Neural Encoding of Olfactory Recognition Memory

Journal of Reproduction and Development, Vol. 51, No. 5, 2005

—Review—

Gabriela SÁNCHEZ-ANDRADE1), Bronwen M JAMES1) and Keith M KENDRICK1)

1)Cognitive and Behavioural Neuroscience, The Babraham Institute, Babraham, Cambridge CB2 4AT, UK

Abstract. Our work with both sheep and mouse models has revealed many of the neural substrates and signalling pathways involved in olfactory recognition memory in the main olfactory system. A distributed neural system is required for initial memory formation and its short-term retention–the olfactory bulb, piriform and entorhinal cortices and hippocampus. Following memory consolidation, after 8 h or so, only the olfactory bulb and piriform cortex appear to be important for effective recall. Similarly, whereas the glutamate–NMDA/AMPA receptor–nitric oxide (NO)–cyclic GMP signalling pathway is important for memory formation it is not involved in recall post-consolidation. Here, within the olfactory bulb, up-regulation of class 1 metabotropic glutamate receptors appears to maintain the enhanced sensitivity at the mitral to granule cell synapses required for effective memory recall. Recently we have investigated whether fluctuating sex hormone levels during the oestrous cycle modulate olfactory recognition memory and the different neural substrates and signalling pathways involved. These studies have used two robust models of social olfactory memory in the mouse which either involve social or non social odours (habituation-dishabituation and social transmission of food preference tasks). In both cases significant improvement of learning retention occurs when original learning takes place during the proestrus phase of the ovarian cycle. This is probably the result of oestrogen changes at this time since transgenic mice lacking functional expression of oestrogen receptors (ERα and ERβ, the two main oestrogen receptor sub-types) have shown problems in social recognition. Therefore, oestrogen appears to act at the level of the olfactory bulb by modulating both noradrenaline and the glutamate/NO signalling pathway. Key words: Olfactory bulb, Olfactory memory, Olfactory recognition, Social learning, Nitric oxide (NO), Oestrous cycle, Sheep, Mouse, Social transmission of food preference (J. Reprod. Dev. 51: 547–558, 2005)

or several decades, olfactory learning has been has well characterised pathways and structural used as a model for the study of memory. This organisation which offers many advantages in is not only because mammalian olfactory learning defining how processing occurs at a systems level. shows a number of features common to the human This makes it feasible to study plasticity at all levels declarative memory system one—trial learning, in the system, from the initial sensory detection of rapid encoding, long lasting memories and a high odourant molecules at the level of the olfactory capacity for information storage–but also because receptors right through to higher cortical olfactory memory models represent a relatively processing. simple form of learning and the olfactory system There are many different forms of olfactory learning and memory in diverse behavioural Accepted for publication: June 2, 2005 Correspondence: K.M. Kendrick contexts involving different mechanisms within the (e-mail: [email protected]) olfactory system. Olfactory learning can be divided 548 SÁNCHEZ-ANDRADE et al. into socially and non-socially motivated tasks–a Social recognition tests in rodents rely on the distinction that has been shown to be important by intrinsic motivation they have for investigating work on the oxytocin knockout mouse [1] where novel conspecifics when introduced into familiar only deficits in social memory have been observed. territory or home cage. Under these circumstances The importance of olfaction and olfactory memory they will investigate a novel conspecific more than on socially mediated behaviours is very a familiar one since they are already acquainted considerable since many mammalian species rely with the odour cues of the latter rather than the upon smell to obtain information about their former [14]. Odour cues such as soiled bedding or environment, particularly their social one. Our urine from familiar animals are sufficient for the work for the past 15 years has focused on the recognition process [15]. In the social recognition involvement of the main olfactory system in social test [16], a rat or mouse is exposed to a novel olfactory learning in the context of lamb-mother stimulus animal (generally juveniles) on two ewe bonding via odour recognition [2, 3] and social occasions, separated by a given time interval. The recognition of individuals [4, 5] and social reduction of investigation time during the second transmission of food preference [6, 7] in rodents. encounter is taken as evidence for social memory. These paradigms represent good ethologically Introduction of a new fresh stimulus animal relevant models for the study of social olfactory triggers an intense investigation bout similar to the memory. Offspring recognition ensures that the initial encounter, proving that the reduced mother will preserve her own genes by limiting investigation is not due to generalised habituation. maternal investment to her own offspring. Social This paradigm has been used mainly as a model of recognition of conspecifics has functional short term memory since it was found to last no consequences in several social contexts [3, 8–11]. It longer than 1 h in studies using individually is crucial for the determination and maintenance of housed rats or mice [16–18]. However, recent social structure in many animal species, studies have shown that housing conditions affect particularly those living in small, stable social retention of these social memories since mice groups. With social transmission of food housed in groups, rather than individually, were preference, after smelling a novel food odour on a shown to remember a familiar juvenile for up to 7 forager’s breath rodents seek out that food days [19]. assuming that it is good to eat and suppressing We have combined two versions of the original their normal neophobic response. These models social recognition paradigm in mice. In the first also offer many advantages in studying olfactory instance a habituation-dishabituation paradigm has memory processes since learning is rapid and been employed [20, 21] where the test animal is robust and long-lasting memories can be formed exposed repeatedly (four trials) to a stimulus that are relatively simple to test experimentally. animal (anaesthetised adult) for 1 min with 10 min In sheep, the maternal ewe is able to recognise its inter-trial intervals. This results in a progressive offspring by identifying the individual body odour general reduction of investigation time of the of her own lamb(s). She forms a selective olfactory stimulus animal during consecutive trials using the memory for each of her offspring within 2–4 h of same stimulus animal (habituation). On a fifth trial, parturition and will consequently reject the a novel stimulus animal is introduced and the approach of any strange lamb [12]. Both the investigation time is restored to the original highest sensitive period for odour learning and the level (dishabituation). The second part of the maternal acceptance behaviour are dependent on paradigm, performed 24 h later, is a social the hormonal environment and are triggered by discrimination test. In this the test animal is given a feedback to the brain from mechanical stimulation binary choice between a previously encountered of the vagina and the cervix which normally occurs familiar and a novel conspecific [22]. A specific during parturition. Thus, a ewe can be induced to recognition memory, assessed by comparing the accept, and form, a new recognition memory for a differential amount of time spent investigating each strange lamb by simply mimicking birth through stimulus animal, is considered to have been formed administering a few minutes of artificial when the novel animal is investigated more than vaginocervical stimulation (VCS) for up to three the unfamiliar one. This second part of the protocol days post-partum [13]. allows the recognition memory test to be REVIEW: ENCODING OF OLFACTORY RECOGNITION MEMORY 549 performed for the familiar versus the unfamiliar one, a liquid-borne chemical substance known as stimulus animal in a single trial. This habituation/ pheromones; the second, a vast number of airborne discrimination social recognition paradigm chemicals with a large variety of structures. This is therefore allows testing both short- and long-term done by segregating odour and pheromone memory with a single experiment. detection using different sensory neurones [34] and A second test we used as a model of social neural pathways [35] in the brain. In the main olfactory memory in mice is the social transmission olfactory system, odours are detected by olfactory of food preference (STFP) paradigm, which uses a receptors (OR) in the olfactory epithelium (OE) of combination of biological and non-biological the nasal cavity where every odour has a unique odours in a social and ethologically relevant combination of responses from up to one thousand context. This ability of socially transmitting food different receptor types [34]. These signals are then preferences has been shown in different rodent relayed to the primary processing region for smell, species such as rats [7], mice (Mus musculus, [23, 24] the main olfactory bulb (MOB) and thence to and gerbils (Meriones unguiculatus [25]. During secondary and tertiary projection areas in the social contact between a recently fed animal cortex, limbic system and thalamus (see Fig. 1). (demonstrator) and a naïve conspecific (observer), Odour signals therefore ultimately reach higher olfactory cues pass from demonstrator to observer cortical areas involved in their conscious subsequently increasing the observer’s preference perception (attention and awareness), as well as for the food its demonstrator ate [7, 26]. Observer limbic areas, such as the amygdala and the animals are then presented with a binary choice of hypothalamus, that are involved in emotional and two equally novel foods. Learning is demonstrated motivational responses. as an increase in probability that the observer will In brief, mitral and tufted cells of the MOB (c.f. select the same food as that eaten by demonstrator, below for detailed description of the MOB) send over other foods [7]. A single brief interaction their axons, via the lateral olfactory tract (LOT), to between two mice or rats is sufficient to induce a the outer layer of the olfactory cortex [36]–the food preference that lasts for more than 6 days. piriform cortex, the olfactory tubercle, anterior This olfactory learning is believed to involve an olfactory nucleus and specific parts of the association between two odours present on the entorhinal cortex and amygdala [37–39]. From the demonstrator’s breath; the odour of the recently olfactory cortex, there is a substantial projection to eaten food and a natural constituent of rat’s breath, the mediodorsal thalamus and the ventral part of carbon disulfide (CS2). The association between the the submedius nucleus (SM), which in turn projects food odour and the carbon disulfide is both to the frontal cortex [40]. Disruption of these necessary and sufficient to support the connections results in deficits in complex but not in development of a food preference [27, 28]. simple odour discrimination tasks [41–43]. All three behavioural tasks, offspring and social Another small projection from the LOT goes to the recognition and STFP are dependent on the main lateral entorhinal cortex [44] and thence via the olfactory system and represent good models for perforant path to the dentate gyrus and CA1/CA3 studying memory formation in the main olfactory of the hippocampus. The olfactory cortex also system. In sheep, sectioning the vomeronasal nerve projects to the hypothalamus and the anterior organ does not disrupt either lamb recognition or cortical amygdala [45]. This connection could be maternal behaviour, whereas interfering with the the one responsible for the regulation of several of olfactory epithelium does [29]. In rodents, lesions the autonomic and neuroendocrine changes that in the olfactory bulb impair basic recognition occur in response to odours [46]. responses in male rats [30, 31]; and chemically induced anosmia can also block individual Learning in the MOS recognition [32, 33]. Odour learning in the main olfactory system, unlike that involving the accessory olfactory The Main Olfactory System (MOS) system [48], is thought to be more distributed, with The mammalian olfactory system is organised in secondary and tertiary olfactory processing regions such a way that it is capable of processing being involved. The activation and involvement of information from two different types of stimuli: these brain areas depends to some extent, on the 550 SÁNCHEZ-ANDRADE et al.

Fig. 1. Schematic diagram showing the major connections of the main olfactory system (not all projections are shown, nor the extensive reciprocal projections between olfactory areas). Olfactory memory involves changes occurring within the olfactory bulb as well as in its secondary (piriform cortex) and tertiary projection sites (entorhinal and orbitofrontal cortices and the hippocampus). Adapted from [47]. nature of the task being performed and on temporal factor (BDNF) and its receptor tyrosine receptor configurations. Dynamic changes occur in both kinase B (trk-B)–a measure of neural plasticity [52]. neural substrates and the molecular involvement in We recently carried out a systematic study of the formation, short- and long-term memory traces. temporal involvement of the major areas in the Studies of brain structure activation and reversible olfactory system, by temporally inactivating them inactivation of some key structures have provided (bilaterally) with either a local anaesthetic some valuable information. Initial memory (tetracaine) or a GABAa receptor agonist formation and its short-term retention seem to (muscimol) directly infused via microdialysis require a distributed neural system, involving the probes. This showed that the olfactory bulb, the olfactory bulb, piriform and entorhinal cortices and piriform and the entorhinal cortices are essential for the hippocampus. Our early work aimed to the formation of a normal selective recognition differentiate neural substrates controlling maternal memory. However, complete disruption, was only behaviour from those involved in olfactory seen with the olfactory bulb and the entorhinal memory formation, by comparing post-parturient cortex [53]. ewes exposed to their lambs for 30 min to There have been fewer studies regarding the hormonally primed animals that received VCS and involvement of neural substrates in social were not exposed to lambs. This showed that recognition in mice. Studies using oxytocin olfactory memory formation involved activation, receptor knockout mice have shown that a brief quantified by measurement of altered mRNA social exposure leads to regional activation of the expression of the immediately-early genes c-fos olfactory system as evidenced by increased c-fOS and/or zif/268, of the MOB, of piriform, entorhinal expression in the olfactory bulb, piriform cortex and orbitofrontal cortices and dentate gyrus [49]. and the medial amygdala [54]. Non-behavioural Studies comparing the response of intact and studies have also shown plasticity changes in both anosmic ewes to their own lamb have also shown the OB and piriform cortex resulting from OB that anosmic ewes, displaying maternal behaviour stimulation. High frequency stimulation of the but not individual lamb recognition, fail to show granule cell layer of the OB results in a selective increased expression in the MOB, piriform and long-term potentiation in both the OB itself and the frontal cortices and cortical amygdala [50, 51]. For piriform cortex [55]. Recently we have also shown subsequent memory consolidation, 4.5 h post- that consecutive 15 min periods of NMDA- partum, the piriform and entorhinal cortices stimulation to the MOB result in a long term seemed to play a key role, as shown by a study potentiation of NMDA-evoked neurotransmitter using expression of the brain derived neurotrophic release in both the olfactory bulb and the piriform REVIEW: ENCODING OF OLFACTORY RECOGNITION MEMORY 551 cortex after 4 h [4, 5]. glomeruli and their secondary dendrites into the Following memory consolidation, after external plexiform layer. Mitral cells use glutamate approximately 8 h, only the olfactory bulb and and possibly aspartate as transmitters and form piriform cortex appear to be important for recall. reciprocal dendro-dendritic synapses with GABA- For olfactory memory recall, there seems to be containing inhibitory interneurones at distinct distinct neural processes for short- and long-term levels. Granule cells, which are the most numerous memory. We have found that for ewes that had neurones in the olfactory bulb, in turn, make formed an olfactory memory of its lamb, synaptic contacts with the lateral secondary inactivation of the entorhinal cortex interfered with dendrites of the mitral cells in the external its recall in the short-term (4 h post-memory plexiform layer and are likely to be involved in the formation) but not in the long-term (12 h to 6 days control of mitral cell output [63, 64]. post-memory formation). However, inactivation of the olfactory bulb disrupted recall at both time Mechanisms of Olfactory Learning points [53]. Consistent with this, it has been shown In early postpartum period (2–4 h) a maternal that the piriform cortex is activated by recognition ewe’s selective recognition of her lamb can be of the lamb after both 4 h and 7 days contact shown by physiological changes in the olfactory durations [50]. In rodents, lesions of the entorhinal bulb that parallel observable behavioural changes. cortex result in deficits of short-term odour Electrophysiological recordings in the mitral cell memory [56–58]. Other studies using a modified layer of the olfactory bulb have shown that several version of the Coolidge effect, another social days postpartum 60% of cells respond olfactory learning test, have also shown that male preferentially to lamb’s odour compared to hamsters with ibotenic acid lesions of the prepartum period where no preferential activity perirhinal-entorhinal cortex have impaired short- was seen [65]. Of the cells that respond term social discrimination memory. Lesions to the significantly to these lamb odours the majority hippocampus, however, have no effect on short- (70%) do so equivalently to own or strange lambs, term recognition memory [59]. however 30% did respond preferentially to the Olfactory recognition memory, therefore, odours of the ewe’s own lamb. In addition, requires a more distributed neural network in the although lamb odours do not stimulate olfactory system for memory formation and short- neurotransmitter release in the olfactory bulb term memory, with the entorhinal cortex playing a prepartum, by 4 h postpartum, all lamb odours key role. As the memory consolidates, this network increase centrifugal transmitter release is reduced, and it seems that only the OB and (noradrenaline (NA) and acetylcholine (ACh)) piriform cortex are needed to maintain a memory while own lamb odours selectively increase trace throughout its existence. intrinsic glutamate and GABA release [12]. Associated with these neurochemical changes is The Main Olfactory Bulb (MOB) evidence for enhanced glutamatergic stimulation of As already discussed, many studies have GABA release from the granule cells following consistently reported that a general characteristic of learning [12]. olfactory memory is that it induces long-lasting How does the reorganisation of the olfactory neural changes at the level of the olfactory bulb. bulb take place? Both centrifugal and intrinsic The main olfactory bulb (MOB) has a simple olfactory bulb pathways play an important role in structure involving three main layered cell types establishing the plasticity changes in the olfactory (Fig. 2): periglomerular, mitral/tufted and granule bulb that lead to olfactory memory formation. cells. The olfactory nerve terminals synapse with Lesions of noradrenergic projections to the mitral and tufted cells in the MOB glomerulus, and olfactory bulb, or direct infusion of β-noradrenergic also with intrinsic periglomerular cells (PG) that antagonists, significantly reduce the number of circumscribe the glomeruli. PG cells (GABAergic animals developing the olfactory recognition or dopaminergic) mediate feedback inhibition memory [66]. Similar effects are seen following within the glomerulus and lateral (feedforward) treatments with cholinergic antagonists such as inhibition between glomeruli [60–62]. Mitral and scopolamine [67]. The result of the changes in NA tufted cells send their primary dendrites into the and ACh in the olfactory bulb is to reduce 552 SÁNCHEZ-ANDRADE et al.

Fig. 2. Left: Schematic representation of cell organisation in the main olfactory bulb (MOB), showing inputs (olfactory receptors, centrifugal pathways) and outputs (mitral cells) to the MOB and its intrinsic organisation and neurotransmitters. Right: Memory formation is associated with enhanced mitral-to- granule and periglomerular cell synapses to bias the network to respond to the learned odour. This involves both, increased acetylcholine (ACh) or noradrenaline (NA) from the centrifugal projections , that in turn reduce GABA release onto mitral cells and glutamate-evoked nitric oxide/cGMP release potentiating glutamate and subsequent GABA release from granule and periglomerular cells. GLU, glutamate; L-arg, L-arginine; L-cit, L-citrulline; nNOS, neuronal nitric oxide synthase; NO, nitric oxide; NMDA-R, NMDA receptor; AMPA/Kainate-R, AMPA/Kainate receptor; mGluR, metabotropic glutamate receptor.

GABAergic feedback inhibition from the granule release onto mitral cells and consequently cells to the mitral cells. The disinhibited mitral cells disinhibits them [12]. With incoming odour- become more active in their response to odours and induced activation of olfactory receptors in the through a glutamate–AMPA/NMDA receptor– presence of this disinhibited state, mitral cells nitric oxide–cyclic GMP signalling cascade there is increase their release of glutamate at their upregulation both of mitral to granule cell synapses reciprocal synapses [69] activating an intracellular as well as of auto-excitation synapses on the mitral molecular cascade that will ultimately release NO cells themselves. Thus pharmacological inhibition from the granule cells. In turn, NO acts as a of NMDA/AMPA receptors, nitric oxide synthase retrograde messenger to stimulate guanylyl cyclase (NOS) or soluble guanylate cyclase all prevent the activity and the formation of cyclic GMP in the pre- formation of an olfactory reccogition memory and synaptic mitral cell, which then facilitates further the ability of the lamb odours to evoke enhanced glutamate release. This glutamate release promotes glutamate and GABA recelease [68]. However once enhanced GABA release from the granule cells a memory has been allowed to form similar thereby increasing feedback inhibition onto the inhibition does not interfere with recall or mitral cells. It also facilitates autoreceptors on the potentiated glutamate and GABA release presynaptic site so that there is enhanced glutamate suggesting that these signalling pathways are not release following exposure to the learned odours. involved in recall of a consolidated recognition In this way learning causes increased sensitivity to memory [61]. glutamate in both mitral cells and at the mitral-to- Thus the overall signalling pathway model we granule cell synapses. This results in both suggest is involved in olfactory memory formation increased excitability of the mitral cells and tighter in the olfactory bulb [3, 68] is as follows (Fig. 2): excitatory/inhibitory coupling between them and Noradrenaline (or acetylcholine) release (triggered their associated granule cells such that learned by attention, arousal or vaginocervical stimulation) odours evoke both a stronger activation of the at the centrifugal projection synapses with mitral cells which are tuned to it and a sharpening GABAergic granule interneurones reduces GABA of their phasic firing discharge pattern caused by a REVIEW: ENCODING OF OLFACTORY RECOGNITION MEMORY 553

Fig. 3. Effects of the AMPA and NMDA receptor antagonists, NBQX (left panel) and MK801 (centre panels) respectively, and NOS inhibitor L-NARG (right panels) on acquisition of social olfactory memories, tested at 24 h with the habituation/ discrimination social recognition task (top panels) and STFP (bottom panels) task. Control mice, injected with saline solution (and/or the inactive enantiomer D-NARG for the L-NARG experiment), showed both a discrimination between familiar and unfamiliar stimulus mice (paired t-test, P<0.05, P<0.01 and P<0.001 vs unfamiliar) and preference for the cued food (paired t-test, * P<0.01 vs cued food). All three treatments resulted in impaired 24 h memory retention in both tasks of social olfactory memory. Taken from [4]. more robust inhibitory feedback response from the these olfactory learning models too disruption of granule cells that follows from each excitatory burst NMDA or NO signalling after memory acquisition [68, 70]. failed to have any influence on subsequent recall This process is not unique to the postpartum [5]. Thus, as with the sheep model, glutamate ewe, as similar changes have been observed in mice acting on ionotropic glutamate receptors and with olfactory conditioning [70], social recognition promoting NO release is required for the initial and STFP [4]. In our model of social olfactory acquisition phase of an olfactory recognition memory using the social recognition habituation/ memory trace but not its recall post-acquisition. discrimination test and the STFP test, long-term (24 In both sheep and mouse models of olfactory h) memory formation can be prevented by blocking induced learning or plasticity there is evidence of either NMDA (using the non-competitive altered sensitivity of class 1 metabotropic antagonist MK801) or AMPA (using the glutamate (mGluR) receptors in mitral and granule competitive antagonist NBQX) receptor activation cells and agonists of this class of receptor are more (Fig. 3). Similarly, preventing NO release by potently able to release glutamate and GABA inhibiting NOS (using the non-selective NOS within the olfactory bulb following consolidation of inhibitor, L-NARG) impaired formation of long- learning in sheep [48,64]. Preliminary term olfactory memory in both models [4]. With pharmacological studies have also shown that 554 SÁNCHEZ-ANDRADE et al. olfactory bulb infusions of antagonists targeting treatment. Ovariectomized mice receiving a four- mGluR1 (AIDA and LY393675) but not mGluR5 week oestrogen-treatment at a high dose (0.72 mg (MPEP) prevent recall of the recognition memory of 17β-oestradiol) showed a long-term (24 h) social after it is consolidated (12–24 h post-partum) but recognition memory whereas OVX and low dose have no impact on memory formation [53, 68, 70]. oestradiol treated animals did not [77]. This makes Therefore, as the olfactory recognition memory it important to determine whether hormonal consolidates there is a shift away from the fluctuations in the normal physiological range can dependence on glutamate acting on NMDA/ have any impact on olfactory memory function, AMPA-NO signalling pathways to a dependence and one obvious place to start is by studying on its actions on mGluR1 receptors. This effectively changes during the course of natural oestrous releases the former signalling pathway to be cycles. utilised for formation of new memory associations. Our early work studied the effect of hormonal fluctuations during the rat oestrous cycle on OB Olfactory Learning and Ovarian Hormones activity evoked by vaginocervical stimulation The sheep offspring recognition model is based (VCS). Such VCS resulting from parturition (or on a biologically distinct event which occurs during mating) enhances NA input to the OB, necessary a short time-window where the ewe is primed for memory formation. In vivo microdialysis appropriately with oestrogen and progesterone experiments have shown no oestrous cycle effects during pregnancy and experiences vaginocervical on basal concentrations of classical transmitters and stimulation (VCS) as a result of giving birth. nitric oxide (NO) in the OB. However, VCS Olfactory memory can even be induced by artificial significantly increases concentrations of glutamate, VCS but only after ewes have been treated with aspartate, GABA, noradrenaline, dopamine and either oestrogen or a combination of oestrogen and nitric oxide (NO) in females during proestrous/ progesterone [13]. This strongly suggests that sex oestrous, but not during metaoestrous/dioestrous hormones play a key role in olfactory memory or following ovariectomy [78]. This suggests OB formation processes described above. However, mitral cells are excited by VCS only when females little has been done to establish the precise effects are at proestrous/oestrous stage. Similar results that gonadal hormones may have on the olfactory were obtained using potassium evoked system and olfactory learning. In male rats, neurochemical release in the olfactory bulb, castration reduces NA concentrations in the OB, confirming that sex hormones changes can indeed but not in the olfactory cortex [71, 72], and reduces act directly at the level of the OB and may be potassium-evoked release of NA in the OB [71, 73]. important for mediating memory related plasticity It also substantially impairs olfactory social changes. recognition [74] and systemic treatment of castrated We decided to test this oestrous cycle effect at a males with L-DOPA restores social recognition to behavioural level using the two tests of social pre-castration levels [75]. Very little work has been olfactory memory in mice, the habituation/ carried out to establish effects of ovarian hormones discrimination and the STFP task. In both tests, on the olfactory system. The first systematic study significant improvement of learning retention relating ovarian hormones to social recognition occurs when original learning takes place during memory in female rats [76] showed a memory- the proestrus phase of the ovarian cycle [5]. No retention improvement effect of oestradiol. differences on either olfactory perception or Ovariectomized (OVX) females had a 30 min, but motivation were seen across the oestrous cycle (Fig. not 120 min memory retention for the familiar rat, 4). Our results suggest that olfactory memory contrasting with oestradiol-treated OVX (OVX+E2) formation is facilitated in females when they are animals that recognised the familiar animal at both reproductively active. This might be because when 30 and 120 min time points. Memory retention females are at a time when they are most likely to impairment did not reappear until 6 weeks after conceive they need to assess the environment for termination of oestradiol treatment, indicating that present dangers and for its suitability for long-term effects of hormones may be involved. A reproductive activities (e.g. sex behaviour, more recent study has shown similar results that pregnancy, and offspring rearing). They must be are dependent on the use of high doses of oestrogen prepared to either engage in sex behaviour when REVIEW: ENCODING OF OLFACTORY RECOGNITION MEMORY 555

Fig. 4. Effect of oestrous cycle on acquisition of social olfactory memories, tested at 24 h with the habituation/discrimination social recognition task (left panels) and STFP (right panels) task. Females showed both a social recognition ( P<0.05, vs unfamiliar) and a food preference (*** P<0.001, vs cued food) at 24 h, only when learning occurred during the proestrous stage of the oestrous cycle. Females during oestrous and dioestrous showed a significantly impaired 24 h olfactory memory retention when compared to females during proestrous (+ P<0.05, ++ P<0.01). There was no difference across the oestrous cycle on the habituation-dishabituation response, all females showed a significant TRIAL effect (P<0.001). Taken from [5]. the environment predicts success–familiar social ovarian hormone actions on olfactory memory may environment will improve reproductive and be via modulation of NO availability since animals rearing success–, or suspend resource-costly lacking the functional expression of neuronal nitric reproductive behaviours until they are more likely oxide synthase (nNOS–/–) do not show oestrous to pay off [79]. cycle dependant effects on memory retention. On Recent studies reporting that female mice lacking the other hand, exogenous treatment with the NO functional oestrogen receptors (ERα, and ERβ) fail donor molsidomine, improves memory retention in to form social memories, suggest that the ovarian dioestrous and oestrous females to the levels seen cycle effects may be mediated via oestrogen acting during proestrous [5]. It would therefore appear on these two oestrogen receptors [80]. Since that oestrogen may be influencing memory oestrogen receptors have been found in the formation in the olfactory bulb both through olfactory bulb [81], we have recently investigated modulation of NA and the subsequently ability of the impact of the oestrous cycle on the NMDA/NO glutamate to stimulate NO release. However the signalling pathway that is critical for olfactory precise mechanisms of action still require further memory formation. Results have shown that investigation.

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