Doctor of Philosophy / Department of Psychology / Stephen W. Kiefer / Methamphetamine is a potent psychomotor stimulant and a major drug of abuse. There is currently no effective medication available for treatment for methamphetamine addiction. The present study investigated the role of the dopamine D3 receptor on IV methamphetamine self-administration and its effect on methamphetamine induced neurochemical changes. Acute administration of the putative D3 receptor antagonists PG-01037 (10, 30 mg/kg, ip) and SB-277011A (12, 24, mg/kg, ip) significantly decreased the break-point for methamphetamine self-administration under a progressive-ratio (PR) schedule by 45 - 70%. Furthermore, both drugs dose dependently attenuated methamphetamine -triggered reinstatement of drug-seeking behavior in the reinstatement model of relapse. As with other drugs of abuse, the rewarding effects of methamphetamine are believed to be mediated by elevated levels of extracellular dopamine in the mesocorticolimbic system. The present study utilized in vivo microdialysis to examine the neurochemical mechanisms modulating the rewarding effects of methamphetamine actions evident in the various animal models of addiction. In the nucleus accumbens and ventral pallidum, acute methamphetamine (1.0 mg/kg, i.p.,) increased extracellular dopamine by 800 - 900% and decreased GABA by 60 – 65 % in the nucleus accumbens and ventral pallidum. Pretreatment with SB-277011A (12, 24 mg/kg) potentiated the methamphetamine induced dopamine increase but attenuated the methamphetamine-induced GABA decrease. Take together these data suggest that D3 selective antagonists’ pharmacotherapeutic potential in the treatment of methamphetamine addiction may involve a GABAergic mechanism.

Les antipsychotiques sont des antagonistes du récepteur dopaminergique de type 2 (DRD2) et constituent le principal traitement pharmacologique de la schizophrénie. Le traitement à long-terme par les antipsychotiques peut causer la tolérance au traitement et le développement de la dyskinésie tardive, dont le mécanisme est mal compris et le traitement, insatisfaisant. Il a été démontré que le traitement chronique aux antipsychotiques induit la surexpression du récepteur DRD2 et le stress oxydatif, deux mécanismes associés à la dyskinésie tardive. Le lien entre ces deux phénomènes est mal déterminé. Précédemment, il a été rapporté, dans mon laboratoire d'accueil, que le stress oxydatif, induit par le peroxyde d'hydrogène (H[indice inférieur 2]O[indice inférieur 2]), augmente le niveau du récepteur DRD2 sur la lignée cellulaire de neuroblastomes humains SH-SY5Y. L'hypothèse de recherche présentée ici est que le stress oxydatif est impliqué dans la surexpression du récepteur DRD2 induite par les antipsychotiques et que l'administration d'un antioxydant peut atténuer cette surexpression. Le projet présenté rapporte ainsi les effets des antipsychotiques sur les niveaux d'expression du récepteur DRD2 et les effets de l'inhibition du stress oxydatif par le traitement de l'acide lipoïque, un antioxydant, sur la lignée cellulaire de neuroblastomes humains SH-SY5Y. L'halopéridol, un antipsychotique de première génération, induit une augmentation des niveaux d'expression du récepteur DRD2, alors que Pamisulpride, un antipsychotique de deuxième génération, n'a pas d'effet significatif. De plus, l'halopéridol induit une plus importante augmentation des biomarqueurs du stress oxydatif (carbonylation des protéines, peroxydation lipidique et production de l'anion superoxyde) que l'amisulpride. L'acide lipoïque atténue la surexpression du récepteur DRD2 et le stress oxydatif induit par l'antipsychotique. L'inhibition de la synthèse de catécholamine par l'alpha-méthyl-DL-tyrosine (AMPT) élève l'expression du DRD2 et prévient sa surexpression par les antipsychotiques. Les résultats suggèrent que la surexpression du récepteur DRD2 induite par l'halopéridol est liée au stress oxydatif et proposent des mécanismes potentiels par lesquels l'acide lipoïque peut être considéré comme un agent thérapeutique pour la prévention et le traitement des effets secondaires reliés à l'utilisation des antipsychotiques.

The neurotransmitters serotonin (5-HT) and dopamine (DA) play roles in eating disorders, mood disorders, such as depression and anxiety, and in the regulation of locomotion. The 5-HT2C receptor is one of fourteen 5-HT receptor subtypes that is expressed in regions of the brain including the hippocampus, amygdala, dorsal striatum, nucleus accumbens (NA) and substantia nigra, and is therefore implicated in behaviours and disorders associated with these regions. 5-HT has been shown to exert both a tonic and phasic inhibitory control, through the 5-HT2C receptor, on the firing rate and bursting activity of DA-containing neurones in the ventral tegmental area which enhances DA release in the NA and prefrontal cortex. In addition, the 5-HT2C receptor is under the control of a monophasic diurnal rhythm and is in a position to alter circadian regulation and behaviour due to its expression in the suprachiasmatic nucleus (the light entrainable oscillator (LEO)). It was hypothesised that elevating expression of the 5-HT2C receptor would have a detrimental effect on mood and cause hypolocomotion while reducing 5-HT2C receptor expression would improve mood, cause hyperphagia, obesity and hyperlocomotion. In order to investigate these hypotheses mouse models that either over- or under-expressed the 5-HT2C receptor were implemented. The 5-HT2C receptor expression pattern and levels were confirmed in all mouse lines. A behavioural phenotype of hypolocomotion and increased anxiety in the 5-HT2C receptor over-expressing mice and hyperphagia, obesity and hyperlocomotion in the 5-HT2C receptor under-expressing mice were found the latter is conistent with current literature. During backcrossing of the mouse lines onto the C57Bl/6 genetic background the abnormal behavioural phenotypes were lost suggesting that 5-HT2C receptor function is particulary sensitive to the genetic background on which it is being expressed. In response to altered expression levels of 5-HT2C receptor, compensatory alterations were found in the 5-HT system, with an inverse relationship existing between both the 5-HT1A receptor mRNA expression levels and 5-HT release in the hippocampus with the expression levels of the 5-HT2C receptor. Over-expression of 5-HT2C receptor appears to inhibit DA release in the cortex. The circadian experiments showed that under-expressing the 5-HT2C receptor did not alter the regulation of the food entrainable oscillator and there was a suggestion that the regulation of the LEO was affected. In summary, these results demonstrate that altered expression of 5-HT2C receptors results in abnormal behaviours consistent with its role in psychiatric disorders, but that the outcome is dependent on the genetic background.

published_or_final_version / abstract / Zoology / Doctoral / Doctor of Philosophy

The neurotransmitter dopamine (DA) represents a neural substrate for positive motivation as its spatiotemporal distribution across the brain is responsible for goaldirected behavior and learning reward associations. The critical determinant of DA release throughout the brain is the firing pattern of DA-producing neurons. Synchronized bursts of spikes can be triggered by sensory stimuli in these neurons, evoking phasic release of DA in target brain areas to drive reward-based reinforcement learning and behavior. These bursts are generated by NMDA-type glutamate receptors (NMDARs). This dissertation reports a novel form of long-term potentiation (LTP) of NMDARmediated excitatory transmission at DA neurons as a putative cellular substrate for changes in DA neuron firing during reward learning. Patch-clamp electrophysiological recording from DA neurons in acute brain slices from young adult rats demonstrated that synaptic NMDARs exhibit LTP in an associative manner, requiring coordinated pre- and postsynaptic burst firing. Ca2+ signals produced by postsynaptic burst firing needed to be amplified by preceding metabotropic neurotransmitter inputs to effectively drive plasticity. Activation of NMDARs themselves was also necessary. These two coincidence detectors governed the timingdependence of NMDAR plasticity in a manner analogous to the timing rule for cuereward learning paradigms in behaving animals. Further mechanistic study revealed that PKA, but not PKC, activity gated LTP induction by regulating the magnitude of Ca2+ signal amplification via the inositol 1,4,5-triphospate (IP3) receptor and release of Ca2+ from intracellular stores. Plasticity of NMDARs was input specific and appeared to be expressed postsynaptically, but was not associated with a change in NMDAR subunit stoichiometry. LTP of NDMARs was DA-independent, and was specific for NMDARs: the same induction protocol produced long-term depression of AMPA receptors. NMDARs that had undergone LTP could be depotentiated in a spike-conditional manner, consistent with active unlearning. Finally, repeated, in vivo amphetamine experience dramatically increased facilitation of spike-evoked Ca2+ signals, which in turn drove enhanced plasticity. NMDAR plasticity thus represents a potential neural substrate for conditioned DA neuron burst responses to environmental stimuli acquired during reward-based learning as well a novel therapeutic target for intervention-based therapy of addictive disorders. / text

The goal of this dissertation was two fold: 1) To relate dopamine responses in the ventral striatum to ethanol preference and consumption, and 2) to investigate the role of the mu opioid receptors in this ethanol induced dopamine release in the ventral striatum. First a two bottle choice experiment established that a substrain of C57BL/6 mice (C57BL/6NCrl) had significantly less preference for and consumption of ethanol than a second substrain of mouse based on the same background (C57BL6/J). The C57BL/6 strain has been extensively used in alcohol drinking studies and is well known for it’s propensity to consume alcohol over water. To determine if differences in ventral striatal dopamine response vii could contribute to this variability in drinking behavior, we characterized the dopamine response in both substrains of mice after intraperitoneal injections of 1.0, 2.0 or 3.0 g/kg ethanol or saline. We found that the acute intraperitoneal ethanol injections in naïve mice caused a significant elevation in dopamine in both substrains at all three doses with a significant difference between substrains at the two highest alcohol doses. Therefore, ethanol induced dopamine release in the ventral striatum may contribute to ethanol preference and consumption. Next, we investigated the effect of acute intraperitoneal ethanol injections on naïve mu opioid receptor knockout mice and in mice pretreated with a mu opioid receptor antagonist. The mice used were all established on the C57BL/6J background. We found that ventral striatal dopamine response was similar in these mice after 1.0, 2.0 and 3.0 g/kg intraperitoneal ethanol injections compared to appropriate controls. As both gene deletion and pharmacological blockade of the mu opioid receptor did not affect ethanol stimulated dopamine release, it points to the conclusion that this receptor may not play a significant role in ethanol induced ventral striatal dopamine release. / text

The current project examined the ability of nicotine to modulate dopamine transporter (DAT) function. Initial experiments determined the dose-response for nicotine to modulate dopamine (DA) clearance in rat striatum and medial prefrontal cortex (MPFC) using in vivo voltammetry and determined if this effect was mediated by nicotinic receptors (nAChRs). In both striatum and MPFC, nicotine increased DA clearance in a mecamylamine-sensitive manner, indicating nAChR-mediation. The effect of acute nornicotine on DAT function was also determined. In contrast to nicotine, nornicotine in a dose-related manner decreased striatal DA clearance in a mecamylamine-sensitive manner, indicating nAChR mediation. To determine if tolerance developed to the nicotine effect nicotine, separate groups of rats were injected once daily for 5 days with nicotine or saline. DA clearance in striatum and MPFC was determined 24 hrs after the last injection. Nicotine increased DA clearance only 10-15% in the group repeatedly administered nicotine, demonstrating that tolerance developed. To determine if nicotine altered striatal DAT efficiency, following nicotine injection, DAT density and maximal velocity of [3H]DA uptake was determined using [3H]GBR12935 binding and saturation analysis of [3H]DA uptake in rat striatum, respectively. Nicotine did not alter the Bmax or Kd of maximal binding of [3H]GBR12935 binding. However, an increase in Vmax was observed at 10 and 40 min following nicotine injection, suggesting that nicotine increases DAT efficiency. To determine if systemic nicotine enhanced DAT function via an action at nAChRs on striatal DA terminals, [3H]DA uptake was determined in striatum in vitro in the absence or presence of nicotine in the buffer. Nicotine did not alter the Vmax for [3H]DA uptake in vitro, suggesting that the nicotine-induced increase in DAT function observed in vivo is mediated by nAChRs on DA cell bodies or another site which indirectly alters DAT function. To determine if the increase in DAT efficiency was due to increased surface expression of striatal DAT, biotinylation and Western blot analyses were performed. Nicotine did not alter striatal DAT, suggesting that the nicotine-induced increase in DA clearance in vivo and DAT efficiency in vitro is not the result of increased trafficking of this protein to the cell surface.

Dans ce travail, nous avons développé le modèle de rat ayant subi une lésion néonatale de l'hippocampe ventral (rat LNHV), un outil reconnu pour l'étude de la schizophrénie. <br />Nous avons validé l'utilisation de l'imagerie anatomique (IRM) comme outil permettant une détection fiable et précoce (avant la puberté) de la taille et la cible des lésions, pour remplacer l'évaluation post mortem.<br />Au niveau comportemental, nous avons montré que le nombre des anomalies qui sont associées à des troubles de la transmission dopaminergique augmente du stade juvénile à l'âge adulte chez les animaux LNHV. <br />Au niveau neurochimique, nous avons montré que les animaux LNHV présentent des altérations des efflux de dopamine au niveau mésocorticolimbique (noyau accumbens et cortex préfrontal) qui n'apparaissent qu'après la puberté. <br />Nos résultats confirment l'implication majeure de la dopamine dans la physiopathologie de la schizophrénie et soulignent l'importance des facteurs qui pourraient précipiter l'émergence de la maladie au moment de l'adolescence.

To test the hypothesis that the hypothalamic tuberoinfundibular dopaminergic neuronal system plays a role in the dynamic regulation of pituitary prolactin secretion, its activity was correlated with experimentally-induced prolactin secretory episodes in the male rat. Direct estimates of tuberoinfundibular neuronal activity were made by measuring its rates of dopamine and norepinephrine synthesis or release. Prolactin secretion was assessed in vivo by measuring radioimmunoassayable prolactin levels in peripheral blood and the pituitary and in vitro by measuring prolactin concentrations released into incubation media. The anesthetic urethane and a substance isolated from the pineal gland were both demonstrated to inhibit prolactin secretion. Significant elevations of newly synthesized tuberoinfundibular dopamine were observed concomitant with this decreased prolactin secretion suggesting that acute increases in tuberoinfundibular dopaminergic neuronal activity were perhaps causally related to acute decreases in prolactin secretion since these substances were without a direct effect on the pituitary in vitro. Conversely, acute decreases in tuberoinfundibular neuronal activity induced by dopamine biosynthesis inhibition or mimicked by pituitary receptor blockade induced acute increases in prolactin secretion. As another prerequisite for its involvement in the dynamic regulation of prolactin secretion, the tuberoinfundibular neuronal system was demonstrated to be involved in the negative feedback control of prolactin over its own secretion. Elevated circulating prolactin levels produced by pituitary homografts transplanted beneath the kidney capsule accelerated tuberoinfundibular dopaminergic neuronal activity. In two unrelated experimental conditions, rats rendered blind and anosmic or hyperprolactinemic, the chronic inhibition of prolactin secretion was not associated with the maintenance of an increased tuberoinfundibular neuronal activity, but rather with a supersensitivity of the anterior pituitary to the prolactin-release-inhibitory action of dopamine. Long-lasting alterations in tuberoinfundibular dopaminergic neuronal activity appeared to induce this pituitary supersensitivity to dopamine. The tuberoinfundibular neuronal system appears to have the capacity to modulate prolactin secretory episodes via the alteration of its dopaminergic activity. Long-lasting alterations in this activity may induce changes in anterior pituitary sensitivity to dopamine essential for the chronic inhibition of pituitary prolactin secretion.

Posttraumatic stress disorder (PTSD) is a chronic disabling condition that results from exposure to traumatic stress. However, although trauma is fairly common, PTSD will only occur in a small proportion of people. This suggests that resilience is a common response to trauma. The neurobiology underlying this adaptive response is thought to involve reward related areas as well as reward functions. This dissertation proposes that reward and reward-related areas have a role in anxiety disorders such as PTSD. This hypothesis was explored using an animal model of PTSD as well as a human mode of psychosical stress. The hypothesis that the ventral tegmental area (VTA), crucial for reward processing, is part of the neural circuitry involved in the symptomatology of PTSD was explored. To assess the role of VTA in PTSD, cells in this area were reversibly inactivated during a single exposure to inescapable foot-shock in a rodent model. Animals that underwent inactivation of VTA neurons decreased avoidance and lowered long-term anxiety-like behaviors in comparison with control groups. To assess short- and long-term electrophysiological effects of trauma on VTA cells, in vivo extracellular recordings were conducted. Results showed that the firing frequency of VTA cells changed both in the short- and long-term, following shock procedures. A human model of psychosical stress was used to test the hypothesis that the ability to respond appropriately to positive stimuli is important for the preservation of positive emotions following stressful events. The results show a positive correlation between trait resilience and trait reward sensitivity. To investigate the link between resilience and reward sensitivity further, the empirical portion of this study used a Monetary Incentive Delay Task (MID) to measure reward sensitivity before and after exposure to a psychosocial stressor. Moreover, behavioral reward sensitivity (as measured by MID and self-report satisfaction after the reward task) also correlated positively with trait and behaviorally measured resilience. The results shown in this dissertation suggest that the neural circuits involved in reward processing and reward function may be involved in resilient responses to stress.