPi/SNr which then reduces the inhibitory influence of the GPi/SNr on excitatory thalamocortical neurons thus facilitating locomotor activity (Bateup et al., 2009). In agreement, D2 receptor agonists increase locomotor activity and antipsychotic drugs, which block D2 receptors, attenuate locomotor activity and at high doses produce catalepsy (Mazurski and Beninger, 1991;

Pijnenburg et al., 1976; Sanberg, 1980).

1.5 Dopamine and Motivation

One of the first researchers to link the DA system to basic motivational behaviors such as hunger and thirst was Ungerstedt (1971a), who showed that 6-OHDA lesions not

only produced akinesia, but also caused severe aphasia and adipsia. Ungerstedt (1971b), by using anatomical immohistofluorescent techniques, also showed that ascending DA axons, arising from cell bodies in the SNpc and VTA passed near several hypothalamic nuclei implicated in the regulation of feeding, drinking and sexual behavior. Similarly, motivational deficits have been observed in people with PD (Shore et al., 2011).

Motivated behaviors can be defined as movements towards biologically relevant stimuli that support survival and away from those stimuli that pose a danger or threat.

Stimuli like food, water, a receptive conspecific, and shelter as well as certain drugs and electrical stimulation of certain brain regions have been shown to elicit approach and are worked for, thus are considered rewarding and motivationally significant (White, 1989).

Based on the above definition, both motivation and motor control constructs overlap considerably requiring the need to discern between motivational versus motoric elements while discussing DA function in behavior.

Expression of motivational processes can be further divided into two classes: preparatory and consummatory (Konorski, 1967). Preparatory behaviors are sets of motor acts that precede contact with biologically significant stimuli. Consummatory behaviors are habitual actions or fixed motor programs that complete the sequence of obtaining the reward such as biting, chewing, and swallowing in the case of food motivation. More importantly, different preparatory patterns can lead to identical consummatory responses.

DA transmission in the ventral striatum appears to be critical for eliciting preparatory behaviors, while DA in the dorsal striatum is necessary for completion of consummatory

acts (Robbins and Everitt, 1992).

Preparatory behaviors can be triggered by presentation of stimuli predictive of desired goal (Bindra, 1974; Bindra and Palfai, 1967; Blackburn et al., 1987; Bolles, 1972;

Breland and Breland, 1961; Weingarten, 1984). First, animals need to learn that a specific stimulus or set of stimuli can predict a motivationally significant event. Konorski (1967) termed this form of associative learning - Pavlovian preparatory conditioning. Unlike consummatory conditioning, preparatory learning reflects the acquisition of responses that are not sensory-specific to the particular unconditional stimulus, but, rather behaviors belonging to its motivational class such as approach or withdrawal.

The reward-predictive stimuli are often referred in the literature as incentive stimuli, conditioned reward stimuli or secondary rewards/reinforcers. The key role of incentive stimuli is to maintain responding in the absence of primary rewards. A number of studies have examined the role of DA on incentive motivation, a state in which behavior selected, initiated and maintained by external incentive stimuli that through learned associations with desired outcomes acquire incentive salience and come to predict the availability and location of primary rewards (Ranaldi and Beninger, 1993;

Taylor and Robbins, 1984; Wyvell and Berridge, 2000)

Blackburn et al. (1987) argued that DA receptor antagonism disrupts the expression of preparatory behaviors before impairing consummatory responses.

Treatment with a DA D2 receptor antagonist significantly decreased the number of receptacle entries after presentation of incentive stimuli, without initially affecting food

consumption (Blackburn et al., 1987; Salamone and Correa, 2002). Wise and colleagues showed that D2 receptor antagonism gradually blocked the expression of motivational and rewarding effects of pro-DA drugs like amphetamine (AMPH; Yokel and Wise,

1975; Yokel and Wise, 1976), food (Wise et al., 1978) and brain stimulation (Corbett and

Wise, 1980). Well-trained animals treated with D2-preferring receptor antagonists pimozide or haloperidol initiate normal responding for desired rewards, however with repeated treatment and testing the animals show an extinction-like decline in responding that cannot be explained simply by motor impairments (Franklin and McCoy, 1979;

Yokel and Wise, 1975; Yokel and Wise, 1976). Franklin and McKoy (1979) showed that extinguished lever-pressing responding under pimozide could be reinstated following a presentation of an incentive stimulus previously associated with the availability of brain stimulation reward. Importantly above results showed that pharmacological DA receptor antagonism interfered with motivational processes and not motor execution (Salamone et al., 1990).

Enhancing DA transmission has been shown to facilitate ongoing motivational processes, independently of general behavioral arousal. Systemic or intracranial treatment with DA agonists has been shown to potentiate responding to stimuli predictive of rewards and impede extinction learning (Nicola et al., 2005; Ranaldi and Beninger, 1993;

Taylor and Robbins, 1984; Wolterink et al., 1993; Wyvell and Berridge, 2000). For example infusions of intra-accumbens AMPH enhance responding on a lever that provides access to an incentive stimulus (Taylor and Robbins, 1984), and enhance the

ability of incentive stimuli to control responding for primary rewards (Wyvell and

Berrdige, 2000). The potentiating effect on responding by AMPH is attenuated by inactivation of DA neurons in the VTA and systemic treatment with antagonists targeting

D2 receptors (and to lesser degree D1-like receptors) (Dickinson et al., 2000; Murschall and Hauber, 2006; Ranaldi and Beninger, 1993).

The role of DA in motivation can also be inferred from correlational studies of changes in DA neuron firing and DA transmission in specific brain areas elicited by motivational stimuli. Neurochemical techniques like brain microdialysis show dynamic changes in extracellular concentrations of DA, operating on a timescale of minutes, represent current motivational state of the animal (Phillips et al., 2008). Extracellular DA levels are considered a summation of many action potential of DA neurons firing phasically (Owesson-White et al., 2012).

Studies have shown that motivationally significant stimuli can elicit DA efflux in the NAc and PFC (Ahn and Phillips, 1999; Bassareo and Di Chiara, 1997; Bassareo and

Di Chiara, 1999; Fiorino et al., 1997; Vacca et al., 2007; Wilson et al., 1995). Wilson et al. (1995) found a significant increase in DA efflux in the NAc in rats during the preparatory phase prior to consumption of food or fluid. This effect was only observed in food or water restricted animals. Bassareo and Di Chiara (1997) showed that rats fed ad libitum on rat chow displayed a significant increase in DA in NAc only when rats were given access to different palatable food. Similarly, a significant increase in DA efflux was observed in anticipation of reward when rats were separated from the food reward or

receptive female by a perforated wire screen. A decline in DA efflux paralleled the development of satiety for both the initial food and copulation with the first receptive female. Furthermore, exposure to a novel food or female and the reinstatement of motivated behavior was accompanied by a rebound increase in DA efflux in both NAc and PFC (Ahn and Phillips, 1999; Fiorino et al., 1997). Thus, enhanced extracellular DA levels in the NAc and PFC may reflect current motivational “state” of the animal.

1.6 Dopamine and Stimulus-Reward Learning

DA plays an important role in motor function and expression of motivated behavior. Nevertheless, motivated behavior is rarely a random search strategy but instead it is an acquired adaptive process. Novel palatable foods require associative learning between the food taste and smell and its post-ingestive consequences before they can become rewarding. However, despite extensive research, the concrete evidence that DA is necessary and sufficient in associative learning remains elusive. The confounding role of DA in non-associative processes such as motor activity and motivation can make behavioral interpretations difficult. Nevertheless, DA function has been shown to play an important role in some forms of learning and synaptic plasticity related to reward processing (Beninger, 1983; Hollerman and Schultz, 1998; Jay, 2003; Kerr and Wickens,

2001; Kim et al., 2012; Parker et al., 2011; Reynolds et al., 2001).

Environmental stimuli can acquire motivational properties through Pavlovian learning mechanisms (Bindra, 1974; Konorski, 1967; Rescorla and Solomon, 1967;

Toates, 1985). Incentive learning theory of motivation proposes that when DAergic

neurons are activated, usually by unexpected rewards, previously neutral environmental stimuli associated with these events can acquire incentive salience and thus the ability to elicit approach and other behaviors in the future (Beninger, 1983; Schmidt and Beninger,

2006). As mentioned earlier, incentive learning is the engine that drives preparatory behaviors and thus expression of incentive motivation. Lesions of the DA system and DA receptor antagonism block the learning of stimuli-reward associations. For example, DA receptor blocking antipsychotic drugs impair the acquisition of active avoidance behavior without affecting escape responses despite similarities between the motor acts required for each response (Blackburn and Phillips, 1989; Posluns, 1962).

Animals will learn to self-administer DA agonists such as AMPH and cocaine, and also develop conditioned preference for a place (CPP) in which they have previously been exposed to such drugs (Spyraki et al., 1982; Wise, 2004). Lesions or pharmacological blockade of DA D1 or D2, but not D3 receptors block the acquisition of these behaviors (Lyness et al., 1979; Roberts and Koob, 1982). D3 receptors antagonists, on the other hand, attenuate the expression of conditioning suggesting that they play a role in the manifestation of learned behaviors (Aujla and Beninger, 2004; Banasikowski et al., 2010; Beninger and Banasikowski, 2008). Coincidental activation of D1 and

NMDA receptors in NAc is necessary for the formation of stimulus-reward associations.

Co-treatment with low doses of D1 + NMDA receptor antagonists blocked the acquisition of responding rewarded by sucrose pellets (Smith-Roe and Kelley, 2000).

DA has been shown to modulate memory processes and enhance retention of

recently acquired behaviors. Amphetamine and D1 or D2 receptor agonists given systemically or into the dorsal striatum facilitate consolidation of recently acquired memories in aversively-motivated tasks where stimuli or places are paired with mild footshock (Carr and White, 1984; Krivanek and McGaugh, 1969; Packard and White,

1989; White et al., 1993; White and Viaud, 1991). On the other hand, post-session treatment with a D1 receptor antagonist impairs the acquisition of Pavlovian approach responses to food (Dalley et al., 2005). Setlow and McGaugh showed that D2 receptors in the ventral and dorsal striatum are necessary for consolidation of long term spatial memories when rats were trained on the Morris water maze escape task (Setlow and

McGaugh, 1998; Setlow and McGaugh, 1999; Setlow and McGaugh, 2000). Also, posttrial systemic, intra-hypothalamic or NAc shell treatment of D1 receptor antagonist impairs memory consolidation of conditioned taste aversion associations (Caulliez et al.,

1996; Fenu et al., 2001).

Electrophysiology studies in monkeys have shown that DA neurons show shifts in burst-firing responses from primary rewards, to reward predicting stimuli during reward learning (Hollerman and Schultz, 1998). Initially, DA neurons are activated by encountering unexpected reward stimuli - primary rewards like food, sexually receptive conspecifics and drugs of abuse. However with extensive training, DA neurons activation is also seen when there is an unexpected presentation of stimuli that have become associated with the rewards – now incentive stimuli (Hollerman and Schultz, 1998;

Glimcher, 2011). Such changes in DA neuron activity appear to result from DA

modulatory processing of limbic and cortical glutamate inputs synapsing onto striatal

MSNs (Kiyatkin and Rebec, 1996).

Studies using genetically modified mice that are not able to synthesize DA question the involvement of DA in associative learning (Cannon and Palmiter, 2003).

However, reward learning observed in these mutants may depend on non-DA compensatory mechanisms. Recent studies from the same lab reported that mice lacking

D1 receptors do not learn Pavlovian approach responses to food (Parker et al., 2010).

Further, mice lacking NMDA receptor subunit NR1 on DA neurons exhibit significant impairments in burst-firing, and phasic DA release (Zweifel et al., 2008). DA NR1 mutants do not acquire conditioned locomotor activity or CPP, spatial learning in a T- maze, instrumental responding for food or aversive conditioning (Zweifel et al., 2011;

Zweifel et al., 2009). Furthermore, mice lacking NR1 subunits on D1 but not D2 receptor-expressing striatal MSNs fail to acquire Pavlovian approach (Parker et al.,

2011). Thus, learning stimulus-reward associations require phasic DA release and concurrent activation of D1 and NMDA receptors on the same cells in the striatum.

Consistent with single-unit recordings discussed by Hollerman and Schultz

(1998), studies using fast-scan voltammetry recording in the NAc during Pavlovian approach revealed initial increase in DA transmission following reward receipt in naïve rats. However, when a stimulus became a reliable predictor of reward delivery, the signal shifted to the predictive stimulus and was no longer present during reward delivery (Day et al., 2007; Sunsay and Rebec, 2008). Microdialysis studies have shown increased

concentrations of extracellular DA in the NAc shell but not core during the initial presentations of unfamiliar palatable foods. This response habituates following initial presentation even though the rat continues to consume food with repeated presentation

(Bassareo and Di Chiara, 1999). With repeated training, stimuli predictive of the food availability consequently produced DA increase in the NAc core. These findings are particularly interesting as they suggest that DA activity in the ventral striatum correlates with acquisition and expression of stimulus-reward associations but in different sub- regions.

1.7 Dopamine and Behavioral Sensitization

The behavioral effects of drugs that act on the brain’s DA system change with repeated exposure to the drug. Psychomotor stimulant drugs such as AMPH or cocaine that augment DA neurotransmission by enhancing release and/or blocking uptake (Giros et al., 1996; Steketee and Kalivas, 2011) produce progressively greater effects, for example, on the stimulation of locomotor responses in rats, with repeated testing. The increased behavioral response to the drug occurs over repeated administration and persists long after drug exposure is discontinued. This phenomenon has been termed,

“psychomotor stimulant sensitization” (Post and Rose, 1976; Segal and Mandell, 1974).

Similarly, antipsychotic drugs that block DA receptors produce progressively greater effects, e.g., on catalepsy responses, termed, “catalepsy sensitization” (Schmidt et al.,

1999).

Anatomical and electrophysiological studies have shown that repeated treatment

with AMPH or cocaine result in long-lasting changes in the DA system (Boudreau and

Wolf, 2005; Ungless et al., 2001; Vezina and Queen, 2000; Badiani et al., 1998; Li et al.,

2004; Paulson and Robinson, 1995; Singer et al., 2009). Behavioral sensitization can be attributed to incentive learning of drug-related stimuli and associated neurophysiological adaptations within the DA system (Beninger, 1983; Hyman, 2005; Lodge and Grace,

2008; Robinson and Berridge, 1993; Vezina and Leyton, 2009). The enhanced behavioral output as well as the enhanced striatal DA release with each repeated psychostimulant treatment (Badiani et al., 1998; Paulson and Robinson, 1995) is context-dependent, i.e., its expression is strongest in the presence of previously drug-paired environmental stimuli (Anagnostaras and Robinson, 1996; Badiani et al., 1998; Pert et al., 1990).

Sensitization to psychomotor stimulants is associated with specific drug- environment effects rather than the drug itself. Environments associated with repeated

AMPH or cocaine treatments elicit conditioned locomotor hyperactivity (Banasikowski et al., 2010; Beninger and Hahn, 1983; Beninger and Herz, 1986; Crombag et al., 2001).

Molecular neuroimaging studies in humans, such as positron emission tomography, confirm the ability of psychomotor stimulant drug-paired cues to produce conditioned

DA release in the absence of the drug itself (Boileau et al., 2007; de la Fuente-Fernandez et al., 2001).

The acquisition of environment-specific conditioned drug effects is often blocked by treatment with D1-like and D2 receptor antagonists (Cervo and Samanin, 1995; Cervo and Samanin, 1996; Dias et al., 2006; Fontana et al., 1993a; Fontana et al., 1993b;

Mazurski and Beninger, 1991). Lodge and Grace (2008) reported that sensitized locomotor responses to AMPH challenge can be reversed by inactivation of ventral HPC, a region that projects to the NAc and plays an important role in contextual learning.

Moreover, the behavioral sensitized response to AMPH was correlated with an increase in ventral HPC activity, as well as increased spontaneous activity of DA neurons in the

VTA. Temporary inactivation of the ventral HPC restored the behavioral response to

AMPH and the DA population activity to baseline levels (Lodge and Grace, 2008).

Extensive lesions of the DA system via the MFB or high doses of DA receptor antagonists produce catalepsy a phenomenon characterized by a loss of voluntary movement and muscular rigidity, quantified by the time a rat remains with its forepaws resting on a suspended horizontal bar (Klemm, 1993; Sanberg et al., 1988). Historically catalepsy has been viewed as an animal model of PD and used to screen for potential extrapyramidal side-effects associated with first-generation antipsychotics (Ossowska,

2002). Rats repeatedly treated with a low, sub-threshold dose of the D2 receptor- preferring antagonist haloperidol do not initially exhibit catalepsy (Schmidt et al., 1999) or a reduction in exploratory behavior in an open field arena (Carey, 1987; Carey and

Kenney, 1987). With repeated drug-environment pairings animals gradually develop day- to-day increases in the catalepsy response and reduction in exploratory behaviors.

Interestingly, when these animals are tested with saline instead of haloperidol they continue to exhibit drug appropriate behaviors. In open-field studies the conditioned effect was only observed in the drug-paired environment but not in a different

environment (Carey and Kenney, 1987). Evidence shows that repeated treatment with antipsychotics can also lead to gradual changes within the DA system (Boye and Rompre,

2000; Grace and Bunney, 1986; Meshul et al., 1992; Valenti and Grace, 2010) and a progressive reduction in incentive properties of reward-related stimuli that may result from a sensitization-like mechanism (Boye and Rompre, 2000; Franklin and McCoy,

1979; Wise, 1982).

Catalepsy sensitization and conditioned catalepsy associated with repeated testing following injection of a DA receptor antagonist has not been as extensively studied. The simplicity of the catalepsy paradigm may further our understanding of behavioral sensitization to psychomotor stimulants. Firstly, is this phenomenon similar or different from psychomotor stimulant sensitization and conditioning? How are different DA receptor subtypes involved in controlling the acquisition and expression of catalepsy- sensitization? Is catalepsy sensitization and conditioning dependent on DA function? If so, does DA play a role in forming drug-environment associations? Or does repeated drug treatment lead to non-associative processes like physiological changes that manifest as motor or motivational impairments, often regarded as performance confounds in reward- related learning?

1.8 Hypothesis

In this thesis I will examine the role of DA D1, D2, and D3 receptors in acquisition and expression of catalepsy sensitization and their specific contribution to drug-environment conditioning. Also, I will examine the different role of associative and

non-associative learning processes in development of catalepsy sensitization. My underlying hypothesis is that catalepsy sensitization is not a simple motor impairment; instead it is a gradually acquired motivational deficit where environmental stimuli lose their ability to engage and control behavior. In the first experimental chapter I will examine the effects of D1-like and D2-receptor preferring antagonist on development of catalepsy sensitization and conditioning. In the second experimental chapter, I will examine the role of D2 and D3 receptors in catalepsy sensitization and acquisition and expression of conditioned catalepsy. In the third experimental chapter, I will examine the interaction between repeated treatment with haloperidol and test environment experience, and whether non-associative processes are involved in the expression of catalepsy sensitization. Furthermore, I will examine how test-environment pre-exposure can influence future sensitization responses. The role of DA and possible mechanisms for how environmental stimuli contribute to catalepsy sensitization will be presented.

1.9 References

Ahn S, Phillips AG (1999). Dopaminergic correlates of sensory-specific satiety in the medial prefrontal cortex and nucleus accumbens of the rat. J Neurosci, 19(19): RC29. Alexander GE, DeLong MR, Strick PL (1986). Parallel organization of functionally segregated circuits linking basal ganglia and cortex. Annu Rev Neurosci, 9: 357- 381. Anagnostaras SG, Robinson TE (1996). Sensitization to the psychomotor stimulant effects of amphetamine: modulation by associative learning. Behav Neurosci, 110(6): 1397-1414. Antelman SM, Rowland NE, Fisher AE (1976). Stress related recovery from lateral hypothalamic aphagia. Brain Res, 102(2): 346-350. Arbuthnott GW, Ingham CA, Wickens JR (1998). Modulation by dopamine of rat corticostriatal input. Adv Pharmacol, 42: 733-736. Aujla H, Beninger RJ (2004). Intra-BLA or intra-NAc infusions of the dopamine D3 receptor partial agonist, BP 897, block intra-NAc amphetamine conditioned activity. Behav Neurosci, 118(6): 1324-1330. Badiani A, Oates MM, Day HE, Watson SJ, et al. (1998). Amphetamine-induced behavior, dopamine release, and c-fos mRNA expression: modulation by environmental novelty. J Neurosci, 18(24): 10579-10593. Banasikowski TJ, Beninger RJ (2010). Conditioned drug effects. In: Stolerman, IP (Ed.), Encyclopedia of Psychopharmacology (pp. 325-331) Heidelberg: Springer. Banasikowski TJ, Bespalov A, Drescher K, Behl B, et al. (2010). Double dissociation of the effects of haloperidol and the dopamine D3 receptor antagonist ABT-127 on acquisition vs. expression of cocaine-conditioned activity in rats. J Pharmacol Exp Ther, 335(2): 506-515. Bassareo V, Di Chiara G (1997). Differential influence of associative and nonassociative learning mechanisms on the responsiveness of prefrontal and accumbal dopamine transmission to food stimuli in rats fed ad libitum. J Neurosci, 17(2): 851-861. Bassareo V, Di Chiara G (1999). Modulation of feeding-induced activation of mesolimbic dopamine transmission by appetitive stimuli and its relation to motivational state. Eur J Neurosci, 11(12): 4389-4397.

Bateup HS, Santini E, Shen W, Birnbaum S, et al. (2009). Distinct subclasses of medium spiny neurons differentially regulate striatal motor behaviors. Proc Natl Acad Sci U S A, 107(33): 14845-14850. Belin D, Everitt BJ (2008). Cocaine seeking habits depend upon dopamine-dependent serial connectivity linking the ventral with the dorsal striatum. Neuron, 57(3): 432-441. Belin D, Jonkman S, Dickinson A, Robbins TW, et al. (2009). Parallel and interactive learning processes within the basal ganglia: relevance for the understanding of addiction. Behav Brain Res, 199(1): 89-102. Beninger RJ (1983). The role of dopamine in locomotor activity and learning. Brain Res, 287(2): 173-196. Beninger RJ, Banasikowski TJ (2008). Dopaminergic mechanism of reward-related incentive learning: focus on the dopamine D(3) receptor. Neurotox Res, 14(1): 57- 70. Beninger RJ, Hahn BL (1983). Pimozide blocks establishment but not expression of amphetamine-produced environment-specific conditioning. Science, 220(4603): 1304-1306. Beninger RJ, Herz RS (1986). Pimozide blocks establishment but not expression of cocaine-produced environment-specific conditioning. Life Sci, 38(15): 1425-1431. Berendse HW, Galis-de Graaf Y, Groenewegen HJ (1992a). Topographical organization and relationship with ventral striatal compartments of prefrontal corticostriatal projections in the rat. J Comp Neurol, 316(3): 314-347. Berendse HW, Groenewegen HJ, Lohman AH (1992b). Compartmental distribution of ventral striatal neurons projecting to the mesencephalon in the rat. J Neurosci, 12(6): 2079-2103. Bindra D (1974). A motivational view of learning, performance, and behavior modification. Psychol Rev, 81(3): 199-213. Bindra D, Palfai T (1967). Nature of positive and negative incentive-motivational effects on general activity. J Comp Physiol Psychol, 63(2): 288-297. Bjorklund A, Dunnett SB (2007). Dopamine neuron systems in the brain: an update. Trends Neurosci, 30(5): 194-202. Blackburn JR, Phillips AG (1989). Blockade of acquisition of one-way conditioned avoidance responding by haloperidol and metoclopramide but not by thioridazine

or clozapine: implications for screening new antipsychotic drugs. Psychopharmacology (Berl), 98(4): 453-459. Blackburn JR, Phillips AG, Fibiger HC (1987). Dopamine and preparatory behavior: I. Effects of pimozide. Behav Neurosci, 101(3): 352-360. Boileau I, Dagher A, Leyton M, Welfeld K, et al. (2007). Conditioned dopamine release in humans: a positron emission tomography [11C]raclopride study with amphetamine. J Neurosci, 27(15): 3998-4003. Bolles R (1972). Reinforcement, expectancy, and learning. Psychological Review, 79(5): 394-409. Boudreau AC, Wolf ME (2005). Behavioral sensitization to cocaine is associated with increased AMPA receptor surface expression in the nucleus accumbens. J Neurosci, 25(40): 9144-9151. Boye SM, Rompre PP (2000). Behavioral evidence of depolarization block of dopamine neurons after chronic treatment with haloperidol and clozapine. J Neurosci, 20(3): 1229-1239. Braak H, Del Tredici K, Bratzke H, Hamm-Clement J, et al. (2002). Staging of the intracerebral inclusion body pathology associated with idiopathic Parkinson's disease (preclinical and clinical stages). J Neurol, 249 Suppl 3: III/1-5. Breland K, Breland M (1961). The misbehaviour of organisms. American Psychologis, 16: 681-684. Bristow LJ, Collinson N, Cook GP, Curtis N, et al. (1997). L-745,870, a subtype selective dopamine D4 receptor antagonist, does not exhibit a neuroleptic-like profile in rodent behavioral tests. J Pharmacol Exp Ther, 283(3): 1256-1263. Cannon CM, Palmiter RD (2003). Reward without dopamine. J Neurosci, 23(34): 10827-10831. Carey RJ (1987). Conditioning and the delayed onset of a haloperidol-induced behavioral effect. Biol Psychiatry, 22(3): 269-277. Carey RJ, Kenney S (1987). Operant conditioning and haloperidol-induced hypokinetic effects. Neuropsychobiology, 18(4): 199-204. Carr GD, White NM (1984). The relationship between stereotypy and memory improvement produced by amphetamine. Psychopharmacology (Berl), 82(3): 203-209.

Caulliez R, Meile MJ, Nicolaidis S (1996). A lateral hypothalamic D1 dopaminergic mechanism in conditioned taste aversion. Brain Res, 729(2): 234-245. Cervo L, Samanin R (1995). Effects of dopaminergic and glutamatergic receptor antagonists on the acquisition and expression of cocaine conditioning place preference. Brain Res, 673(2): 242-250. Cervo L, Samanin R (1996). Effects of dopaminergic and glutamatergic receptor antagonists on the establishment and expression of conditioned locomotion to cocaine in rats. Brain Res, 731(1-2): 31-38. Corbett D, Wise RA (1980). Intracranial self-stimulation in relation to the ascending dopaminergic systems of the midbrain: a moveable electrode mapping study. Brain Res, 185(1): 1-15. Costall B, Olley JE (1971). Cholinergic- and neuroleptic-induced catalepsy: modification by lesions in the caudate-putamen. Neuropharmacology, 10(3): 297- 306. Crombag HS, Badiani A, Chan J, Dell'Orco J, et al. (2001). The ability of environmental context to facilitate psychomotor sensitization to amphetamine can be dissociated from its effect on acute drug responsiveness and on conditioned responding. Neuropsychopharmacology, 24(6): 680-690. Dalley JW, Laane K, Theobald DE, Armstrong HC, et al. (2005). Time-limited modulation of appetitive Pavlovian memory by D1 and NMDA receptors in the nucleus accumbens. Proc Natl Acad Sci U S A, 102(17): 6189-6194. Day JJ, Roitman MF, Wightman RM, Carelli RM (2007). Associative learning mediates dynamic shifts in dopamine signaling in the nucleus accumbens. Nat Neurosci, 10(8): 1020-1028. de la Fuente-Fernandez R, Ruth TJ, Sossi V, Schulzer M, et al. (2001). Expectation and dopamine release: mechanism of the placebo effect in Parkinson's disease. Science, 293(5532): 1164-1166. Desai RI, Terry P, Katz JL (2005). A comparison of the locomotor stimulant effects of D1-like receptor agonists in mice. Pharmacol Biochem Behav, 81(4): 843-848. Dias FR, Carey RJ, Carrera MP (2006). Conditioned locomotion induced by unilateral intrastriatal administration of apomorphine: D(2) receptor activation is critical but not the expression of the unconditioned response. Brain Res, 1083(1): 85-95.

Dickinson A, Smith J, Mirenowicz J (2000). Dissociation of Pavlovian and instrumental incentive learning under dopamine antagonists. Behav Neurosci, 114(3): 468-483. Fenu S, Acquas E, Di Chiara G (2001). Role of striatal acetylcholine on dopamine D1 receptor agonist-induced turning behavior in 6-hydroxydopamine lesioned rats: a microdialysis-behavioral study. Neurol Sci, 22(1): 63-64. Fiorino DF, Coury A, Phillips AG (1997). Dynamic changes in nucleus accumbens dopamine efflux during the Coolidge effect in male rats. J Neurosci, 17(12): 4849-4855. Floresco SB, West AR, Ash B, Moore H, et al. (2003). Afferent modulation of dopamine neuron firing differentially regulates tonic and phasic dopamine transmission. Nat Neurosci, 6(9): 968-973. Fog R, Randrup A, Pakkenberg H (1970). Lesions in corpus striatum and cortex of rat brains and the effect on pharmacologically induced stereotyped, aggressive and cataleptic behaviour. Psychopharmacologia, 18(4): 346-356. Fontana D, Post RM, Weiss SR, Pert A (1993a). The role of D1 and D2 dopamine receptors in the acquisition and expression of cocaine-induced conditioned increases in locomotor behavior. Behav Pharmacol, 4(4): 375-387. Fontana DJ, Post RM, Pert A (1993b). Conditioned increases in mesolimbic dopamine overflow by stimuli associated with cocaine. Brain Res, 629(1): 31-39. Franklin KB, McCoy SN (1979). Pimozide-induced extinction in rats: stimulus control of responding rules out motor deficit. Pharmacol Biochem Behav, 11(1): 71-75. Fuxe K, Hokfelt T, Olson L, Ungerstedt U (1977). Central monoaminergic pathways with emphasis on their relation to the so called 'extrapyramidal motor system'. Pharmacol Ther B, 3(2): 169-210. Gerfen CR (1988). Synaptic organization of the striatum. J Electron Microsc Tech, 10(3): 265-281. Gerfen CR, Young WS, 3rd (1988). Distribution of striatonigral and striatopallidal peptidergic neurons in both patch and matrix compartments: an in situ hybridization histochemistry and fluorescent retrograde tracing study. Brain Res, 460(1): 161-167. Giros B, Jaber M, Jones SR, Wightman RM, et al. (1996). Hyperlocomotion and indifference to cocaine and amphetamine in mice lacking the dopamine transporter. Nature, 379(6566): 606-612.

Glimcher PW (2011). Understanding dopamine and reinforcement learning: the dopamine reward prediction error hypothesis. Proc Natl Acad Sci U S A, 108 Suppl 3: 15647-15654. Grace AA, Bunney BS (1984). The control of firing pattern in nigral dopamine neurons: burst firing. J Neurosci, 4(11): 2877-2890. Grace AA, Bunney BS (1986). Induction of depolarization block in midbrain dopamine neurons by repeated administration of haloperidol: analysis using in vivo intracellular recording. J Pharmacol Exp Ther, 238(3): 1092-1100. Grace AA, Floresco SB, Goto Y, Lodge DJ (2007). Regulation of firing of dopaminergic neurons and control of goal-directed behaviors. Trends Neurosci, 30(5): 220-227. Haber SN, Fudge JL, McFarland NR (2000). Striatonigrostriatal pathways in primates form an ascending spiral from the shell to the dorsolateral striatum. J Neurosci, 20(6): 2369-2382. Hollerman JR, Schultz W (1998). Dopamine neurons report an error in the temporal prediction of reward during learning. Nat Neurosci, 1(4): 304-309. Hong S, Hikosaka O (2011). Dopamine-mediated learning and switching in corticostriatal circuit explain behavioral changes in reinforcement learning. Front Behav Neurosci, 5: 15. Hornykiewicz O (1973). Parkinson's disease: from brain homogenate to treatment. Fed Proc, 32(2): 183-190. Hornykiewicz O (1975). Parkinsonism induced by dopaminergic antagonists. Adv Neurol, 9: 155-164. Hyman SE (2005). Addiction: a disease of learning and memory. Am J Psychiatry, 162(8): 1414-1422. Jaber M, Robinson SW, Missale C, Caron MG (1996). Dopamine receptors and brain function. Neuropharmacology, 35(11): 1503-1519. Jay TM (2003). Dopamine: a potential substrate for synaptic plasticity and memory mechanisms. Prog Neurobiol, 69(6): 375-390. Keefe KA, Salamone JD, Zigmond MJ, Stricker EM (1989). Paradoxical kinesia in parkinsonism is not caused by dopamine release. Studies in an animal model. Arch Neurol, 46(10): 1070-1075.

Kerr JN, Wickens JR (2001). Dopamine D-1/D-5 receptor activation is required for long-term potentiation in the rat neostriatum in vitro. J Neurophysiol, 85(1): 117- 124. Kim KM, Baratta MV, Yang A, Lee D, et al. (2012). Optogenetic mimicry of the transient activation of dopamine neurons by natural reward is sufficient for operant reinforcement. PLoS One, 7(4): e33612. Kiyatkin EA, Rebec GV (1996). Dopaminergic modulation of glutamate-induced excitations of neurons in the neostriatum and nucleus accumbens of awake, unrestrained rats. J Neurophysiol, 75(1): 142-153. Klemm WR (1993). The catalepsy of blocked dopaminergic receptors. Psychopharmacology (Berl), 111(2): 251-255. Konorski J (1967). Integrative activity of the brain. An interdisciplinary approach. Chicago: University of Chicago Press. Kravitz AV, Freeze BS, Parker PR, Kay K, et al. (2010). Regulation of parkinsonian motor behaviours by optogenetic control of basal ganglia circuitry. Nature, 466(7306): 622-626. Krivanek JA, McGaugh JL (1969). Facilitating effects of pre- and posttrial amphetamine administration on discrimination learning in mice. Agents Actions, 1(2): 36-42. Levant B (1997). The D3 dopamine receptor: neurobiology and potential clinical relevance. Pharmacol Rev, 49(3): 231-252. Levant B, Grigoriadis DE, De Souza EB (1995). Relative affinities of dopaminergic drugs at dopamine D2 and D3 receptors. Eur J Pharmacol, 278(3): 243-247. Li Y, Acerbo MJ, Robinson TE (2004). The induction of behavioural sensitization is associated with cocaine-induced structural plasticity in the core (but not shell) of the nucleus accumbens. Eur J Neurosci, 20(6): 1647-1654. Lim SY, Fox SH, Lang AE (2009). Overview of the extranigral aspects of Parkinson disease. Arch Neurol, 66(2): 167-172. Lodge DJ, Grace AA (2008). Amphetamine activation of hippocampal drive of mesolimbic dopamine neurons: a mechanism of behavioral sensitization. J Neurosci, 28(31): 7876-7882. Lynch MR, Carey RJ (1987). Environmental stimulation promotes recovery from haloperidol-induced extinction of open field behavior in rats. Psychopharmacology (Berl), 92(2): 206-209. 29

Lyness WH, Friedle NM, Moore KE (1979). Destruction of dopaminergic nerve terminals in nucleus accumbens: effect on d-amphetamine self-administration. Pharmacol Biochem Behav, 11(5): 553-556. Marshall JF, Levitan D, Stricker EM (1976). Activation-induced restoration of sensorimotor functions in rats with dopamine-depleting brain lesions. J Comp Physiol Psychol, 90(6): 536-546. Martin JP (1967). The basal ganglia and posture. Philadelphia: Lippincott. Matamales M, Bertran-Gonzalez J, Salomon L, Degos B, et al. (2009). Striatal medium-sized spiny neurons: identification by nuclear staining and study of neuronal subpopulations in BAC transgenic mice. PLoS One, 4(3): e4770. Mazurski EJ, Beninger RJ (1991). Effects of selective drugs for dopaminergic D1 and D2 receptors on conditioned locomotion in rats. Psychopharmacology (Berl), 105(1): 107-112. McGeer PL, McGeer EG, Hattori T (1977). Dopamine-acetylcholine-GABA neuronal linkages in the extrapyramidal and limbic systems. Adv Biochem Psychopharmacol, 16: 397-402. Meshul CK, Janowsky A, Casey DE, Stallbaumer RK, et al. (1992). Effect of haloperidol and clozapine on the density of "perforated" synapses in caudate, nucleus accumbens, and medial prefrontal cortex. Psychopharmacology (Berl), 106(1): 45-52. Mink JW (1996). The basal ganglia: focused selection and inhibition of competing motor programs. Prog Neurobiol, 50(4): 381-425. Missale C, Nash SR, Robinson SW, Jaber M, et al. (1998). Dopamine receptors: from structure to function. Physiol Rev, 78(1): 189-225. Murschall A, Hauber W (2006). Inactivation of the ventral tegmental area abolished the general excitatory influence of Pavlovian cues on instrumental performance. Learn Mem, 13(2): 123-126. Nicola SM, Taha SA, Kim SW, Fields HL (2005). Nucleus accumbens dopamine release is necessary and sufficient to promote the behavioral response to reward- predictive cues. Neuroscience, 135(4): 1025-1033. Ossowska K (2002). Neuronal basis of neuroleptic-induced extrapyramidal side effects. Pol J Pharmacol, 54(4): 299-312.

Owesson-White CA, Roitman MF, Sombers LA, Belle AM, et al. (2012). Sources contributing to the average extracellular concentration of dopamine in the nucleus accumbens. J Neurochem, 121(2): 252-262. Packard MG, White NM (1989). Memory facilitation produced by dopamine agonists: role of receptor subtype and mnemonic requirements. Pharmacol Biochem Behav, 33(3): 511-518. Papeschi R (1972). Dopamine, extrapyramidal system, and psychomotor function. Psychiatr Neurol Neurochir, 75(1): 13-48. Parker JG, Beutler LR, Palmiter RD (2011). The contribution of NMDA receptor signaling in the corticobasal ganglia reward network to appetitive Pavlovian learning. J Neurosci, 31(31): 11362-11369. Parker JG, Zweifel LS, Clark JJ, Evans SB, et al. (2010). Absence of NMDA receptors in dopamine neurons attenuates dopamine release but not conditioned approach during Pavlovian conditioning. Proc Natl Acad Sci U S A, 107(30): 13491-13496. Paulson PE, Robinson TE (1995). Amphetamine-induced time-dependent sensitization of dopamine neurotransmission in the dorsal and ventral striatum: a microdialysis study in behaving rats. Synapse, 19(1): 56-65. Pert A, Post R, Weiss SR (1990). Conditioning as a critical determinant of sensitization induced by psychomotor stimulants. NIDA Res Monogr, 97: 208-241. Phillips AG, Vacca G, Ahn S (2008). A top-down perspective on dopamine, motivation and memory. Pharmacol Biochem Behav, 90(2): 236-249. Pijnenburg AJ, Honig WM, Van der Heyden JA, Van Rossum JM (1976). Effects of chemical stimulation of the mesolimbic dopamine system upon locomotor activity. Eur J Pharmacol, 35(1): 45-58. Pijnenburg AJ, Honig WM, Van Rossum JM (1975). Inhibition of d-amphetamine- induced locomotor activity by injection of haloperidol into the nucleus accumbens of the rat. Psychopharmacologia, 41(2): 87-95. Posluns D (1962). An analysis of chlorpromazine-induced suppression of the avoidance response. Psychopharmacologia, 3: 361-373. Post RM, Rose H (1976). Increasing effects of repetitive cocaine administration in the rat. Nature, 260(5553): 731-732.

Ranaldi R, Beninger RJ (1993). Dopamine D1 and D2 antagonists attenuate amphetamine-produced enhancement of responding for conditioned reward in rats. Psychopharmacology (Berl), 113(1): 110-118. Rankin ML, Hazelwood LA, Free RB, Namkung Y, et al. (2010). Molecular Pharmacology of Dopamine Receptors. In: Iversen, LI, S. Dunnett, S., Bjorklund, A (Eds.) Dopamine Handbook New York: Oxford University Press. Remy P, Doder M, Lees A, Turjanski N, et al. (2005). Depression in Parkinson's disease: loss of dopamine and noradrenaline innervation in the limbic system. Brain, 128(Pt 6): 1314-1322. Rescorla RA, Solomon RL (1967). Two-process learning theory: Relationships between Pavlovian conditioning and instrumental learning. Psychol Rev, 74(3): 151-182. Reynolds JN, Hyland BI, Wickens JR (2001). A cellular mechanism of reward-related learning. Nature, 413(6851): 67-70. Richfield EK, Penney JB, Young AB (1989a). Anatomical and affinity state comparisons between dopamine D1 and D2 receptors in the rat central nervous system. Neuroscience, 30(3): 767-777. Richfield EK, Young AB, Penney JB (1989b). Comparative distributions of dopamine D-1 and D-2 receptors in the cerebral cortex of rats, cats, and monkeys. J Comp Neurol, 286(4): 409-426. Roberts DC, Koob GF (1982). Disruption of cocaine self-administration following 6- hydroxydopamine lesions of the ventral tegmental area in rats. Pharmacol Biochem Behav, 17(5): 901-904. Robinson TE, Berridge KC (1993). The neural basis of drug craving: an incentive- sensitization theory of addiction. Brain Res Brain Res Rev, 18(3): 247-291. Roth BL, Tandra S, Burgess LH, Sibley DR, et al. (1995). D4 dopamine receptor binding affinity does not distinguish between typical and atypical antipsychotic drugs. Psychopharmacology (Berl), 120(3): 365-368. Salamone JD, Correa M (2002). Motivational views of reinforcement: implications for understanding the behavioral functions of nucleus accumbens dopamine. Behav Brain Res, 137(1-2): 3-25. Salamone JD, Zigmond MJ, Stricker EM (1990). Characterization of the impaired feeding behavior in rats given haloperidol or dopamine-depleting brain lesions. Neuroscience, 39(1): 17-24. 32

Sanberg PR (1980). Haloperidol-induced catalepsy is mediated by postsynaptic dopamine receptors. Nature, 284(5755): 472-473. Sanberg PR, Bunsey MD, Giordano M, Norman AB (1988). The catalepsy test: its ups and downs. Behav Neurosci, 102(5): 748-759. Schmidt WJ, Beninger RJ (2006). Behavioural sensitization in addiction, schizophrenia, Parkinson's disease and dyskinesia. Neurotox Res, 10(2): 161-166. Schmidt WJ, Tzschentke TM, Kretschmer BD (1999). State-dependent blockade of haloperidol-induced sensitization of catalepsy by MK-801. Eur J Neurosci, 11(9): 3365-3368. Schwab RS, Zieper I (1965). Effects of mood, motivation, stress and alertness on the performance in Parkinson's disease. Psychiatr Neurol (Basel), 150(6): 345-357. Segal DS, Mandell AJ (1974). Long-term administration of d-amphetamine: progressive augmentation of motor activity and stereotypy. Pharmacol Biochem Behav, 2(2): 249-255. Sesack SR, Grace AA (2010). Cortico-Basal Ganglia reward network: microcircuitry. Neuropsychopharmacology, 35(1): 27-47. Setlow B, McGaugh JL (1998). Sulpiride infused into the nucleus accumbens posttraining impairs memory of spatial water maze training. Behav Neurosci, 112(3): 603-610. Setlow B, McGaugh JL (1999). Involvement of the posteroventral caudate-putamen in memory consolidation in the Morris water maze. Neurobiol Learn Mem, 71(2): 240-247. Setlow B, McGaugh JL (2000). D2 dopamine receptor blockade immediately post- training enhances retention in hidden and visible platform versions of the water maze. Learn Mem, 7(3): 187-191. Shore DM, Rafal R, Parkinson JA (2011). Appetitive motivational deficits in individuals with Parkinson's disease. Mov Disord, 26(10): 1887-1892. Singer BF, Tanabe LM, Gorny G, Jake-Matthews C, et al. (2009). Amphetamine- induced changes in dendritic morphology in rat forebrain correspond to associative drug conditioning rather than nonassociative drug sensitization. Biol Psychiatry, 65(10): 835-840. Smith-Roe SL, Kelley AE (2000). Coincident activation of NMDA and dopamine D1 receptors within the nucleus accumbens core is required for appetitive instrumental learning. J Neurosci, 20(20): 7737-7742. 33

Souques A (1921). Rapports sure les syndromes parkinsonien. Revista de Neurologia (Paris), 37: 534-715. Spyraki C, Fibiger HC, Phillips AG (1982). Dopaminergic substrates of amphetamine- induced place preference conditioning. Brain Res, 253(1-2): 185-193. Steketee JD, Kalivas PW (2011). Drug wanting: behavioral sensitization and relapse to drug-seeking behavior. Pharmacol Rev, 63(2): 348-365. Stoof JC, Kebabian JW (1981). Opposing roles for D-1 and D-2 dopamine receptors in efflux of cyclic AMP from rat neostriatum. Nature, 294(5839): 366-368. Sunsay C, Rebec GV (2008). Real-time dopamine efflux in the nucleus accumbens core during Pavlovian conditioning. Behav Neurosci, 122(2): 358-367. Taylor JR, Robbins TW (1984). Enhanced behavioural control by conditioned reinforcers following microinjections of d-amphetamine into the nucleus accumbens. Psychopharmacology (Berl), 84(3): 405-412. Tepper JM, Bolam JP (2004). Functional diversity and specificity of neostriatal interneurons. Curr Opin Neurobiol, 14(6): 685-692. Toates F (1985). Psychobiology: the neurobiology of motivation and reward. Science, 229(4717): 962-963. Tran AH, Tamura R, Uwano T, Kobayashi T, et al. (2002). Altered accumbens neural response to prediction of reward associated with place in dopamine D2 receptor knockout mice. Proc Natl Acad Sci U S A, 99(13): 8986-8991. Tran AH, Tamura R, Uwano T, Kobayashi T, et al. (2005). Dopamine D1 receptors involved in locomotor activity and accumbens neural responses to prediction of reward associated with place. Proc Natl Acad Sci U S A, 102(6): 2117-2122. Tran AH, Uwano T, Kimura T, Hori E, et al. (2008). Dopamine D1 receptor modulates hippocampal representation plasticity to spatial novelty. J Neurosci, 28(50): 13390-13400. Ungerstedt U (1971). Adipsia and aphagia after 6-hydroxydopamine induced degeneration of the nigro-striatal dopamine system. Acta Physiol Scand Suppl, 367: 95-122. Ungless MA, Whistler JL, Malenka RC, Bonci A (2001). Single cocaine exposure in vivo induces long-term potentiation in dopamine neurons. Nature, 411(6837): 583-587.

Vacca G, Ahn S, Phillips AG (2007). Effects of short-term abstinence from escalating doses of D-amphetamine on drug and sucrose-evoked dopamine efflux in the rat nucleus accumbens. Neuropsychopharmacology, 32(4): 932-939. Valenti O, Grace AA (2010). Antipsychotic drug-induced increases in ventral tegmental area dopamine neuron population activity via activation of the nucleus accumbens-ventral pallidum pathway. Int J Neuropsychopharmacol, 13(7): 845- 860. Vallone D, Picetti R, Borrelli E (2000). Structure and function of dopamine receptors. Neurosci Biobehav Rev, 24(1): 125-132. Van Tol HH, Bunzow JR, Guan HC, Sunahara RK, et al. (1991). Cloning of the gene for a human dopamine D4 receptor with high affinity for the antipsychotic clozapine. Nature, 350(6319): 610-614. Vezina P, Leyton M (2009). Conditioned cues and the expression of stimulant sensitization in animals and humans. Neuropharmacology, 56 Suppl 1: 160-168. Vezina P, Queen AL (2000). Induction of locomotor sensitization by amphetamine requires the activation of NMDA receptors in the rat ventral tegmental area. Psychopharmacology (Berl), 151(2-3): 184-191. Weingarten HP (1984). Meal initiation controlled by learned cues: basic behavioral properties. Appetite, 5(2): 147-158. Whishaw IQ, Mittleman G, Evenden JL (1989). Training-dependent decay in performance produced by the neuroleptic cis(Z)-flupentixol on spatial navigation by rats in a swimming pool. Pharmacol Biochem Behav, 32(1): 211-220. White NM (1989). Reward or reinforcement: what's the difference? Neurosci Biobehav Rev, 13(2-3): 181-186. White NM, Packard MG, Seamans J (1993). Memory enhancement by post-training peripheral administration of low doses of dopamine agonists: possible autoreceptor effect. Behav Neural Biol, 59(3): 230-241. White NM, Viaud M (1991). Localized intracaudate dopamine D2 receptor activation during the post-training period improves memory for visual or olfactory conditioned emotional responses in rats. Behav Neural Biol, 55(3): 255-269. Wickens JR, Budd CS, Hyland BI, Arbuthnott GW (2007). Striatal contributions to reward and decision making: making sense of regional variations in a reiterated processing matrix. Ann N Y Acad Sci, 1104: 192-212.

Wilson C, Nomikos GG, Collu M, Fibiger HC (1995). Dopaminergic correlates of motivated behavior: importance of drive. J Neurosci, 15(7 Pt 2): 5169-5178. Wise RA (1982). Neuroleptics and operant behavior: The anhedonia hypothesis. Behavioral and Brain Sciences, 5(01): 39-53. Wise RA (2004). Dopamine, learning and motivation. Nat Rev Neurosci, 5(6): 483-494. Wise RA (2009). Roles for nigrostriatal--not just mesocorticolimbic--dopamine in reward and addiction. Trends Neurosci, 32(10): 517-524. Wise RA, Spindler J, deWit H, Gerberg GJ (1978). Neuroleptic-induced "anhedonia" in rats: pimozide blocks reward quality of food. Science, 201(4352): 262-264. Wolterink G, Phillips G, Cador M, Donselaar-Wolterink I, et al. (1993). Relative roles of ventral striatal D1 and D2 dopamine receptors in responding with conditioned reinforcement. Psychopharmacology (Berl), 110(3): 355-364. Wyvell CL, Berridge KC (2000). Intra-accumbens amphetamine increases the conditioned incentive salience of sucrose reward: enhancement of reward "wanting" without enhanced "liking" or response reinforcement. J Neurosci, 20(21): 8122-8130. Yokel RA, Wise RA (1975). Increased lever pressing for amphetamine after pimozide in rats: implications for a dopamine theory of reward. Science, 187(4176): 547-549. Yokel RA, Wise RA (1976). Attenuation of intravenous amphetamine reinforcement by central dopamine blockade in rats. Psychopharmacology (Berl), 48(3): 311-318. Zhou QY, Palmiter RD (1995). Dopamine-deficient mice are severely hypoactive, adipsic, and aphagic. Cell, 83(7): 1197-1209. Zweifel LS, Argilli E, Bonci A, Palmiter RD (2008). Role of NMDA receptors in dopamine neurons for plasticity and addictive behaviors. Neuron, 59(3): 486-496. Zweifel LS, Fadok JP, Argilli E, Garelick MG, et al. (2011). Activation of dopamine neurons is critical for aversive conditioning and prevention of generalized anxiety. Nat Neurosci, 14(5): 620-626. Zweifel LS, Parker JG, Lobb CJ, Rainwater A, et al. (2009). Disruption of NMDAR- dependent burst firing by dopamine neurons provides selective assessment of phasic dopamine-dependent behavior. Proc Natl Acad Sci U S A, 106(18): 7281- 7288.

Chapter 2

Haloperidol Conditioned Catalepsy in Rats: A Possible Role for D1-like

Receptors

2.1 Abstract

Decreases in brain dopamine (DA) lead to catalepsy, quantified by the time a rat remains with its forepaws resting on a suspended horizontal bar. Low doses of the DA D2 receptor-preferring antagonist haloperidol repeatedly injected in a particular environment lead to gradual day-to-day increases in catalepsy (catalepsy sensitization) and subsequent testing following an injection of saline reveal conditioned catalepsy. We tested the hypothesis that D1-like and D2 receptors play different roles in catalepsy sensitization and in acquisition and expression of conditioned catalepsy. Rats were repeatedly treated with either the DA D1-like receptor antagonist SCH 23990 (0.05, 0.1 and 0.25 mg/kg; i.p.), the D2 receptor-preferring antagonist haloperidol (0.1, 0.25 and 0.5 mg/kg; i.p.) or a combination of the two drugs and tested for catalepsy each day in the same environment.

Following 10 drug treatment days rats were injected with saline and tested for conditioned catalepsy in the previously drug-paired environment. Haloperidol did not elicit cataleptic responses in the initial session; however rats developed sensitization with repeated testing. Significant catalepsy sensitization was not observed in rats repeatedly tested with SCH 23390. When rats were injected and tested with saline following haloperidol sensitization they exhibited conditioned catalepsy in the test environment; conditioned catalepsy was not seen following SCH 23390. Rats treated with SCH 23390

0.05 mg/kg + haloperidol 0.25 mg/kg showed catalepsy sensitization but failed to show conditioned catalepsy. Conversely, SCH 23390 (0.05 mg/kg) given on the test day after sensitization to haloperidol (0.25 mg/kg) failed to block conditioned catalepsy. Repeated

antagonism of D2 receptors leads to catalepsy sensitization with repeated testing in a specific environment. Conditioned catalepsy requires intact D1-like receptor function during sensitization sessions but not during test sessions. In conclusion, repeated antagonism of D2 receptors leads to catalepsy sensitization and conditioning in specific environment that appears to be conditional on functional D1-like receptor activity.

2.2 Introduction

Catalepsy is observed when animals are placed in abnormal or unusual postures and maintain these postured for a period of time. A normal animal will correct its position within seconds and explore its environment but a cataleptic animal will maintain this externally imposed posture for a prolonged period of time (Miller et al., 1990; Ossowska et al., 1990; Sanberg et al., 1988). Catalepsy is thought to share similarities with symptoms of human neuropsychiatric diseases such as Parkinson’s disease and damage involving parts of the basal ganglia (Crocker and Hemsley, 2001; Elliott et al., 1990;

Hornykiewicz, 1973; Papeschi, 1972). Decreased DA transmission at post-synaptic D2 receptors has been implicated in catalepsy produced by antipsychotic drugs (Klemm,

1985b; Wadenberg et al., 2000). Mice lacking post-synaptic D2 receptors exhibit profound Parkinson-like motor deficits (Baik et al., 1995; Centonze et al., 2004; Usiello et al., 2000). Disrupting DA transmission with low doses of a potent D2 receptor- preferring antagonist like haloperidol (Lanis and Schmidt, 2001; Schmidt et al., 1999) or with partial bilateral 6-hydroxydopamine (6-OHDA) lesions of the striatum (Klein and

Schmidt, 2003) or the medial forebrain bundle (Srinivasan and Schmidt, 2004) leads to

the development of catalepsy sensitization in rats. Importantly, these animals do not show catalepsy responses on the first couple of test sessions, however with repeated testing catalepsy gradually develops. This sensitization is context-dependent; testing the animal in a different context abolishes catalepsy (Klein and Schmidt, 2003). When rats are given saline in the context previously associated with haloperidol they exhibit conditioned catalepsy (Amtage and Schmidt, 2003; Banasikowski and Beninger, 2010).

Conditioned catalepsy appears to share common learning characteristics such as sensitization, context dependency, and extinction with other more established pavlovian and instrumental learning paradigms (Amtage and Schmidt, 2003; Banasikowski and

Beninger, 2010; Schmidt and Beninger, 2006). DA D1-like receptors play an important role in acquisition of learned behaviors like Pavlovian conditioned approach (Choi et al.,

2005; Dalley et al., 2005), conditioned activity (Mazurski and Beninger, 1991), conditioned place preference (Cervo and Samanin, 1995), sensitization (Vezina, 1996).

The current study examined the interactive role of D1-like and D2 receptor antagonism in catalepsy sensitization and acquisition and expression of conditioned catalepsy in rats.

The first experiment examined the hypothesis that catalepsy sensitization and conditioned catalepsy based on haloperidol is dose dependent. In the second experiment we examined the hypothesis that treating rats with the D1-like receptor antagonist, SCH

23390, will also result in catalepsy sensitization and conditioned catalepsy. In the third experiment, we examined the hypothesis that a combination of haloperidol and SCH

23390 will lead to the development of catalepsy sensitization and conditioned catalepsy.

2.3 Materials and Methods

2.3.1 Subjects

Experimentally naïve male albino Wistar rats (N = 108) weighing 200-225 g upon arrival from Charles River Canada (St. Constant QC) were housed in pairs or threes in clear Plexiglas cages (45.0 x 25.0 x 22.0 cm). Average temperature in the colony was 21º

C, humidity 70% with reversed light–dark cycle (lights off from 0700 to 1900 hr). Rats were maintained with food (LabDiet 5001, PMI Nutrition International, Brentwood, MO,

USA) and water continuously available. Treatment of rats was in accordance with the guidelines of the Animals for Research Act, the Canadian Council on Animal Care, and was approved by the Queen’s University Animal Care Committee.

2.3.2 Drugs

Haloperidol, (4-[4-(4-chlorophenyl)-4-hydroxy-1-piperidyl]-1-(4-fluorophenyl)- butan-1-one (Sigma, St. Louis, MO, USA) and SCH 23390, (R)-(+)-7-chloro-8-hydroxy-

3-methyl-1-phenyl-2,3,4,5-te trahydro-1H-3-benzazepine hydrochloride (Tocris,

Oakville, ON) were prepared in a 0.3% distilled-water solution of tartaric acid. Injections were administered intraperitoneally (i.p.) in a volume of 1.0 ml/kg.

2.3.3 Behavioral Testing

Rats were randomly assigned to treatment (paired) and control groups (unpaired).

Animals were subsequently injected i.p. with drug or vehicle outside the testing room and put back into their cages. Thirty min (SCH 23390) and 60 min (haloperidol) later, they

were tested on a horizontal bar (1.6 cm diam. threaded or smooth rod with end bolts attached to Plexiglas supports, 10 cm above the surface), by gently placing both forepaws on the bar. Descent latency was measured as the time span from placing the animal on the bar until first active paw movement. A cut-off time of 180 s was used, i.e., the trial was terminated when the animal did not make an active paw movement within that time.

2.3.4 Experimental Design

The general experimental protocol consisted of two phases: conditioning (10 sessions) and test (1 session). All sessions occurred during the dark phase. The test apparatus consisted of two distinct (one with three black sides and one with three white sides) rectangular Plexiglas catalepsy chambers (32 x 42 x 30 cm). Each chamber was equipped with a distinct horizontal bar. The black chamber was equipped with a 1.6 cm diam. smooth rod with end bolts and clear Plexiglas legs (1 cm thick, 10 cm high), and the white chamber was equipped with a 1.6 cm diam. threaded rod with end bolts and black Plexiglas legs (0.5 cm thick and 10 cm high). Surgical paper was used as the floor covering in both chambers. Rats were randomly assigned to one of the two test chambers.

Each rat was always placed in the same chamber during the haloperidol sensitization phase and all treatment groups included rats sensitized in each chamber. We observed no relationship between the magnitude of the catalepsy effect and the testing chambers, and no effect of testing chambers on catalepsy sensitization or conditioning.

Experiment 1: Haloperidol Catalepsy

Rats (N = 36) were used to examine the effects of different doses of haloperidol

on catalepsy. Animals in the ‘paired’ condition were injected with haloperidol (0.1, 0.25 or 0.5 mg/kg; ns = 9) outside the testing room for 10 consecutive days, 60 min before catalepsy test. One hour after the catalepsy test, rats were administered vehicle in their colony room (homecage injection). The ‘unpaired’ group (n = 9) received vehicle 1 hour before and haloperidol (0.5 mg/kg) 1 hour after the catalepsy test. This design allowed for all animals to experience the same drug history but in different environments. On day 11 all animals received saline 60 min prior to the conditioned catalepsy test.

Experiment 2: SCH 23390 Catalepsy

Rats (N = 37) were used to examine catalepsy sensitization and conditioning with different doses of SCH 23390. Animals in the ‘paired’ condition were injected with SCH

23390 (0.05, 0.1 or 0.25 mg/kg; ns = 9/10/9, respectively) outside the testing room for 10 consecutive days, 30 min before the catalepsy test. One hour after the catalepsy test, rats were given homecage injection of vehicle. The ‘unpaired’ group (n = 9) received vehicle

30 min before and SCH 23390 (0.05 mg/kg) 1 hour after the catalepsy test. On day 11 all animals received saline 30 min prior to the catalepsy test.

Experiment 3: Haloperidol + SCH 23390 Catalepsy

One group (n = 9) examined the effects of co-treatment of haloperidol and SCH

23390 on catalepsy. Rats received haloperidol (0.25 mg/kg) 60 min plus SCH 23390

(0.05 mg/kg) 30 min before the catalepsy test for 10 consecutive days. On day 11 animals were injected with saline 1 hour prior to the catalepsy test. A second group (n = 9) was

treated with haloperidol (0.25 mg/kg) 60 min before the catalepsy test for 10 consecutive days. On day 11, rats were given SCH 23390 (0.05 mg/kg) 30 minutes before test.

2.3.5 Statistics

All statistical analysis was performed using IBM SPSS 19.0. All data are presented as means ± standard error. Data were submitted to both parametric and nonparametric tests and each type of analysis yielded similar results. Only the parametric tests are reported in most cases. They involved two-way and one-way analyses of variance (ANOVA). Hypotheses tests were completed using α = 0.05 and pair-wise comparisons were made using Dunnett’s test and where specified, Fisher’s LSD test.

2.4 Results

2.4.1 Experiment 1: Haloperidol Catalepsy

Across conditioning days 1 to 10 (Fig. 1A), a two-way ANOVA revealed a significant main effect of day [F(9, 288) = 10.49, p < 0.001] , treatment [F(3,32) = 7.34, p

< 0.01] and interaction [F(27,320 = 1.61, p < 0.05]. Post-hoc comparison of groups averaged over days found that rats treated with haloperidol 0.1, 0.25 and 0.5 mg/kg spent significantly more time with both paws on the bar than the unpaired hal 0.5/sal control group (p < 0.05). Rats treated with the high dose of haloperidol (0.5 mg/kg) spent significantly (LSD test) more time on the bar than rats given 0.1, or 0.25 mg/kg haloperidol for the 10 conditioning trials combined. This finding suggests that magnitude of catalepsy sensitization may dependent on haloperidol dose. To further analyze the

interaction, simple effects tests of group were performed on the first and last day of conditioning. No significant group differences were obtained on conditioning day 1 [F

(3,32) = 0.42, p = 0.729] but a significant group difference was seen on conditioning day

10 [F (3,32) = 7.02, p < 0.01]. Post-hoc pair-wise comparisons for day 10 found that rats treated with haloperidol 0.1, 0.25 and 0.5 mg/kg spent significantly more time with both paws on the bar than the ‘unpaired’ control group (p < 0.05), and the hal 0.5/sal group spent significantly more time on the bar than the hal 0.1/sal and hal 0.25/sal (LSD test; p

< 0.05) groups.

On the saline test day, ANOVA revealed a significant difference among groups

[F(3,32) = 4.36, p < 0.05; Fig. 1B]. Post-hoc comparisons found that on test day rats previously treated with haloperidol 0.1, 0.25 and 0.5 mg/kg during conditioning spent significantly more time with both paws on the bar than the unpaired hal 0.5/sal control group (p < 0.05). We found no significant difference in latency among the three haloperidol groups suggesting that the magnitude of conditioning expression was independent of haloperidol dose [F(2,24) = 0.16, p > 0.05].

Figure 1

Mean (± SEM) descent latency (s) in groups of rats receiving haloperidol (hal) in doses of 0.1, 0.25 or 0.5 mg/kg 60 min prior to each conditioning day or 0.5 mg/kg 60 min following conditioning days (unpaired) (A) and saline (sal) 60 min prior to the test session (B). * significantly different from unpaired hal 0.5/sal in pairwise comparison (Dunnett’s test) following significant group effects in analysis of variance; † significantly different from hal 0.1/sal and hal 0.25/sal in pairwise comparisons (Fisher’s LSD).

2.4.2 Experiment 2: SCH 23390 Catalepsy

Across conditioning days 1 to 10, a two-way ANOVA revealed a significant main effect of day [F(9, 297) = 3.02, p < 0.01] and treatment [F(3,33) = 4.01, p < 0.05; Fig.

2A]. Post-hoc comparison of groups averaged over days found that rats treated with SCH

23390 (0.05 0.1, and 0.25 mg/kg) spent significantly more time with both paws on the bar than the unpaired SCH 0.05/sal control group (p < 0.05). The significant day effect shows that all groups combined showed increased catalepsy time over conditioning days.

Although the interaction was not significant [F(27,297) = 1.05, p = 0.410 ], examination of Fig. 2A suggests that the day effect occurred in groups treated with SCH 23390 during conditioning. However, there was no further statistical evidence for increased catalepsy over days in the paired SCH 23390 groups. Thus tests of simple effects of day for each group failed to reveal a significant day effects in any of the groups.

On the saline test day, ANOVA revealed no significant difference between

‘paired’ and ‘unpaired’ groups [F(3,33) = 0.96, p = 0.424; Fig. 2B].

2.4.3 Experiment 3: Haloperidol + SCH 23390 Catalepsy

Rats treated with haloperidol + SCH 23390 spent more time with both paws on the bar than the haloperidol-alone group. Across conditioning days 1 to 10, a two-way

ANOVA revealed a significant main effect of day, both groups showing catalepsy sensitization [F(9,144) = 7.33, p < 0.01] and a treatment effect [F(1,16) = 43.00, p <

0.001; Fig. 3A]. Because we used a cutoff catalepsy score of 180 s, the distribution of mean descent latencies for some of the days may have violated the homogeneity of 47

variance assumption of ANOVA. We therefore re-analyzed these data for each group over days using the Friedman nonparametric test and found a significant day effect for each group (p < 0.01). We used the Wilcoxon signed-rank test to compare groups on day

1 and on day 10 and observed a significant group effect in both comparisons (p < 0.01).

To compare the increased catalepsy seen in the co-treatment group with the effect in each group alone, we summed catalepsy scores of the hal 0.25/SCH 0.05 group (hal

0.25 alone) with the SCH 0.05/sal group from experiment 2 (SCH 0.05 alone). As can be seen in Fig. 3A the co-treatment group exhibited a strong synergistic catalepsy effect starting on day 1 that continued for the remainder of conditioning.

On the saline test day conditioned catalepsy was observed in the hal 0.25/SCH

0.05 group but not the hal 0.25 + SCH 0.05/sal group. One-way ANOVA revealed a significant group effect [F (1,16) = 15.95, p < 0.01; Fig. 3B].

2.5 Discussion

The results can be summarized as follows: In experiment 1, rats treated with haloperidol and tested for catalepsy in a specific environment showed no initial catalepsy but the cataleptic response increased from day-to-day across subsequent conditioning days. This day-to-day sensitization in catalepsy was seen with each dose of haloperidol tested but appeared to be strongest with the highest dose of haloperidol (0.5 mg/kg). Rats previously given haloperidol and tested with saline in the drug-paired environment exhibited conditioned catalepsy; all doses of haloperidol resulted in a similar amount of

Figure 2

Mean (± SEM) descent latency (s) in groups of rats receiving SCH 23390 (SCH) in doses 0.05, 0.1 or 0.25 mg/kg 30 min prior to each conditioning day or 0.05 mg/kg 60 min following conditioning days (unpaired) (A) and saline (sal) 30 min prior to the test session (B). * significantly different from unpaired SCH 0.05/sal in pairwise comparison (Dunnett’s test) following significant group effects in analysis of variance.

Figure 3

Mean (± SEM) descent latency (s) in groups of rats receiving 0.25 mg/kg haloperidol (hal) 60 min, or co- treated with 0.25 mg/kg haloperidol (60 min) and 0.05 mg/kg SCH 23390 (SCH) 30 min prior to each conditioning day (A) and saline (sal) 60 min or 0.05 mg/kg SCH 23390 30 min prior to test session (B). * significantly different from hal 0.25 + SCH 0.05/sal in analysis of variance. Note that the hal 0.25/SCH 0.05 + SCH 0.05 group (shown in broken line) is a visual representation only for the sum of group means for each conditioning day for the haloperidol- alone group and the SCH 23390- alone group from experiment 2.

conditioned catalepsy. In experiment 2, rats treated with SCH 23390 exhibited catalepsy but did not show significant sensitization. No conditioned catalepsy was observed on the saline test day following conditioning with SCH 23390 at all doses tested.

In experiment 3, co-treatment with SCH 23390 and haloperidol resulted in a synergistic catalepsy effect. Both drugs independently failed to induce catalepsy on the first conditioning day, however, when administered together the catalepsy curve was shifted to the left, and a significant catalepsy was observed on the first test. This catalepsy effect showed sensitization and approached the maximum cut-off time of 180 s on and after conditioning day 4. On the saline test day, rats previously co-treated with

SCH 23390 and haloperidol failed to exhibit conditioned catalepsy. This result supports our initial finding in experiment 2, and suggests that functional D1-like receptors are critical for acquisition of conditioned catalepsy. On the other hand, rats conditioned with haloperidol and given SCH 23390 on the test day continued to exhibit conditioned catalepsy. The above finding suggests that D1-like receptors antagonism does not block expression but only acquisition of conditioned catalepsy in rats.

Our findings that haloperidol, a D2 receptor-preferring antagonist, produces day- to-day catalepsy sensitization is in agreement with previous reports (Schmidt et al., 1999;

Lanis and Schmidt, 2001; Klein and Schmidt, 2003). Schmidt and colleagues have shown that a daily low dose of haloperidol failed to produce catalepsy during the initial test session however, day-to-day increases in the catalepsy response were acquired with

repeated pairing of haloperidol with a particular testing environment (Klein and Schmidt,

2003; Schmidt and Beninger, 2006). On the other hand, a number of studies have reported environment-specific tolerance to haloperidol catalepsy (Barnes et al., 1990;

Hinson et al., 1982; Lappalainen et al., 1989; Poulos and Hinson, 1982). This difference in results has been attributed to dose of the drug (others used supra-threshold catalepsy doses), behavioral testing conditions (others used multiple within-session catalepsy tests) and administration frequency (others used chronic treatment with exponentially higher doses of haloperidol).

When rats were injected with saline instead of haloperidol in the test environment, they continued to exhibit catalepsy - now conditioned. This phenomenon has previously been reported but its mechanisms have not been clearly investigated (Banasikowski and

Beninger, 2010; de Sousa Moreira et al., 1982; Sanberg et al., 1988; Sanberg et al.,

1980). In the present study, control groups that received environment–saline pairing and also received haloperidol on conditioning days in their homecage (colony room) environment did not exhibit increased catalepsy during the test session. Therefore the effect of increased catalepsy in rats having received environment-haloperidol pairing can be attributed to the association of environmental stimuli with haloperidol rather than previous history of haloperidol treatment. Furthermore, Klein and Schmidt (2003) showed that following repeated haloperidol-environment pairings, if the same dose of haloperidol is injected and catalepsy tested in another different environment, animals fail to express catalepsy. Results suggest that haloperidol catalepsy is specific to the

environmental stimuli associated with haloperidol. Results also make an interpretation based on physiological effect of repeated injection (e.g., depolarization block of DA neuron activity) unlikely mechanisms to explain catalepsy sensitization and conditioning.

Catalepsy can also be produced by administration of the D1-like receptor antagonist SCH 23390 (Meller et al., 1985; Morelli and Di Chiara, 1985; Ogren and

Fuxe, 1988). We found that rats treated with SCH 23390 exhibited little catalepsy initially but with repeated treatment and testing catalepsy was observed. When rats were injected with saline instead of SCH 23390 in the test environment, they did not show conditioned catalepsy. The lack of significant catalepsy sensitization seen with SCH

23390 compared to haloperidol cannot explain the lack of conditioned catalepsy, as rats co-treated with both antagonists failed to acquire conditioned catalepsy despite showing a strong catalepsy response following drug treatment. These results dissociate D1-like and

D2 receptors in conditioned catalepsy, where D1-like receptor activity appears to be critical for conditioning to develop.

Synergy between D1-like and D2 antagonists producing catalepsy has been previously reported (Klemm, 1993; Klemm and Block, 1988; Parashos et al., 1989;

Wanibuchi and Usuda, 1990). We found that co-treatment with a dose of SCH 23390 and a dose of haloperidol that failed to produce catalepsy on initial test resulted in a strong synergistic catalepsy response. Further analysis showed that the co-treated group exhibited a significantly greater catalepsy score than the sum of each drug given alone across conditioning days, suggesting that D1-like and D2 receptors may synergize to

produce decreases in initiating voluntary movements.

The mechanism by which both D1-like and D2 receptor antagonists produce extrapyramidal symptoms has previously been reviewed by Miller et al. (1990). In acute studies it has been shown that haloperidol produces catalepsy by interactive effects with striatal cholinergic tone (Costall and Naylor, 1974; Costall et al., 1972; Costall and Olley,

1971; Klemm, 1983; Klemm, 1985a) probably due to disinhibition of the cholinergic interneurons via D2 receptor antagonism (Miller, 2009). On the other hand, D1-like receptor antagonists do not directly influence acetylcholine transmission in the striatum, yet these compounds induce catalepsy. Furthermore catalepsy caused by both haloperidol and SCH 23390 can be rescued with anticholinergic agents (Costall and Naylor, 1974;

Morelli and Di Chiara, 1985; Undie and Friedman, 1988) implying that both D1-like and

D2 receptor antagonists produce catalepsy by increasing cholinergic tone. Also it has be revealed that D2 receptor agonists (Klemm, 1993; Meller et al., 1985; Morelli and Di

Chiara, 1985) but not D1-like receptor agonists (Wanibuchi and Usuda, 1990) can lift catalepsy induced by SCH 23390 suggesting that perhaps the final pathway by which

SCH 23390 produces catalepsy is by its indirect action on D2 receptors and cholinergic tone within the striatum (Miller et al., 1990). Perhaps the observation that co-treatment with SCH 23390 + haloperidol resulted in a synergistic and not additive catalepsy response is related to the combined action of both drugs on D2 receptors.

Alternatevely, haloperidol catalepsy sensitization may result from alterations in the strength of glutamatergic synaptic connections between the cortex and striatum

(Wiecki et al., 2009). Dopamine receptor activation has been shown to play a complex role in controlling corticostriatal plasticity at medium-spiny neurons (MSNs; Reynolds and Wickens, 2000, 2002; Shen et al., 2008). Neurocomputational model developed by

Frank and colleagues (2009) proposes that haloperidol catalepsy sensitization develops as a result of plasticity within the striatopallidal pathway (Wiecki et al., 2009), a major downstream output pathway of the striatum which predominantly expresses D2 receptors

(Berendse et al., 1992; Gerfen and Young, 1988). Indeed DA concentration within the striatum have been implicated in determining the sign of plastic change in both striatopallidal and striatonigral pathways. Under low DA concentration the striatopallidal neurons express long-term potentiation (LTP), and conversely long-term depression

(LTD) under high or normal concentrations (Centonze et al., 2004). The opposite appears to be true for the striatonigral pathway where low DA concentrations lead to LTD, and high DA concentrations lead to LTP (Shen et al., 2008).

It may be that plasticity at corticostriatal synapses relies on cholinergic interneurons as they are believed to play a critical role in integrating both coriticostriatal and DA-ergic activities (Calabresi et al., 2000; Tozzi et al., 2011). Recent evidence shows that induction of LTD within the striatum is critically dependent on the activity of

DA D2 receptors located on cholinergic interneurons (Tozzi et al., 2011; Wang et al.,

2006).

In accordance with the model proposed by Wiecki et al. (2009) context-dependent catalepsy sensitization could result from over-activation of the striatopallidal neurons via

D2 receptor antagonism and subsequent LTP in this pathway (Centonze et al., 2004,

Wiecki et al., 2009). However, as the D2 receptors are found both pre -and post synaptically, initial treatments with haloperidol produce profound increase in synaptic

DA concentrations due to blockade of the pre-synaptic D2 autoreceptors (Bunney and

Grace, 1978; Chiodo and Bunney, 1983; Lidsky and Banerjee, 1993). Augmented DA activity in the striatum can further result from environment novelty and exploratory behavior (Alttoa et al., 2007; Gruen et al., 1990; Redgrave and Gurney, 2006). Thus, enhanced striatal DA transmission due to D2 receptor occupancy and environmental exploration and novelty may lead to higher D1-like receptor stimulation and increased

LTP within the striatonigral pathway (Surmeier et al., 2007). Based on our current behavioral findings, concurrent LTP in the striatonigral pathway (via D1-like receptors activation) as well as in the striatopallidal pathway (via D2 receptor antagonism) may be necessary for development of catalepsy sensitization and conditioning.

The medial prefrontal cortex (mPFC) and hippocampus (HPC) have been shown to influence haloperidol catalepsy (Dijk et al., 1991; Klockgether et al., 1988; Tucci et al.,

1994). Ibotenic acid lesions of the mPFC or the ventral HPC significantly attenuate haloperidol catalepsy in rats (Lipska et al., 1995). However, these studies used a cataleptogenic dose of haloperidol in acute tests. The mPFC and ventral HPC have been shown to be critical in the development of context-dependent behavioral sensitization to psychostimulants (Cador et al., 1999; Lodge and Grace, 2008; Wolf et al., 1995). The possible contribution of these structures to catalepsy sensitization and conditioning awaits

further study.

Context-dependent sensitization (Klein and Schmidt, 2003) and conditioning

(Amtage et al., 2003; de Sousa Moreira et al., 1982; Sanberg, 1988) of haloperidol induced catalepsy have been reported in the past but their mechanism has not been elucidated. Our finding that D1-like receptors are necessary for development of catalepsy sensitization and conditioning is in agreement with previous reports implicating D1-like receptors in locomotor sensitization to psychostimulants (El-Ghundi et al., 2010; Vezina,

1996) and in acquisition of many conditioned behaviors such as conditioned place preference (Cervo and Samanin, 1995), conditioned activity (Mazurski and Beninger,

1991), conditioned approach (Choi et al., 2005; Dalley et al., 2005) and instrumental responding (Smith-Roe and Kelley, 2000).

Dopamine receptor blockade has been implicated in reductions in motivation and goal-directed movements in both animals (Salamone and Correa, 2002; Wise, 2004) and humans (Stern et al., 2005). Animals given a low dose of haloperidol are not cataleptic during initial drug-environment pairings instead exhibiting similar exploratory behaviors to control animals. It is the progressive loss of exploratory behaviors due to decreased engagement with their environment that eventually manifests as catalepsy.

Thus context-dependent catalepsy sensitization and conditioning are intriguing phenomena that allow us to study how environmental stimuli experienced under altered

DA transmission may contribute to certain pathologies in humans, particularly psychostimulant dependence, Parkinson’s disease (PD) and schizophrenia. Our finding

that catalepsy sensitization and conditioned catalepsy is dependent on D1-like receptors supports studies examining psychostimulant sensitization (Vezina, 1996; Schmidt and

Beninger, 2006) and context conditioning of psychostimulant effects in rats

(Banasikowski and Beninger, 2010; Cervo and Samanin, 1995; Mazurski and Beninger,

1991). Results suggest that these types of learning are more closely related than previously thought. Altered DA levels in PD may change the way environmental stimuli control behavior perhaps contributing to the progression of PD symptoms. Furthermore, understanding antipsychotic drug-environment interactions may shed light on the slow therapeutic benefit of antipsychotic drugs in schizophrenia (Agid et al., 2003; Beninger,

1988; Grace, 1997; Valenti et al., 2011) and why certain schizophrenic patients relapse into a delusional state once they have been released from the hospital (Ayuso-Gutierrez and del Rio Vega, 1997; Fallon and Dursun, 2010; Gaebel and Pietzcker, 1985). Our findings identify possible mechanisms related to the role of DA in influencing the ability of environmental stimuli to control behavior that may be relevant to these observations.

2.6 References

Agid O, Kapur S, Arenovich T, Zipursky RB (2003). Delayed-onset hypothesis of antipsychotic action: a hypothesis tested and rejected. Archives of General Psychiatry, 60(12), 1228-1235. Alttoa A, Eller M, Herm L, Rinken A, Harro J (2007) Amphetamine-induced locomotion, behavioral sensitization to amphetamine, and striatal D2 receptor function in rats with high or low spontaneous exploratory activity: Differences in the role of locus coeruleus. Brain Research, 1131, 138-148. Amtage J, Schmidt WJ (2003). Context-dependent catalepsy intensification is due to classical conditioning and sensitization. Behavioural Pharmacology, 14(7), 563- 567. Ayuso-Gutierrez JL, del Rio Vega JM (1997). Factors influencing relapse in the long- term course of schizophrenia. Schizophrenia Research, 28(2-3), 199-206. Baik JH, Picetti R, Saiardi A, Thiriet G, et al. (1995). Parkinsonian-like locomotor impairment in mice lacking dopamine D2 receptors. Nature, 377(6548), 424-428. Banasikowski TJ, Beninger RJ (2010) Conditioned drug effects. In: Stolerman IP (Ed.) Encyclopedia of Psychopharmacology (pp. 325-331) Heidelberg: Springer. Barnes DE, Robinson B, Csernansky JG, Bellows EP (1990). Sensitization versus tolerance to haloperidol-induced catalepsy: multiple determinants. Pharmacology Biochemistry and Behaviour, 36(4), 883-887. Beninger RJ (1988). The slow therapeutic action of antipsychotic drugs: a possible mechanism involving the role of dopamine in incentive learning. In: Simon P, Soubrié P, Wildlocher D (Eds.), Animal Models of Psychiatric Disorders Vol 1: Selected Models of Anxiety, Depression and Psychosis (pp 36-51). Basal: Karger. Berendse HW, Groenewegen HJ, Lohman AH (1992). Compartmental distribution of ventral striatal neurons projecting to the mesencephalon in the rat. Journal of Neuroscience, 12(6), 2079-2103. Bunney BS, Grace AA (1978) Acute and chronic haloperidol treatment: comparison of effects on nigral dopaminergic cell activity. Life Science, 23, 1715-1727. Cador M, Bjijou Y, Cailhol S, Stinus L (1999). D-amphetamine-induced behavioral sensitization: implication of a glutamatergic medial prefrontal cortex-ventral tegmental area innervation. Neuroscience, 94(3): 705-721.

Calabresi P, Centonze D, Gubellini P, Pisani A, et al. (2000). Acetylcholine-mediated modulation of striatal function. Trends in Neuroscience, 23(3), 120-126. Centonze D, Usiello A, Costa C, Picconi B, et al. (2004). Chronic haloperidol promotes corticostriatal long-term potentiation by targeting dopamine D2L receptors. Journal of Neuroscience, 24(38), 8214-8222. Cervo L, Samanin R (1995). Effects of dopaminergic and glutamatergic receptor antagonists on the acquisition and expression of cocaine conditioning place preference. Brain Research, 673(2), 242-250. Chiodo LA, Bunney BS (1983) Typical and atypical neuroleptics: differential effects of chronic administration on the activity of A9 and A10 midbrain dopaminergic neurons. Journal of Neuroscience, 3,1607-1619. Choi WY, Balsam PD, Horvitz JC (2005). Extended habit training reduces dopamine mediation of appetitive response expression. Journal of Neuroscience, 25(29), 6729-6733. Costall B, Naylor RJ (1974). On catalepsy and catatonia and the predictability of the catalepsy test for neuroleptic activity. Psychopharmacologia, 34(3), 233-241. Costall B, Naylor RJ, Olley JE (1972). On the involvement of the caudate-putamen, globus pallidus and substantia nigra with neuroleptic and cholinergic modification of locomotor activity. Neuropharmacology, 11(3), 317-330. Costall B, Olley JE (1971). Cholinergic and neuroleptic induced catalepsy: modification by lesions in the globus pallidus and substantia nigra. Neuropharmacology, 10(5), 581-594. Crocker AD, Hemsley KM (2001). An animal model of extrapyramidal side effects induced by antipsychotic drugs: relationship with D2 dopamine receptor occupancy. Progress in Neuropsychopharmacology and Biological Psychiatry, 25(3), 573-590. Dalley JW, Laane K, Theobald DE, Armstrong HC, et al. (2005). Time-limited modulation of appetitive Pavlovian memory by D1 and NMDA receptors in the nucleus accumbens. Proceedings of the National Academy of Science U S A, 102(17), 6189-6194. de Sousa Moreira LF, da Conceição Camargo Pinheiro M, Masur J (1982). Catatonic Behavior Induced by Haloperidol, Increased by Retesting and Elicited without Drug in Rats. Pharmacology, 25(1), 1.

Dijk S, Krugers HJ, Korf J (1991). The effect of theophylline and immobilization stress on haloperidol-induced catalepsy and on metabolism in the striatum and hippocampus, studied with lactography. Neuropharmacology, 30(5), 469-473. El-Ghundi MB, Fan T, Karasinska JM, Yeung J, Zhou M, O'Dowd BF, George SR (2010) Restoration of amphetamine-induced locomotor sensitization in dopamine D1 receptor-deficient mice. Psychopharmacology (Berl), 207,599-618. Elliott PJ, Close SP, Walsh DM, Hayes AG, et al. (1990). Neuroleptic-induced catalepsy as a model of Parkinson's disease. I. Effect of dopaminergic agents. Journal of Neural Transmission Parkinson’s Disease and Dementia Section, 2(2), 79-89. Fallon P, Dursun SM (2010). A naturalistic controlled study of relapsing schizophrenic patients with tardive dyskinesia and supersensitivity psychosis. Journal of Psychopharmacology, 25(6), 755-762. Gaebel W, Pietzcker A (1985). Multidimensional study of the outcome of schizophrenic patients 1 year after clinic discharge. Predictors and influence of neuroleptic treatment. European Archive in Psychiatry and Neurological Sciences, 235(1), 45-52. Gerfen CR, Young WS (1988) Distribution of striatonigral and striatopallidal peptidergic neurons in both patch and matrix compartments: an in situ hybridization histochemistry and fluorescent retrograde tracing study. Brain Research, 460,161-167. Grace AA, Bunney BS, Moore H, Todd CL (1997). Dopamine-cell depolarization block as a model for the therapeutic actions of antipsychotic drugs. Trends in Neuroscience, 20(1), 31-37. Graybiel AM (2008) Habits, rituals, and the evaluative brain. Annual Reviews in Neuroscience, 31, 359-387. Gruen RJ, Deutch AY, Roth RH (1990) Perinatal diazepam exposure: Alterations in exploratory behavior and mesolimbic dopamine turnover. Pharmacology Biochemistry and Behavior, 36,169-175. Hinson RE, Poulos CX, Thomas WL (1982). Learning in tolerance to haloperidol- induced catalepsy. Progress in Neuropsychopharmacology and Biological Psychiatry, 6(4-6): 395-398. Hornykiewicz O (1973). Parkinson's disease: from brain homogenate to treatment. Federation Proceedings, 32(2), 183-190.

Klein A, Schmidt WJ (2003). Catalepsy intensifies context-dependently irrespective of whether it is induced by intermittent or chronic dopamine deficiency. Behavioural Pharmacology, 14(1), 49-53. Klemm WR (1983). Cholinergic-dopaminergic interactions in experimental catalepsy. Psychopharmacology (Berl), 81(1), 24-27. Klemm WR (1985a). Evidence for a cholinergic role in haloperidol-induced catalepsy. Psychopharmacology (Berl,, 85(2), 139-142. Klemm WR (1985b). Neuroleptic-induced catalepsy: a D2 blockade phenomenon? Pharmacology Biochemistry and Behaviour, 23(6), 911-915. Klemm WR (1993). The catalepsy of blocked dopaminergic receptors. Psychopharmacology (Berl), 111(2), 251-255. Klemm WR, Block H (1988). D-1 and D-2 receptor blockade have additive cataleptic effects in mice, but receptor effects may interact in opposite ways. Pharmacology Biochemistry and Behaviour, 29(2), 223-229. Klockgether T, Schwarz M, Turski L, Sontag KH (1988). Catalepsy after microinjection of haloperidol into the rat medial prefrontal cortex. Experimental Brain Research, 70(2), 445-447. Lanis A, Schmidt WJ (2001). NMDA receptor antagonists do not block the development of sensitization of catalepsy, but make its expression state- dependent. Behavioural Pharmacology, 12(2), 143-149. Lappalainen J, Hietala J, Syvalahti E (1989). Differential tolerance to cataleptic effects of SCH 23390 and haloperidol after repeated administration. Psychopharmacology (Berl), 98(4), 472-475. Lidsky TI, Banerjee SP (1993) Acute administration of haloperidol enhances dopaminergic transmission. Journal of Pharmacology and Experimental Therapeutics, 265, 1193-1198. Lipska BK, Jaskiw GE, Braun AR, Weinberger DR (1995). Prefrontal cortical and hippocampal modulation of haloperidol-induced catalepsy and apomorphine- induced stereotypic behaviors in the rat. Biological Psychiatry, 38(4), 255-262. Lodge DJ, Grace AA (2008). Amphetamine activation of hippocampal drive of mesolimbic dopamine neurons: a mechanism of behavioral sensitization. Journal of Neuroscience, 28(31), 7876-7882. Matamales M, Bertran-Gonzalez J, Salomon L, Degos B, Deniau JM, Valjent E, Herve D, Girault JA (2009) Striatal medium-sized spiny neurons: identification 62

by nuclear staining and study of neuronal subpopulations in BAC transgenic mice. PLoS One, 4, e4770. Mazurski EJ, Beninger RJ (1991). Effects of selective drugs for dopaminergic D1 and D2 receptors on conditioned locomotion in rats. Psychopharmacology (Berl), 105(1), 107-112. Meller E, Kuga S, Friedhoff AJ, Goldstein M (1985). Selective D2 dopamine receptor agonists prevent catalepsy induced by SCH 23390, a selective D1 antagonist. Life Science, 36(19), 1857-1864. Miller R (2009) Mechanisms of Action of Antipsychotic Drugs of Different Classes, Refractoriness to Therapeutic Effects of Classical Neuroleptics, and Individual Variation in Sensitivity to their Actions: PART I. Current Neuropharmacolology, 7, 302-314. Miller R, Wickens JR, Beninger RJ (1990). Dopamine D-1 and D-2 receptors in relation to reward and performance: a case for the D-1 receptor as a primary site of therapeutic action of neuroleptic drugs. Progress in Neurobiology, 34(2), 143- 183. Morelli M, Di Chiara G (1985). Catalepsy induced by SCH 23390 in rats. European Journal of Pharmacology, 117(2), 179-185. Ogren SO, Fuxe K (1988). D1- and D2-receptor antagonists induce catalepsy via different efferent striatal pathways. Neuroscience Letters, 85(3), 333-338. Ossowska K, Karcz M, Wardas J, Wolfarth S (1990). Striatal and nucleus accumbens D1/D2 dopamine receptors in neuroleptic catalepsy. European Journal of Pharmacology, 182(2), 327-334. Papeschi R (1972). Dopamine, extrapyramidal system, and psychomotor function. Psychiatra Neurologia Neurochirurgia, 75(1), 13-48. Parashos SA, Marin C, Chase TN (1989). Synergy between a selective D1 antagonist and a selective D2 antagonist in the induction of catalepsy. Neuroscience Letters, 105(1-2), 169-173. Poulos CX, Hinson R (1982). Pavlovian conditional tolerance to haloperidol catalepsy: evidence of dynamic adaptation in the dopaminergic system. Science, 218(4571), 491-492. Redgrave P, Gurney K (2006) The short-latency dopamine signal: a role in discovering novel actions? Nature Reviews Neuroscience, 7, 967-975.

Reynolds JN, Wickens JR (2000) Substantia nigra dopamine regulates synaptic plasticity and membrane potential fluctuations in the rat neostriatum, in vivo. Neuroscience, 99, 199-203. Reynolds JN, Wickens JR (2002) Dopamine-dependent plasticity of corticostriatal synapses. Neural Networks, 15, 507-521. Riedinger K, Kulak A, Schmidt WJ, von Ameln-Mayerhofer A (2011) The role of NMDA and AMPA/Kainate receptors in the consolidation of catalepsy sensitization. Behavioural Brain Research, 218, 194-199. Salamone JD, Correa M (2002). Motivational views of reinforcement: implications for understanding the behavioral functions of nucleus accumbens dopamine. Behavioural Brain Research, 137(1-2), 3-25. Sanberg PR, Bunsey MD, Giordano M, Norman AB (1988). The catalepsy test: its ups and downs. Behavioural Neuroscience, 102(5), 748-759. Sanberg PR, Pisa M, Faulks IJ, Fibiger HC (1980). Experimental influences on catalepsy. Psychopharmacology (Berl), 69(2), 225-226. Shen W, Flajolet M, Greengard P, Surmeier DJ (2008) Dichotomous dopaminergic control of striatal synaptic plasticity. Science, 321, 848-851. Schmidt WJ, Beninger RJ (2006). Behavioural sensitization in addiction, schizophrenia, Parkinson's disease and dyskinesia. Neurotoxicity Research, 10(2), 161-166. Schmidt WJ, Tzschentke TM, Kretschmer BD (1999). State-dependent blockade of haloperidol-induced sensitization of catalepsy by MK-801. European Jouranl of Neuroscience, 11(9), 3365-3368. Smith-Roe SL, Kelley AE (2000). Coincident activation of NMDA and dopamine D1 receptors within the nucleus accumbens core is required for appetitive instrumental learning. Journal of Neuroscience, 20(20), 7737-7742. Srinivasan J, Schmidt WJ (2004). Intensification of cataleptic response in 6- hydroxydopamine-induced neurodegeneration of substantia nigra is not dependent on the degree of dopamine depletion. Synapse, 51(3), 213-218. Stern ER, Horvitz JC, Cote LJ, Mangels JA (2005). Maintenance of response readiness in patients with Parkinson's disease: evidence from a simple reaction time task. Neuropsychology, 19(1), 54-65.

Surmeier DJ, Ding J, Day M, Wang Z, Shen W (2007) D1 and D2 dopamine-receptor modulation of striatal glutamatergic signaling in striatal medium spiny neurons. Trends in Neuroscience, 30, 228-235. Tozzi A, de Iure A, Di Filippo M, Tantucci M, et al. (2011). The distinct role of medium spiny neurons and cholinergic interneurons in the D/AA receptor interaction in the striatum: implications for Parkinson's disease. Journal of Neuroscience, 31(5), 1850-1862. Tucci S, Fernandez R, Baptista T, Murzi E, et al. (1994). Dopamine increase in the prefrontal cortex correlates with reversal of haloperidol-induced catalepsy in rats. Brain Research Bulletin, 35(2), 125-133. Undie AS, Friedman E (1988). Differences in the cataleptogenic actions of SCH23390 and selected classical neuroleptics. Psychopharmacology (Berl), 96(3), 311-316. Usiello A, Baik JH, Rouge-Pont F, Picetti R, et al. (2000). Distinct functions of the two isoforms of dopamine D2 receptors. Nature, 408(6809), 199-203. Valenti O, Cifelli P, Gill KM, Grace AA (2011). Antipsychotic drugs rapidly induce dopamine neuron depolarization block in a developmental rat model of schizophrenia. Journal of Neuroscience, 31(34), 12330-12338. Vezina P (1996). D1 dopamine receptor activation is necessary for the induction of sensitization by amphetamine in the ventral tegmental area. Journal of Neuroscience, 16(7), 2411-2420. Wang Z, Kai L, Day M, Ronesi J, et al. (2006). Dopaminergic control of corticostriatal long-term synaptic depression in medium spiny neurons is mediated by cholinergic interneurons. Neuron, 50(3): 443-452. Wadenberg ML, Kapur S, Soliman A, Jones C, et al. (2000). Dopamine D2 receptor occupancy predicts catalepsy and the suppression of conditioned avoidance response behavior in rats. Psychopharmacology (Berl), 150(4), 422-429. Wanibuchi F, Usuda S (1990). Synergistic effects between D-1 and D-2 dopamine antagonists on catalepsy in rats. Psychopharmacology (Berl), 102(3), 339-342. Wiecki TV, Riedinger K, von Ameln-Mayerhofer A, Schmidt WJ, Frank MJ (2009) A neurocomputational account of catalepsy sensitization induced by D2 receptor blockade in rats: context dependency, extinction, and renewal. Psychopharmacology (Berl) 204, 265-277. Wise RA (2004). Dopamine, learning and motivation. Nature Reviews Neuroscience, 5(6), 483-494.

Wolf ME, Dahlin SL, Hu XT, Xue CJ, et al. (1995). Effects of lesions of prefrontal cortex, amygdala, or fornix on behavioral sensitization to amphetamine: comparison with N-methyl-D-aspartate antagonists. Neuroscience, 69(2): 417- 439.

Chapter 3

Reduced Expression of Haloperidol Conditioned Catalepsy in Rats by

the Dopamine D3 Receptor Antagonists Nafadotride and NGB 2904

3.1 Abstract

Haloperidol, a dopamine (DA) D2 receptor-preferring antagonist, produces catalepsy whereby animals maintain awkward posture for a period of time. Sub-threshold doses of haloperidol fail to produce catalepsy initially, however, when the drug is given repeatedly in the same test environment, gradual day-to-day increases in catalepsy are observed. More importantly, if sensitized rats are injected with saline instead of haloperidol they continue to be cataleptic in the test environment suggesting that environment-drug associations may play a role. DA D3 receptors have been implicated in a number of conditioned behaviors. We were interested if DA D3 receptors contribute to catalepsy sensitization and conditioning in rats. We tested this hypothesis using the DA

D3 receptor-selective antagonist NGB 2904 (0.5, 1.8 mg/kg) and the DA D3 receptor- preferring antagonist nafadotride (0.1, 0.5 mg/kg). For 10 consecutive conditioning days rats were treated with one of the D3 receptor antagonists alone or in combination with haloperidol (0.25 mg/kg) and tested for catalepsy, quantified by the time a rat remained with its forepaws on a horizontal bar. On test day (day 11), rats were injected with saline or the D3 receptor antagonist and tested for conditioned catalepsy in the previously drug- paired environment. Rats treated with NGB 2904 or nafadotride alone did not develop catalepsy. Rats treated with haloperidol or haloperidol plus NGB 2904 or nafadotride developed catalepsy sensitization with repeated conditioning. When injected with saline they continued to exhibit catalepsy in the test environment – now conditioned. On the other hand, NGB 2904 (1.8 mg/kg) or nafadotride (0.5 mg/kg) given on the test day (after

sensitization to haloperidol) significantly attenuated the expression of conditioned catalepsy. Our data suggest that the D3 receptor antagonist NGB 2904 (1.8 mg/kg) and nafadotride (0.5 mg/kg) significantly attenuate conditioned catalepsy in rats when given in test but not when given during sensitization. Results implicate DA D3 receptors in regulating the expression of conditioned catalepsy.

3.2 Introduction

Alterations in DA neurotransmission observed in a number of neuropsychiatric conditions such as schizophrenia, Parkinson’s disease (PD) and substance abuse have been associated with changes in DA D3 receptor function. Initially, postmortem studies found elevated D3 receptor levels in the ventral striatum in drug-free schizophrenia patients, but not in patients on medication at the time of death (Guillin et al., 2001;

Gurevich et al., 1997). Recent studies using positron emission tomography (PET) found an increase in D3 receptor binding using [11C]-(+)-PHNO in the globus pallidus, substantia nigra and a decrease in the ventral striatum after initiation of repeated antipsychotic treatment in previously drug-naïve schizophrenia patients (Mizrahi et al.,

2011). On the other hand, PET studies in drug-naïve PD patients found a decrease in D3 receptor binding in the globus pallidus and ventral striatum (Boileau et al., 2009).

Postmortem studies in people who overdosed on cocaine found increased D3 receptor binding and mRNA in the ventral striatum when compared to age-matched cocaine-free subjects (Segal et al., 1997; Staley and Mash, 1996).

Changes in D3 receptor mRNA and/or expression levels in the ventral striatum

have been identified in experimental animals treated with drugs that directly or indirectly activate the DA system such as levodopa (Bezard et al., 2003; Bordet et al., 1997), amphetamine (Chiang et al., 2003), cocaine (Le Foll et al., 2002; Neisewander et al.,

2004), nicotine (Le Foll et al., 2003) and morphine (Liang et al., 2011). The animals that showed changes in D3 receptor levels also exhibited behavioral changes associated with the repeated drug treatment such as sensitization (Bordet et al., 1997; Chiang et al., 2003;

Le Foll et al., 2003) and drug-environment conditioning (Le Foll et al., 2002; Liang et al.,

2011; Neisewander et al., 2004). Further studies using pharmacological manipulations have shown that D3 receptor antagonists block the expression of cocaine (Banasikowski et al., 2010; Le Foll et al., 2002) and nicotine (Pak et al., 2006) conditioned activity as well as the expression of conditioned place preference (CPP) based on cocaine (Vorel et al., 2002)and nicotine (Le Foll et al., 2005).

Recently it has been shown that repeated dopamine receptor antagonism

(haloperidol) can produce catalepsy sensitization in a context-dependent manner

(Banasikowski and Beninger, 2012a; Riedinger et al., 2011; Schmidt et al., 1999; Wiecki et al., 2009) and conditioned catalepsy (Amtage and Schmidt, 2003; Banasikowski and

Beninger, 2012a)when tested drug-free in the drug-paired environment. Similarly to sensitization and conditioning with drugs that increase DA concentrations, haloperidol catalepsy sensitization and conditioning is dependent on the environmental stimuli present during conditioning and test (Schmidt and Beninger, 2006).

As D3 receptors play a significant role in the expression of conditioned behaviors

based on drugs that activate the DA system (i.e., conditioned activity, CPP), it is important to test whether D3 receptors are involved in conditioning to drugs that reduce

DA activity (i.e., conditioned catalepsy). Our studies examined the role of D3 receptors in haloperidol catalepsy sensitization, and in acquisition and expression of conditioned catalepsy. We tested the hypothesis that the D3 receptor-preferring antagonist nafadotride and the D3 receptor-selective antagonist NGB 2904 will attenuate the expression of haloperidol conditioned catalepsy at doses that will fail to attenuate its acquisition.

3.3 Materials and Methods

3.3.1 Subjects Experimentally naïve male albino Wistar rats (N = 135, Charles River Canada, St.

Constant QC) weighing 320-340 g upon the start of experiments were housed in pairs or threes in clear Plexiglas cages (45.0 x 25.0 x 22.0 cm). Average temperature in the colony was 21º C, humidity 70% with reversed light–dark cycle (lights off from 0700 to

1900 hr). Rats were maintained with food (LabDiet 5001, PMI Nutrition International,

Brentwood, MO, USA) and water continuously available. Treatment of rats was in accordance with the guidelines of the Animals for Research Act, the Canadian Council on

Animal Care, and was approved by the Queen’s University Animal Care Committee.

3.3.2 Drugs Haloperidol, (4-[4-(4-chlorophenyl)-4-hydroxy-1-piperidyl]-1-(4-fluorophenyl)- butan-1-one (Sigma, St. Louis, MO, USA) was prepared in a 0.3 % distilled water solution of tartaric acid. Nafadotride, N-[(1-Butyl-2-pyrrolidinyl)methyl]-4-cyano-1- 71

methoxy-2-naphthalenecarboxamide and NGB 2904, N-[4-[4-(2,3-Dichlorophenyl)-1- piperazinyl]butyl]-9H-fluorene-2-carboxamide (Tocris, Oakville, ON) was prepared in a

10% distilled water solution of 2-hydroxypropyl-β-cyclodextrin. Injections were administered intraperitoneally (i.p.) in a volume of 1 ml/kg.

3.3.3 Behavioral Testing

Rats were randomly assigned to treatment (paired) and control groups (unpaired).

Animals were removed from the colony room and injected i.p. with drug or saline and put back into their cages in a hallway lit by 34-watt fluorescent tubes. Thirty minutes

[nafadotride (NAF), NGB 2904 (NGB)] or 60 min [haloperidol (hal)] later, they were tested, in a small testing room also illuminated by two 34-watt fluorescent tubes, on a horizontal bar (1.6 cm diam. threaded or smooth rod with end bolts attached to Plexiglas supports, 10 cm above the surface), by gently placing both forepaws on the bar. Descent latency was measured as the time span from placing the animal on the bar until the first active paw movement. A cut-off time of 180 s was used, i.e., the trial was terminated when the animal did not make an active paw movement within that time.

3.3.4 Experimental Design

equipped with a distinct horizontal bar. The black chamber was equipped with a 1.6 cm diam. smooth rod bolted to clear Plexiglass supports (1 cm thick, 10 cm high), and the white chamber was equipped with a 1.6 cm diam. threaded rod bolted to black Plexiglass supports (0.5 cm thick and 10 cm high). Surgical paper was used as the floor covering in both chambers. Rats were randomly assigned to one of the two test chambers. Each rat was always placed in the same chamber during the haloperidol sensitization phase and all treatment groups included rats sensitized in each chamber. We observed no relationship between the magnitude of the catalepsy effect and the testing chambers, and no effect of testing chambers on catalepsy sensitization or conditioning.

Experiment 1: Nafadotride Effects on Acquisition and Expression of Catalepsy

Two groups of rats were used to examine haloperidol (0.25 mg/kg) catalepsy sensitization and conditioned catalepsy; the 0.25 mg/kg dose was used in previous conditioned catalepsy studies by Schmidt and coworkers (1999) and by us (Banasikowski et al., 2012). Animals in the ‘paired’ condition were injected with haloperidol outside the testing room for 10 consecutive days, 60 min before the catalepsy test. Sixty min after the catalepsy test, rats were administered saline in their colony room (homecage injection).

This was the hal 0.25 paired/sal group (n = 9). The ‘unpaired’ group received saline 60 min before and haloperidol 60 min after the catalepsy test. This was the hal 0.25 unpaired/sal group (n = 9). This design allowed for all animals to experience the same drug history but in different environments and at different time intervals (before test versus after tests). On day 11 all animals received saline 60 min prior to the conditioned

catalepsy test and no homecage injection.

Two groups of rats received ‘paired’ (n = 8) and ‘unpaired’ (n = 10) treatment with nafadotride (0.5 mg/kg).

Two groups of rats examined the effect of nafadotride on haloperidol catalepsy sensitization and acquisition of conditioned catalepsy. Rats received haloperidol (0.25 mg/kg) 60 min before and nafadotride [0.1 mg/kg (n = 10) or 0.5 mg/kg (n = 9)] 30 min before each catalepsy test. These doses of nafadotride were used previously in behavioral studies of amphetamine-conditioned place preference (Banasikowski et al., 2012). On day

11 all animals received saline 60 min prior to the catalepsy test and no homecage injection.

Two groups of rats examined the effects of nafadotride on expression of haloperidol conditioned catalepsy. Rats received haloperidol (0.25 mg/kg) for 10 consecutive conditioning days 60 min before each catalepsy test. On the saline test day rats were injected with nafadotride [0.1 mg/kg (n = 10) or 0.5 mg/kg (n =8)] 30 min before the catalepsy test with saline.

Experiment 2: NGB 2904 Effects on Acquisition and Expression of Catalepsy

The drug treatment and behavioral testing protocol was based on procedures used in experiment 1. Two groups of rats, ‘paired’ (n = 10) and ‘unpaired’ (n = 9), were injected with haloperidol (0.25 mg/kg) or saline for 10 consecutive conditioning days, 60 min before the catalepsy test, and received saline or haloperidol (0.25 mg/kg), respectively, 60 min after testing in their colony room (homecage injection). A group of

rats (n = 8) injected with NGB 2904 (1.8 mg/kg) was used to examine whether NGB 2904 alone produced catalepsy. Two groups examined the effect of NGB 2904 on haloperidol catalepsy sensitization and acquisition of conditioned catalepsy. NGB 2904 doses were based on those used previously in stimulant self-administration studies (Gilbert et al.,

2005; Martelle et al., 2007). Groups hal 0.25 + NGB 0.5/sal (n = 9) and hal 0.25 + NGB

1.8/sal (n = 9) received haloperidol (0.25 mg/kg) 60 min and NGB (0.5 or 1.8 mg/kg, respectively) 30 min before the catalepsy test. On day 11 all animals received saline 60 min prior to catalepsy test and no homecage injection.

Two groups examined the effects of NGB 2904 on expression of haloperidol conditioned catalepsy. Groups hal 0.25/NGB 0.5 (n = 8) and hal 0.25/NGB 1.8 (n =9) were injected with haloperidol (0.25 mg/kg) for 10 conditioning days. On day 11 rats were injected with NGB 2904 (0.5 or 1.8 mg/kg) 30 min before the saline catalepsy test.

3.3.5 Statistics All statistical analysis was performed using IBM SPSS 19.0. All data are presented as means ± standard error. Data were submitted to both parametric and nonparametric tests and each type of analysis yielded similar results. Only the parametric tests are reported in most cases. They involved two-way and one-way analyses of variance (ANOVA). Hypotheses tests were completed using α = 0.05 and pair-wise comparisons were made using Dunnett’s test and where specified, Fisher’s LSD test.

3.4 Results

3.4.1 Experiment 1: Nafadotride Effects on Acquisition and Expression of Catalepsy 75

Across conditioning days 1 to 10 all group receiving haloperidol prior to testing showed increasing catalepsy over days (Fig. 4A); catalepsy was not observed in the

‘unpaired’ or ‘paired’ nafadotride-alone groups. A two-way ANOVA revealed a significant main effect of day [F(9, 585) = 13.34, p < 0.001] , treatment [F(7,65) = 8.70, p

< 0.001] and interaction [F(63,585 = 1.74, p < 0.01]. For the main effect of group, post- hoc comparison found that rats treated with haloperidol (hal 0.25 paired/sal, hal

0.25/NAF 0.1, hal 0.25/NAF 0.5) or co-treated with haloperidol and nafadotride (hal 0.25

+ NAF 0.1/sal, hal 0.25 + NAF 0.5/sal) spent significantly more time with both paws on the bar (descent latency) than hal 0.25 unpaired/sal controls (p < 0.05). Nafadotride given alone in sensitization sessions did not produce catalepsy.

To further analyze the interaction, simple effects tests of group were performed on the first and last day of conditioning. No significant group differences were obtained on conditioning day 1 [F(7,65) = 1.85, p = 0.92] but a significant group difference was seen on conditioning day 10 [F(7,65) = 4.36, p < 0.01]. Post-hoc comparisons for day 10 found that groups conditioned with haloperidol alone, hal 0.25 + NAF 0.1/sal, or hal 0.25

+ NAF 0.5/sal had longer descent latencies than the hal 0.25 unpaired/sal control group (p

< 0.05). The groups showing catalepsy did not significantly differ from one another (p >

0.05; Fisher’s LSD).

On the saline test day rats previously sensitized with haloperidol or haloperidol plus nafadotride continued to be cataleptic in the test environment. The 0.5 mg/kg dose of nafadotride given during the test reduced the conditioned catalepsy effect. One-way

Figure 4

Mean (± SEM) descent latency (s) in groups of rats receiving 0.25 mg/kg haloperidol (hal) 60 min, or co-treated with 0.25 mg/kg haloperidol (60 min) and 0.1 or 0.5 mg/kg nafadotride (NAF) 30 min prior to each conditioning day (A) and saline (sal) 60 min or 0.1 or 0.5 mg/kg NAF 30 min prior to test session (B). * significantly different from hal 0.25 unpaired/sal in pairwise comparison (Dunnett’s test) following significant group effects in analysis of variance. # significantly different from hal 0.25 paired/sal in pairwise comparison (Fisher’s LSD test) following significant group effects in analysis of variance. † significantly different from NAF 0.5 unpaired/sal.

ANOVA revealed a significant difference among groups [F(7,65) = 4.54, p <

0.001; Fig. 4B]. Post-hoc comparisons found that groups hal 0.25 paired/sal, hal 0.25 +

NAF 0.1/sal, hal 0.25 + NAF 0.5/sal and hal 0.25/NAF 0.1, but not hal 0.25/NAF 0.5, had longer descent latencies than hal 0.25 unpaired/sal (p < 0.05). Further analysis found that the hal 0.25/NAF 0.5 but not hal 0.25 + NAF 0.5/sal group was significantly different from the hal 0.25 paired/sal group (p < 0.05; Fisher’s LSD) suggesting that a nafadotride dose of 0.5 mg/kg reduced expression but not acquisition of conditioned catalepsy.

A separate 1-way ANOVA revealed that rats in the NAF 0.5 paired/sal group showed significant conditioned catalepsy when compared to the NAF 0.5 unpaired/sal group [F(1, 16) = 5.86, p < 0.05]. Similarly, the NAF 0.5 paired/sal group differed from the hal 0.25 unpaired/sal group [F(1,16) = 8.74, p < 0.05]; the NAF 0.5 unpaired/sal and hal 0.25 unpaired/sal groups did not differ significantly from one another [F(1,17) = 0.55, p > 0.05]. This finding suggests that although repeated treatment with nafadotride in the test environment fails to produce catalepsy or catalepsy sensitization, it is capable of producing a low level of conditioned catalepsy.

3.4.2 Experiment 2: NGB 2904 Effects on Acquisition and Expression of Catalepsy

Across conditioning days 1 to 10 a two-way ANOVA revealed a significant main effect of day [F(9, 495) = 10.73, p < 0.001], treatment [F(6,55) = 12.14, p < 0.001; Fig.

5A] and interaction [F(54, 495) = 1.69, p < 0.01]. Post-hoc comparisons for the main effect of treatment found that all rats given haloperidol during sensitization exhibited

significantly longer descent latencies than the hal 0.25 unpaired/sal control group (p <

0.05). NGB 2904 (1.8 mg/kg) given alone during sensitization sessions did not produce catalepsy.

To further analyze the interaction, simple effects tests of group were performed on conditioning day1, day 5 and day 10. No significant group differences were obtained on conditioning day 1 [F (6,55) = 1.97, p = 0.86] but a significant group effect was seen on conditioning day 5 [F(6,55) = 12.78, p < 0.001] and conditioning day 10 [F (6,55) =

4.10, p < 0.01]. Post-hoc comparisons found that group hal 0.25 + NGB 1.8/sal exhibited significantly higher catalepsy scores from other haloperidol-treated animals on day 5 (p <

0.05; Fisher’s LSD) but not on day 10. Further post-hoc comparisons on day 10 found that groups hal 0.25 paired/sal, hal 0.25 + NGB 0.5/sal, hal 0.25 + NGB 1.8/sal, hal

0.25/NGB 0.5, and hal 0.25/NGB 1.8 developed catalepsy sensitization and spent significantly more time with both paws on the bar than the hal 0.25 unpaired/sal group (p

< 0.05).

On the saline test day, 1-way ANOVA revealed a significant difference among groups [F(6,55) = 3.46, p < 0.01; Fig. 5B]. Post-hoc comparisons found that groups hal

0.25 paired/sal, hal 0.25 + NGB 0.5/sal, hal 0.25 + NGB 1.8/sal, and hal 0.25/NGB 0.5 but not hal 0.25/NGB 1.8 or NGB 1.8 paired/sal had significantly longer descent latencies than the hal 0.25 unpaired/sal (p < 0.05). Further comparisons revealed that the NGB 1.8 paired/sal group was significantly different from hal 0.25 paired/sal (p < 0.05; Fisher’s

LSD) and did not differ significantly from the hal 0.25 unpaired/sal group. Similar to the

Figure 5

Mean (± SEM) descent latency (s) in groups of rats receiving 0.25 mg/kg haloperidol (hal) 60 min, or co-treated with 0.25 mg/kg haloperidol (60 min) and 0.5 or 1.8 mg/kg NGB 2904 (NGB) 30 min prior to each conditioning day (A) and saline (sal) 60 min or 0.5 or 1.8 mg/kg NGB 30 min prior to test session (B). * significantly different from hal 0.25 unpaired/sal in pairwise comparison (Dunnett’s test) following significant group effects in analysis of variance. # significantly different from hal 0.25 paired/sal in pairwise comparison (Fisher’s LSD test) following significant group effects in analysis of variance.

findings for nafadotride in experiment 1, the high dose of NGB 2904 (1.8 mg/kg) given in test following haloperidol sensitization reduced expression but not acquisition of conditioned catalepsy.

3.5 Discussion

The results can be summarized as follows: In experiment 1, all rats treated with haloperidol or haloperidol + nafadotride did not show catalepsy initially, but gradually catalepsy sensitization was observed. Rats previously sensitized with haloperidol or haloperidol plus nafadotride exhibited conditioned catalepsy when tested with saline in the test environment. When nafadotride (0.5 mg/kg) was given on the test day to rats previously sensitized with haloperidol, a significant reduction of the catalepsy response was seen. The same dose of nafadotride given during haloperidol sensitization sessions did not reduce conditioned catalepsy acquisition. Nafadotride treatment in the test environment did not produce catalepsy sensitization, however, when rats were tested with saline a small but significant conditioned catalepsy emerged. As this effect was not seen in the nafadotride ‘unpaired’ rats, our results make an interpretation based on physiological effect of repeated injection (e.g., D3 receptor upregulation) an unlikely mechanism. Instead, this catalepsy is specific to environmental stimuli associated with previous nafadotride treatment. However, this interpretation should be taken with caution as the group conditioned with haloperidol plus this dose of nafadotride did not show an elevated conditioned catalepsy response compared to the group conditioned with haloperidol alone, as might be expected if conditioning with nafadotride led to

conditioned catalepsy. The possible mechanisms contributing to this finding await further study.

In experiment 2, rats treated with haloperidol or haloperidol plus NGB 2904 showed gradual catalepsy sensitization with repeated testing. Rats previously sensitized with haloperidol or haloperidol plus NGB 2904 continued to be cataleptic when tested with saline in the drug environment. When NGB 2904 (1.8 mg/kg) was given on the test day to previously sensitized rats, it significantly attenuated the conditioned catalepsy response. The same dose of NGB 2904 given during haloperidol sensitization did not affect conditioned catalepsy acquisition. The findings in experiment 1 and 2 suggest that a DA D3 receptor-preferring or a D3 receptor-selective antagonist appear to reduce expression of conditioned catalepsy at doses that fail to affect acquisition.

Our findings that haloperidol, a D2 receptor-preferring antagonist, produces catalepsy sensitization is in agreement with previous reports (Lanis and Schmidt, 2001;

Schmidt et al., 1999). Schmidt and colleagues have shown that a daily low dose of haloperidol (0.25 mg/kg) failed to produce catalepsy during the initial test session, however, day-to-day increases in the catalepsy response were observed with repeated haloperidol-environment pairings (Banasikowski and Beninger, 2012a; Klein and

Schmidt, 2003; Riedinger et al., 2011). In the present study, control groups that received environment-saline pairing and also received haloperidol in their homecage (colony room) environment did not exhibit increased catalepsy during the test session.

Furthermore, following catalepsy sensitization, if the same dose of haloperidol is injected

and catalepsy tested in another different environment, animals fail to express catalepsy

(Klein and Schmidt, 2003; Wiecki et al., 2009). Therefore the effect of increased catalepsy in rats having received environment-haloperidol pairing can be attributed to the association of environmental stimuli with haloperidol rather than the previous history of haloperidol treatment (Banasikowski and Beninger, 2010).

The D3 receptor-preferring antagonist nafadotride and the D3 receptor-selective antagonist NGB 2904 failed to induce catalepsy when given alone. Our findings are consistent with previous studies using nafadotride (doses comparable to the current study) and NGB 2904 (Audinot et al., 1998; Martelle et al., 2007; Sautel et al., 1995; Xi and Gardner, 2007). Other D3 receptor antagonists including S 14297 (Millan et al.,

1997), SB 277011 and S 33084 (Gyertyan and Saghy, 2007) also failed to produce catalepsy in rats.

A number of studies have implicated D3 receptors in motor control. Acute treatment with D3 receptor antagonists has been shown to reduce haloperidol catalepsy in rats (Gyertyan and Saghy, 2007; Millan et al., 1997). Also D3 receptor antagonists appear to alleviate a number of parkinsonian symptoms (Mela et al., 2010; Silverdale et al.,

2004) in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 6-hydroxydopamine

(6-OHDA) lesioned animals. Comparison with the present results is difficult to make as the dose of haloperidol used in our study is too low to elicit catalepsy on the first test day.

Only after repeated haloperidol-environment pairings does catalepsy sensitization appear.

Thus, our design makes it difficult to assess the role of acute D3 receptor antagonism in

the reduction of motor impairments.

The finding that nafadotride and NGB 2904 reduced expression of haloperidol conditioned catalepsy at doses that failed to block acquisition is consistent with previous reports examining the role of D3 receptor antagonists in many drug conditioned behaviors (for a review see Beninger and Banasikowski, 2008). For example, in self- administration studies, the D3 receptor antagonists NGB 2904 and SB 277011, decrease reinstating effects of cues previously paired with cocaine (Cervo et al., 2007; Gilbert et al., 2005). SB 277011 or the D3 receptor partial agonist BP 897 (that would also act as a partial antagonist) reduced response suppression in the presence of conditioned stimuli

(Swain et al., 2008). We previously showed that the D3 receptor antagonist ABT 127 blocked expression but not acquisition of cocaine conditioned activity (Banasikowski et al., 2010). SB 277011 blocked expression of cocaine conditioned activity (Le Foll et al.,

2002) and the expression of cocaine (Vorel et al., 2002) and nicotine (Le Foll et al., 2005)

CPP. On the other hand, the effects of D3 receptor antagonists on the acquisition of CPP based on psychostimulants (Cervo et al., 2005; Gyertyan and Gal, 2003; Vorel et al.,

2002) often were non-significant.

Following drug-environment pairings, a change in D3 receptor expression in behaviorally conditioned animals has been reported. For example, rats with a history of cocaine self-administration in a particular environment showed increased D3 receptor binding following a reinstatement test (Neisewander et al., 2004). Le Foll et al. (2002) reported a significant increase in D3 (but not D1 or D2) receptor mRNA and D3

receptors in the ventral striatum in mice tested with saline in an environment previously associated with cocaine. More importantly, the up-regulation of D3 receptors was absent in mice that received saline-environment pairings and cocaine homecage injections.

Similarly, rats repeatedly injected with morphine in a particular context developed environment-dependent locomotor sensitization which was associated with increases in

D3 receptor mRNA in the ventral striatum. Rats treated with morphine in the homecage environment did not develop sensitization and showed no change in D3 receptor mRNA

(Liang et al., 2011). Thus, the observations of increased D3 receptors are dependent on the drug-environment association; perhaps similar mechanism may play a role in conditioned catalepsy.

Based on limited studies in mice, it could be argued that the decrease in conditioned catalepsy expression observed following treatment with nafadotride or NGB

2904 could result from an increase in spontaneous locomotor activity in the animals on the test day. Nafadotride (Sautel et al., 1995) and NGB 2904 (Pritchard et al., 2007) have been reported to increase locomotor activity in extensively habituated mice. However, studies in rats using other D3 receptor antagonists such as SB 277011 (Reavill et al.,

2000) and S 33084 (Millan et al., 2004) failed to report similar effects. Most importantly,

Gyertyan and Saghy (2004) found increased spontaneous activity with SB 277011only in mice but not in rats, suggesting that there may be a species-specific locomotor response to D3 receptor antagonism (Gyertyan and Saghy, 2004). We failed to observe a decrease in catalepsy sensitization in haloperidol groups co-treated with nafadotride or NGB 2904

during conditioning, thus we have no reason to suspect that the effect seen on expression of conditioned catalepsy was simply due to locomotor-activating drug properties. Instead our data suggests that D3 receptors play an important role in mediating the expression of conditioned responses to environmental stimuli associated with haloperidol treatment, and that nafadotride and NGB 2904 attenuates the ability of environmental stimuli to elicit conditioned catalepsy in rats.

DA levels increase initially following haloperidol treatment. Could elevated levels of DA inhibit the effects of D3 receptor antagonists during the catalepsy sensitization phase by competing for D3 receptors (see Xi and Gardner, 2007)? During the test phase with saline, these elevations in DA levels would not occur; could the observed effects of D3 receptor antagonists in the conditioned catalepsy test occur because DA levels are no longer elevated? It is possible that endogenous DA and D3 receptor antagonists may compete for binding to D3 receptors during initial sensitization sessions. However, with repeated haloperidol treatments DA neurons undergo physiological adaptations (i.e., depolarization block) whereby they gradually shift from a hyperactive to a hypoactive state (Boye and Rompre, 2000; Chiodo and Bunney, 1983;

Grace and Bunney, 1986; Grace et al., 1997; Valenti and Grace, 2010). Accordingly, an effect of D3 receptor antagonists on catalepsy sensitization should emerge over trials but no such effect was observed. We conclude that the significant effect of D3 receptor antagonists on the expression of catalepsy conditioning but not on the acquisition of catalepsy sensitization cannot be attributed to differential levels of endogenous dopamine

competing with D3 receptors for DA.

In aggregate, we have shown for the first time that the D3 receptor antagonists nafadotride or NGB 2904 attenuate expression of conditioned catalepsy at doses that fail to affect acquisition. This finding is consistent with previous reports showing that D3 receptor activity is more critical for the expression than acquisition of drug conditioning effects to stimulants (Banasikowski et al., 2010; Banasikowski et al., 2012; Cervo and

Samanin, 1995; Cervo and Samanin, 1996; Wang et al., 2006) and opiates (Liang et al.,

2011). We advance this notion by providing evidence for a role of D3 receptors in haloperidol conditioned catalepsy. Our findings suggest that conditioning to drugs that either increase or decrease dopaminergic neurotransmission may rely on similar mechanisms.

3.6 References

Amtage J, Schmidt WJ (2003). Context-dependent catalepsy intensification is due to classical conditioning and sensitization. Behav Pharmacol, 14(7): 563-567. Audinot V, Newman-Tancredi A, Gobert A, Rivet JM, et al. (1998). A comparative in vitro and in vivo pharmacological characterization of the novel dopamine D3 receptor antagonists (+)-S 14297, nafadotride, GR 103,691 and U 99194. J Pharmacol Exp Ther, 287(1): 187-197. Banasikowski TJ, Beninger RJ (2010). Conditioned drug effects. In: Stolerman, IP (Ed.), Encyclopedia of Psychopharmacology (pp. 325-331) Heidelberg: Springer. Banasikowski TJ, Beninger RJ (2012). Haloperidol conditioned catalepsy in rats: a possible role for D1-like receptors. The International Journal of Neuropsychopharmacology, FirstView: 1-10. Banasikowski TJ, Bespalov A, Drescher K, Behl B, et al. (2010). Double dissociation of the effects of haloperidol and the dopamine D3 receptor antagonist ABT-127 on acquisition vs. expression of cocaine-conditioned activity in rats. J Pharmacol Exp Ther, 335(2): 506-515. Banasikowski TJ, MacLeod LS, Beninger RJ (2012). Comparison of nafadotride, CNQX, and haloperidol on acquisition versus expression of amphetamine- conditioned place preference in rats. Behav Pharmacol, 23(1): 89-97. Bezard E, Ferry S, Mach U, Stark H, et al. (2003). Attenuation of levodopa-induced dyskinesia by normalizing dopamine D3 receptor function. Nat Med, 9(6): 762- 767. Boileau I, Guttman M, Rusjan P, Adams JR, et al. (2009). Decreased binding of the D3 dopamine receptor-preferring ligand [11C]-(+)-PHNO in drug-naive Parkinson's disease. Brain, 132(Pt 5): 1366-1375. Bordet R, Ridray S, Carboni S, Diaz J, et al. (1997). Induction of dopamine D3 receptor expression as a mechanism of behavioral sensitization to levodopa. Proc Natl Acad Sci U S A, 94(7): 3363-3367. Boye SM, Rompre PP (2000). Behavioral evidence of depolarization block of dopamine neurons after chronic treatment with haloperidol and clozapine. J Neurosci, 20(3): 1229-1239. Cervo L, Burbassi S, Colovic M, Caccia S (2005). Selective antagonist at D3 receptors, but not non-selective partial agonists, influences the expression of cocaine- 88

induced conditioned place preference in free-feeding rats. Pharmacol Biochem Behav, 82(4): 727-734. Cervo L, Cocco A, Petrella C, Heidbreder CA (2007). Selective antagonism at dopamine D3 receptors attenuates cocaine-seeking behaviour in the rat. Int J Neuropsychopharmacol, 10(2): 167-181. Cervo L, Samanin R (1995). Effects of dopaminergic and glutamatergic receptor antagonists on the acquisition and expression of cocaine conditioning place preference. Brain Res, 673(2): 242-250. Cervo L, Samanin R (1996). Effects of dopaminergic and glutamatergic receptor antagonists on the establishment and expression of conditioned locomotion to cocaine in rats. Brain Res, 731(1-2): 31-38. Chiang Y-C, Chen P-C, Chen J-C (2003). D3 dopamine receptors are down-regulated in amphetamine sensitized rats and their putative antagonists modulate the locomotor sensitization to amphetamine. Brain Research, 972(1-2): 159. Chiodo LA, Bunney BS (1983). Typical and atypical neuroleptics: differential effects of chronic administration on the activity of A9 and A10 midbrain dopaminergic neurons. J Neurosci, 3(8): 1607-1619. Gilbert JG, Newman AH, Gardner EL, Ashby CR, Jr., et al. (2005). Acute administration of SB-277011A, NGB 2904, or BP 897 inhibits cocaine cue- induced reinstatement of drug-seeking behavior in rats: role of dopamine D3 receptors. Synapse, 57(1): 17-28. Grace AA, Bunney BS (1986). Induction of depolarization block in midbrain dopamine neurons by repeated administration of haloperidol: analysis using in vivo intracellular recording. J Pharmacol Exp Ther, 238(3): 1092-1100. Grace AA, Bunney BS, Moore H, Todd CL (1997). Dopamine-cell depolarization block as a model for the therapeutic actions of antipsychotic drugs. Trends Neurosci, 20(1): 31-37. Guillin O, Diaz J, Carroll P, Griffon N, et al. (2001). BDNF controls dopamine D3 receptor expression and triggers behavioural sensitization. Nature, 411(6833): 86- 89. Gurevich EV, Bordelon Y, Shapiro RM, Arnold SE, et al. (1997). Mesolimbic dopamine D3 receptors and use of antipsychotics in patients with schizophrenia. A postmortem study. Arch Gen Psychiatry, 54(3): 225-232.

Gyertyan I, Gal K (2003). Dopamine D3 receptor ligands show place conditioning effect but do not influence cocaine-induced place preference. Neuroreport, 14(1): 93-98. Gyertyan I, Saghy K (2004). Effects of dopamine D3 receptor antagonists on spontaneous and agonist-reduced motor activity in NMRI mice and Wistar rats: comparative study with nafadotride, U 99194A and SB 277011. Behav Pharmacol, 15(4): 253-262. Gyertyan I, Saghy K (2007). The selective dopamine D3 receptor antagonists, SB 277011-A and S 33084 block haloperidol-induced catalepsy in rats. Eur J Pharmacol, 572(2-3): 171-174. Klein A, Schmidt WJ (2003). Catalepsy intensifies context-dependently irrespective of whether it is induced by intermittent or chronic dopamine deficiency. Behav Pharmacol, 14(1): 49-53. Lanis A, Schmidt WJ (2001). NMDA receptor antagonists do not block the development of sensitization of catalepsy, but make its expression state- dependent. Behav Pharmacol, 12(2): 143-149. Le Foll B, Diaz J, Sokoloff P (2003). Increased dopamine D3 receptor expression accompanying behavioral sensitization to nicotine in rats. Synapse, 47(3): 176- 183. Le Foll B, Frances H, Diaz J, Schwartz JC, et al. (2002). Role of the dopamine D3 receptor in reactivity to cocaine-associated cues in mice. Eur J Neurosci, 15(12): 2016-2026. Le Foll B, Sokoloff P, Stark H, Goldberg SR (2005). Dopamine D3 receptor ligands block nicotine-induced conditioned place preferences through a mechanism that does not involve discriminative-stimulus or antidepressant-like effects. Neuropsychopharmacology, 30(4): 720-730. Liang J, Zheng X, Chen J, Li Y, et al. (2011). Roles of BDNF, dopamine D3 receptors, and their interactions in the expression of morphine-induced context-specific locomotor sensitization. European Neuropsychopharmacology, In Press, Corrected Proof. Martelle JL, Claytor R, Ross JT, Reboussin BA, et al. (2007). Effects of two novel D3-selective compounds, NGB 2904 [N-(4-(4-(2,3-dichlorophenyl)piperazin-1- yl)butyl)-9H-fluorene-2-carboxami de] and CJB 090 [N-(4-(4-(2,3- dichlorophenyl)piperazin-1-yl)butyl)-4-(pyridin-2-yl)benzami de], on the reinforcing and discriminative stimulus effects of cocaine in rhesus monkeys. J Pharmacol Exp Ther, 321(2): 573-582. 90

Mela F, Millan MJ, Brocco M, Morari M (2010). The selective D(3) receptor antagonist, S33084, improves parkinsonian-like motor dysfunction but does not affect L-DOPA-induced dyskinesia in 6-hydroxydopamine hemi-lesioned rats. Neuropharmacology, 58(2): 528-536. Millan MJ, Gressier H, Brocco M (1997). The dopamine D3 receptor antagonist, (+)-S 14297, blocks the cataleptic properties of haloperidol in rats. Eur J Pharmacol, 321(3): R7-9. Millan MJ, Seguin L, Gobert A, Cussac D, et al. (2004). The role of dopamine D3 compared with D2 receptors in the control of locomotor activity: a combined behavioural and neurochemical analysis with novel, selective antagonists in rats. Psychopharmacology (Berl), 174(3): 341-357. Mizrahi R, Agid O, Borlido C, Suridjan I, et al. (2011). Effects of antipsychotics on D3 receptors: a clinical PET study in first episode antipsychotic naive patients with schizophrenia using [11C]-(+)-PHNO. Schizophr Res, 131(1-3): 63-68. Neisewander JL, Fuchs RA, Tran-Nguyen LT, Weber SM, et al. (2004). Increases in dopamine D3 receptor binding in rats receiving a cocaine challenge at various time points after cocaine self-administration: implications for cocaine-seeking behavior. Neuropsychopharmacology, 29(8): 1479-1487. Pak AC, Ashby CR, Jr., Heidbreder CA, Pilla M, et al. (2006). The selective dopamine D3 receptor antagonist SB-277011A reduces nicotine-enhanced brain reward and nicotine-paired environmental cue functions. Int J Neuropsychopharmacol, 9(5): 585-602. Pritchard LM, Newman AH, McNamara RK, Logue AD, et al. (2007). The dopamine D3 receptor antagonist NGB 2904 increases spontaneous and amphetamine- stimulated locomotion. Pharmacol Biochem Behav, 86(4): 718-726. Reavill C, Taylor SG, Wood MD, Ashmeade T, et al. (2000). Pharmacological actions of a novel, high-affinity, and selective human dopamine D(3) receptor antagonist, SB-277011-A. J Pharmacol Exp Ther, 294(3): 1154-1165. Riedinger K, Kulak A, Schmidt WJ, von Ameln-Mayerhofer A (2011). The role of NMDA and AMPA/Kainate receptors in the consolidation of catalepsy sensitization. Behav Brain Res, 218(1): 194-199. Sautel F, Griffon N, Sokoloff P, Schwartz JC, et al. (1995). Nafadotride, a potent preferential dopamine D3 receptor antagonist, activates locomotion in rodents. J Pharmacol Exp Ther, 275(3): 1239-1246.

Schmidt WJ, Beninger RJ (2006). Behavioural sensitization in addiction, schizophrenia, Parkinson's disease and dyskinesia. Neurotox Res, 10(2): 161-166. Schmidt WJ, Tzschentke TM, Kretschmer BD (1999). State-dependent blockade of haloperidol-induced sensitization of catalepsy by MK-801. Eur J Neurosci, 11(9): 3365-3368. Segal DM, Moraes CT, Mash DC (1997). Up-regulation of D3 dopamine receptor mRNA in the nucleus accumbens of human cocaine fatalities. Brain Res Mol Brain Res, 45(2): 335-339. Silverdale MA, Nicholson SL, Ravenscroft P, Crossman AR, et al. (2004). Selective blockade of D(3) dopamine receptors enhances the anti-parkinsonian properties of ropinirole and levodopa in the MPTP-lesioned primate. Exp Neurol, 188(1): 128- 138. Staley JK, Mash DC (1996). Adaptive increase in D3 dopamine receptors in the brain reward circuits of human cocaine fatalities. J Neurosci, 16(19): 6100-6106. Swain SN, Beuk J, Heidbreder CA, Beninger RJ (2008). Role of dopamine D3 receptors in the expression of conditioned fear in rats. Eur J Pharmacol, 579(1-3): 167-176. Valenti O, Grace AA (2010). Antipsychotic drug-induced increases in ventral tegmental area dopamine neuron population activity via activation of the nucleus accumbens-ventral pallidum pathway. Int J Neuropsychopharmacol, 13(7): 845- 860. Vorel SR, Ashby CR, Jr., Paul M, Liu X, et al. (2002). Dopamine D3 receptor antagonism inhibits cocaine-seeking and cocaine-enhanced brain reward in rats. J Neurosci, 22(21): 9595-9603. Wang Z, Kai L, Day M, Ronesi J, et al. (2006). Dopaminergic control of corticostriatal long-term synaptic depression in medium spiny neurons is mediated by cholinergic interneurons. Neuron, 50(3): 443-452. Wiecki TV, Riedinger K, von Ameln-Mayerhofer A, Schmidt WJ, et al. (2009). A neurocomputational account of catalepsy sensitization induced by D2 receptor blockade in rats: context dependency, extinction, and renewal. Psychopharmacology (Berl), 204(2): 265-277. Xi ZX, Gardner EL (2007). Pharmacological actions of NGB 2904, a selective dopamine D3 receptor antagonist, in animal models of drug addiction. CNS Drug Rev, 13(2): 240-259.

Chapter 4

Drug-Environment Interaction Controls the Acquisition and Expression

of Haloperidol Catalepsy Sensitization and Conditioning

4.1 Abstract

Dopamine (DA) plays a critical role in our ability to move and in some forms of learning. DA drugs used to treat Parkinson’s disease and schizophrenia produce progressively greater effects on behavior with repeated exposures (i.e., behavioral sensitization). Low doses of the DA D2-receptor preferring antagonist haloperidol (0.25 mg/kg, i.p.), repeatedly injected in a particular environment, produce day-to-day increases in catalepsy, quantified by time spent on a horizontal bar without active movement. In experiment 1, we observed that expression of catalepsy sensitization is environment-specific, as it is not observed when testing is performed in different environments despite usual haloperidol treatment. Furthermore, places associated with repeated haloperidol injections and catalepsy tests also elicit catalepsy, now conditioned.

This was observed when saline was substituted for haloperidol and rats were tested in their regular, but not in a novel test environment. Following repeated saline-environment pairings, conditioned catalepsy was extinguished. Post-extinction challenge with haloperidol produced a small catalepsy response that was now independent of the test environment, suggesting that the haloperidol drug cue alone can elicit conditioned catalepsy. In experiment 2, we show that expression of the sensitization response is dependent on a drug-environment interaction, as animals with a previous history of drug treatment in a different environment failed to exhibit significant catalepsy when given haloperidol in the test environment. In experiment 3, we show for the first time that habituation to the test environment prior to commencing haloperidol treatment

significantly enhanced the rate of catalepsy sensitization. This was not observed when habituation was performed in a different environment. Thus, acquisition and expression of catalepsy sensitization to haloperidol is conditional on the interaction between the drug and test environment. Results suggest that the therapeutic effects of antipsychotic drugs depend on the environment associated with their use.

4.2 Introduction

The behavioral effects of drugs that act on the brain’s DA system change with repeated exposure to the drug. This may be particularly important in understanding the response to treatment of psychosis, a clinical hallmark of schizophrenia (Abi-Dargham et al., 2000; Abi-Dargham et al., 2009; Grace, 2000; Kegeles et al., 2010; Lodge and Grace,

2011; Wadenberg et al., 2001). The time course of the therapeutic effects of antipsychotics is variable; some people with schizophrenia exhibit a delayed onset of action, where psychosis may persist sometimes for weeks after the initiation of treatment

(Emsley et al., 2006; Grace et al., 1997; Miller, 1987).

Psychosis is believed to result from a sensitization-like learning processes in which DA system dysregulation (Carlsson and Lindqvist, 1963; Kapur and Remington,

2001; Seeman et al., 1976; Wadenberg et al., 2001) provides a neurochemical milieu for acquisition of psychotic symptoms (Featherstone et al., 2007; Lodge and Grace, 2008;

Lodge and Grace, 2012; Peleg-Raibstein et al., 2008). Accordingly, enhanced striatal DA release to acute amphetamine treatment observed in people with schizophrenia (Kegeles

et al., 2010; Laruelle, 2000; Laruelle et al., 1996) resembles that of healthy volunteers with a history of repeated amphetamine treatments (Boileau et al., 2006).

Psychomotor stimulant drugs like amphetamine produce progressively greater effects on behavior with repeated exposures [i.e., behavioral sensitization; (Post and

Rose, 1976; Segal and Mandell, 1974)]. The enhanced behavioral output as well as the enhanced striatal DA release with each repeated drug treatment (Badiani et al., 1998;

Paulson and Robinson, 1995) is context-dependent, i.e., its expression is strongest in the presence of previously drug-paired environmental stimuli (Anagnostaras and Robinson,

1996; Badiani et al., 1998; Pert et al., 1990). Similarly, antipsychotic drugs, that block

DA receptors, produce progressively greater effects on behavior with repeated testing

(Carey, 1987; Li et al., 2007; Schmidt and Beninger, 2006).

The present experiments tested the interaction between the drug and test environment using the haloperidol catalepsy sensitization model (Amtage and Schmidt,

2003; Banasikowski and Beninger, 2012a; Banasikowski and Beninger, 2012b; Klein and

Schmidt, 2003; Lanis and Schmidt, 2001; Riedinger et al., 2011; Schmidt et al., 1999).

Unlike animals treated with suprathreshold doses of haloperidol (> 1.0 mg/kg), rats given a low dose (≤ 0.5 mg/kg) do not exhibit catalepsy during initial drug-environment pairings. After repeated drug-environment pairings animals show gradual day-to-day increases in the catalepsy response, quantified by the time a rat remains with its forepaws resting on a suspended horizontal bar (Banasikowski and Beninger, 2012a). Moreover, when the animals are tested with saline instead of haloperidol they continue to exhibit 96

catalepsy in the drug-paired environment (Amtage and Schmidt, 2003; Banasikowski and

Beninger, 2012a; Banasikowski and Beninger, 2012b). It appears that the sensitization effect is due to a progressive loss of exploratory behaviors as a consequence of decreased engagement with environmental stimuli, which eventually manifests as catalepsy.

In comparison to psychomotorstimulant drugs that facilitate and enhance DA transmission, sensitization and conditioning induced by treatments that reduce the brain’s

DA activity have not been extensively studied. Environmental stimuli appear to control antipsychotic-produced catalepsy sensitization from the observation that sensitized responding is lost when rats are injected with haloperidol and tested in a different environment (Klein and Schmidt, 2003; Lynch and Carey, 1987; Wiecki et al., 2009). In experiments 1and 2, we carried out a systematic test of the hypothesis that environmental stimuli control the expression of catalepsy sensitization and conditioning. Experiment 1 examined the contribution of environmental stimuli (i.e., exteroceptive cues) to expression of sensitization and conditioned catalepsy, the extinction of conditioned catalepsy responding, and how haloperidol (interoceptive) drug cues (Carey and Gui,

1998) may contribute to the conditioned catalepsy response. Experiment 2 examined the independent roles of test environment history and drug history in expression of catalepsy sensitization.

To further understand the relationship between catalepsy sensitization and environmental influences, experiment 3 tested the hypothesis that pre-exposure

(habituation) to the ‘to-be’ test environment will accelerate the rate of catalepsy 97

sensitization compared to pre-exposure to in a different environment. This hypothesis was based on the observation that novel stimuli activate DA systems (Ahn and Phillips,

1999; Bardo et al., 1990; Rebec et al., 1997) and that experience with these stimuli reduces DA activation (Ahn and Phillips, 1999). If catalepsy sensitization results from repeated association of environmental stimuli with attenuated DA neurotransmission, then environmental stimuli producing less DA release should augment the rate of catalepsy sensitization.

4.3 Materials and Methods

4.3.1 Subjects

Experimentally naïve male albino Wistar rats (N = 89) weighing 200-225 g upon arrival from Charles River Canada (St. Constant, QC) were housed in pairs or threes in clear Plexiglas cages (45.0 x 25.0 x 22.0 cm). Average temperature in the colony was 21º

C, humidity 70% with reversed light–dark cycle (lights off from 0700 to 1900 hr). Rats were maintained with food (LabDiet 5001, PMI Nutrition International, Brentwood, MO,

4.3.2 Drugs

Haloperidol, (4-[4-(4-chlorophenyl)-4-hydroxy-1-piperidyl]-1-(4-fluorophenyl)- butan-1-one (Sigma, St. Louis, MO, USA) was prepared in a 0.3% distilled-water

solution of tartaric acid. Injections were administered intraperitoneally (i.p.) in a volume of 1.0 ml/kg.

4.3.3 Behavioral Testing

Rats were randomly assigned to treatment (paired) and control groups (unpaired or saline). The catalepsy sensitization procedure consisted of injecting rats (i.p.) with haloperidol or saline outside the testing room and putting them back into their cages.

Sixty minutes later, they were tested on a horizontal bar [1.6 cm diam. threaded or smooth rod with end bolts attached to Plexiglas supports, 10 cm above the surface (see below)], by gently placing both forepaws on the bar. Descent latency was measured as the time span from placing the animal on the bar until the first active paw movement. A cut-off time of 180 s was used, i.e., the trial was terminated when the animal did not make an active paw movement within that time.

4.3.4 Experimental Design

All catalepsy test sessions occurred between 0900 and 1700, during the dark phase of the light-dark cycle. The test apparatus consisted of two distinct (one with three black sides and one with three white sides) rectangular Plexiglas catalepsy chambers (32 x 42 x 30 cm). Each chamber was equipped with a distinct horizontal bar. The black chamber was equipped with a 1.6 cm diam. smooth rod bolted to clear Plexiglas supports

(1 cm thick, 10 cm high), and the white chamber was equipped with a 1.6 cm diam. threaded rod bolted to black Plexiglas supports (0.5 cm thick and 10 cm high). Surgical

paper was used as the floor covering in both chambers. Rats were randomly assigned to one of the two test chambers. Each rat was always placed in the same chamber during the haloperidol sensitization phase and all treatment groups included rats sensitized in each chamber. We observed no relationship between the magnitude of the catalepsy effect and the testing chambers, and no effect of testing chambers on catalepsy sensitization or conditioning.

Experiment 1: Environment-Specific Expression of Haloperidol Catalepsy

Sensitization and Conditioning.

Four groups of rats were used to examine the acquisition of haloperidol (0.25 mg/kg) catalepsy sensitization and its subsequent expression in a familiar or novel test environment. The 0.25 mg/kg dose was used in previous conditioned catalepsy studies by

Schmidt et al. (1999) and by us (Banasikowski and Beninger, 2012a; Banasikowski and

Beninger, 2012b). The general protocol consisted of two phases: a series of sensitization sessions in one test environment (Context A), followed by a drug challenge session in

Context A or an unfamiliar test environment (Context B).

Animals in the ‘paired’ condition were injected with haloperidol outside the testing room (Context A) for 10 consecutive days, 60 min before the catalepsy test. Two hrs after the catalepsy test, rats were administered saline in their colony room (homecage injection). These were the Hal 0.25 / Context A and Hal 0.25 / Context B groups (ns = 9).

The term after the slash indicates the context on the test day. The ‘unpaired’ groups received saline 60 min before and haloperidol 2 hr after the catalepsy test. These were the 100

Unpaired Hal 0.25 / Context A (n = 9) and Unpaired Hal 0.25 / Context B group (n = 10).

This design allowed for all animals to experience the same drug history but in different environments and at different times. On day 11 animals in groups Hal 0.25 / Context B and Unpaired Hal 0.25 / Context B were injected with haloperidol or saline, respectively, in an unfamiliar hallway and tested in an unfamiliar catalepsy chamber (black vs. white) located in an unfamiliar testing room (i.e., Context B). On the other hand, the Hal 0.25 /

Context A and Unpaired Hal 0.25 / Context A groups were injected with haloperidol or saline, respectively, and tested in their usual test environment (Context A). No homecage injections were given during this test phase. After day 11 rats in groups Hal 0.25 /

Context B and Unpaired Hal 0.25 / Context B were no longer tested.

In the next experimental phase, days 12-13, we examined the role of environmental conditioning on catalepsy responses. On day 12, five animals from the Hal

0.25 / Context A and five animals from the Unpaired Hal 0.25 / Context A groups were injected with saline 60 min before and tested for catalepsy in their usual test environment

(Context A). The remaining animals were injected with saline and tested for catalepsy in an unfamiliar testing environment (Context B). The next day, another saline test was administered, with the test environments reversed. Thus, over the two test days all animals were tested for conditioned catalepsy in an environment previously paired with haloperidol and in an unfamiliar environment.

During the extinction procedure, days 14-17, rats were administered saline 60 min before the catalepsy test in Context A. No homecage injections were given during this

101

phase. On day 18, five animals from Hal 0.25 / Context A and five animals from the

Unpaired Hal 0.25 / Context A groups were injected with haloperidol 60 min before and tested for catalepsy in context A. The remaining animals were injected with haloperidol and tested for catalepsy in context B. On day 19, another haloperidol test was administered, with the test environments from day 18 reversed. Thus, all animals were tested for catalepsy in an environment previously paired with haloperidol (Context A) and an environment never paired with haloperidol (Context B).

Experiment 2: Associative or Non-Associative Processes Involved in Expression of

Haloperidol Catalepsy Sensitization.

We tested the hypothesis that repeated treatment with haloperidol outside of the test environment (unpaired group) would produce an augmented catalepsy response compared to saline control animals when haloperidol is given to both groups for the first time 60 min prior to placement in the test environment. The general procedure consisted of two phases: sensitization (15 days) and test (1 day). On the test day (day 16), all rats were challenged with haloperidol (0.25 mg/kg) 60 min before the catalepsy test.

The Hal 0.25 / Hal 0.25 group (n = 10) was treated with haloperidol (0.25 mg/kg)

60 min before the catalepsy test for 15 consecutive days. Two hrs after the catalepsy test, these rats were administered saline in their colony room (homecage injection). The term after the slash indicates the treatment on the test day. The unpaired Hal 0.25 / Hal 0.25 group (n = 10) was treated with saline 60 min before catalepsy test for 15 days, and was given homecage injections of haloperidol (0.25 mg/kg) 2 hrs later. The Saline / Hal 0.25

102

group (n = 9) was treated with saline 60 min before catalepsy test, and again with saline 2 hrs after the test.

Experiment 3: Environment Pre-Exposure and Acquisition of Future Haloperidol

Catalepsy Sensitization.

Two groups of rats (N = 24) examined the effect of test environment pre-exposure on the subsequent acquisition of haloperidol catalepsy sensitization. The general protocol consisted of two phases: pre-exposure (6 days) and sensitization (13 days). During the pre-exposure phase rats in the Pre-exposure context A / Hal 0.25 group (n = 12) were injected with saline and placed in the ‘to-be’ test environment (Context A) for 5 min/day for six consecutive days (30 min total). The Pre-exposure context B / Hal 0.25 group (n

=12) was injected with saline and placed in an alternate test environment (Context B) for

5 min/day for six consecutive days (30 min total). Context B was physically different from context A as it consisted of a dark-lit testing room with different testing apparatus

(black vs. white catalepsy chambers) and was located in a different part of the building.

On day 7 all rats began the sensitization phase, and were injected with haloperidol outside the testing room (Context A) for 13 consecutive days, 60 min before the catalepsy test.

4.3.5 Statistics

Statistical analyses were performed using IBM SPSS 19.0. All data are presented as means ± standard error. Data were submitted to both parametric and nonparametric tests and each type of analysis yielded similar results. Only the parametric tests are 103

reported. They involved two-way and one-way analyses of variance (ANOVA).

Hypotheses tests were completed using α = 0.05 and pair-wise comparisons were made using Dunnett’s test and where specified, Tukey’s HSD test.

4.4 Results

4.4.1 Experiment 1: Environment-Specific Expression of Haloperidol Catalepsy

Sensitization and Conditioning.

Across conditioning days 1 to 10 the groups receiving haloperidol prior to testing showed increasing catalepsy over days (Fig. 6); catalepsy was not observed in the

‘unpaired’ groups. A two-way ANOVA revealed a significant main effect of day [F(9,

297) = 9.00, p < 0.001], treatment [F(3,33) = 13.75, p < 0.001] and interaction [F(27,297

= 3.16, p < 0.001]. For the main effect of treatment, post-hoc comparison found that the drug-paired Hal 0.25 / Context A and Hal 0.25 / Context B groups spent significantly more time with both paws on the bar (descent latency) than the two unpaired control groups (p < 0.001; Tukey’s HSD).

On day 11 (the context challenge test day), animals in the Hal 0.25 / Context A group continued to exhibit catalepsy in the test environment; on the other hand, rats previously sensitized in context A but tested in a novel test environment (Hal 0.25 /

Context B) did not. This observation was statistically supported with an ANOVA

[F(3,33) = 25.95, p < 0.001]. Post-hoc comparisons revealed a significant difference

104

Figure 6

Environment-specific expression of haloperidol catalepsy sensitization and conditioning. On days 1-10 groups of rats received either 0.25 mg/kg haloperidol (Hal; paired) or saline (sal; unpaired) 60 min prior to each conditioning day. Two hrs after catalepsy test, rats in the paired groups were administered saline, and rats in the unpaired groups were given haloperidol (0.25 mg/kg) in their colony room. On day 11, rats in the paired groups were given haloperidol and rats in the unpaired groups were given saline 60 min before and tested in either the familiar environment (Context A) or a novel environment (Context B). No colony room injections were given from this point on in the experiment and only the Hal 0.25 / Context A and Unpaired Hal 0.25 / Context A groups continued to be tested. On day 12, half of the animals in the paired and unpaired groups were injected with saline 60 min before and tested for catalepsy in Context A. The remaining animals were injected with saline and tested in Context B. On day 13, another saline test was administered, with the test environments from day 12 reversed. On days 14-17, rats were administered saline before the catalepsy test in Context A. On day 18, half of the animals from the paired and unpaired groups were injected with haloperidol 60 min before and tested for catalepsy in Context A. The remaining animals were injected with haloperidol and tested for catalepsy in Context B. On day 19, another haloperidol test was given, with the test environments from day 18 reversed. All data are presented as Mean (± SEM). ## (p < 0.01) significantly different from Unpaired Hal 0.25 / Context A or Unpaired Hal 0.25 / Context B in pairwise comparison (Tukey’s HSD test) following significant group effects in analysis of variance (ANOVA); *** (p < 0.001) significantly different from Hal 0.25 / Context B, Unpaired Hal 0.25 / Context A or Unpaired Hal 0.25 / Context B in pairwise comparison (Dunnett’s Test) following significant group effects in ANOVA; ** (p < 0.01), * (p < 0.05) significantly different from Unpaired Hal 0.25 / Context A.

105

between Hal 0.25 / Context A group and the other three groups (p < 0.001). The expression of catalepsy sensitization was dependent on the presence of stimuli previously associated with haloperidol treatment.

Two groups, Hal 0.25 / Context A and Unpaired Hal 0.25 / Context A were subsequently tested for environmental conditioning effects in the regular test environment

(Context A) and in a novel test environment (Context B). When given saline, the Hal 0.25

/ Context A rats exhibited conditioned catalepsy, i.e., significantly longer descent latencies in the test environment associated with haloperidol (Context A), but not in an unfamiliar environment (Context B), i.e., conditioned catalepsy. This observation was supported by a two-way ANOVA which revealed a significant main effect of context

[F(1, 16) = 9.43, p < 0.01], treatment [F(1,16) = 22.49, p < 0.001] and interaction [F(1,16

= 6.76, p < 0.05].

Animals continued to receive saline in the test environment (Context A) on days

14-17. ANOVA revealed a significant main effect of day [F(3,48) = 4.52, p < 0.01], main effect of treatment [F(1, 16) = 13.03, p < 0.01] and interaction [F(3, 48) = 6.96, p < 0.01].

To further analyze the interaction, simple effects tests of group were performed on each extinction day. Significant group differences between the Hal 0.25 / Context A and

Unpaired Hal 0.25 / Context A groups were observed on extinction day 1 [F(1,17) =

15.55, p < 0.01] and extinction day 2 [F(1,17) = 10.23, p < 0.01], but not days 3 and 4.

When challenged with haloperidol on days 18 and 19, rats in the Hal 0.25 /

Context A group exhibited significant catalepsy in the regular test environment (Context

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A) as well as in Context B. This observation was supported by a two-way ANOVA that revealed a significant main effect of group [F(1,16) = 5.99, p < 0.05]. The above findings suggest that following extinction in Context A, the sensitized catalepsy response induced by haloperidol has shifted from being environment-specific to environment-independent, but only in the previously sensitized animals.

4.4.2 Experiment 2: Associative or Non-Associative Processes Involved in

Expression of Haloperidol Catalepsy Sensitization

Across sensitization days 1 to 15, a two-way ANOVA revealed a significant main effect of day [F(14, 364) = 2.68, p < 0.001], treatment [F(2,26) = 14.95, p < 0.001] and interaction [F(28,364) = 2.03, p < 0.05; Fig. 7A]. Post-hoc comparisons for the main effect of treatment found that rats given haloperidol prior to the catalepsy test (Paired Hal

0.25 / Hal 0.25) had significantly longer descent latencies than the Unpaired Hal 0.25 /

Hal 0.25 or the Saline / Hal 0.25 groups (p < 0.001). Simple effects analysis revealed a day effect in the Paired Hal 0.25 / Hal 0.25 group [F(14,126) = 2.36, p < 0.05], but not the other two groups.

On the test day, all rats were challenged with 0.25 mg/kg of haloperidol and tested for catalepsy. ANOVA revealed a significant difference among treatment conditions

[F(2,26) = 8.637, p < 0.01; Fig. 7B]. Post-hoc comparison found that the Paired Hal 0.25

/ Hal 0.25 group exhibited significantly longer descent latencies than the Unpaired Hal

0.25 / Hal 0.25 or Saline / Hal 0.25 groups (p < 0.01). When the previously unpaired

(Unpaired Hal 0.25 / Hal 0.25) and saline (Saline / Hal 0.25 groups) treated animals were

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injected with haloperidol for the first time prior to catalepsy testing in the test environment we found no significant difference in their catalepsy scores (p > 0.05;

Tukey’s HSD or student t-test). Expression of the catalepsy response was conditional on the presence of environmental stimuli previously associated with the haloperidol treatment.

4.4.3 Experiment 3: Environment Pre-Exposure and Acquisition of Future

Haloperidol Catalepsy Sensitization

Following the six-session pre-exposure phase in either the ‘to-be’ test environment (Context A), or the alternative environment (Context B), all rats tested with haloperidol showed an increased day-to-day catalepsy response (Fig. 8). However, rats previously exposed to the ‘to-be’ test environment exhibited a steeper catalepsy sensitization curve compared to the Pre-exposure context B / Hal 0.25 group. This observation was supported statistically with a two-way ANOVA that revealed a significant main effect of day [F(12, 264) = 14.73, p < 0.001], treatment [F(1,22) = 5.36, p < 0.05] and interaction [F(12,264) = 2.23, p < 0.05]. Thus, test environment pre- exposures can modify future effects of repeated haloperidol treatment.

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Figure 7

Associative or non-associative processes involved in expression of haloperidol catalepsy sensitization. (A) Mean (± SEM) descent latency (s) in groups of rats receiving either saline (Saline / Hal 0.25, Unpaired Hal 0.25 / Hal 0.25) or 0.25 mg/kg haloperidol (Paired Hal 0.25 / Hal 0.25) 60 min prior to each conditioning day. Two hrs after the catalepsy test, rats in the paired (Paired Hal 0.25 / Hal 0.25) and saline (Saline / Hal 0.25) groups were administered saline, and rats in the unpaired group (Unpaired Hal 0.25 / Hal 0.25) were given haloperidol (0.25 mg/kg) in their colony room. (B) On the test day (day 16) all animals were administered 0.25 mg/kg haloperidol (Hal) 60 min prior to the test session. *** (p<0.001), ** (p< 0.01) significantly different from Unpaired Hal 0.25 / Hal 0.25 and Saline / Hal 0.25 in pairwise comparison (Dunnett’s test) following significant group effects in analysis of variance.

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Figure 8 Environment pre-exposure and acquisition of future haloperidol catalepsy sensitization. Mean (± SEM) descent latency (s) in groups of rats previously pre-exposed to either the test environment (Context A) or alternative environment (Context B) receiving 0.25 mg/kg haloperidol (Hal) 60 min prior to testing. * significantly different from pre-exposure Context B / Hal 0.25 by analysis of variance.

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4.5 Discussion

Rats treated with a low dose of haloperidol and tested for catalepsy in a specific environment showed no initial catalepsy but the cataleptic response increased from day- to-day across subsequent test days. This catalepsy sensitization is a behavioral conditioning phenomenon. Thus, (i) paired but not unpaired groups showed catalepsy sensitization. This was seen in experiments 1, 2 and 3 and is in agreement with a number of previous findings (Amtage and Schmidt, 2003; Banasikowski and Beninger, 2012a;

Banasikowski and Beninger, 2012b) (ii) Development of catalepsy sensitization in one environment did not transfer to another environment, as shown on day 11 in experiment

1. This finding is in agreement with Klein and Schmidt (2003). In addition, rats with a history of haloperidol treatments outside of the test environment (unpaired group) did not exhibit significant catalepsy when given haloperidol for the first time prior to catalepsy testing, as shown on day 16 in experiment 2. (iii) Conditioned catalepsy was observed in the drug-paired environment but not in a different environment. This was seen on days 12 and 13 in experiment 1and has been reported previously by us (Banasikowski and

Beninger, 2012a; Banasikowski and Beninger, 2012b) (iv) Conditioned catalepsy showed gradual extinction over days 14-17 of experiment 1. Similar extinction of conditioned catalepsy has been reported by Amtage and Schmidt (2003). These results provide compelling evidence that catalepsy sensitization is a conditioning phenomenon.

Haloperidol produced a small but significant catalepsy effect in both Context A and B on test days 18 and 19 in experiment 1. This effect was observed in Context A

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after conditioned catalepsy had been extinguished in Context A and might be taken as evidence that a 0.25 mg/kg dose of haloperidol was producing an unconditioned catalepsy effect. Yet this dose of haloperidol did not produce a significant catalepsy effect in Context B on day 11 in experiment 1, nor did it produce significant catalepsy on day 1 of testing in experiment 1, 2 or 3, arguing against an unconditioned catalepsy effect. One possibility is that the small but significant catalepsy effect of haloperidol in

Context A on days 18 and 19 was due to cue properties of haloperidol that had not been extinguished. Haloperidol has been shown to serve as a cue in drug discrimination experiments (Cohen et al., 1997; Gauvin et al., 1994; Goudie and Smith, 1999; McElroy et al., 1989). Psychomotor stimulants also possess strong discriminative stimulus properties (Colpaert et al., 1978; Wise et al., 2008) and similarly have been observed to retain enhanced stimulant effects after extinction (Carey and Gui, 1998; Stewart and

Vezina, 1991).

A remaining question is: why does 0.25 mg/kg of haloperidol produce a small but significant catalepsy effect in Context B on days 18 and 19 in experiment 1 after extinction training in Context A when the same dose of haloperidol produced no significant catalepsy effect in Context B on day 11? The answer may lie in the effect of extinction training in Context A over days 14 to 17. Prior to extinction training, the cues of Context A plus the putative cue properties of haloperidol may have controlled conditioned catalepsy. After extinction of the exteroceptive Context A cues, the only remaining (putative) cue controlling catalepsy would be the interoceptive drug cue of

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haloperidol itself as it had not been extinguished. Since the cues of Context A had been extinguished by the time haloperidol was again tested on days 18 and 19, the only remaining (putative) cue controlling conditioned catalepsy was the drug cue.

The expression of behavioral sensitization may depend collectively on a parallel and balanced conditioning between the drug, exteroceptive and interoceptive cues (Carey and Gui, 1998). The fact that catalepsy was observed on days 18 and 19 only in the animals previously given haloperidol-test environment pairings but not in the unpaired group supports this idea. Previous sensitization studies have suggested that the test environment may serve as an ‘occasion-setter’ for previously conditioned drug cues

(Anagnostaras and Robinson, 1996; Badiani et al., 1998; Bouton and Swartzentruber,

1986). In other words, a drug can acquire a specific meaning and elicit an appropriate conditioned response but only in that particular environment. We expand this view and propose that extinction of exteroceptive stimuli may also result in a shift in the balance between the strength of conditioned exteroceptive and interoceptive cues. This may ultimately contribute to a shift from environment-specific catalepsy toward environment- independent catalepsy, now solely maintained by drug cues. Thus, following extinction training in Context A, haloperidol 0.25 mg/kg produced a small but significant catalepsy response in both context A and B. Further studies are needed to evaluate this interpretation.

We showed for the first time in experiment 3 that previous exposure to the catalepsy test environment led to a more rapid development of catalepsy sensitization in 113

experiment 3. Novel environmental stimuli have been shown to activate the DA system

(Badiani et al., 1998; Bardo et al., 1990; Bromberg-Martin et al., 2010; Hooks and

Kalivas, 1995; Horvitz, 2000; Redgrave and Gurney, 2006; Redgrave et al., 2008) and habituation to these conditions reduces such activation (Legault and Wise, 2001). As haloperidol is a competitive DA D2 receptor antagonist, its effect is influenced by the concentration of endogenous DA. Thus, under conditions where DA release is low, i.e., in habituated animals, haloperidol will be more effective at further reducing DA transmission to a point where it shifts the catalepsy sensitization curve to the left.

Haloperidol treatment has been shown to facilitate different types of inhibitory learning like habituation of exploratory behavior (Bespalov et al., 2007; Lynch and Carey, 1986), extinction of operant responding (Gray and Wise, 1980; Phillips and Fibiger, 1979), and latent inhibition (Ruob et al., 1998; Weiner and Feldon, 1987) further supporting this view. Therefore, repeated haloperidol treatments in test environments with lower incentive motivational properties (e.g., habituated environments) can produce an augmented catalepsy sensitization response in comparison to similar treatments in less familiar testing environments.

Previous research has shown that repeated antipsychotic treatment can affect motivational processes (Salamone and Correa, 2002; Wise, 1982; Wise, 2004). Wise and colleagues were the first to show that DA function is critical for conditioned incentive stimuli to maintain operant responding for stimulant drugs (Yokel and Wise, 1975; Yokel and Wise, 1976), food (Wise et al., 1978) and brain stimulation (Corbett and Wise, 1980).

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These studies showed that well-trained animals treated with the D2-preferring receptor antagonists pimozide or haloperidol normally initiated responding for available rewards, however with repeated treatment and testing the animals showed an extinction-like decline in responding that could not be explained by performance impairments (Franklin and McCoy, 1979; Yokel and Wise, 1975; Yokel and Wise, 1976).

Sensitization to the effects of antipsychotics has also been shown in expression of conditioned avoidance behavior (Boivin and Beninger, 2008; Li et al., 2007). In these studies, first-generation (haloperidol) or second-generation (olanzapine, risperidone) antipsychotics produced a gradual between-session decline in avoidance responses to an auditory stimulus predictive of an aversive foot shock. According to Carey (1987) behavioral sensitization and conditioning are critical to the variable onset of therapeutic action of antipsychotics. Studies from his lab have shown that animals repeatedly treated with haloperidol exhibited a more rapid extinction-like decline in exploratory behavior controlled by unconditioned incentive stimuli such as environmental novelty or entrance of the dark compartment of a light/dark apparatus (Carey, 1987; Carey and Kenney,

1987). In each case, when saline was substituted for haloperidol, animals showed drug- appropriate conditioned responses but only in the haloperidol-paired test environment and not when tested in a different environment (Lynch and Carey, 1987). Results implicate

DA neurotransmission in the regulation of responses to novel and habituated unconditioned incentive stimuli, as well as conditioned incentive stimuli.

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Excessive activation of the DA system in people with schizophrenia (Abi-

Dargham et al., 2009; Laruelle et al., 1999; Lodge and Grace, 2011) or following repeated psychomotor stimulant intake (Li et al., 2004; Robinson and Berridge, 1993;

Singer et al., 2009) can lead to excessive incentive learning about environmental stimuli that normally would not elicit approach and other responses (Beninger, 1983; Crow,

1979; Gray et al., 1991; Kapur, 2003; McKenna, 1987; Miller, 1976; Murray, 2010;

Schmidt and Beninger, 2006)). This aberrant incentive learning is believed to be involved in the development of psychosis (Beninger, 1983; Miller, 1976). On the other hand, antipsychotic treatment can reduce incentive motivational properties of both conditioned and unconditioned stimuli, resulting in a gradual loss of the ability of those stimuli to attract, engage and maintain behavior as shown here and by others (Banasikowski and

Beninger, 2012a; Beninger, 1983; Carey, 1987; Salamone and Correa, 2002; Wise, 1982;

Wise, 2004).

In aggregate, similar to the behavioral sensitization produced by psychostimulant drugs, the acquisition and expression of sensitization to haloperidol is also conditional on the presence of drug-associated environmental stimuli. Our findings provide further insight into the current understanding of learning processes involved in the action of antipsychotic drugs. Thus, the dependence of antipsychotic drugs on environmental stimuli for their action may provide a basis for understanding the variable-onset of therapeutic benefit (Emsley et al., 2006) and why the schizophrenia outpatient population remains vulnerable to psychosis relapse despite medication adherence (Chen et al., 2010;

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Crow et al., 1986; Hogarty and Ulrich, 1998; Robinson, 2011).

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4.6 References

Abi-Dargham A, Rodenhiser J, Printz D, Zea-Ponce Y, et al. (2000). Increased baseline occupancy of D2 receptors by dopamine in schizophrenia. Proc Natl Acad Sci U S A, 97(14): 8104-8109. Abi-Dargham A, van de Giessen E, Slifstein M, Kegeles LS, et al. (2009). Baseline and amphetamine-stimulated dopamine activity are related in drug-naive schizophrenic subjects. Biol Psychiatry, 65(12): 1091-1093. Ahn S, Phillips AG (1999). Dopaminergic correlates of sensory-specific satiety in the medial prefrontal cortex and nucleus accumbens of the rat. J Neurosci, 19(19): RC29. Amtage J, Schmidt WJ (2003). Context-dependent catalepsy intensification is due to classical conditioning and sensitization. Behav Pharmacol, 14(7): 563-567. Anagnostaras SG, Robinson TE (1996). Sensitization to the psychomotor stimulant effects of amphetamine: modulation by associative learning. Behav Neurosci, 110(6): 1397-1414. Badiani A, Oates MM, Day HE, Watson SJ, et al. (1998). Amphetamine-induced behavior, dopamine release, and c-fos mRNA expression: modulation by environmental novelty. J Neurosci, 18(24): 10579-10593. Banasikowski TJ, Beninger RJ (2012a). Haloperidol conditioned catalepsy in rats: a possible role for D1-like receptors. The International Journal of Neuropsychopharmacology, FirstView: 1-10. Banasikowski TJ, Beninger RJ (2012b). Reduced expression of haloperidol conditioned catalepsy in rats by the dopamine D3 receptor antagonists nafadotride and NGB 2904. European Neuropsychopharmacology, (0). Bardo MT, Bowling SL, Pierce RC (1990). Changes in locomotion and dopamine neurotransmission following amphetamine, haloperidol, and exposure to novel environmental stimuli. Psychopharmacology (Berl), 101(3): 338-343. Beninger RJ (1983). The role of dopamine in locomotor activity and learning. Brain Res, 287(2): 173-196. Bespalov A, Jongen-Relo AL, van Gaalen M, Harich S, et al. (2007). Habituation deficits induced by metabotropic glutamate receptors 2/3 receptor blockade in mice: reversal by antipsychotic drugs. J Pharmacol Exp Ther, 320(2): 944-950.

118

Boileau I, Dagher A, Leyton M, Gunn RN, et al. (2006). Modeling sensitization to stimulants in humans: an [11C]raclopride/positron emission tomography study in healthy men. Arch Gen Psychiatry, 63(12): 1386-1395. Boivin GA, Beninger RJ (2008). Differential effects of dopamine and AMPA receptor antagonists on the expression of conditioned avoidance responding in rats. Behav Neurosci, 122(2): 377-384. Bouton ME, Swartzentruber D (1986). Analysis of the associative and occasion-setting properties of contexts participating in a Pavlovian discrimination. Journal of Experimental Psychology: Animal Behavior Processes, 12(4): 333. Bromberg-Martin ES, Matsumoto M, Hikosaka O (2010). Dopamine in motivational control: rewarding, aversive, and alerting. Neuron, 68(5): 815-834. Carey RJ (1987). Conditioning and the delayed onset of a haloperidol-induced behavioral effect. Biol Psychiatry, 22(3): 269-277. Carey RJ, Gui J (1998). Cocaine conditioning and cocaine sensitization: what is the relationship? Behav Brain Res, 92(1): 67-76. Carey RJ, Kenney S (1987). Operant conditioning and haloperidol-induced hypokinetic effects. Neuropsychobiology, 18(4): 199-204. Carlsson A, Lindqvist M (1963). Effect Of Chlorpromazine Or Haloperidol On Formation Of 3methoxytyramine And Normetanephrine In Mouse Brain. Acta Pharmacol Toxicol (Copenh), 20: 140-144. Chen EY, Hui CL, Lam MM, Chiu CP, et al. (2010). Maintenance treatment with quetiapine versus discontinuation after one year of treatment in patients with remitted first episode psychosis: randomised controlled trial. Bmj, 341: c4024. Cohen C, Sanger DJ, Perrault G (1997). Characterization of the discriminative stimulus produced by the dopamine antagonist tiapride. J Pharmacol Exp Ther, 283(2): 566-573. Colpaert FC, Niemegeers CJ, Janssen PA (1978). Discriminative stimulus properties of cocaine and d-amphetamine, and antagonism by haloperidol: a comparative study. Neuropharmacology, 17(11): 937-942. Corbett D, Wise RA (1980). Intracranial self-stimulation in relation to the ascending dopaminergic systems of the midbrain: a moveable electrode mapping study. Brain Res, 185(1): 1-15.

119

Crow TJ (1979). Catecholamine reward pathways and schizophrenia: the mechanism of the antipsychotic effect and the site of the primary disturbance. Fed Proc, 38(11): 2462-2467. Crow TJ, MacMillan JF, Johnson AL, Johnstone EC (1986). A randomised controlled trial of prophylactic neuroleptic treatment. Br J Psychiatry, 148: 120-127. Emsley R, Rabinowitz J, Medori R (2006). Time course for antipsychotic treatment response in first-episode schizophrenia. Am J Psychiatry, 163(4): 743-745. Featherstone RE, Kapur S, Fletcher PJ (2007). The amphetamine-induced sensitized state as a model of schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry, 31(8): 1556-1571. Franklin KB, McCoy SN (1979). Pimozide-induced extinction in rats: stimulus control of responding rules out motor deficit. Pharmacol Biochem Behav, 11(1): 71-75. Gauvin DV, Goulden KL, Holloway FA (1994). A three-choice haloperidol-saline- cocaine drug discrimination task in rats. Pharmacol Biochem Behav, 49(1): 223- 227. Goudie AJ, Smith JA (1999). Discriminative stimulus properties of antipsychotics. Pharmacol Biochem Behav, 64(2): 193-201. Grace AA (2000). Gating of information flow within the limbic system and the pathophysiology of schizophrenia. Brain Res Brain Res Rev, 31(2-3): 330-341. Grace AA, Bunney BS, Moore H, Todd CL (1997). Dopamine-cell depolarization block as a model for the therapeutic actions of antipsychotic drugs. Trends Neurosci, 20(1): 31-37. Gray JA, Feldon J, Rawlins JNP, Hemsley DR, et al. (1991). The neuropsychology of schizophrenia. Behavioral and Brain Sciences, 14(01): 1-20. Gray T, Wise RA (1980). Effects of pimozide on lever pressing behavior maintained on an intermittent reinforcement schedule. Pharmacol Biochem Behav, 12(6): 931- 935. Hogarty GE, Ulrich RF (1998). The limitations of antipsychotic medication on schizophrenia relapse and adjustment and the contributions of psychosocial treatment. J Psychiatr Res, 32(3-4): 243-250. Hooks MS, Kalivas PW (1995). The role of mesoaccumbens--pallidal circuitry in novelty-induced behavioral activation. Neuroscience, 64(3): 587-597.

120

Horvitz JC (2000). Mesolimbocortical and nigrostriatal dopamine responses to salient non-reward events. Neuroscience, 96(4): 651-656. Kapur S (2003). Psychosis as a state of aberrant salience: a framework linking biology, phenomenology, and pharmacology in schizophrenia. Am J Psychiatry, 160(1): 13-23. Kapur S, Remington G (2001). Dopamine D(2) receptors and their role in atypical antipsychotic action: still necessary and may even be sufficient. Biol Psychiatry, 50(11): 873-883. Kegeles LS, Abi-Dargham A, Frankle WG, Gil R, et al. (2010). Increased synaptic dopamine function in associative regions of the striatum in schizophrenia. Arch Gen Psychiatry, 67(3): 231-239. Klein A, Schmidt WJ (2003). Catalepsy intensifies context-dependently irrespective of whether it is induced by intermittent or chronic dopamine deficiency. Behav Pharmacol, 14(1): 49-53. Lanis A, Schmidt WJ (2001). NMDA receptor antagonists do not block the development of sensitization of catalepsy, but make its expression state- dependent. Behav Pharmacol, 12(2): 143-149. Laruelle M (2000). The role of endogenous sensitization in the pathophysiology of schizophrenia: implications from recent brain imaging studies. Brain Res Brain Res Rev, 31(2-3): 371-384. Laruelle M, Abi-Dargham A, Gil R, Kegeles L, et al. (1999). Increased dopamine transmission in schizophrenia: relationship to illness phases. Biol Psychiatry, 46(1): 56-72. Laruelle M, Abi-Dargham A, van Dyck CH, Gil R, et al. (1996). Single photon emission computerized tomography imaging of amphetamine-induced dopamine release in drug-free schizophrenic subjects. Proc Natl Acad Sci U S A, 93(17): 9235-9240. Legault M, Wise RA (2001). Novelty-evoked elevations of nucleus accumbens dopamine: dependence on impulse flow from the ventral subiculum and glutamatergic neurotransmission in the ventral tegmental area. Eur J Neurosci, 13(4): 819-828. Li M, Fletcher PJ, Kapur S (2007). Time course of the antipsychotic effect and the underlying behavioral mechanisms. Neuropsychopharmacology, 32(2): 263-272.

121

Li Y, Acerbo MJ, Robinson TE (2004). The induction of behavioural sensitization is associated with cocaine-induced structural plasticity in the core (but not shell) of the nucleus accumbens. Eur J Neurosci, 20(6): 1647-1654. Lodge DJ, Grace AA (2008). Amphetamine activation of hippocampal drive of mesolimbic dopamine neurons: a mechanism of behavioral sensitization. J Neurosci, 28(31): 7876-7882. Lodge DJ, Grace AA (2011). Hippocampal dysregulation of dopamine system function and the pathophysiology of schizophrenia. Trends Pharmacol Sci, 32(9): 507-513. Lodge DJ, Grace AA (2012). Divergent activation of ventromedial and ventrolateral dopamine systems in animal models of amphetamine sensitization and schizophrenia. Int J Neuropsychopharmacol: 1-8. Lynch MR, Carey RJ (1986). Within-session data. Biol Psychiatry, 21(5-6): 573-574. Lynch MR, Carey RJ (1987). Environmental stimulation promotes recovery from haloperidol-induced extinction of open field behavior in rats. Psychopharmacology (Berl), 92(2): 206-209. McElroy JF, Stimmel JJ, O'Donnell JM (1989). Discriminative stimulus properties of haloperidol. Drug Development Research, 18(1): 47. McKenna PJ (1987). Pathology, phenomenology and the dopamine hypothesis of schizophrenia. Br J Psychiatry, 151: 288-301. Miller R (1976). Schizophrenic psychology, associative learning and the role of forebrain dopamine. Med Hypotheses, 2(5): 203-211. Miller R (1987). The time course of neuroleptic therapy for psychosis: role of learning processes and implications for concepts of psychotic illness. Psychopharmacology (Berl), 92(4): 405-415. Murray GK (2010). The emerging biology of delusions. Psychol Med, 41(1): 7-13. Paulson PE, Robinson TE (1995). Amphetamine-induced time-dependent sensitization of dopamine neurotransmission in the dorsal and ventral striatum: a microdialysis study in behaving rats. Synapse, 19(1): 56-65. Peleg-Raibstein D, Knuesel I, Feldon J (2008). Amphetamine sensitization in rats as an animal model of schizophrenia. Behav Brain Res, 191(2): 190-201. Pert A, Post R, Weiss SR (1990). Conditioning as a critical determinant of sensitization induced by psychomotor stimulants. NIDA Res Monogr, 97: 208-241.

122

Phillips AG, Fibiger HC (1979). Decreased resistance to extinction after haloperidol: implications for the role of dopamine in reinforcement. Pharmacol Biochem Behav, 10(5): 751-760. Post RM, Rose H (1976). Increasing effects of repetitive cocaine administration in the rat. Nature, 260(5553): 731-732. Rebec GV, Christensen JR, Guerra C, Bardo MT (1997). Regional and temporal differences in real-time dopamine efflux in the nucleus accumbens during free- choice novelty. Brain Res, 776(1-2): 61-67. Redgrave P, Gurney K (2006). The short-latency dopamine signal: a role in discovering novel actions? Nat Rev Neurosci, 7(12): 967-975. Redgrave P, Gurney K, Reynolds J (2008). What is reinforced by phasic dopamine signals? Brain Res Rev, 58(2): 322-339. Riedinger K, Kulak A, Schmidt WJ, von Ameln-Mayerhofer A (2011). The role of NMDA and AMPA/Kainate receptors in the consolidation of catalepsy sensitization. Behav Brain Res, 218(1): 194-199. Robinson DG (2011). Medication adherence and relapse in recent-onset psychosis. Am J Psychiatry, 168(3): 240-242. Robinson TE, Berridge KC (1993). The neural basis of drug craving: an incentive- sensitization theory of addiction. Brain Res Brain Res Rev, 18(3): 247-291. Ruob C, Weiner I, Feldon J (1998). Haloperidol-induced potentiation of latent inhibition: interaction with parameters of conditioning. Behav Pharmacol, 9(3): 245-253. Salamone JD, Correa M (2002). Motivational views of reinforcement: implications for understanding the behavioral functions of nucleus accumbens dopamine. Behav Brain Res, 137(1-2): 3-25. Schmidt WJ, Beninger RJ (2006). Behavioural sensitization in addiction, schizophrenia, Parkinson's disease and dyskinesia. Neurotox Res, 10(2): 161-166. Schmidt WJ, Tzschentke TM, Kretschmer BD (1999). State-dependent blockade of haloperidol-induced sensitization of catalepsy by MK-801. Eur J Neurosci, 11(9): 3365-3368. Seeman P, Lee T, Chau-Wong M, Wong K (1976). Antipsychotic drug doses and neuroleptic/dopamine receptors. Nature, 261(5562): 717-719.

123

Segal DS, Mandell AJ (1974). Long-term administration of d-amphetamine: progressive augmentation of motor activity and stereotypy. Pharmacol Biochem Behav, 2(2): 249-255. Singer BF, Tanabe LM, Gorny G, Jake-Matthews C, et al. (2009). Amphetamine- induced changes in dendritic morphology in rat forebrain correspond to associative drug conditioning rather than nonassociative drug sensitization. Biol Psychiatry, 65(10): 835-840. Stewart J, Vezina P (1991). Extinction procedures abolish conditioned stimulus control but spare sensitized responding to amphetamine. Behav Pharmacol, 2(1): 65-71. Wadenberg ML, Soliman A, VanderSpek SC, Kapur S (2001). Dopamine D(2) receptor occupancy is a common mechanism underlying animal models of antipsychotics and their clinical effects. Neuropsychopharmacology, 25(5): 633- 641. Weiner I, Feldon J (1987). Facilitation of latent inhibition by haloperidol in rats. Psychopharmacology (Berl), 91(2): 248-253. Wiecki TV, Riedinger K, von Ameln-Mayerhofer A, Schmidt WJ, et al. (2009). A neurocomputational account of catalepsy sensitization induced by D2 receptor blockade in rats: context dependency, extinction, and renewal. Psychopharmacology (Berl), 204(2): 265-277. Wise RA (1982). Neuroleptics and operant behavior: The anhedonia hypothesis. Behavioral and Brain Sciences, 5(01): 39-53. Wise RA (2004). Dopamine, learning and motivation. Nat Rev Neurosci, 5(6): 483-494. Wise RA, Spindler J, deWit H, Gerberg GJ (1978). Neuroleptic-induced "anhedonia" in rats: pimozide blocks reward quality of food. Science, 201(4352): 262-264. Wise RA, Wang B, You ZB (2008). Cocaine serves as a peripheral interoceptive conditioned stimulus for central glutamate and dopamine release. PLoS One, 3(8): e2846. Yokel RA, Wise RA (1975). Increased lever pressing for amphetamine after pimozide in rats: implications for a dopamine theory of reward. Science, 187(4176): 547-549. Yokel RA, Wise RA (1976). Attenuation of intravenous amphetamine reinforcement by central dopamine blockade in rats. Psychopharmacology (Berl), 48(3): 311-318.

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Chapter 5

General Discussion

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5.1 Experimental Summary

The results from the three experimental chapters can be summarized as follows:

Rats treated with a low dose of the D2 receptor-preferring antagonist haloperidol and tested for catalepsy in a specific environment showed no initial catalepsy but the catalepsy response emerged and increased from day-to-day across subsequent test days.

This effect was observed in experimental chapters 1, 2 and 3. Day-to-day sensitization of catalepsy was seen with each dose of haloperidol tested but was strongest with the highest dose of haloperidol (0.5 mg/kg).

Development of catalepsy sensitization in one environment did not transfer to another environment. Similarly, rats with a history of haloperidol treatments outside of the test environment (unpaired group) did not exhibit significant catalepsy when given haloperidol for the first time prior to catalepsy testing suggesting that a history of drug- environment interaction controls catalepsy expression. Further, rats previously given haloperidol and tested with saline in the drug environment exhibited conditioned catalepsy. The magnitude of conditioned catalepsy observed was not dose-dependent as all doses of haloperidol resulted in a similar amount of conditioned catalepsy.

Importantly, conditioned catalepsy was observed in the drug-paired environment but not in a different environment suggesting that drug-paired stimuli acquire the ability to elicit catalepsy.

Conditioned catalepsy showed gradual day-to-day extinction with repeated saline treatment in the previously haloperidol-paired environment. It appears that following

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extinction the performance of sensitized responding to haloperidol shifted from environment-specific toward environment-independent expression. Subsequent haloperidol challenges produced a small but significant catalepsy effect in both Context

A and B, but only in previously sensitized animals (paired group) but not in the unpaired group. As the conditioned environmental cues were extinguished in Context A, it was hypothesized that the catalepsy response in Context A and B was now produced by putative conditioned drug cue properties of haloperidol.

Rats treated with the D1-like receptor antagonist SCH 23390 exhibited some catalepsy but failed to develop significant day-to-day sensitization as seen with haloperidol. Repeated treatment with SCH 23390 did not produce conditioned catalepsy.

Co-treatment with SCH 23390 and haloperidol resulted in a synergistic catalepsy effect and a shift of the catalepsy sensitization curve to the left. When co-treated rats were challenged with saline and tested in the drug-paired environment they failed to exhibit conditioned catalepsy. On the other hand, rats previously sensitized with haloperidol alone and given SCH 23390 before the saline test showed conditioned catalepsy. These results suggest that functional D1-like receptors are critical for acquisition of conditioned catalepsy, but not for its expression.

Treatment with the D3 receptor antagonists, nafadotride or NGB 2904 did not produce catalepsy. Rats previously sensitized with haloperidol alone and treated with nafadotride or NGB 2904 before the saline test day showed an attenuated conditioned catalepsy response. The same doses of nafadotride or NGB 2904 given with haloperidol

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during sensitization sessions did not affect conditioned catalepsy acquisition. These results suggest that functional D3 receptors play a role in expression of conditioned catalepsy, but not for its acquisition.

Previous exposure to the catalepsy test environment led to a more rapid development of catalepsy sensitization. It is believed that environmental novelty can elicit unconditioned incentive motivational responding that is manifested as approach and other exploratory behavior. Moreover, novel environmental stimuli activate the DA system (Badiani et al., 1998; Badiani et al., 1999; Bardo et al., 1990; Bromberg-Martin et al., 2010; Hooks and Kalivas, 1995; Horvitz, 2000; Redgrave et al., 2008) and habituation to these conditions reduces such activation (Legault and Wise, 2001). A reduction in the level of DA activity in environment following habituation may have led to an augmentation of the effects of haloperidol and a shift of the catalepsy sensitization curve to the left.

5.2 Dopamine Receptor Subtypes and Behavior

5.2.1 Behavioral Sensitization

DA plays an important role in motor function, expression of motivated behavior and certain types of learning (Beninger, 1983; Carey, 1987; Wise, 1982). The behavioral effects of drugs that act on the brain’s DA system change with repeated exposure to the drug. Moreover, different DA receptor sub-types may contribute in a circumstance- specific way to various aspects of DA-mediated behaviors (Banasikowski et al., 2010;

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Banasikowski et al., 2012). Although sensitization to the effects of antipsychotics has been shown in the past (Amtage and Schmidt, 2003; Boivin and Beninger, 2008; Carey,

1987; Li et al., 2007; Wiecki et al., 2009), the mechanisms responsible for this phenomenon have not been elucidated. The current results show that repeated treatment with a D2, but not D1-like or D3 receptor-preferring antagonists in a particular test environment produces catalepsy sensitization. Moreover, acquisition of conditioned catalepsy is dependent on D1-like receptors, while its expression is dependent on D3 receptors.

Catalepsy sensitization is controlled by a drug-environment interaction. Control groups that received environment-saline pairing and also received haloperidol in their homecage (colony room) environment did not exhibit increased catalepsy during the test sessions. Moreover, following catalepsy sensitization, if the same dose of haloperidol was injected and catalepsy tested in a different environment, animals failed to express catalepsy, an effect shown here and by others (Klein and Schmidt, 2003; Wiecki et al.,

2009). Similarly, rats with a history of haloperidol treatments outside of the test environment (unpaired group) did not exhibit significant catalepsy when given haloperidol for the first time prior to catalepsy testing. Such results make an interpretation based on non-associative processes such as motor or sensory impairments resulting from physiological effect of repeated haloperidol injections (e.g., cumulative drug effects) an unlikely mechanism to explain catalepsy sensitization and conditioning.

Thus the effect of increased catalepsy in rats that have received environment-haloperidol

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pairing can be attributed to the association of environmental stimuli with haloperidol rather than a previous history of treatment with a D2 receptor antagonist.

The sensitization and conditioning effects previously reported for psychomotor stimulants also have been shown to rely on their association with environmental stimuli.

Repeated treatment and testing with psychomotor stimulant drugs that enhance DA transmission, e.g., amphetamine (AMPH) or cocaine, produces a day-to-day increase in locomotor responses in rats (Kalivas and Stewart, 1991; Pert et al., 1990; Post and Rose,

1976; Segal and Mandell, 1974). A number of results suggest that the magnitude of various electorphysiological and neurochemical responses to these drugs, including the level of extracellular DA in the dorsal and ventral striatum, increases with repeated treatments (Badiani et al., 1999; Goto and Grace, 2005; Li et al., 2004; Paulson and

Robinson, 1995; Singer et al., 2009; Ungless et al., 2001; Vezina and Queen, 2000). The enhanced locomotor response as well as the enhanced striatal DA release with each repeated AMPH treatment (Badiani et al., 1998; Badiani et al., 1999) is context- dependent, i.e., its expression is strongest in the presence of previously drug-paired environmental stimuli (Anagnostaras and Robinson, 1996; Badiani et al., 1998; Pert et al., 1990). Co-treatment with either D1-like or D2 receptor antagonists plus the DA agonist has been shown to block drug-induced day-to-day increases in locomotor activity

(Kuribara, 1995; Vezina, 1996; Vezina and Stewart, 1989; Wise and Carlezon, 1994).

However, when the DA agonist is subsequently given alone without the DA antagonists, behavioral sensitization has been reported in rats previously treated with a D2 but not D1-

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like receptor antagonist (Vezina and Stewart, 1989) suggesting a critical role of D1-like receptors in the development of psychomotor stimulant sensitization (Vezina, 1996).

Development of behavioral sensitization to drugs that directly or indirectly activate the DA system is accompanied by an increase in D3 receptor mRNA and/or expression in the striatum (Bezard et al., 2003; Bordet et al., 1997; Chiang et al., 2003;

Le Foll et al., 2003; Liang et al., 2011; Neisewander et al., 2004). The expression of behavioral sensitization is also attenuated with D3 receptor antagonism (Liang et al.,

2011; Bordet et al., 1997). Furthermore, pre-treatment with the D1-like receptor antagonist SCH 23390 actually prevents the induction of D3 receptor changes correlated with behavioral sensitization to L-DOPA (Bordet et al., 1997).

When viewed together, results from sensitization and conditioning studies with

DA receptor antagonists (usually haloperidol or pimozide) and those from similar studies with DA receptor agonists (usually AMPH or cocaine) reveal a number of striking parallels. Thus, both classes of drugs produce behavioral sensitization with repeated testing that depends on the test environment. Both produce conditioned responses to the drug-associated environment. The development of behavioral sensitization and conditioning to DA receptor antagonists or to DA receptor agonists depends on intact D1- like receptors. The expression of conditioned responses to both classes of drugs depends on intact D3 receptors. These parallel observations imply that the same DA receptor mechanisms may be involved in the development of sensitized and conditioned responses to DA antagonists and to DA agonists.

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5.2.2 Incentive (Stimulus – Outcome) Learning and Motivation

Beninger (1983) was one of the first to argue that DA plays a critical role in incentive learning. Rats repeatedly treated with pro-DA drugs like cocaine and AMPH in a particular environment are observed to be more active than control animals when injected with saline and tested in the drug-paired environment (Banasikowski et al., 2010;

Beninger, 1983; Beninger and Hahn, 1983; Beninger and Herz, 1986). From the incentive learning point of view, the environment stimuli paired with a drug like AMPH or cocaine acquire incentive salience and thus an increased ability to elicit approach and other responses during conditioning that manifests as conditioned activity during drug-free testing. The acquisition of environment-specific conditioned drug effects is blocked by treatment with D1-like and D2, but not D3 receptor antagonists (Banasikowski et al.,

2010; Banasikowski et al., 2012; Beninger and Hahn, 1983; Cervo and Samanin, 1995;

Dias et al., 2006; Fontana et al., 1993a; Mazurski and Beninger, 1991), while initial expression of conditioning is attenuated by D3 receptor but not D1-like or D2 receptor antagonists (Banasikowski et al., 2010; Banasikowski et al., 2012; Beninger and Hahn,

1983; Cervo and Samanin, 1995; Fontana et al., 1993a; Franklin and McCoy, 1979;

Ljungberg, 1990; Wise et al., 1978). Following drug-environment pairings, a change in

D3 receptor expression in behaviorally conditioned animals has been reported (Le Foll et al., 2002; Neisewander et al., 2004). More importantly, the up-regulation of D3 receptors was absent in animals that received saline-environment pairings and drug treatment in the homecage environment (Le Foll et al., 2002; Liang et al., 2011) suggesting that a mere

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history of drug treatment is not enough to elicit this physiological change.

In the catalepsy sensitization model, when rats were injected with saline instead of haloperidol in the test environment, they continued to exhibit catalepsy - now conditioned. Co-treatment with a D1-like but not D3 receptor antagonist during pairing sessions blocked the acquisition of haloperidol conditioned catalepsy suggesting that this form of learning requires functional D1-like receptors. D1-like receptor activity has been implicated in encoding different forms of learning (Dalley et al., 2005; El-Ghundi et al.,

1999; Fenu et al., 2001; Parker et al., 2010; Smith-Roe and Kelley, 2000), while D2 receptor antagonism has been shown to facilitate inhibitory learning, whereby stimulus exposure extinguishes its prediction of a potential consequence (Bespalov et al., 2007;

Bouton, 2004; Ruob et al., 1998; Trimble et al., 2002; Weiner and Feldon, 1987; Wise et al., 1978). The ‘stimulus-no outcome’ relationship observed in latent inhibition or the omission of expected outcome seen in extinction training share certain similarity with catalepsy sensitization and conditioning. In agreement with this observation, post-trial injections with SCH 23390 impaired acquisition of extinction to stimuli previously associated with cocaine (Fricks-Gleason et al., 2012). Thus, catalepsy sensitization and conditioning resulting from a progressive loss of engagement with drug-paired stimuli is dependent on D1-like receptor activity.

In agreement with the previously reported role of D3 receptors in conditioned behaviors, the expression of conditioned catalepsy was attenuated by D3 (but not D1- like) receptors antagonism (Banasikowski et al., 2010; Banasikowski et al., 2012;

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Beninger and Banasikowski, 2008). Both D2 and D3 receptor have been implicated in the expression of incentive motivation, especially when conditioned behavior is maintained only by reward-predictive incentive stimuli (Beninger and Ranaldi, 1992; Cervo et al.,

2003; Cervo et al., 2007; Di Ciano et al., 2003; Gilbert et al., 2005; Hitchcott and

Phillips, 1998). Supporting this observation, it has been shown that D2 and D3 receptors interact physically and functionally with one another forming D2/D3 heterodimers

(Maggio et al., 2009; Scarselli et al., 2001). Both D2 and D3 receptor antagonists reduce lever press responding for conditioned rewards (Cervo et al., 2007; Di Ciano et al., 2003), however there are apparent differences between animals treated with D2 and D3 receptor antagonists. For example, in well-trained animals, D2 receptor antagonism produces an extinction-like decline of continuously reinforced responding after the animal has repeated experience with reward under D2 receptor blockade (Wise, 2004). On the other hand, D3 receptor antagonism does not affect behavior on continuously rewarded trials

(Di Ciano et al., 2003; Gilbert et al., 2005). Further, recent work from our lab has shown that while D3 receptor antagonism dose-dependently attenuates expression of cocaine conditioned activity and AMPH conditioned place preference, D2 receptor antagonism does not attenuate expression of these conditioned behaviors during initial extinction training (Banasikowski et al., 2010; Banasikowski et al., 2012).

The current finding that treatment with a D2, but not D3 or D1-like receptor- preferring antagonist in a particular test environment produce catalepsy sensitization, while acquisition of conditioned catalepsy is dependent on D1-like receptors, and its

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expression is dependent on D3 receptors advances considerably the current thinking of the role of DA receptor function in stages of incentive learning and expression of incentive motivation. Repeated D2 receptor agonism or antagonism and subsequent changes in DA levels in a specific context may change the way environmental stimuli are perceived resulting in a progressive gain or loss, respectively, of engagement with those environmental stimuli. Further examination of the mechanisms involved in this form of learning reveals essential, independent roles, with D2 receptors "setting" the incentive value of reward-related stimuli, while D1-like receptors control their encoding, and D3 receptors control the retrieval of incentive value assigned to them.

5.3 Dopamine and Striatal Plasticity

Dopamine receptor activation has been shown to play a complex role in controlling corticostriatal plasticity at medium-spiny neurons (MSNs) (Reynolds and

Wickens, 2002; Shen et al., 2008). DA neurons are activated by encountering unexpected reward stimuli - primary rewards like food, water, a receptive conspecific, shelter and drugs of abuse. Activation is also seen when there is an unexpected presentation of stimuli that have become associated with the rewards – now conditioned incentive stimuli

(Glimcher, 2011; Hollerman and Schultz, 1998). The sudden firing of DA neurons in response to these stimuli appears to modulate the processing of the concurrent glutamate inputs to the MSNs (Kiyatkin and Rebec, 1996).

Both antipsychotics and psychomotor stimulants activate expression of immediate early gene (IEG) transcription factors c-fos and zif268 in the ventral and dorsal striatum

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(Badiani et al., 1998; Bertran-Gonzalez et al., 2008; Bhat et al., 1992; Graybiel et al.,

1990; LaHoste et al., 2000). IEG activation represents an indirect marker of neuronal activity and possibly the onset of neuronal plasticity (Dragunow et al., 1989; Dragunow and Robertson, 1988). Activation of IEGs in the striatal MSNs occurs with concurrent treatment of D1-like and D2 receptor agonists, but not when either drug is given alone

(Capper-Loup et al., 2002; LaHoste et al., 1993). Cocaine- and AMPH-induced expression of IEGs is observed in both D1- expressing striatonigral and D2-expressing striatopallidal projecting MSNs when treatment was performed in a novel test environment (Badiani et al., 1999; Bertran-Gonzalez et al., 2008). Antipsychotic drug - induced striatal IEG is also environment-dependent (Murphy and Feldon, 2001) and is reversed by cocaine or AMPH treatment (Graybiel et al., 1990; Robertson et al., 1991).

Enhanced DA transmission produced by psychomotor stimulants or incentive stimuli increases activity at D1 receptor-expressing MSNs and inhibits activity at D2 receptor-expressing MSNs (Surmeier et al., 2007) thus promoting locomotion by its action on both pathways (Bateup et al., 2010). Similarly, activity of MSNs expressing either D1 or D2 receptors has been shown to have opposing roles in mediating behavioral sensitization effects produced by AMPH (Beutler et al., 2011; Ferguson et al., 2011;

Pascoli et al., 2012, Hikida et al., 2010). Inactivation of neuronal excitability of striatopallidal neurons facilitates the development of AMPH behavioral sensitization

(Ferguson et al., 2011), while disruption of activity of striatonigral neurons abolishes stimulant effects of AMPH, development of sensitization, and conditioning to stimuli

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predictive of the drug (Beutler et al., 2011; Hikida et al., 2010).

Similarly haloperidol catalepsy sensitization may result from alterations in the strength of glutamatergic synaptic connections between the cortex and striatum

(Centonze et al., 2004). A neurocomputational model of haloperidol catalepsy sensitization developed by Frank and colleagues (2009) suggested that plasticity within the striatopallidal pathway alone is responsible for environment-specific catalepsy sensitization and conditioning (Wiecki et al., 2009). However, current evidence at hand requires the need to expand this hypothesis to include plasticity at both striatopallidal and striatonigral MSNs for catalepsy sensitization and conditioning to be manifested. Many behavioral, electrophysiological and gene-activating DA-mediated functions require concomitant activation of D1-like and D2 receptors, a phenomenon known as ‘requisite

D1/D2 synergism’ (Capper-Loup et al., 2002; LaHoste et al., 1993; Walters et al., 1987).

Moreover, D1-like receptor activity plays a critical role in acquisition of many conditioned behaviors that require encoding of contextual information (Cervo and

Samanin, 1995; El-Ghundi et al., 1999; Granado et al., 2008; Lemon and Manahan-

Vaughan, 2006; O'Carroll and Morris, 2004).

As D2 receptors are found both pre-and post-synaptically it is likely that initial treatments with haloperidol produce profound increase in synaptic DA concentrations due to blockade of the pre-synaptic D2 autoreceptors (Bunney and Grace, 1978; Chiodo and

Bunney, 1983; Grace and Bunney, 1984; Grace and Bunney, 1986; Lidsky and Banerjee,

1993). Augmented DA activity in the striatum can further result from environment

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novelty and exploratory behavior (Legault and Wise, 2001). Thus, enhanced striatal DA transmission due to D2 autoreceptor occupancy, environmental novelty and exploration may lead to higher D1-like receptor stimulation and thus increased excitability within the striatonigral pathway (Surmeier et al., 2007). Based on our current behavioral findings, concurrent plasticity in the striatonigral pathway (via D1-like receptors activation) as well as in the striatopallidal pathway (via D2 receptor antagonism) may be necessary for development of catalepsy sensitization and conditioning.

5.4 Dopamine and the Action of Antipsychotic Drugs

Dysregulation of the DA system is implicated in the development of psychosis, a clinical hallmark of schizophrenia (Abi-Dargham et al., 2000; Abi-Dargham et al., 2009;

Grace, 2000; Kegeles et al., 2010; Lodge and Grace, 2011; Wadenberg et al., 2001).

Drugs effective in treating psychosis block the D2 receptors (Carlsson and Lindqvist,

1963; Kapur and Remington, 2001; Seeman et al., 1976; Wadenberg et al., 2001); psychomotor stimulant drugs like AMPH that facilitate the release of DA and increase

DA levels in the brain, can produce symptoms that mimic an acute psychotic state in normal subjects and can exacerbate psychosis in people with schizophrenia (Angrist et al., 1974; Friedman and Sienkiewicz, 1991); and drug-naïve subjects diagnosed with schizophrenia exhibit greater striatal DA release (measured by enhanced displacement of

D2 receptor ligands) in response to an acute AMPH challenge (Kegeles et al., 2010;

Laruelle, 2000; Laruelle et al., 1996).

The time course of the therapeutic action of antipsychotic drugs in people with

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schizophrenia is variable, with psychotic symptoms persisting sometimes for weeks depending on the length of previously untreated symptoms (Emsley et al., 2006; Grace et al., 1997; Miller, 1987). Thus, it may be more appropriate to think of psychosis, not as a direct result of a hyperactive DA system, but as a result of sensitization-like learning processes in which DA system dysregulation provides a symptomological milieu for acquisition of psychosis (Featherstone et al., 2007; Lodge and Grace, 2008; Lodge and

Grace, 2012; Peleg-Raibstein et al., 2008). This notion is supported by the findings that the DA response to acute AMPH observed in people with schizophrenia resembles that of healthy volunteers with a history of multiple psychomotor stimulant exposures

(Boileau et al., 2006).

Endogenous and drug-induced psychosis are not instantaneous phenomena but instead are slow and progressive in nature possibly due to gradual alterations produced by the DA system (Laruelle, 2000; Lieberman et al., 1997; Yung and McGorry, 1996).

Elevated DA may allow for irrelevant stimuli to acquire excessive incentive value leading to psychotic symptoms (Beninger, 1983; Crow, 1979; Gray et al., 1991; Kapur, 2003;

McKenna, 1987; Miller, 1976; Murray, 2010; Schmidt and Beninger, 2006). A sensitization-like mechanism may also be responsible for the variable-onset of therapeutic action associated with antipsychotics. Evidence shows that repeated treatment with antipsychotics leads to gradual changes within the DA system (Grace and Bunney,

1986; Meshul et al., 1992; Valenti et al., 2011; Valenti and Grace, 2010) and a progressive reduction in incentive motivational properties of reward-related stimuli

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(Boye and Rompre, 2000; Franklin and McCoy, 1979; Wise, 1982). Therefore, repeated treatment with antipsychotic drugs in people experiencing psychosis may result in a loss of incentive value previously attributed to irrelevant stimuli leading to a gradual de- construction of psychotic symptoms (Beninger, 1983; Miller, 1976).

In conclusion, similar to the behavioral sensitization produced by psychomotor stimulant drugs, the acquisition and expression of sensitization to haloperidol is also conditional on the presence of drug-associated environmental stimuli. Our findings provide further insight into the current understanding of learning processes involved in the action of antipsychotic drugs and the dissociable roles of D1-like, D2 and D3 receptors in controlling their effects. Interestingly, the dependence of antipsychotic drugs on environmental stimuli for their action may provide a basis for understanding the variable-onset of therapeutic benefit (Emsley et al., 2006) and why the schizophrenia outpatient population remains vulnerable to psychosis relapse despite medication adherence (Chen et al., 2010; Crow et al., 1986; Hogarty and Ulrich, 1998; Robinson,

2011).

The End.

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5.5 References

Abi-Dargham A, Rodenhiser J, Printz D, Zea-Ponce Y, et al. (2000). Increased baseline occupancy of D2 receptors by dopamine in schizophrenia. Proc Natl Acad Sci U S A, 97(14): 8104-8109. Abi-Dargham A, van de Giessen E, Slifstein M, Kegeles LS, et al. (2009). Baseline and amphetamine-stimulated dopamine activity are related in drug-naive schizophrenic subjects. Biol Psychiatry, 65(12): 1091-1093. Amtage J, Schmidt WJ (2003). Context-dependent catalepsy intensification is due to classical conditioning and sensitization. Behav Pharmacol, 14(7): 563-567. Anagnostaras SG, Robinson TE (1996). Sensitization to the psychomotor stimulant effects of amphetamine: modulation by associative learning. Behav Neurosci, 110(6): 1397-1414. Angrist B, Sathananthan G, Wilk S, Gershon S (1974). Amphetamine psychosis: behavioral and biochemical aspects. J Psychiatr Res, 11: 13-23. Badiani A, Oates MM, Day HE, Watson SJ, et al. (1998). Amphetamine-induced behavior, dopamine release, and c-fos mRNA expression: modulation by environmental novelty. J Neurosci, 18(24): 10579-10593. Badiani A, Oates MM, Day HE, Watson SJ, et al. (1999). Environmental modulation of amphetamine-induced c-fos expression in D1 versus D2 striatal neurons. Behav Brain Res, 103(2): 203-209. Banasikowski TJ, Bespalov A, Drescher K, Behl B, et al. (2010). Double dissociation of the effects of haloperidol and the dopamine D3 receptor antagonist ABT-127 on acquisition vs. expression of cocaine-conditioned activity in rats. J Pharmacol Exp Ther, 335(2): 506-515. Banasikowski TJ, MacLeod LS, Beninger RJ (2012). Comparison of nafadotride, CNQX, and haloperidol on acquisition versus expression of amphetamine- conditioned place preference in rats. Behav Pharmacol, 23(1): 89-97. Bardo MT, Bowling SL, Pierce RC (1990). Changes in locomotion and dopamine neurotransmission following amphetamine, haloperidol, and exposure to novel environmental stimuli. Psychopharmacology (Berl), 101(3): 338-343. Bateup HS, Santini E, Shen W, Birnbaum S, et al. (2010). Distinct subclasses of medium spiny neurons differentially regulate striatal motor behaviors. Proc Natl Acad Sci U S A, 107(33): 14845-14850. 141

Beninger RJ (1983). The role of dopamine in locomotor activity and learning. Brain Res, 287(2): 173-196. Beninger RJ, Banasikowski TJ (2008). Dopaminergic mechanism of reward-related incentive learning: focus on the dopamine D(3) receptor. Neurotox Res, 14(1): 57- 70. Beninger RJ, Hahn BL (1983). Pimozide blocks establishment but not expression of amphetamine-produced environment-specific conditioning. Science, 220(4603): 1304-1306. Beninger RJ, Herz RS (1986). Pimozide blocks establishment but not expression of cocaine-produced environment-specific conditioning. Life Sci, 38(15): 1425-1431. Beninger RJ, Ranaldi R (1992). The effects of amphetamine, apomorphine, SKF 38393, quinpirole and bromocriptine on responding for conditioned reward in rats. Behav Pharmacol, 3(2): 155-163. Bertran-Gonzalez J, Bosch C, Maroteaux M, Matamales M, et al. (2008). Opposing patterns of signaling activation in dopamine D1 and D2 receptor-expressing striatal neurons in response to cocaine and haloperidol. J Neurosci, 28(22): 5671- 5685. Bespalov A, Jongen-Relo AL, van Gaalen M, Harich S, et al. (2007). Habituation deficits induced by metabotropic glutamate receptors 2/3 receptor blockade in mice: reversal by antipsychotic drugs. J Pharmacol Exp Ther, 320(2): 944-950. Beutler LR, Wanat MJ, Quintana A, Sanz E, et al. (2011). Balanced NMDA receptor activity in dopamine D1 receptor (D1R)- and D2R-expressing medium spiny neurons is required for amphetamine sensitization. Proc Natl Acad Sci U S A, 108(10): 4206-4211. Bezard E, Ferry S, Mach U, Stark H, et al. (2003). Attenuation of levodopa-induced dyskinesia by normalizing dopamine D3 receptor function. Nat Med, 9(6): 762- 767. Bhat RV, Cole AJ, Baraban JM (1992). Role of monoamine systems in activation of zif268 by cocaine. J Psychiatry Neurosci, 17(3): 94-102. Boileau I, Dagher A, Leyton M, Gunn RN, et al. (2006). Modeling sensitization to stimulants in humans: an [11C]raclopride/positron emission tomography study in healthy men. Arch Gen Psychiatry, 63(12): 1386-1395.

142

Boivin GA, Beninger RJ (2008). Differential effects of dopamine and AMPA receptor antagonists on the expression of conditioned avoidance responding in rats. Behav Neurosci, 122(2): 377-384. Bordet R, Ridray S, Carboni S, Diaz J, et al. (1997). Induction of dopamine D3 receptor expression as a mechanism of behavioral sensitization to levodopa. Proc Natl Acad Sci U S A, 94(7): 3363-3367. Bouton ME (2004). Context and behavioral processes in extinction. Learn Mem, 11(5): 485-494. Boye SM, Rompre PP (2000). Behavioral evidence of depolarization block of dopamine neurons after chronic treatment with haloperidol and clozapine. J Neurosci, 20(3): 1229-1239. Bromberg-Martin ES, Matsumoto M, Hikosaka O (2010). Dopamine in motivational control: rewarding, aversive, and alerting. Neuron, 68(5): 815-834. Bunney BS, Grace AA (1978). Acute and chronic haloperidol treatment: comparison of effects on nigral dopaminergic cell activity. Life Sci, 23(16): 1715-1727. Capper-Loup C, Canales JJ, Kadaba N, Graybiel AM (2002). Concurrent activation of dopamine D1 and D2 receptors is required to evoke neural and behavioral phenotypes of cocaine sensitization. J Neurosci, 22(14): 6218-6227. Carey RJ (1987). Conditioning and the delayed onset of a haloperidol-induced behavioral effect. Biol Psychiatry, 22(3): 269-277. Carlsson A, Lindqvist M (1963). Effect Of Chlorpromazine Or Haloperidol On Formation Of 3methoxytyramine And Normetanephrine In Mouse Brain. Acta Pharmacol Toxicol (Copenh), 20: 140-144. Centonze D, Usiello A, Costa C, Picconi B, et al. (2004). Chronic haloperidol promotes corticostriatal long-term potentiation by targeting dopamine D2L receptors. J Neurosci, 24(38): 8214-8222. Cervo L, Carnovali F, Stark JA, Mennini T (2003). Cocaine-seeking behavior in response to drug-associated stimuli in rats: involvement of D3 and D2 dopamine receptors. Neuropsychopharmacology, 28(6): 1150-1159. Cervo L, Cocco A, Petrella C, Heidbreder CA (2007). Selective antagonism at dopamine D3 receptors attenuates cocaine-seeking behaviour in the rat. Int J Neuropsychopharmacol, 10(2): 167-181.

143

Cervo L, Samanin R (1995). Effects of dopaminergic and glutamatergic receptor antagonists on the acquisition and expression of cocaine conditioning place preference. Brain Res, 673(2): 242-250. Chen EY, Hui CL, Lam MM, Chiu CP, et al. (2010). Maintenance treatment with quetiapine versus discontinuation after one year of treatment in patients with remitted first episode psychosis: randomised controlled trial. Bmj, 341: c4024. Chiang Y-C, Chen P-C, Chen J-C (2003). D3 dopamine receptors are down-regulated in amphetamine sensitized rats and their putative antagonists modulate the locomotor sensitization to amphetamine. Brain Research, 972(1-2): 159. Chiodo LA, Bunney BS (1983). Typical and atypical neuroleptics: differential effects of chronic administration on the activity of A9 and A10 midbrain dopaminergic neurons. J Neurosci, 3(8): 1607-1619. Crow TJ (1979). Catecholamine reward pathways and schizophrenia: the mechanism of the antipsychotic effect and the site of the primary disturbance. Fed Proc, 38(11): 2462-2467. Crow TJ, MacMillan JF, Johnson AL, Johnstone EC (1986). A randomised controlled trial of prophylactic neuroleptic treatment. Br J Psychiatry, 148: 120-127. Dalley JW, Laane K, Theobald DE, Armstrong HC, et al. (2005). Time-limited modulation of appetitive Pavlovian memory by D1 and NMDA receptors in the nucleus accumbens. Proc Natl Acad Sci U S A, 102(17): 6189-6194. Di Ciano P, Underwood RJ, Hagan JJ, Everitt BJ (2003). Attenuation of cue- controlled cocaine-seeking by a selective D3 dopamine receptor antagonist SB- 277011-A. Neuropsychopharmacology, 28(2): 329-338. Dias FR, Carey RJ, Carrera MP (2006). Conditioned locomotion induced by unilateral intrastriatal administration of apomorphine: D(2) receptor activation is critical but not the expression of the unconditioned response. Brain Res, 1083(1): 85-95. Dragunow M, Currie RW, Faull RL, Robertson HA, et al. (1989). Immediate-early genes, kindling and long-term potentiation. Neurosci Biobehav Rev, 13(4): 301- 313. Dragunow M, Robertson HA (1988). Localization and induction of c-fos protein-like immunoreactive material in the nuclei of adult mammalian neurons. Brain Res, 440(2): 252-260. El-Ghundi M, Fletcher PJ, Drago J, Sibley DR, et al. (1999). Spatial learning deficit in dopamine D(1) receptor knockout mice. Eur J Pharmacol, 383(2): 95-106.

144

Emsley R, Rabinowitz J, Medori R (2006). Time course for antipsychotic treatment response in first-episode schizophrenia. Am J Psychiatry, 163(4): 743-745. Featherstone RE, Kapur S, Fletcher PJ (2007). The amphetamine-induced sensitized state as a model of schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry, 31(8): 1556-1571. Fenu S, Acquas E, Di Chiara G (2001). Role of striatal acetylcholine on dopamine D1 receptor agonist-induced turning behavior in 6-hydroxydopamine lesioned rats: a microdialysis-behavioral study. Neurol Sci, 22(1): 63-64. Ferguson SM, Eskenazi D, Ishikawa M, Wanat MJ, et al. (2011). Transient neuronal inhibition reveals opposing roles of indirect and direct pathways in sensitization. Nat Neurosci, 14(1): 22-24. Fontana D, Post RM, Weiss SR, Pert A (1993). The role of D1 and D2 dopamine receptors in the acquisition and expression of cocaine-induced conditioned increases in locomotor behavior. Behav Pharmacol, 4(4): 375-387. Franklin KB, McCoy SN (1979). Pimozide-induced extinction in rats: stimulus control of responding rules out motor deficit. Pharmacol Biochem Behav, 11(1): 71-75. Fricks-Gleason AN, Khalaj AJ, Marshall JF (2012). Dopamine D1 receptor antagonism impairs extinction of cocaine-cue memories. Behav Brain Res, 226(1): 357-360. Friedman A, Sienkiewicz J (1991). Psychotic complications of long-term levodopa treatment of Parkinson's disease. Acta Neurol Scand, 84(2): 111-113. Gilbert JG, Newman AH, Gardner EL, Ashby CR, Jr., et al. (2005). Acute administration of SB-277011A, NGB 2904, or BP 897 inhibits cocaine cue- induced reinstatement of drug-seeking behavior in rats: role of dopamine D3 receptors. Synapse, 57(1): 17-28. Glimcher PW (2011). Understanding dopamine and reinforcement learning: the dopamine reward prediction error hypothesis. Proc Natl Acad Sci U S A, 108 Suppl 3: 15647-15654. Goto Y, Grace AA (2005). Dopamine-dependent interactions between limbic and prefrontal cortical plasticity in the nucleus accumbens: disruption by cocaine sensitization. Neuron, 47(2): 255-266. Grace AA (2000). Gating of information flow within the limbic system and the pathophysiology of schizophrenia. Brain Res Brain Res Rev, 31(2-3): 330-341.

145

Grace AA, Bunney BS (1984). The control of firing pattern in nigral dopamine neurons: burst firing. J Neurosci, 4(11): 2877-2890. Grace AA, Bunney BS (1986). Induction of depolarization block in midbrain dopamine neurons by repeated administration of haloperidol: analysis using in vivo intracellular recording. J Pharmacol Exp Ther, 238(3): 1092-1100. Grace AA, Bunney BS, Moore H, Todd CL (1997). Dopamine-cell depolarization block as a model for the therapeutic actions of antipsychotic drugs. Trends Neurosci, 20(1): 31-37. Granado N, Ortiz O, Suarez LM, Martin ED, et al. (2008). D1 but not D5 dopamine receptors are critical for LTP, spatial learning, and LTP-Induced arc and zif268 expression in the hippocampus. Cereb Cortex, 18(1): 1-12. Gray JA, Feldon J, Rawlins JNP, Hemsley DR, et al. (1991). The neuropsychology of schizophrenia. Behavioral and Brain Sciences, 14(01): 1-20. Graybiel AM, Moratalla R, Robertson HA (1990). Amphetamine and cocaine induce drug-specific activation of the c-fos gene in striosome-matrix compartments and limbic subdivisions of the striatum. Proc Natl Acad Sci U S A, 87(17): 6912-6916. Hitchcott PK, Phillips GD (1998). Effects of intra-amygdala R(+) 7-OH-DPAT on intra-accumbens d-amphetamine-associated learning. II. Instrumental conditioning. Psychopharmacology (Berl), 140(3): 310-318. Hogarty GE, Ulrich RF (1998). The limitations of antipsychotic medication on schizophrenia relapse and adjustment and the contributions of psychosocial treatment. J Psychiatr Res, 32(3-4): 243-250. Hollerman JR, Schultz W (1998). Dopamine neurons report an error in the temporal prediction of reward during learning. Nat Neurosci, 1(4): 304-309. Hooks MS, Kalivas PW (1995). The role of mesoaccumbens--pallidal circuitry in novelty-induced behavioral activation. Neuroscience, 64(3): 587-597. Horvitz JC (2000). Mesolimbocortical and nigrostriatal dopamine responses to salient non-reward events. Neuroscience, 96(4): 651-656. Kalivas PW, Stewart J (1991). Dopamine transmission in the initiation and expression of drug- and stress-induced sensitization of motor activity. Brain Res Brain Res Rev, 16(3): 223-244. Kapur S (2003). Psychosis as a state of aberrant salience: a framework linking biology, phenomenology, and pharmacology in schizophrenia. Am J Psychiatry, 160(1): 13-23. 146

Kapur S, Remington G (2001). Dopamine D(2) receptors and their role in atypical antipsychotic action: still necessary and may even be sufficient. Biol Psychiatry, 50(11): 873-883. Kegeles LS, Abi-Dargham A, Frankle WG, Gil R, et al. (2010). Increased synaptic dopamine function in associative regions of the striatum in schizophrenia. Arch Gen Psychiatry, 67(3): 231-239. Kiyatkin EA, Rebec GV (1996). Dopaminergic modulation of glutamate-induced excitations of neurons in the neostriatum and nucleus accumbens of awake, unrestrained rats. J Neurophysiol, 75(1): 142-153. Klein A, Schmidt WJ (2003). Catalepsy intensifies context-dependently irrespective of whether it is induced by intermittent or chronic dopamine deficiency. Behav Pharmacol, 14(1): 49-53. Kuribara H (1995). Dopamine D1 receptor antagonist SCH 23390 retards methamphetamine sensitization in both combined administration and early posttreatment schedules in mice. Pharmacol Biochem Behav, 52(4): 759-763. LaHoste GJ, Henry BL, Marshall JF (2000). Dopamine D1 receptors synergize with D2, but not D3 or D4, receptors in the striatum without the involvement of action potentials. J Neurosci, 20(17): 6666-6671. LaHoste GJ, Yu J, Marshall JF (1993). Striatal Fos expression is indicative of dopamine D1/D2 synergism and receptor supersensitivity. Proc Natl Acad Sci U S A, 90(16): 7451-7455. Laruelle M (2000). The role of endogenous sensitization in the pathophysiology of schizophrenia: implications from recent brain imaging studies. Brain Res Brain Res Rev, 31(2-3): 371-384. Laruelle M, Abi-Dargham A, van Dyck CH, Gil R, et al. (1996). Single photon emission computerized tomography imaging of amphetamine-induced dopamine release in drug-free schizophrenic subjects. Proc Natl Acad Sci U S A, 93(17): 9235-9240. Le Foll B, Diaz J, Sokoloff P (2003). Increased dopamine D3 receptor expression accompanying behavioral sensitization to nicotine in rats. Synapse, 47(3): 176- 183. Le Foll B, Frances H, Diaz J, Schwartz JC, et al. (2002). Role of the dopamine D3 receptor in reactivity to cocaine-associated cues in mice. Eur J Neurosci, 15(12): 2016-2026.

147

Legault M, Wise RA (2001). Novelty-evoked elevations of nucleus accumbens dopamine: dependence on impulse flow from the ventral subiculum and glutamatergic neurotransmission in the ventral tegmental area. Eur J Neurosci, 13(4): 819-828. Lemon N, Manahan-Vaughan D (2006). Dopamine D1/D5 receptors gate the acquisition of novel information through hippocampal long-term potentiation and long-term depression. J Neurosci, 26(29): 7723-7729. Li M, Fletcher PJ, Kapur S (2007). Time course of the antipsychotic effect and the underlying behavioral mechanisms. Neuropsychopharmacology, 32(2): 263-272. Li Y, Acerbo MJ, Robinson TE (2004). The induction of behavioural sensitization is associated with cocaine-induced structural plasticity in the core (but not shell) of the nucleus accumbens. Eur J Neurosci, 20(6): 1647-1654. Liang J, Zheng X, Chen J, Li Y, et al. (2011). Roles of BDNF, dopamine D3 receptors, and their interactions in the expression of morphine-induced context-specific locomotor sensitization. European Neuropsychopharmacology, In Press, Corrected Proof. Lidsky TI, Banerjee SP (1993). Acute administration of haloperidol enhances dopaminergic transmission. J Pharmacol Exp Ther, 265(3): 1193-1198. Lieberman JA, Sheitman BB, Kinon BJ (1997). Neurochemical sensitization in the pathophysiology of schizophrenia: deficits and dysfunction in neuronal regulation and plasticity. Neuropsychopharmacology, 17(4): 205-229. Ljungberg T (1990). Differential attenuation of water intake and water-rewarded operant responding by repeated administration of haloperidol and SCH 23390 in the rat. Pharmacol Biochem Behav, 35(1): 111-115. Lodge DJ, Grace AA (2008). Amphetamine activation of hippocampal drive of mesolimbic dopamine neurons: a mechanism of behavioral sensitization. J Neurosci, 28(31): 7876-7882. Lodge DJ, Grace AA (2011). Hippocampal dysregulation of dopamine system function and the pathophysiology of schizophrenia. Trends Pharmacol Sci, 32(9): 507-513. Lodge DJ, Grace AA (2012). Divergent activation of ventromedial and ventrolateral dopamine systems in animal models of amphetamine sensitization and schizophrenia. Int J Neuropsychopharmacol: 1-8.

148

Maggio R, Aloisi G, Silvano E, Rossi M, et al. (2009). Heterodimerization of dopamine receptors: new insights into functional and therapeutic significance. Parkinsonism Relat Disord, 15 Suppl 4: S2-7. Mazurski EJ, Beninger RJ (1991). Effects of selective drugs for dopaminergic D1 and D2 receptors on conditioned locomotion in rats. Psychopharmacology (Berl), 105(1): 107-112. McKenna PJ (1987). Pathology, phenomenology and the dopamine hypothesis of schizophrenia. Br J Psychiatry, 151: 288-301. Meshul CK, Janowsky A, Casey DE, Stallbaumer RK, et al. (1992). Effect of haloperidol and clozapine on the density of "perforated" synapses in caudate, nucleus accumbens, and medial prefrontal cortex. Psychopharmacology (Berl), 106(1): 45-52. Miller R (1976). Schizophrenic psychology, associative learning and the role of forebrain dopamine. Med Hypotheses, 2(5): 203-211. Miller R (1987). The time course of neuroleptic therapy for psychosis: role of learning processes and implications for concepts of psychotic illness. Psychopharmacology (Berl), 92(4): 405-415. Murphy CA, Feldon J (2001). Interactions between environmental stimulation and antipsychotic drug effects on forebrain c-fos activation. Neuroscience, 104(3): 717-730. Murray GK (2010). The emerging biology of delusions. Psychol Med, 41(1): 7-13. Neisewander JL, Fuchs RA, Tran-Nguyen LT, Weber SM, et al. (2004). Increases in dopamine D3 receptor binding in rats receiving a cocaine challenge at various time points after cocaine self-administration: implications for cocaine-seeking behavior. Neuropsychopharmacology, 29(8): 1479-1487. O'Carroll CM, Morris RG (2004). Heterosynaptic co-activation of glutamatergic and dopaminergic afferents is required to induce persistent long-term potentiation. Neuropharmacology, 47(3): 324-332. Parker JG, Zweifel LS, Clark JJ, Evans SB, et al. (2010). Absence of NMDA receptors in dopamine neurons attenuates dopamine release but not conditioned approach during Pavlovian conditioning. Proc Natl Acad Sci U S A, 107(30): 13491-13496. Pascoli V, Turiault M, Luscher C (2012). Reversal of cocaine-evoked synaptic potentiation resets drug-induced adaptive behaviour. Nature, 481(7379): 71.

149

Paulson PE, Robinson TE (1995). Amphetamine-induced time-dependent sensitization of dopamine neurotransmission in the dorsal and ventral striatum: a microdialysis study in behaving rats. Synapse, 19(1): 56-65. Peleg-Raibstein D, Knuesel I, Feldon J (2008). Amphetamine sensitization in rats as an animal model of schizophrenia. Behav Brain Res, 191(2): 190-201. Pert A, Post R, Weiss SR (1990). Conditioning as a critical determinant of sensitization induced by psychomotor stimulants. NIDA Res Monogr, 97: 208-241. Post RM, Rose H (1976). Increasing effects of repetitive cocaine administration in the rat. Nature, 260(5553): 731-732. Redgrave P, Gurney K, Reynolds J (2008). What is reinforced by phasic dopamine signals? Brain Res Rev, 58(2): 322-339. Reynolds JN, Wickens JR (2002). Dopamine-dependent plasticity of corticostriatal synapses. Neural Netw, 15(4-6): 507-521. Robertson HA, Paul ML, Moratalla R, Graybiel AM (1991). Expression of the immediate early gene c-fos in basal ganglia: induction by dopaminergic drugs. Can J Neurol Sci, 18(3 Suppl): 380-383. Robinson DG (2011). Medication adherence and relapse in recent-onset psychosis. Am J Psychiatry, 168(3): 240-242. Ruob C, Weiner I, Feldon J (1998). Haloperidol-induced potentiation of latent inhibition: interaction with parameters of conditioning. Behav Pharmacol, 9(3): 245-253. Scarselli M, Novi F, Schallmach E, Lin R, et al. (2001). D2/D3 dopamine receptor heterodimers exhibit unique functional properties. J Biol Chem, 276(32): 30308- 30314. Schmidt WJ, Beninger RJ (2006). Behavioural sensitization in addiction, schizophrenia, Parkinson's disease and dyskinesia. Neurotox Res, 10(2): 161-166. Seeman P, Lee T, Chau-Wong M, Wong K (1976). Antipsychotic drug doses and neuroleptic/dopamine receptors. Nature, 261(5562): 717-719. Segal DS, Mandell AJ (1974). Long-term administration of d-amphetamine: progressive augmentation of motor activity and stereotypy. Pharmacol Biochem Behav, 2(2): 249-255. Shen W, Flajolet M, Greengard P, Surmeier DJ (2008). Dichotomous dopaminergic control of striatal synaptic plasticity. Science, 321(5890): 848-851. 150

Singer BF, Tanabe LM, Gorny G, Jake-Matthews C, et al. (2009). Amphetamine- induced changes in dendritic morphology in rat forebrain correspond to associative drug conditioning rather than nonassociative drug sensitization. Biol Psychiatry, 65(10): 835-840. Smith-Roe SL, Kelley AE (2000). Coincident activation of NMDA and dopamine D1 receptors within the nucleus accumbens core is required for appetitive instrumental learning. J Neurosci, 20(20): 7737-7742. Surmeier DJ, Ding J, Day M, Wang Z, et al. (2007). D1 and D2 dopamine-receptor modulation of striatal glutamatergic signaling in striatal medium spiny neurons. Trends Neurosci, 30(5): 228-235. Trimble KM, Bell R, King DJ (2002). Effects of the selective dopamine D(1) antagonists NNC 01-0112 and SCH 39166 on latent inhibition in the rat. Physiol Behav, 77(1): 115-123. Ungless MA, Whistler JL, Malenka RC, Bonci A (2001). Single cocaine exposure in vivo induces long-term potentiation in dopamine neurons. Nature, 411(6837): 583-587. Valenti O, Cifelli P, Gill KM, Grace AA (2011). Antipsychotic drugs rapidly induce dopamine neuron depolarization block in a developmental rat model of schizophrenia. J Neurosci, 31(34): 12330-12338. Valenti O, Grace AA (2010). Antipsychotic drug-induced increases in ventral tegmental area dopamine neuron population activity via activation of the nucleus accumbens-ventral pallidum pathway. Int J Neuropsychopharmacol, 13(7): 845- 860. Vezina P (1996). D1 dopamine receptor activation is necessary for the induction of sensitization by amphetamine in the ventral tegmental area. J Neurosci, 16(7): 2411-2420. Vezina P, Queen AL (2000). Induction of locomotor sensitization by amphetamine requires the activation of NMDA receptors in the rat ventral tegmental area. Psychopharmacology (Berl), 151(2-3): 184-191. Vezina P, Stewart J (1989). The effect of dopamine receptor blockade on the development of sensitization to the locomotor activating effects of amphetamine and morphine. Brain Res, 499(1): 108-120. Wadenberg ML, Soliman A, VanderSpek SC, Kapur S (2001). Dopamine D(2) receptor occupancy is a common mechanism underlying animal models of

151

antipsychotics and their clinical effects. Neuropsychopharmacology, 25(5): 633- 641. Walters JR, Bergstrom DA, Carlson JH, Chase TN, et al. (1987). D1 dopamine receptor activation required for postsynaptic expression of D2 agonist effects. Science, 236(4802): 719-722. Weiner I, Feldon J (1987). Facilitation of latent inhibition by haloperidol in rats. Psychopharmacology (Berl), 91(2): 248-253. Wiecki TV, Riedinger K, von Ameln-Mayerhofer A, Schmidt WJ, et al. (2009). A neurocomputational account of catalepsy sensitization induced by D2 receptor blockade in rats: context dependency, extinction, and renewal. Psychopharmacology (Berl), 204(2): 265-277. Wise RA (1982). Neuroleptics and operant behavior: The anhedonia hypothesis. Behavioral and Brain Sciences, 5(01): 39-53. Wise RA (2004). Dopamine, learning and motivation. Nat Rev Neurosci, 5(6): 483-494. Wise RA, Carlezon WA, Jr. (1994). Attenuation of the locomotor-sensitizing effects of the D2 dopamine agonist bromocriptine by either the D1 antagonist SCH 23390 or the D2 antagonist raclopride. Synapse, 17(3): 155-159. Wise RA, Spindler J, deWit H, Gerberg GJ (1978). Neuroleptic-induced "anhedonia" in rats: pimozide blocks reward quality of food. Science, 201(4352): 262-264. Yung AR, McGorry PD (1996). The prodromal phase of first-episode psychosis: past and current conceptualizations. Schizophr Bull, 22(2): 353-370.

152