Endogenous neurosteroids influence synaptic GABAA receptors during postnatal development.

GABA plays a key role in both embryonic and neonatal brain development. For example, during early neonatal nervous system maturation, synaptic transmission, mediated by GABAA receptors (GABAA Rs), undergoes a temporally specific form of synaptic plasticity to accommodate the changing requirements of maturing neural networks. Specifically, the duration of miniature inhibitory postsynaptic currents (mIPSCs), resulting from vesicular GABA activating synaptic GABAA Rs, is reduced, permitting neurones to appropriately influence the window for postsynaptic excitation. Conventionally, programmed expression changes to the subtype of synaptic GABAA R are primarily implicated in this plasticity. However, it is now evident that, in developing thalamic and cortical principal- and inter-neurones, an endogenous neurosteroid tone (eg, allopregnanolone) enhances synaptic GABAA R function. Furthermore, a cessation of steroidogenesis, as a result of a lack of substrate, or a co-factor, appears to be primarily responsible for early neonatal changes to GABAergic synaptic transmission, followed by further refinement, which results from subsequent alterations of the GABAA R subtype. The timing of this cessation of neurosteroid influence is neurone-specific, occurring by postnatal day (P)10 in the thalamus but approximately 1 week later in the cortex. Neurosteroid levels are not static and change dynamically in a variety of physiological and pathophysiological scenarios. Given that GABA plays an important role in brain development, abnormal perturbations of neonatal GABAA R-active neurosteroids may have not only a considerable immediate, but also a longer-term impact upon neural network activity. Here, we review recent evidence indicating that changes in neurosteroidogenesis substantially influence neonatal GABAergic synaptic transmission. We discuss the physiological relevance of these findings and how the interference of neurosteroid-GABAA R interaction early in life may contribute to psychiatric conditions later in life.

Immunohistochemical assessment of mesotelencephalic dopamine activity during the acquisition and expression of Pavlovian versus instrumental behaviours.

Dopaminergic activity during Pavlovian or instrumental learning in key target regions of the mesotelencephalic dopamine system was investigated immunohistochemically using antibodies raised against glutaraldehyde-conjugated dopamine. Experiment 1 examined dopamine immunoreactivity during acquisition of a Pavlovian conditioned-approach response. Observations were taken at three stages of learning: initial, intermediate and asymptotic; each with a conditioned stimulus+ (CS+) group for whom visual or auditory stimuli immediately preceded an unconditioned stimulus (sucrose), and a conditioned stimulus- (CS-) group for whom stimuli and the unconditioned stimulus were unpaired. Animals learned to approach the alcove during CS+ presentations, whilst approach behaviour of the CS- group remained low. In general, target regions exhibiting a dopaminergic reaction responded maximally during the intermediate stage of acquisition, and were less responsive initially, and not responsive at all at asymptote. Specifically, the pattern of dopaminergic response was: shell more than core of the nucleus accumbens; prefrontal cortex, central and basolateral nuclei of the amygdala also significantly responsive. Mediodorsal and laterodorsal striatal regions were reactive only very early in training. Experiment 2 examined dopaminergic reaction following acquisition of a novel conditioned instrumental response. The conditioned response+ (CR+) group responded at a much higher rate on the lever for which unconditioned stimulus-associated stimuli were presented, than on the control lever. The conditioned response- (CR-) group responded at a low rate on both levers. In contrast with experiment 1, the most responsive regions were the core of the nucleus accumbens, medial prefrontal cortex and basolateral area of the amygdala. Thus, the acquisition, but not expression of Pavlovian associations activated dopamine within several key target regions of the mesotelencephalic dopamine system, and preferentially within the shell rather than core of the nucleus accumbens. By contrast, acquisition of a novel instrumental response preferentially activated the core of the nucleus accumbens, and basolateral area of the amygdala. These data carry significant implications for the potential role of these regions in learning and memory.

An immunohistochemical examination of the effects of sensitisation on the mesotelencephalic dopaminergic response to d-amphetamine.

The dopaminergic response to d-amphetamine with or without prior repeated experience with the drug was investigated immunohistochemically in key target regions of the mesotelencephalic dopamine system using antibodies raised against glutaraldehyde-conjugated dopamine. This methodology permitted the unambiguous determination of dopaminergic activity within specific subregions of structures implicated in the behavioural effects of psychomotor stimulants drugs, and in the expression of behavioural sensitisation. Experiment 1 examined dopamine immunoreactivity in central or basolateral amygdala, shell or core of the nucleus accumbens, medial and lateral caudate-putamen and medial prefrontal cortex following the administration of various doses of d-amphetamine to drug-naïve rats. Whilst dose-related increases in dopaminergic activity were detected in all regions examined, a regional heterogeneity was clearly evident. For example, d-amphetamine enhanced dopaminergic activity preferentially within the shell subregion of the nucleus accumbens both with respect to the core subregion and to other striatal and non-striatal areas. Experiment 2 examined changes in dopaminergic activity following the administration of a low dose of d-amphetamine to d-amphetamine-sensitised rats and saline-pretreated control animals. Regional heterogeneity both between and within terminal areas was again detected. Thus, there was evidence of a preferential increase in dopaminergic activity within the shell of the nucleus accumbens of sensitised rats. Moreover, sensitisation to d-amphetamine increased the dopaminergic response to acute administration of d-amphetamine within all striatal and non-striatal areas examined. Comparison of this effect across subterritories of the areas under investigation revealed that in sensitised rats, acute administration of d-amphetamine elevated dopaminergic activity within the shell of the nucleus accumbens to a greater extent than within the core. These data therefore indicate that systemic administration of d-amphetamine is associated with regionally heterogeneous changes in dopaminergic activity within terminal regions of the mesotelencephalic dopamine system in both sensitised and unsensitised rats. Moreover, the present methodology permitted resolution of these changes at an anatomical level beyond that of conventional approaches.

Enhanced dopamine efflux in the amygdala by a predictive, but not a non-predictive, stimulus: facilitation by prior repeated D-amphetamine.

Extracellular levels of dopamine within the amygdala were monitored using in vivo microdialysis during performance of an appetitive Pavlovian conditioning task in sensitized rats and unsensitized controls. Animals received exposure either to D-amphetamine or to vehicle for seven consecutive days (2 mg/kg/day, i.p.) in the home cage. Training began following a further seven injection-free days. Animals were exposed to two session types: during conditioning sessions, a stimulus (tone or light) immediately preceded sucrose pellet delivery. During control sessions, the alternative stimulus was also presented, but not in temporal proximity to an otherwise identical schedule of pellet delivery. There was a total of three alternating presentations of each session type during training. Sensitization enhanced Pavlovian conditioned approach behaviour to the stimulus predictive of imminent pellet delivery, and was without effect upon approach behaviours either to the food pellets themselves or to the control stimulus. Extracellular levels of dopamine within the amygdala were assessed during the fourth conditioning and control sessions. Mesoamygdaloid dopamine efflux increased significantly during the conditioning test session, but not during the control session, and this dopaminergic response was more marked in rats with prior repeated D-amphetamine experience. Hence, these results add to evidence suggesting a role for amygdaloid dopamine in appetitive Pavlovian conditioning, and in the facilitation of associative learning following prior experience of D-amphetamine.

Enhanced conditioned inhibition following repeated pretreatment with d-amphetamine.

We have shown that prior repeated exposure to d-amphetamine facilitates appetitive Pavlovian conditioning. However, animals sensitised in this manner also show elevated levels of stimulated activity. To investigate whether enhanced conditioning was dependent upon increased activity, a conditioned inhibition task was employed in the present study. Rats received d-amphetamine (2 mg/kg, i.p.) or vehicle once per day for 7 days. After a 7-day drug-free period, an activity assay confirmed that repeated d-amphetamine treatment markedly elevated the locomotor response to a subsequent challenge with 0.5 mg/kg d-amphetamine. Conditioning began 6 days later. In phase 1, stimulus A+ (light or tone) immediately preceded sucrose availability (excitatory conditioning). In phase 2, sucrose again was presented after A+ alone, but not after presentation of a compound of A+ with a second stimulus (AB-). Sensitisation enhanced the acquisition of conditioned approach behaviour to the excitatory stimulus A+ in phase 1. Furthermore, acquisition of conditioned inhibition to the stimulus compound, AB-, was also facilitated. Thus, sensitised rats showed reduced levels of responding to the stimulus compound far sooner than controls. Finally, a retardation test was carried out in stage 3, in which the inhibitory stimulus B- was paired alone with sucrose reward. Sensitised rats initially showed retarded acquisition of excitatory conditioned responding relative to controls, suggesting that B possessed stronger inhibitory associations in these animals. However, sensitised animals again exhibited higher levels of responding in later sessions, consistent with the enhanced excitatory conditioning shown in phase 1. These findings suggest that prior repeated d-amphetamine enhanced the acquisition of inhibitory and excitatory Pavlovian associations; a propensity not readily attributable to stimulated locomotor hyperactivity.

Post-session intra-perifornical region quinpirole infusions retard the consolidation of an appetitive differential conditioning task.

We have shown previously that post-session infusions of the D2/D3 dopamine receptor antagonist sulpiride within the perifornical region of the lateral hypothalamus (PFH) greatly facilitated the acquisition of an appetitive differential conditioning task. Here, we have examined the effects of the D2/D3 dopamine receptor agonist quinpirole on acquisition of this task. Conditioning sessions consisted of an initially neutral stimulus (CS+) preceding availability of a 10% sucrose reward (US), and a control stimulus (CS-), never associated with the US, were presented, followed by bilateral intra-PFH infusions of quinpirole (0, 0.1, 10 nmol). The acquisition of a conditioned approach response during the CS+ period was impaired by quinpirole administration in a dose-dependent manner. No other behavioural measure was affected. All groups subsequently acquired the conditioned approach response, following cessation of the post-session infusion procedure. The present data extends previous findings, suggesting that the level of activation of dopamine receptors in the vicinity of the PFH significantly influences the consolidation of appetitive Pavlovian associations.

A novel 96-well scintillation proximity assay for the measurement of apoptosis.

The translocation of phospholipids across the plasma membrane has been widely documented as one of the earliest measurable biochemical events of apoptosis. Using fluorescently labelled annexin V, which preferentially binds phosphatidylserine (PS) in the presence of Ca(2+), the externalization of PS can be measured and apoptosis quantified using flow cytometry. Conventional detection methods utilizing annexin V, while faster than in situ DNA end-labelling or DNA laddering, require extensive sample preparation which may compromise samples and makes rapid, high volume screening prohibitive. This paper describes a novel assay for the measurement of apoptosis based upon binding of radiolabelled annexin V to apoptotic cells attached to the growth surface of a 96-well scintillating microplate (Cytostar-T(R)). We compared measurements of apoptosis made by flow cytometry to those obtained with the scintillating microplate in three model systems, treatment of: mouse connective tissue (L-M) cells with lymphotoxin (LT), human lung carcinoma (H460) cells with Apo-2 ligand and human umbilical vein endothelial (HUVE) cells with staurosporine. In this assay, we compare both direct and indirect labelling methods by utilizing either iodinated annexin V or biotinylated annexin V/[(35)S] streptavidin to radiolabel apoptotic cells. The signal detected is a direct consequence of the binding of annexin V to externalized PS on apoptotic cells and the proximity of the label to the base of the plate. Using this method, separation of bound and unbound radiolabel signal occurs directly within the well resulting in a sensitive assay that requires minimal manipulation and can accomodate a large number of samples.

Interactions between intra-perifornical region sulpiride and intra-ventral tegmental area AP5 on measures of locomotor activity and conditioned place preference.

Infusions of sulpiride, a dopamine D2/D3 receptor antagonist within the perifornical region of the lateral hypothalamus have been shown previously to exhibit a behavioural profile generally attributed specifically to activation of the mesoaccumbens dopamine projection. Experiment 1 confirmed previous work showing that repeated homecage pretreatment with sulpiride (5 microg) in the perifornical region of the lateral hypothalamus resulted subsequently in an enhanced locomotor response to a d-amphetamine challenge. Experiment 2 examined the possibility that the observed behavioural changes were due to stimulation of the mesoaccumbens dopamine projection via the ventral tegmental area. Thus, repeated intra-perifornical infusions with sulpiride were without effect initially, but resulted in a gradual increase in locomotor activity during subsequent sessions. Intra-ventral tegmental area infusions of the NMDA receptor antagonist AP5 (0.3, 1.0 nmol) were without intrinsic effect upon locomotor activity at any time. However, AP5 blocked the ability of repeated sulpiride infusions to increase locomotor activity, and the ability of intra-perifornical sulpiride to support the acquisition of a conditioned place preference. AP5-sulpiride co-infusions also increased locomotor activity in a non-incremental manner. These data suggest there to be a functionally significant projection from the perifornical region of the lateral hypothalamus to the ventral tegmental area in the control over locomotor activity and rewarded behaviour.

Isolation rearing enhances the rate of acquisition of a discriminative approach task but does not affect the efficacy of a conditioned reward.

Male Lister hooded rats were reared from weaning either alone (isolation reared) or in groups of five (socially reared controls). Experiments began at 18-weeks postweaning. Subjects were trained to associate an arbitrary stimulus with 10% sucrose reward. Trials (VT30sec) consisted of a 5-s light stimulus (houselight off, wall lights on), followed by a 5-s period of access to the sucrose reward. Alcove approach between trials delayed the next trial by the duration of approach plus 3 s. Activity not associated directly with approach behaviour was also recorded. Isolation rearing enhanced the rate of acquisition of the discriminative approach response. Acquisition of both conditioned approach behaviour during trials and conditioned avoidance between trials was more rapid in isolates. In general, effects were most evident early in training, with asymptotic performance least affected. Horizontal and vertical activity extraneous to approach behaviour was enhanced in isolates during the first training session and increased further relative to social controls after several training sessions. Subsequently, two novel levers were presented: a response on one lever resulted in a 0.5-s presentation of the conditioned stimulus (CS) (probability 0.5), whereas the second lever had no programmed consequences. Sucrose reward was not available at any time. Both groups of animals showed a preference for the CS-associated lever, although rates of response by isolates were higher than social controls on both levers. The proportion of responses emitted upon the active lever, by comparison with the total number of responses recorded upon both levers, was, however, unaffected by isolation rearing.

Double dissociation of the behavioural effects of R(+) 7-OH-DPAT infusions in the central and basolateral amygdala nuclei upon Pavlovian and instrumental conditioned appetitive behaviours.

Dopaminergic cell bodies located within the ventral mesencephalon innervate the amygdaloid complex, a region critically involved in the attribution of affective significance to environmental stimuli. Recently, we have shown that post-session intra-amygdala administration of a D3 dopamine receptor agonist enhances selectively the acquisition of an appetitive conditioned response. In the present study, we have investigated the potential involvement of the central nucleus and the basolateral nuclei of the amygdala in mediating this effect. Thus, rats were trained to associate an arbitrary stimulus (CS+) with the availability of 10% sucrose reward. Post-session infusions of the D3 receptor-preferring agonist, R(+) 7-OH-DPAT, were made into either the central nucleus or basolateral nuclei. Acquisition of a conditioned approach response was enhanced by R(+) 7-OH-DPAT infusions within the central nucleus, but not within the basolateral nuclei. Drug infusions into either region failed to affect approach behaviour elicited by presentation of a control stimulus (CS-), explicitly unpaired with sucrose reward. The effects of pre-test infusions of R(+) 7-OH-DPAT on the instrumental properties of the stimuli were then determined. Rats were presented with two novel levers, depression of one lever resulted in presentation of the CS+, while presentation of the CS- was contingent upon depression of the other lever. Rates of response upon each lever as well as the ability of the conditioned stimuli subsequently to elicit conditioned approach behaviour were recorded. Data revealed a double dissociation of the effects of R(+) 7-OH-DPAT on the expression of the Pavlovian and instrumental properties of the reward-related stimulus. Thus, within the central nucleus R(+) 7-OH-DPAT dose-dependently attenuated expression of the conditioned approach response, but had no effect upon instrumental responding maintained by the conditioned reward. In contrast, within the basolateral nuclei, R(+) 7-OH-DPAT had no effect upon expression of conditioned approach behaviour, but abolished selectively the ability of the reward-associated stimulus to support the acquisition of a novel instrumental response. Hence, these data indicate that distinct regions of the amygdaloid complex process distinct aspects of conditioned appetitive behaviours.

Effects of intra-amygdala R(+) 7-OH-DPAT on intra-accumbens d-amphetamine-associated learning. I. Pavlovian conditioning.

We have previously obtained evidence that the mesoamygdaloid dopamine projection modulates the acquisition of a conditioned response (CR) elicited by presentation of a conditioned stimulus (CS) predicting the availability of a natural (sucrose) reward. This property was found to be dependent upon D3, but not D1 or D2, dopamine receptor activation. The aim of the present study was to determine whether the mesoamygdaloid dopamine projection is similarly involved in the acquisition of a drug-associated CR. Thus, two groups of rats with guide cannulae aimed at the nucleus accumbens and amygdala were trained using a Pavlovian conditioning procedure in which an initially neutral CS was paired with a computer-controlled, bilateral intraaccumbens infusion of d-amphetamine (the unconditioned stimulus: US). Conditioning sessions were conducted in standard operant chambers, with each session consisting of a single CS-US trial. For one group of rats, CS presentation was positively correlated with the drug US (Paired group), while for the second group CS and US presentations were negatively correlated (Unpaired group). During training, locomotor activity was recorded and was utilised as the measure both of the unconditioned (UR) and conditioned response (CR). A within-subjects design was utilised to investigate the effect of post-session bilateral intraamygdala administration of R(+) 7-OH-DPAT on the development of the drug-associated CR. Hence, both Paired and Unpaired groups were exposed to two different CSs which were presented on alternate sessions. Post-session bilateral intra-amygdala administration of R(+) 7-OH-DPAT (10 nmol) followed sessions in which one CS was presented, while intra-amygdala vehicle followed sessions in which the alternate CS was presented. The development of a CR occurred only in the presence of a CS that had been positively correlated with presentation of the drug US. Post-session, intra-amygdala administration of R(+) 7-OH-DPAT enhanced the acquisition of this CR. However, R(+) 7-OH-DPAT was without effect upon the unconditioned response to intra-accumbens d-amphetamine. Our previous data indicate a comparable effect of R(+) 7-OH-DPAT on conditioning to a CS associated with a non-drug, natural reward. Therefore, taken together, these findings suggest that D3 dopamine receptors within the amygdala modulate specifically the acquisition of Pavlovian conditioned responses, regardless of whether drug or natural rewards constitute the US.

Effects of intra-amygdala R(+) 7-OH-DPAT on intra-accumbens d-amphetamine-associated learning. II. Instrumental conditioning.

Rats were trained to associate an initially neutral conditioned stimulus (CS) with a response-independent, intra-accumbens infusion of d-amphetamine (the unconditioned stimulus; US). Elsewhere, we have reported that as a result of this training, presentations of the CS alone elicited a conditioned response consisting of increased locomotor activity and that acquisition of this conditioned response was enhanced by post-session, intra-amygdala infusion of the dopamine D3 receptor preferring agonist, R(+) 7-OH-DPAT. Here, in this same group of animals, we have examined the conditioned rewarding properties of the drug-associated CS by determining its ability to support the acquisition of a novel instrumental response in the absence of drug reward. Thus, rats were presented with two novel levers. Presentation of the drug-associated CS was made contingent upon depression of one of the levers (CR lever), while responding upon the other lever (NCR lever) had no programmed consequences. Preferential responding upon the lever delivering the drug-associated CS was observed despite a 6-week interval between CS-US training and the conditioned reward test. Intra-accumbens administration of d-amphetamine (0-20 microg) increased the control over behaviour exerted by the CS, increasing CR. but not NCR lever responding. In contrast, rats that received three pairings of an intra-accumbens infusion of d-amphetamine in combination with intra-amygdala infusion of R(+) 7-OH-DPAT, 3 weeks prior to testing, displayed similar rates of response upon both levers and were insensitive to the potentiation of responding for conditioned reward following intra-accumbens d-amphetamine. However, intra-accumbens d-amphetamine stimulated locomotor activity in a similar, dose-related manner in both groups. In this way, rats that had received intra-accumbens infusion of d-amphetamine in combination with intra-amygdala infusion of R(+) 7-OH-DPAT appeared exactly like control group rats, for which the CS had been paired with intra-accumbens d-amphetamine on a negative basis only. A locomotor activity test indicated that one behavioural consequence of intra-amygdala administration of R(+) 7-OH-DPAT was the reduction of the unconditioned locomotor response resulting from intra-accumbens administration of d-amphetamine. Hence, the present data demonstrate that the conditioned rewarding properties of a drug-associated CS are specific to the CS-US association and are relatively insensitive to decay over time. However, the rewarding properties of a drug-associated CS were selectively abolished following activation of amygdala D3 receptors during presentation of the drug reward. Potential explanations for this effect are discussed, including the possibility that intra-amygdala R(+) 7-OH-DPAT reduced the incentive value of the US.

Post-session sulpiride infusions within the perifornical region of the lateral hypothalamus enhance consolidation of associative learning.

Whilst neurons within the lateral hypothalamus are well known to be responsive to the presentation of previously learned associative stimuli, the consolidation of a Pavlovian association is thought to depend in large part upon other brain regions, including the amygdala. The present study addressed this assumption directly, by examining the effect of post-session infusions of sulpiride within the lateral hypothalamus upon the acquisition of a conditioned approach response in an appetitive differential conditioning task. Subjects were exposed to an initially neutral stimulus (CS+), which immediately preceded the availability of a 10% sucrose reward (US). A second, control stimulus (CS ) was also presented. but never in close temporal proximity to the US. The number and duration of alcove approaches were recorded. Immediately following each training session, subjects were infused bilaterally with sulpiride (0, 0.5, 5 microg) in the vicinity of the perifornical region of the lateral hypothalamus. Sulpiride dose-dependently enhanced the rate of acquisition of a conditioned approach response to presentation of the CS+, but was without affect upon approach behaviour during CS(-) or US presentations. Thus, 0.5 microg sulpiride facilitated at an early stage (session 2 onwards) the number of alcove approaches to the CS+, while 5 microg sulpiride enhanced to a greater extent the duration of conditioned approach, particularly during later sessions. A subsequent locomotor test using 0.5 mg/kg d-amphetamine indicated that repeated infusions of the higher dose sulpiride (5 microg), but not the lower dose (0.5 microg), resulted in behavioural sensitisation to administration of the psychomotor stimulant. Acquisition of a novel conditioned instrumental response was not affected by previous exposure to sulpiride. These data suggest that dopamine-sensitive neurons within the lateral hypothalamus may play a significant role in the acquisition of appetitive Pavlovian associations.

Isolation rearing enhances acquisition in a conditioned inhibition paradigm.

Isolation rearing from weaning has been reported to enhance excitatory conditioning. The present study employed a conditioned inhibition procedure to examine whether this result was attributable to locomotor hyperactivity. Rats were raised from 21 days old in isolation or in groups of five. In Phase 1, presentation of stimulus A+ was followed immediately by sucrose availability (excitatory conditioning). In Phase 2, sucrose was again presented after A+ alone, but not after a compound presentation of A+ with a second stimulus, B-. Thus, B is believed to acquire conditioned inhibitory properties, countering the excitatory impact of A, and reducing responding specifically to this stimulus compound. Isolates showed enhanced excitatory conditioning in Phase 1. Furthermore, acquisition of conditioned inhibition in Phase 2 was also facilitated by isolation rearing. In Phase 3, B- was paired with a period of sucrose availability. Isolation rearing initially retarded responding to B-, confirming that this stimulus possessed a greater degree of behavioural inhibition in these animals. Later in training, isolates showed enhanced excitatory conditioning to B-, as shown previously to A+ in Stage 1. These results suggest that isolation rearing enhances the acquisition of appetitive Pavlovian associations, independently of locomotor hyperactivity.

Enhanced appetitive conditioning following repeated pretreatment with d-amphetamine.

The behavioural response to psychomotor stimulants is augmented with repeated exposure to these drugs. Enhanced stimulated dopamine overflow within the nucleus accumbens and amygdala has been found to accompany this behavioural sensitization. In the present experiment, rats received 2 mg/kg d-amphetamine or 1 ml/kg physiological saline once per day for 5 days. Five days later, a behavioural assay confirmed that prior repeated d-amphetamine treatment markedly enhanced the locomotor activating effects of a d-amphetamine (0.5 mg/kg, i.p.) challenge. Training on a Pavlovian conditioning task began six days subsequently. In Stage 1, a stimulus (light or tone, S-) was presented negatively correlated with a sucrose reward. In Stage 2, presentation of the alternative counterbalanced stimulus (light or tone, S+) was paired with the availability of a 10% sucrose solution. There were no differences between the two groups in their response to the the S- stimulus. However, sensitized animals showed a selective enhancement in the acquisition of conditioned responding to S+, relative to vehicle-injected controls. No differences in behaviour were recorded during the prestimulus periods, nor during presentations of sucrose. Levels of activity within the operant chamber extraneous to alcove approach were also similar in both groups of animals. The conditioned instrumental efficacy of S+, relative to S- was assessed in Stage 3, in which stimulus availability was made contingent on a novel lever-pressing response. Both groups showed a similar preference for the S+ over the S- stimulus. Hence, rats sensitized by prior repeated d-amphetamine showed enhanced appetitive Pavlovian conditioning, without subsequent effect on conditioned reward efficacy. These data are discussed in light of possible changes in mesoamygdaloid dopamine functioning.

Interactions between sulpiride infusions within the perifornical region of the lateral hypothalamus and the nucleus accumbens on measures of locomotor activity and conditioned place preference.

There is growing evidence of a functionally significant link between the perifornical region of the lateral hypothalamus (PFH) and the mesoaccumbens dopamine projection. The present study assessed the ability of intra-accumbens dopamine receptor blockade, firstly to impair the locomotor stimulant properties of intra-PFH sulpiride, and secondly the ability of intra-PFH sulpiride to support a conditioned place preference. Locomotor activity and conditioned place preference training were assessed using an apparatus consisting of three distinctive compartments. Following a pre-exposure session in which groups showed equivalent preference for the three compartments, conditioned place preference training was carried out over five drug (0, 0.03 or 0.1 microg intra-accumbens sulpiride; 0 or 5 microg intra-PFH sulpiride) and five vehicle sessions. For each animal, drug and vehicle treatments were randomly paired with one of the two outermost compartments. Locomotor activity was monitored during each session. Intra-PFH sulpiride alone increased locomotor activity during later sessions, and this gradual increase in locomotor response, relative to vehicle infusions, was blocked by intra-accumbens sulpiride co-infusion. On the final drug-free test day for conditioned place preference, animals were again permitted free access to all three compartments, and the time spent in each was compared with that on initial exposure. Intra-PFH sulpiride alone supported a conditioned place preference. Intra-accumbens sulpiride was without intrinsic effect, but nevertheless blocked the ability of intra-PFH sulpiride to support a conditioned place preference. These data provide further support for a functionally significant link between the PFH and mesoaccumbens dopamine projection, with regard to the locomotor stimulant properties and ability of intra-PFH infusions of sulpiride to support a conditioned place preference.

Isolation rearing enhances the locomotor stimulant properties of intra-perifornical sulpiride, but impairs the acquisition of a conditioned place preference.

Previous data indicated that infusions of the D2/D3 dopamine receptor antagonist sulpiride within the perifornical region of the lateral hypothalamus may engage neural circuitry relevant to activation of the mesoaccumbens dopamine projection. The present work examined this proposition further. Experiment 1 examined the ability of intra-perifornical sulpiride to induce a conditioned place preference, using an unbiased conditioning procedure. Thus, bilateral guide cannulae were implanted to gain access to the perifornical region of the lateral hypothalamus. Following recovery, animals were subjected to an initial exposure to the place preference apparatus. The apparatus consisted of three distinctive compartments, the central compartment allowing access to the two outermost compartments. Initial exposure indicated equal preference for each. Then, in alternating sessions, animals received infusions of sulpiride (5, 10 or 20 micrograms) before being placed in one of the two outermost compartments, and infusions of vehicle before being placed in the alternate compartment. Compartment-drug pairings were counterbalanced across animals. Four drug, and four saline sessions were completed, each being separated by at least 2 full days. On the final test day, animals were allowed free access to compartments, and the time spent in each was compared with that of initial exposure. Results showed that intra-perifornical sulpiride increased activity during drug-conditioning sessions in an incremental fashion, and supported dose-dependently the acquisition of a conditioned place preference. Experiment 2 examined the effects of isolation rearing upon the locomotor stimulant properties of intra-perifornical sulpiride, and the acquisition of a conditioned place preference. Rats were raised from weaning either alone (isolation-reared) or in groups of five (socially-reared controls) until 4 months of age. Consistent with previous reports of the effects of isolation rearing upon psychomotor stimulant responsivity, here isolates were found to be more responsive to the locomotor stimulant properties of intra-perifornical sulpiride, but were less responsive to the ability of intra-perifornical sulpiride to support the acquisition of a conditioned place preference. These data were suggested to provide further support for the proposition that blockade of dopamine receptors of the D2 family within the perifornical region of the lateral hypothalamus results in the activation of the mesoaccumbens dopamine projection, via the ventral tegmental area.

Enhanced stimulus-reward learning by intra-amygdala administration of a D3 dopamine receptor agonist.

The amygdala is considered to be a critical neural substrate underlying the formation of stimulus-reward associations, and is known to receive substantial innervation from dopaminergic neurons located within the ventral mesencephalon. However, relatively little is known about the function of the mesoamygdaloid dopamine projection in stimulus-reward learning. Recently, we have found post-session intra-amygdala microinjections of d-amphetamine to enhance appetitive Pavlovian conditioning as assessed in a discriminative approach task. In the present study, we have examined the effects of dopamine receptor agonists possessing relative selectivity for the D1, D2 and D3 receptor subtypes in order to examine more fully the role of the mesoamygdaloid dopamine projection in stimulus-reward learning. Thus, subjects were trained to associated an initially neutral stimulus (CS+) with 10% sucrose reward (US). A second, control stimulus (CS-) was also presented but never paired with sucrose reward. In order to measure specifically the conditioned response to CS+/CS- presentation, responding during CS and US presentations was measured separately. Immediately following each training session, subjects received bilateral intra-amygdala infusion of 0.1, 1 or 10 nmol/side of SKF-38393, quinpirole or 7-OH-DPAT. Infusions of SKF-38393 or quinpirole were without effect on CS+ approach. However, post-session intra-amygdala infusions of 7-OH-DPAT enhanced selectively CS+ approach in a dose-dependent fashion. No dose of any drug affected CS- approach, US behaviours, or measures of extraneous behaviour. Subsequent acquisition of a novel conditioned instrumental response was also unaffected. Thus, the present data indicate a selective involvement of the D3 dopamine receptor subtype in the modulation of stimulus-reward learning by the mesoamygdaloid dopamine projection.

Cross-sensitisation with d-amphetamine following repeated intra-perifornical sulpiride infusions.

Infusions of the dopamine D2/D3 receptor antagonist sulpiride within the perifornical region of the lateral hypothalamus have been reported to increase locomotor activity. The current investigation examined the effect of repeated lateral hypothalamic sulpiride infusions. In experiment la, rats were placed repeatedly in an activity chamber either prior to, or following an infusion of 10 micrograms sulpiride or vehicle. Repeated infusions of sulpiride prior to, but not following exposure to the activity chamber increased locomotor activity during subsequent sessions. In experiment 1b, repeated pretreatment with intra-perifornical sulpiride prior to placement within the activity chamber was found to engender a significant increase in conditioned activity when placed subsequently within the same chamber drug-free. Alternatively, pretreatment with sulpiride in the home cage was found subsequently to engender a significant increase in locomotor activity during a test session with intra-perifornical sulpiride. In experiment 2, repeated pretreatment with intra-perifornical sulpiride significantly increased the locomotor response to a subsequent systemic challenge with d-amphetamine. Animals pretreated in the home cage exhibited a moderate increase in activity over vehicle controls, while animals repeatedly pretreated immediately prior to placement in the activity chamber exhibited the largest response subsequently to d-amphetamine of any group. Experiment 3 showed that repeated sulpiride infusions either 1 mm anterior or 1 mm posterior to the perifornical region were without effect upon locomotor activity. These data are suggested to reflect an indirect action of intra-perifornical sulpiride upon the mesoaccumbens dopamine projection, via the level of the ventral tegmental area. Precise neural mechanisms are under current investigation.