Neuroscience 153 (2008) 1213–1224 DIFFERENTIAL BEHAVIORAL EFFECTS OF PARTIAL BILATERAL LESIONS OF VENTRAL TEGMENTAL AREA OR SUBSTANTIA NIGRA PARS COMPACTA IN RATS E. Y. PIOLI,a W. MEISSNER,a,b R. SOHR,c C. E. GROSS,a occur across three distinct disorders: parkinsonism, de- E. BEZARDa AND B. H. BIOULACa* pression and negative symptoms of schizophreniaDe ( aUniversite Victor Segalen, Bordeaux 2, Centre National de la Recher-Ajuriaguerra, 1975; Bermanzohn and Siris,). 1992The che Scienctifique, Bordeaux Institute of Neuroscience, UMR 5227, occurrence146 of akinetic symptoms in diverse psychiatric rue Léo Saignat, 33076 Bordeaux Cedex, France disorders supports the notion that akinesia involves not bDepartment of Neurology, CHU de Bordeaux, Hôpital du Haut-only motor behavior but also cognitive and motivational Lévêque Bordeaux, France processes. All these functions are known to be modulated c Institute of Pharmacology and Toxicology, Charité Campus Mitte,by dopamine (DA). Indeed, DA in the basal ganglia con- Humboldt Universitaet, Berlin, Germany tributes to the regulation of motor and limbic processes and DA at the cortical level is correlated with cognitive Abstract—Akinesia (or absence of movement) is a prominentfunctions Nieoullon,( 2002; Nieoullon and Coquerel,). 2003 feature of Parkinson’s disease. Akinetic symptoms, however,In this view, clinical evidence shows that motor symptoms are also observed in depression and schizophrenia, whichin neurodegenerative disorders such as PD are accompa- support the hypothesis that akinesia involves more than only nied by cognitive deficitsBrown ( and Marsden, 1990; motor behavior. A common feature of these disorders is the disruption of dopamine homeostasis in the CNS. HereBrown we and Jahanshahi, ).1996 aimed at relating the respective involvement of the nigrostri-Anatomically, the substantia nigra pars compacta atal and mesocortical dopaminergic pathways to akinesia.(SNc) mainly projects to the caudate nucleus and the We investigated in the rat the relative effects of selectiveputamen. By contrast, the ventral tegmental area (VTA) bilateral partial lesions of substantia nigra pars compactamainly innervates the frontal cortex, the ventral striatum (SNc) or ventral tegmental area (VTA) which did not andaffect the hippocampal formationFuxe, ( 1965; Lindvall and locomotion, on fine motor, motivational and cognitive behav- iors. Motor impairments were measured by the evaluationBjorklund, of 1974; Fallon et al., 1978; Fallon and Moore, fine motor control in the stepping test and in the paw1978a,b; reach- Haber and Fudge, ). 1997While SNc/VTA projec- ing test. Cognitive functions were assessed by various tionspar- to the forebrain are segregated, a degree of overlap adigms: spontaneous alternation in the Y maze and objectexists in other cortical terminal fields and some subcortical exploration task. Motivational behavior was evaluated by areas.the In particular, neurons projecting to the frontal cor- 100-pellets test. The results suggested that specific behav-tex, the cingulate cortex, the septum rostro-medial caudate ioral impairments are obtained following selective lesions of either SNc or VTA. SNc-lesioned rats exhibited deficits inand fine the nucleus accumbens are located in the medial SNc motor functions as previously described in animal modelsas ofwell as the lateralFallon VTA and( Moore, 1978b). Parkinson’s disease, whereas VTA-lesioned rats demon- Bilateral DA-containing neuron degeneration of the strated traits of perseveration without significant motorSNc is the main characteristic of human PD. Although impairments. © 2008 IBRO. Published by Elsevier Ltd.damage All to the SNc primarily causes PD, the VTA is also rights reserved. modestly affected, even in early stages Damierof PD ( Key words: akinesia, dopamine, 6-OHDA. et al., 1999a,b), rendering it difficult to attribute a specific deficit to a given nucleus. To solve this ambiguity, we induced selective partial Akinesia is considered to be a principal feature of bilateralParkin- 6-hydroxydopamine (6-OHDA) lesions of either son’s disease (PD) along with rigidity and tremor. theIt lateralis SNc or the medial VTA Pioliin theet al.,rat ( defined as poverty of initiation and execution of willed2004 and) . The rats were subjected to various motor, motivational associated movements in the absence of Lee,paralysis (and cognitive behavioral experiments. Motor impairments 1989). Akinesia represents a clinical syndrome which werecan estimated by the measure of fine motor control in the *Corresponding author. Tel: ϩ33-557-571-551; fax: ϩ33-556-901-421. stepping test and in the paw reaching test. The motiva- E-mail address: [email protected] (B. H. Bioulac). tional behavior was evaluated by the 100 pellets test. Two Abbreviations: ANOVA, analysis of variance; DA, dopamine; DOPAC, paradigms were employed to assess cognitive impair- 3,4-dihydroxyphenylacetic acid; EDTA, ethylenediaminetetraacetic acid; HPLC, high-pressure liquid chromatography; HVA, homovanillic ments: spontaneous alternation in the Y maze and an acid; NE, northeast; NW, northwest; PD, Parkinson’s disease; SE, object exploration task. Object exploration is associated southeast; S.E.M., standard error of the mean; SNc, substantia nigra with an index of cognitive activity, especially in rodents. pars compacta; SW, southwest; TH, tyrosine hydroxylase; TH-IR, tyrosine hydroxylase immunoreactive; VTA, ventral tegmental area; Both paradigms (in which rats explore without contingent 6-OHDA, 6-hydroxydopamine. reinforcement) allow the study of various behavioral func- 0306-4522/08$32.00ϩ0.00 © 2008 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2008.01.084 1213 1214 E. Y. Pioli et al. / Neuroscience 153 (2008) 1213–1224 tions, such as attention, memory, and reactions to change in the environment. EXPERIMENTAL PROCEDURES Animals Twenty-nine Wistar rats (Depré, St. Doulchard, France) weighing 450–480 g were housed under controlled conditions of light (12-h light/dark cycle, i.e. lights on from 07:00–19:00 h), temperature (22 °C) and humidity. Food and water were available ad libitum. All experiments were carried out in accordance with the European Committee Council Directive of 24 November 1986 (86/609/EEC) and were approved by the local ethical committee. All reasonable efforts were made to minimize animal suffering and to use the minimum number of animals necessary to perform statistically valid analyses. 6-OHDA procedure Rats were anesthetized with 4% chloral hydrate (400 mg/kg i.p.; Sigma, St. Quentin Fallavier, France) and placed in a stereotaxic apparatus as previously described (Pioli et al., 2004). Animals were pre-treated with desipramine (25 mg/kg i.p.; Sigma) 30 min prior to the 6-OHDA injection in order to protect noradrenergic neurons. 6-OHDA (2 ␮g/␮l in 0.9% NaCl containing 0.01% ascor- bic acid; Sigma) was injected first into the right and 1 week later into the left hemisphere. The following coordinates were employed for the SNc injection (4 ␮l, 0.5 ␮l/min): 5.2 mm posterior to bregma; Ϯ2.2 mm from midline; Ϫ7.7 mm from dura; and for the Fig. 1. Experimental schedule. ␮ ␮ VTA injection (1 l, 0.125 l/min): 5.2 mm posterior to bregma; field, a frame placed at 3 cm height with 16 photocell beams per Ϯ Ϫ 0.8 mm from midline; 7.6 mm from dura. After each injection, side ensured movement detection. Total motor activity was re- the needle was left in position for 10 min to allow absorption of corded over a period of 7 min on three consecutive days. Animals the injected solution and to minimize spread of the toxin along the from sham and lesioned groups were tested in random order. After needle tract. Sham-operated controls received an identical proce- each test, the open field was cleaned with a moistened sponge. dure with the exception that no toxin was infused but only vehicle Results are expressed as mean activity of the 3 daysϮstandard (0.9% NaCl containing 0.01% ascorbic acid). This procedure was error of the mean (S.E.M.) and analyzed using a one-way analysis selected based on numerous pilot studies which reliably demon- of variance (ANOVA) with group as independent variable. strated selective lesions of the target region. Stepping adjustments. Stepping adjustments were mea- Experimental protocol sured as described by Olsson et al. (1995). Briefly, the experi- menter took the rat with one hand holding both hind limbs and the The animals were allowed to recover for 3 weeks before behav- other hand holding one of the forelimbs. The free paw was placed ioral testing commenced. This period of recovery has been de- in contact with a flat surface. The experimenter then moved scribed to produce stable 6-OHDA lesions (Schwarting and Hus- the animal slowly sideways in forehand and backhand directions. ton, 1996a,b). All rats were subjected to a sequence of behavioral The number of adjusting steps was counted for both paws in the tests as summarized in Fig. 1. After completion of the behavioral backhand and forehand directions. The test was repeated twice a evaluations, the rats were killed with an anesthetic overdose of day for three consecutive days. Animals from sham and lesioned urethane (Sigma) and the brains were rapidly removed. groups were tested in random order. Results are expressed as the The selectivity of the 6-OHDA lesions was confirmed by ste- mean number of adjusting steps during the six sessionsϮS.E.M. reological counting of mesencephalic neurons characterized by The means were subjected to a two-way ANOVA with group as tyrosine hydroxylase (TH) immunochemistry and by measurement the independent variable and paw as the dependent factor. When of DA and 5-HT content and their metabolites in cortical and appropriate, ANOVAs were followed by post hoc t-tests corrected striatal areas. For this purpose, the cortex and striatum were for multiple comparisons by the method of Bonferroni. dissected, flash-frozen on dry ice and stored at Ϫ80 °C. The caudal part of the brain containing the mesencephalon was post- Food consumption test. Food consumption was assessed fixed in PFA 4% for 2 days at 4 °C and transferred to 20% sucrose as the time taken to eat 100 pellets as described by Baunez et al.