Differential involvement of dopamine in the anterior and posterior parts of the dorsal striatum in latent inhibition

Neuroscience 118 (2003) 233–241

DIFFERENTIAL INVOLVEMENT OF DOPAMINE IN THE ANTERIOR AND POSTERIOR PARTS OF THE DORSAL STRIATUM IN LATENT INHIBITION J. JEANBLANC, A. HOELTZEL AND A. LOUILOT*

Elsevier Science Ltd. All rights reserved.

INSERM U 405 and Institute of Physiology, University Louis Pasteur, Faculty of Medicine, 11 rue Humann, 67085 Strasbourg Cedex, France

Key words: conditioned olfactory aversion, affective processes, cognitive processes, schizophrenia, in vivo voltammetry, entorhinal cortex.

Abstract—The involvement of mesostriatal dopaminergic neurons in cognitive operations is not well understood, and needs to be further clarified. The use of latent inhibition paradigms is a means of investigating cognitive processes. In this study, we investigated the involvement in latent inhibition of dopaminergic inputs in the anterior part and posterior part of the dorsal striatum. The latent inhibition phenomenon was observed in a conditioned olfactory aversion paradigm. Changes in extracellular dopamine levels induced by the conditioned olfactory stimulus (banana odor) were monitored in the two parts of the dorsal striatum in the left hemisphere after pre-exposure to the olfactory stimulus using in vivo voltammetry in freely moving rats. During the conditioning session animals received either an i.p. injection of NaCl (0.9%) (control groups) or an i.p. injection of LiCl (0.15 M) (conditioned groups). Dopamine variations and place preference or aversion toward the stimulus were analyzed simultaneously in pre-exposed and non-pre-exposed animals. Data collected during the retention (test) session were as follows. Where the anterior part of the striatum was concerned, similar enhancements in dopamine levels (ⴙ100%) were obtained in pre-exposed and non-pre-exposed control animals, as well as in the pre-exposed experimental animals. In contrast, dopamine levels in the non-pre-exposed experimental group (conditioned animals) remained fairly consistently close to the baseline after the presentation of the olfactory stimulus. Where the posterior part of the striatum was concerned, increases in extracellular dopamine levels were similar (ⴙ50%) for the different groups. The present results suggested that dopaminergic neurons reaching the anterior part of the dorsal striatum are implicated in the latent inhibition phenomenon and affective perception, whereas dopaminergic terminals in the posterior part of the dorsal striatum appeared to be involved neither in latent inhibition nor in affective perception of the stimulus, seeming only to be affected by the intrinsic properties of the stimulus. Cognitive as well as affective deficits have been reported in patients with schizophrenia. Thus the present data may be considered in the context of the pathophysiology of schizophrenic psychoses. © 2003 IBRO. Published by

Dopaminergic (DAergic) neurons originating from the ventral mesencephalon are organized in several subunits which reach numerous forebrain structures separately (for reviews see Swanson, 1982; Bjo¨rklund and Lindvall, 1986; Oades and Halliday, 1987). These mesencephalic neurons appear to facilitate the integrative processing at the level of the innervated structures by enabling the switching between different inputs into a dopamine (DA)-receptive brain region (see Oades, 1985 for discussion). Coordination of the specific integrations occurring at the level of the DA-innervated structures may be assured by the existence of an interdependent functioning between different mesencephalic DAergic subunits (Louilot et al., 1985b, 1987b, 1989a,b; Louilot and Le Moal, 1994; Louilot and Choulli, 1997). All these properties may allow the mesencephalic DAergic neurons to be involved in the various processes contributing to the emergence of the behavioral responses related to the different characteristics (environmental, behavioral, motivational, emotional, cognitive) of the context. Thus, as assessed with appetitive or aversive conditioned stimuli, recent data obtained using microdialysis, and voltammetric and electrophysiological approaches (Bassareo and Di Chiara, 1999; Besson and Louilot, 1995, 1997; Louilot and Besson, 2000; Mirenowicz and Schultz, 1996; Schultz et al., 1997; Rosenkranz and Grace, 2002) suggested that DAergic neurons originating from the midbrain are involved in positive and negative affective processes, i.e. processes contributing to the attribution of an attractive or an aversive value to a given stimulus, previously associated with a positive or negative reinforcement. DAergic responses in the nucleus accumbens (ACC), monitored using in vivo methods, have also been found to be affected by pre-exposure to the conditioned stimulus (CS) before the aversive conditioning session in sensory preconditioning (Young et al., 1998) and latent inhibition (LI) paradigms, suggesting a possible involvement of these mesencephalic DAergic neurons in cognitive processes (Young et al., 1993; Murphy et al., 2000, Jeanblanc et al., 2002). Historically, LI has been defined as a phenomenon whereby prior exposure to a CS without negative or positive reinforcement retards the subsequent generation of conditioned responses when the CS is paired with the

*Corresponding author. Tel: ⫹33-3-9024-3253; fax: ⫹33-3-90243256. E-mail address: [email protected] (A. Louilot). Abbreviations: ACC, nucleus accumbens; ANOVA, analysis of variance; AP, anterior to the interaural line; CS, conditioned stimulus; DA, dopamine; DAergic, dopaminergic; DNPV, differential normal pulse voltammetry; DOPAC, 3,4-dihydroxyphenyl acetic acid; H, below the cortical surface; L, lateral to the midline; LI, latent inhibition; NPE, non-pre-exposed; PE, pre-exposed; ST, striatum; US, unconditioned stimulus.

0306-4522/03$30.00⫹0.00 © 2003 IBRO. Published by Elsevier Science Ltd. All rights reserved. doi:10.1016/S0306-4522(02)00823-0

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unconditioned stimulus (US), suggesting that the non-reinforced pre-exposure to the CS inhibits the learning of an association between the CS and the US (Lubow and Moore, 1959). Since then, it has been demonstrated that LI is a ubiquitous phenomenon occurring in numerous animal species, including humans, and in diverse kinds of conditioning preparations (for review see Lubow, 1989). Despite its relative simplicity, theoretical explanations of the LI phenomenon have proven difficult. Thus LI has been interpreted variously as reflecting a lack of attention to the pre-exposed CS, a decrease in the acquisition of conditioning or an alteration in the expression of the conditioning (for reviews see Lubow, 1989; Oades and Sartory, 1997; Escobar et al., 2002). However, recent data obtained in a conditioned olfactory aversion paradigm lend much credence to the view, also held by Escobar et al. (2002), that LI corresponds to an impairment of the behavioral expression of the effects of conditioning, but not to a deficit in the acquisition of conditioning (Jeanblanc et al., 2002). More precisely, the results obtained in this voltammetric study suggested that DAergic neurons innervating three subregions (core, dorsomedial shell, ventromedial shell) of ACC are differentially involved in LI, as DAergic neurons that reach the core and dorsomedial shell parts of the ACC were involved in LI, whereas those innervating the ventromedial shell part of the ACC, appeared only to be involved in the perception of the affective value (positive or negative) of the stimulus. Recent conceptualizations relating to the neural substrates of LI phenomenon do not include striatal DAergic innervation, in contrast to the DAergic connection of the ACC (Gray et al., 1997; Weiner and Feldon, 1997). However, whereas studies carried out with local microinjections of D-amphetamine initially suggested that DAergic neurons innervating the striatum (ST) are not involved in LI (Solomon and Staton, 1982), more recent data suggested the reverse (Ellenbroek et al., 1997). One interpretation could be that the paradigms used are different since Solomon and Staton (1982) measured LI using a conditioned avoidance-response paradigm whereas the LI experiments of Ellenbroek et al. (1997) were carried out in a conditioned taste-aversion paradigm, which involved a different temporal association between CS and US. However, in view of the regionalization that has been observed for the ACC (Jeanblanc et al., 2002), the discrepancy between the results obtained in the two studies (Solomon and Staton, 1982; Ellenbroek et al., 1997) may be similarly related to a differential implication in LI of the striatal subregions chosen in each of the previous studies. In the course of this study, we investigated, using a conditioned olfactory aversion paradigm, the involvement in LI of DAergic projections in anterior and posterior ST subregions. Changes in extracellular ST DA levels were measured, using in vivo voltammetry in freely moving rats, in the left hemisphere of animals pre-exposed (PE) and non-pre-exposed (NPE) to the conditional olfactory stimulus. The voltammetric measurements of DA release in PE and NPE animals were assessed in parallel with place preference or aversion toward the stimulus.

EXPERIMENTAL PROCEDURES Male Sprague–Dawley rats (Janvier, Le Genest, France) weighing 400⫾15 g at the time of surgery were individually housed at 22⫾2 °C with free access to food and water and maintained on a 12-h light/dark cycle (lights off at 11 a.m.), at least 1 week before the surgical procedure. A total of 68 rats were operated on. All the animals were anesthetized using chloral hydrate (400 mg/kg i.p.) and placed on a stereotaxic frame (incisor bar 3.3 mm below the interaural line). They were then implanted with a specially designed microsystem allowing the periodic replacement of the working electrodes (Unime´canique, Epinay/Seine, France) (Louilot et al., 1987a). The stereotaxic coordinates were adjusted for the rat strain we used in accordance with the atlas of Paxinos and Watson (1986). For the left anterior part of the ST they were 10.2 mm anterior to the interaural line (AP), 1.8 mm lateral to the midline (L) and 5.9 mm below the cortical surface (H); and for the left posterior part of the ST they were 9.4 mm (AP), 1.8 mm (L) and 5.5 mm (H). Animals were allowed 1 week to recover after surgery. Experiments were performed in accordance with guidelines for care and use of experimental animals issued by the European Communities Council Directive (86/609/EEC) and the French Ministry of Agriculture. Every effort was made to limit the number of animals used and their suffering. Electrochemical procedures were similar to those previously described (Gonzalez-Mora et al., 1991; Louilot et al., 1991). A classical three-electrode potentiostatic setting was used with working, reference and auxiliary electrodes. The working electrodes used were pyrolytic carbon fiber microelectrodes (12-␮m diameter, 500-␮m length, ref AGT 8000, SEROFIM, Gennevilliers, France) electrochemically pretreated according to Gonon et al. (1984). For both in vitro and in vivo measurements, the reference electrode was an AgCl-coated silver wire (ref. AG10T, Medwire Corporation, Mt. Vernon, NY, USA). The auxiliary electrode used for in vitro measurements was a platinum wire while a stainless steel screw was used for in vivo determinations. Differential normal pulse voltammetry (DNPV) combined with computerized waveform analysis of the catechol peak was used to establish the extracellular levels of DA selectively (Gonzalez-Mora et al., 1991; see also O’Neill et al., 1998). Briefly, the waveform analysis method takes into account the small but consistent difference (approximately 36 mV) between the oxidation potentials of 3,4dihydroxyphenyl acetic acid (DOPAC) and DA; the parameters of each substance (DOPAC and DA) are determined separately in vitro for each carbon fiber electrode. The voltammogram is modeled as a sum of normal probability distributions corresponding to the relevant electroactive species; the relative contribution of DA to the mixed catechol peak is calculated using the least squares techniques (Gonzalez-Mora et al., 1991). DNP voltammograms were recorded every minute. The voltammetric apparatus (Biopulse, Tacussel, Villeurbanne, France) was set up with the following parameters: scan range ⫺240 mV, ⫹200 mV; scan rate 10 mV/s; potential step 4 mV; pulse period 400 ms; prepulse duration ⫹120 ms; pulse duration 40 ms; pulse amplitude 40 mV. The protocol used for aversive conditioning was as follows: animals were placed for 1 h in the experimental cage (24⫻27⫻44 cm) without being exposed to the olfactory stimulus. They were then exposed for 1 h to a new olfactory stimulus, i.e. the banana odor used in previous studies (Louilot and Besson, 2000, Jeanblanc et al., 2002). At the end of the second hour, animals received either an i.p. injection of NaCl (0.9%) or an i.p. injection of a toxin inducing nausea (LiCl 0.15 M) (Garcia et al., 1985), and then remained in the experimental cage for a further hour with the olfactory stimulus still present; 72 h later, the animals were exposed for a further hour to the conditional olfactory stimulus. The LI procedure followed exactly the same protocol except

J. Jeanblanc et al. / Neuroscience 118 (2003) 233–241 that animals were submitted to a supplementary pre-exposure session 72 h before the aversive conditioning session. Pre-exposed (PE) animals were placed in the experimental cage for 1 h without olfactory stimulus and were then exposed for 2 h to banana odor alone (Jeanblanc et al., 2002). Animals were tested during the dark period of the light/dark cycle. The position of the animals in the experimental cage was monitored using a small infrared camera (CA-H34C, ref 51.8050 Selectronic, Lille, France) installed in the roof of the cage and connected to a monitor and a tape recorder. The olfactory stimulus was fed into the cage through a hole in the wall adjacent to the door. The cage was divided into two virtual parts, one part containing the hole consisting of the largest possible half-circle that could be drawn around it (with a radius equal to half the width and a surface area covering 35% of the cage), the second part consisting of the remaining area outside the half-circle. Behavioral analysis was carried out over 10-min periods. We considered that if animals moved randomly around in the cage they should spend 35% (210 s) of the 10-min period in the part containing the hole. Preference or aversion toward the olfactory stimulus was evaluated by measuring the time spent in the part containing the hole over 10-min periods, before and after the presentation of the olfactory stimulus. For each experiment the mean of the 10 last measures of DA recorded over the control period (at least 30 min) during which the signal varied less than 10% was calculated and taken as the 100% value. Results are expressed as percentages (mean⫾S.E.M.) relative to the mean pre-test values. Statistical analysis of the results was performed using a two-way analysis of variance (ANOVA) with repeated measures. Only between-group ANOVAs are shown unless otherwise indicated. As regards behavioral data, post hoc comparisons between the different groups were made using Bonferroni’s t-test (unpaired data). Statistical significance was set at P⬍0.05 for all analyses. The recording sites were visualized at the end of the experiment by electrocoagulation as previously described (Louilot et al., 1985a). In all cases, the location of the recording site was checked histologically by Thionin Blue staining of sections with reference to the atlas of Paxinos and Watson (1986) (Fig. 1).

RESULTS Behavioral study (Fig. 2) In the retention session (test session), before presentation of the stimulus, no statistical difference was observed between the different groups in the time spent near the hole shown by the pre-exposure⫻conditioning interaction (F[1, 64]⫽0.36; n.s.). During the hour following the presentation of the olfactory stimulus, a significant preexposure⫻conditioning effect (F[1, 64]⫽8.66; P⬍0.005) was observed. This effect was even more obvious during the first 30 min following the presentation of the stimulus (F[1, 64]⫽18.29; P⬍0.0001). As far as PE animals were concerned, after the presentation of the olfactory stimulus, the time spent near the hole remained close to the neutral value in both the PE control group (maximal value 265.5 s) and the PE conditioned group (maximal value 276.5 s). No significant differences were observed between the two PE groups (F[1, 31]⫽0.02; n.s.) throughout the hour of the test session. Concerning NPE animals, after the last presentation of the stimulus, an increase in the time spent near the hole was observed in the NPE control group (maximal value 389.4 s) whereas a decrease in this parameter was obtained in the

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Fig. 1. Typical recording sites in the left anterior part of ST (A) and in the left posterior part of ST (B) are indicated by arrows. Recording sites were visualized by electrocoagulation at the end of the experiment. In all cases, the location of the recording sites was checked histologically by Thionin Blue staining of sections. Scale bar⫽2 mm.

NPE experimental group (minimal value 71.1 s). Detailed statistical analysis showed that the time spent near the hole following presentation of the stimulus was significantly greater in the NPE control group than in the NPE conditioned group for the first 30 min (F[1, 33]⫽60.57; P⬍0.000001) and during the total 60-min period of the test (F[1, 33]⫽23.92; P⬍0.0001). In other respects no statistical differences were observed between PE and NPE control groups (F[1, 30]⫽1.77; n.s.) whereas the time spent near the hole was found to be significantly different in PE and NPE conditioned groups for the first 30 min (F[1, 34]⫽17.4; P⬍0.0005) and the whole hour (F[1, 34]⫽9.10; P⬍0.005). Percentages of time spent near the hole during the first 30 min/total time were 40.2⫾5.7% (PE–NaCl) and 43.8%⫾6.5% (PE–LiCl) in PE groups and 55.9⫾4.9% (NPE–NaCl) and 15.9⫾2.1% (NPE–LiCl) in NPE groups respectively. Values obtained in PE–LiCl and NPE–LiCl were significantly different (t34⫽4.173, P⬍0.01). Percentages in NPE–NaCl and NPE–LiCl were also statistically different (t33⫽7.783, P⬍0.00001). Voltammetric study (Fig. 3) Only animals presenting clear implantations in either the anterior part (Fig. 1A) or posterior part of ST (Fig. 1B) were considered for the voltammetric study.

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30 min following the presentation of banana odor with values of about ⫹9% at 30 min and ⫹43% at 60 min respectively (F[1, 11]⫽7.22; P⬍0.05). Time courses of DA responses in the two PE and NPE conditioned groups were found to be statistically different (F[29, 319]⫽2.28; P⬍0.0005). In all other respects, in NPE animals only, a significant conditioning effect (NaCl/LiCl) was observed (F[1, 8]⫽10.39; P⬍0.05). However, time courses of DA changes in control and conditioned groups were found to be significantly different both for PE animals (F[29, 377]⫽2.15; P⬍0.001) and NPE animals (F[29, 232]⫽2.45; P⬍0.0005). Variations in the posterior ST

Fig. 2. Time spent near the hole in animals exposed to the conditional olfactory stimulus. Seventy-two hours after the aversive conditioning, PE animals were exposed for the third time to the banana odor whereas NPE animals were exposed for the second time to the olfactory stimulus (arrow). The insert represents the percentage of total time spent near the hole during the first 30 min following presentation of the stimulus. In control animals (white) the banana odor has been associated with the i.p. injection of NaCl (0.9%) whereas in conditioned animals (black) the banana odor has been associated with the same volume of LiCl (0.15 M) during the conditioning session. The cage was divided into two virtual parts. One part contained the hole and was the largest possible half-circle drawn around the hole, covering about 35% of the total surface area, while the other part was the remaining surface area outside the half-circle. Behavioral analysis was carried out over periods of 10 min. The dotted line represents 35% of one period of 10 min (210 s) and corresponds to a neutral distribution of the animals in the cage. n represents the number of rats per group (PE and NPE): PE–NaCl⫽16, PE–LiCl⫽17, NPE–NaCl⫽16, NPE– LiCl⫽19. Statistical significance between groups, **P⬍0.01, ****P⬍0.0001, *****P⬍0.00001 (Bonferroni’s t test).

Variations in the anterior part of the ST In the retention session, for the hour following presentation of the olfactory stimulus, a significant pre-exposure⫻ conditioning effect was observed (F[1, 21]⫽11.34; P⬍0.005). More precisely, extracellular DA changes observed in PE and NPE control animals were similar in magnitude and direction, whereas distinct DA variations were obtained in PE and NPE conditioned groups (Fig. 3A). Thus, after exposure to the banana odor, DA levels rose progressively to reach, by the end of the hour, the maximum values of about 62% and 102% above basal levels in PE and NPE control animals respectively (F[1, 10]⫽4.30; n.s.). Time courses of DA responses, however, in the two PE and NPE control groups were found to be statistically different (F[29, 290]⫽2.91; P⬍0.00001). In the conditioned animals, the PE group showed a progressive increase in DA levels reaching about 72% and 113% above the baseline 30 min and 60 min respectively after the stimulus presentation. By contrast, in the NPE conditioned group DA levels remained close to the baseline during the first 30 min and increased slightly during the last

In the retention session, after presentation of the olfactory stimulus, no differences between the PE and NPE groups were observed (F[1, 16]⫽0.33; n.s.) and no significant conditioning effect (NaCl/LiCl) was observed (F[1, 16]⫽0.09; n.s.) (Fig. 3B). However, time courses were statistically different in PE and NPE groups (preexposure⫻time interaction) during the hour following the presentation of the olfactory stimulus (F[29, 424]⫽2.75; P⬍0.00001). By contrast no significant conditioning⫻time interaction was observed (F[29, 424]⫽0.98; n.s.). Similar DA changes were observed in PE and NPE control animals. DA levels increased slightly following presentation of the banana odor reaching maximum values of about 48.5% and 59.4% above the baseline in PE and NPE control groups respectively (F[1, 7]⫽0.24; n.s.). DA variations in PE and NPE conditioned groups were moderate and did not differ markedly; maximum values were about 34.9% above the baseline in PE conditioned animals and 67.5% in NPE conditioned animals (F[1, 9]⫽0.83; n.s.). Time courses in PE and NPE conditioned animals were statistically different for the hour following presentation of the banana odor (F[29, 261]⫽2.23; P⬍0.0005); more detailed analysis showed that the difference between PE and NPE time courses was statistically significant only during the last 30 min (F[14, 126]⫽2.37; P⬍0.01). Finally, no significant conditioning effect was observed either for the PE animals (F[1, 6]⫽5.51; n.s.) or the NPE animals (F[1, 10]⫽0.18; n.s.).

DISCUSSION The results of the present study suggest that DAergic innervations of the anterior and posterior parts of the ST are not equally involved in LI processes. More precisely, DAergic neurons innervating the anterior part of the ST were found to be involved in LI obtained in a conditioned olfactory-aversion paradigm, whereas those reaching the posterior ST appeared to be involved neither in the LI phenomenon nor in the affective perception of the stimulus. DAergic terminals in the posterior part of ST seemed to be affected only by the presentation of the olfactory stimulus, since similar DA variations were observed in this ST subregion in the various situations, whether or not animals were PE or conditioned to the stimulus.

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Fig. 3. Changes in DA release in the left anterior part of ST (A) and in the left posterior part of ST (B) of male rats exposed to the olfactory stimulus (banana odor) after conditioning. PE animals were exposed for the third time to the banana odor whereas NPE animals were exposed for the second time to the olfactory stimulus. Seventy-two hours after aversive conditioning (LiCl 0.15 M injection), animals were placed in the experimental cage for 1 h and the olfactory stimulus was presented again (arrow). Extracellular levels of DA were assessed using DNPV and computer-assisted numerical analysis in freely moving rats. Voltammograms were recorded every minute. For each experiment the mean of the last 10 measures during the control period (at least 30 min) was calculated and taken as the 100% value. Only mean values and S.E.M. corresponding to two scans are shown; where no S.E.M. is indicated the size is less than the radius of the symbol. The arrow corresponds to the presentation of the olfactory stimulus. n represents the number of rats per group. Results were analyzed by factorial analysis of variance (ANOVA).

DA changes observed in the anterior ST in NPE animals were consistent with those previously reported for the ST after the presentation of the conditional olfactory stimulus, i.e. a DA increase in control animals and a lack of variations of extracellular DA levels in conditioned animals (Besson and Louilot, 1997; Louilot and Besson, 2000). However, we did not observe in this study the delayed decrease in DA levels obtained in conditioned animals in the two previous studies. This discrepancy may correspond to a difference in the ST regions investigated, since the stereotaxic coordinates for the rat strain used in the two previous studies and that used in the present study did not match exactly, suggesting a divergence

in the brain structure conformation in the two rat strains. From a behavioral point of view, place attraction or aversion observed in NPE rats depending on the positive or aversive value of the stimulus were similar to the behavioral changes reported previously in NPE animals (Louilot and Besson, 2000; Jeanblanc et al., 2002). Behavioral data in PE rats, showing that performances of control and conditioned animals did not differ, were also consistent with responses previously obtained in the same paradigm (Jeanblanc et al., 2002). The present study provides evidence of a differentiation in the DA responses in the anterior and posterior ST in

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a behavioral context. Thus, as far as the posterior ST was concerned, presentation of the olfactory stimulus was followed by a DA increase irrespective of conditioning and LI phenomenon, suggesting that DA terminals in the posterior ST are only affected by the intrinsic properties of the stimulus. However, the olfactory system has not been reported to send direct projections to the posterior ST, precluding a direct stimulation of striatal DAergic terminals by olfactory entries. The olfactory system may be connected indirectly to the posterior ST by the entorhinal cortex which receives direct olfactory inputs (Witter et al., 1989) reaching the ST by entorhinofugal pathways (Swanson and Ko¨hler, 1986; Witter et al., 1989; Totterdell and Meredith, 1997). However, entorhinal efferents in the posterior ST appear to relate almost exclusively to the borders of the ventricle and the edge of the corpus callosum (Totterdell and Meredith, 1997). Thus, a DA regulation by these entorhinostriatal efferents seems unlikely. Another possibility is that DA changes in the posterior ST may be related to an activation of DA neurons at the level of the mesencephalon, since some nigral DA-like neurons were found to be stimulated by olfactory stimuli (Chiodo et al., 1980). Anyhow, as discussed previously, posterior ST did not appear to receive direct or indirect olfactory information. Therefore the DA increases we observed in posterior ST may not contribute to integrative processes concerning olfaction, but in fact may facilitate sensory motor integrations for other modalities, thereby allowing the animal to pay attention to the olfactory stimulus, irrespective of the acquired value of the stimulus during pre-exposure or conditioning. This suggestion is consistent with data reported several years ago showing that unilateral destruction of the nigrostriatal DAergic system was followed by a deficit of sensory orientation toward stimuli of all modalities, olfaction included (Ljungberg and Ungerstedt, 1976). As regards the anterior ST, the results of the present study provide the first demonstration of a direct involvement of striatal DAergic terminals in LI phenomenon. Previous data showing that local microinjections of d-amphetamine in the dorsal ST lead to a disruption of LI do suggest that DAergic neurons are involved in this phenomenon, but they are difficult to compare with the present data since local microinjections of the indirect DA agonist were carried out before the pre-exposure and the conditioning sessions (Ellenbroek et al., 1997). Considered in the light of our data, the lack of effects on LI observed by Solomon and Staton (1982) with intra-caudate d-amphetamine injections could be explained by the fact that the striatal region investigated in their study did not correspond to the anterior ST. Another explanation could be that Solomon and Staton (1982) studied LI phenomenon in a conditioned avoidance-response paradigm where a tone was paired with footshocks while Ellenbroek et al. (1997) and we studied LI in conditioned-aversion paradigms where taste and olfaction were associated with internal malaise induced by an LiCl injection. However, it seems unlikely that the discrepancy between the results obtained in the different studies may be related to the behavioral paradigms used. Indeed intra-ACC d-amphetamine injections were

found to disrupt LI in conditioned-avoidance response by Solomon and Staton (1982) and to have no effects on LI in conditioned taste aversion by Ellenbroek et al. (1997) whereas, using another conditioned-aversion paradigm, namely conditioned olfactory aversion, we observed in the ACC, as in the anterior ST (present study), specific LI DA responses (Jeanblanc et al., 2002). This suggests that a regionalization in the involvement of DA in LI phenomenon, both in the dorsal and ventral ST, is the key explanation for the discrepancies reported in the literature concerning neural substrates of LI. DA variations observed in the anterior ST in both PE and NPE animals possessed the same profile as those obtained in the core part of the ACC (Jeanblanc et al., 2002), which was reminiscent of the anatomical similarities reported between the two striatal regions (Heimer et al., 1991; Zahm, 1992; Zahm and Brog, 1992). However, DA changes in the anterior ST in NPE animals, in contrast to DA variations observed in the core of ACC, have not been found to be influenced by the basolateral nucleus of the amygdala, either in control or in experimental animals (Louilot and Besson, 2000). Neither the anterior ST nor the posterior ST has been reported to receive direct olfactory information, suggesting that DA responses observed in the anterior ST during this study are dependent on at least one other forebrain structure. Although the whole ST of the rat is innervated by afferents from the entire cortical mantle (McGeorge and Faull, 1989), the anterior ST has been shown, using the 2-deoxyglucose method, more specifically to be functionally related to the anteromedian prefrontal cortex (Divac and Diemer, 1980). To be more precise, the anterior striatal region selected in our study is reached by direct projections from the dorsal prelimbic/ventral orbital areas (Beckstead, 1979; Sesack et al., 1989; Berendse et al., 1992; Montaron et al., 1996). These anteromedian prefrontal cortex sectors receive olfactory information (Price et al., 1991) and the ventral orbital prefrontal cortex has been found to be involved in affective processes in monkeys (Dias et al., 1996) and humans (Perlstein et al., 2002). Moreover, it has been shown that ibotenic lesions of the anteromedian prefrontal cortex including the prelimbic/orbital regions induced an increase in DA turnover in the anteromedian ST (Jaskiw et al., 1990). Therefore, bearing in mind all the foregoing aspects, it may be tentatively suggested that the DA responses we observed in NPE animals, in the anterior ST, were dependent at least in part on the anteromedian prefrontal cortex. Ibotenic lesions or pharmacological manipulation of the DAergic transmission in the anteromedian prefrontal cortex did not seem to affect LI (Ellenbroek et al., 1996; Lacroix et al., 2000a,b). However, it is important to mention that frontal DA depletions have been found to affect, at the same time, striatal DA activity and blocking responses in the conditioned blocking paradigm closely related to LI (Oades et al., 1987). Moving on now to the PE animals, we should begin by mentioning that the similar DA responses obtained previously in NPE and PE animals (Jeanblanc et al., 2002), in the ventromedial shell part of ACC, argue for the estab-

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lishment of an aversive conditioning in PE animals. Hence, as with the dorsomedial shell and core parts of ACC, it may be suggested that DA changes in the anterior ST in PE animals correspond to the influence of a recognition memory system interacting competitively with a system processing the aversive value of the stimulus; as mentioned above, the anteromedian prefrontal cortex may be part of the latter system. Since intra-caudate microinjections of d-amphetamine during pre-exposure have been found to disrupt LI (Ellenbroek et al., 1997) it seems reasonable to suggest that the competitive interaction occurs at the ST DA terminals level. It could also be suggested that the recognition system exerts its regulatory influence on DA transmission in the anterior ST at least in part through the involvement of the entorhinal cortex. Indeed this structure has been reported to play a crucial role in recognition memory, especially of olfactory stimuli in the rat (Suzuki and Eichenbaum, 2000). Moreover, our suggestion is consistent with the anatomical data showing that entorhinal innervates the anteromedian ST more densely than the posterior ST (Swanson and Ko¨hler, 1986; Witter et al., 1989; Totterdell and Meredith, 1997) and with recent behavioral data suggesting that the entorhinal cortex is the only parahippocampal region involved in LI (Coutureau et al., 1999). In conclusion, irrespective of the exact nature of the neural systems involved, the data collected during this study lend major credence to the view that DA projections in the anterior ST subregion are functionally related to networks processing the cognitive and affective values of environmental information in parallel, thus contributing to DA changes observed in LI phenomenon and conditioned responses. In this context it is important to notice that the suggestion that DA terminals are involved in the anterior ST in cognitive processes is borne out further by data obtained in humans showing that the head of the caudate is involved in some kind of cognitive operations (Abdullaev and Melnichuk, 1997; Abdullaev et al., 1998). Finally, the results of the present study might be considered in the context of the pathophysiology of schizophrenia. A reduction of LI in never-treated patients with schizophrenia has been reported by several authors (Baruch et al., 1988; Gray et al., 1992, 1995; Lubow et al., 2000; Rascle et al., 2001). In other respects, it is generally accepted that there is an unknown DAergic functional perturbation in schizophrenia (Swerdlow and Koob, 1987; Harrison, 1999; Carlsson et al., 2001). Recently, it has been shown that striatal DA release induced by d-amphetamine is increased in patients with schizophrenia (Laruelle et al., 1996; Breier et al., 1997). This effect appears to result from an abnormal glutamatergic regulation of ST DAergic neurons (Kegeles et al., 2000), suggesting that the control of DA release by glutamatergic projections originating from one or several forebrain structures, yet to be determined, may be impaired in certain tasks. Recognition memory has been found to be deficient in schizophrenics (Huron et al., 1995; Danion et al., 1999). Therefore, it is tempting to propose that a functional impairment of the neural system involved in recognition memory may contribute to an abnormal regulation of ST DA release and

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(Accepted 14 October 2002)