Selective genetic disruption of dopaminergic, serotonergic and noradrenergic neurotransmission: insights into motor, emotional and addictive behaviour.

BACKGROUND: The monoaminergic transmitters dopamine (DA), noradrenaline (NE) and serotonin (5-HT) modulate cerebral functions via their extensive effects in the brain. Investigating their roles has led to the creation of vesicular monoaminergic transporter-2 (VMAT2) knockout (KO) mice. While this mutation results in postnatal death, VMAT2-heterozygous (HET) mice are viable and show a complex behavioural phenotype. However, the simultaneous alteration of the 3 systems prevents investigations into their individual functions. METHODS: To assess the specific role of NE, 5-HT and DA, we genetically disrupted their neurotransmission by creating conditional VMAT2-KO mice with targeted recombination. These specific recombinations were obtained by breeding VMAT2(lox/lox) mice with DBHcre, SERTcre and DATcre mice, respectively. We conducted a complete neurochemical and behavioural characterization of VMAT2-HET animals in each system. RESULTS: Conditional VMAT2-KO mice revealed an absence of VMAT2 expression, and a specific decrease in the whole brain levels of each monoamine. Although NE- and 5-HT-depleted mice are viable into adulthood, DA depletion results in postnatal death before weaning. Interestingly, alteration of the DA transmission fully accounted for the increased amphetamine response formerly observed in the VMAT2-HET mice, whereas alteration of the 5-HT system was solely responsible for the increase in cocaine response. LIMITATIONS: We used VMAT2-HET mice that displayed a mild phenotype. Because the VMAT2-KO in DA neurons is lethal, it precluded a straightforward comparison of the full KOs in the 3 systems. CONCLUSION: Given the intermingled functions of NE, 5-HT and DA in regulating cognitive and affective functions, this model will enhance understanding of their respective roles in the pathophysiology of psychiatric disorders.

Chronic stress alters the dendritic morphology of callosal neurons and the acute glutamate stress response in the rat medial prefrontal cortex.

We have previously reported that interhemispheric regulation of medial prefrontal cortex (PFC)-mediated stress responses is subserved by glutamate (GLU)- containing callosal neurons. Evidence of chronic stress-induced dendritic and spine atrophy among PFC pyramidal neurons led us to examine how chronic restraint stress (CRS) might alter the apical dendritic morphology of callosal neurons and the acute GLU stress responses in the left versus right PFC. Morphometric analyses of retrogradely labeled, dye-filled PFC callosal neurons revealed hemisphere-specific CRS-induced dendritic retraction; whereas significant dendritic atrophy occurred primarily within the distal arbor of left PFC neurons, it was observed within both the proximal and distal arbor of right PFC neurons. Overall, CRS also significantly reduced spine densities in both hemispheres with the greatest loss occurring among left PFC neurons, mostly at the distal extent of the arbor. While much of the overall decrease in dendritic spine density was accounted by the loss of thin spines, the density of mushroom-shaped spines, despite being fewer in number, was halved. Using microdialysis we found that, compared to controls, basal PFC GLU levels were significantly reduced in both hemispheres of CRS animals and that their GLU response to 30 min of tail-pinch stress was significantly prolonged in the left, but not the right PFC. Together, these findings show that a history of chronic stress alters the dendritic morphology and spine density of PFC callosal neurons and suggest a mechanism by which this might disrupt the interhemispheric regulation of PFC-mediated responses to subsequent stressors.

Uninterrupted in vivo cerebral microdialysis measures of the acute neurochemical response to a single or repeated concussion in a rat model combining force and rotation.

Altered extracellular amino acid concentrations following concussion or mild traumatic brain injury can result in delayed neuronal damage through overactivation of NMDA glutamatergic receptors. However, the consequences of repeated concussions prior to complete recovery are not well understood. In this study, we utilized in vivo cerebral microdialysis and a weight-drop model to investigate the acute neurochemical response to single and repeated concussions in adult rats that were fully conscious. A microdialysis probe was inserted into the hippocampus and remained in place during impact. Primary outcomes included concentrations of glutamate, GABA, taurine, glycine, glutamine, and serine, while secondary outcomes were righting times and excitotoxic indices. Compared to sham injury, the first concussion resulted in significant increases in glutamate, GABA, taurine, and glycine levels, longer righting times, and higher excitotoxic indices. Following the second concussion, righting times were significantly longer, suggesting cumulative effects of repeated concussion while only partial increases were observed in glutamate and taurine levels. GABA and glycine levels, and excitotoxic indices were comparable to sham injury. These findings suggest that single and repeated concussions may induce acute increases in several amino acids, while repeated concussions could exacerbate neurological symptoms despite less pronounced neurochemical changes.

Exposure to high fat during early development impairs adaptations in dopamine and neuroendocrine responses to repeated stress.

Perturbations in the perinatal environment have been shown to significantly alter mesolimbic dopamine (DA) and hypothalamic-pituitary-adrenal (HPA) responses to stressors in adulthood. We have previously demonstrated that adult offspring exposed to high fat during the last week of gestation and throughout lactation display permanent alterations in mesolimbic DA function and behavior. The goal of the present study was to investigate nucleus accumbens (NAc) DA and HPA responses to acute and repeated stress in high fat exposed (HFD, 30% fat) and control (CD, 5% fat) offspring. Using microdialysis to monitor extracellular DA, we report that adult HFD offspring show an enhanced NAc DA response to acute tail-pinch compared to CD offspring. With repeated tail-pinch, the response of the HFD animals remains unchanged while CD offspring exhibit a sensitized DA response. The pattern of the DA response to both acute and repeated stress is also significantly altered by early diet exposure with an earlier peak and faster return to baseline levels in CD compared with HFD offspring. Similarly, neuroendocrine adaptations to repeated tail-pinch are observed in CD animals, but not in HFD animals. While controls display a habituated adrenocorticotropic hormone (ACTH) response to repeated tail-pinch, and an exacerbated ACTH response to a novel stressor, this effect was not observed in the HFD offspring. Together, our data demonstrate that exposure to high fat during early development impairs adaptations in NAc DA and HPA responses usually observed with repeated stress.

Reduced dopamine release in Dcc haploinsufficiency male mice abolishes the rewarding effects of cocaine but not those of morphine and ethanol.

RATIONALE: The Netrin-1/DCC guidance cue pathway is critically involved in the adolescent organization of the mesocorticolimbic dopamine circuitry. Adult mice heterozygous for Dcc show reduced dopamine release in the nucleus accumbens in response to amphetamine and, in turn, blunted sensitivity to the rewarding effects of this drug. OBJECTIVE: Here, we tested whether the protective effects of Dcc haploinsufficiency are specific to stimulant drugs of abuse or instead extrapolate to opioids and ethanol. METHODS: We used the place preference paradigm to measure the rewarding effects of cocaine (20 mg/kg), morphine (5 or 10 mg/Kg), or ethanol (20%) in adult (PND 75) male Dcc haploinsufficient mice or their wild-type litter mates. In a second experiment, we compared in these two genotypes, in vivo dopamine release in the nucleus accumbens after a single i.p. injection of morphine (10 mg/kg). RESULTS: We found reduced morphine-induced dopamine release in the nucleus accumbens of Dcc haploinsufficient male mice, but, contrary to the effects of stimulant drugs, there is no effect of genotype on morphine-induced conditioned preference. CONCLUSION: These findings show that reduced drug-induced mesolimbic dopamine in Dcc haploinsufficient male mice protects specifically against the rewarding effects of stimulant drugs, but not against the rewarding properties of morphine and ethanol. These results suggest that these drugs exert their rewarding effect via different brain circuits.

Investigating the "two-hit hypothesis": Effects of prenatal maternal immune activation and adolescent cannabis use on neurodevelopment in mice.

Prenatal exposure to maternal immune activation (MIA) and chronic adolescent cannabis use are both risk factors for neuropsychiatric disorders. However, exposure to a single risk factor may not result in major mental illness, indicating that multiple exposures may be required for illness onset. Here, we examine whether combined exposure to prenatal MIA and adolescent delta-9-tetrahydrocannabinol (THC), the main psychoactive component of cannabis, lead to enduring neuroanatomical and behavioural changes in adulthood. Mice were prenatally exposed to viral mimetic, poly I:C (5 mg/kg), or vehicle at gestational day (GD) 9, and postnatally exposed to chronic THC (5 mg/kg, intraperitoneal) or vehicle during adolescence (postnatal day [PND]28-45). Longitudinal magnetic resonance imaging (MRI) was performed pre-treatment, PND 25, post-treatment, PND 50, and in adulthood, PND85, followed by behavioural tests for anxiety-like, social, and sensorimotor gating. Post-mortem assessment of cannabinoid (CB)1 and 2 receptor expressing cells was performed in altered regions identified by MRI (anterior cingulate and somatosensory cortices, striatum, and hippocampus). Subtle deviations in neurodevelopmental trajectory and subthreshold anxiety-like behaviours were observed in mice exposed to both risk factors. Sex-dependent effects were observed in patterns of shared brain-behaviour covariation, indicative of potential sex differences in response to MIA and THC. Density of CB1 and CB2 receptor positive cells was significantly decreased in all mice exposed to MIA, THC, or both. These findings suggest that there may be a cumulative effect of risk factor exposure on gross neuroanatomical development, and that the endocannabinoid system may be sensitive to both prenatal MIA, adolescent THC, or the combination.

Efficacy of prophylactic versus therapeutic administration of the NMDA receptor antagonist MK-801 on the acute neurochemical response to a concussion in a rat model combining force and rotation.

OBJECTIVE: Alterations in amino acid concentrations are a major contributor to the persistent neurological and behavioral effects induced by concussions and mild traumatic brain injuries (TBIs). Glutamate, the most abundant excitatory amino acid in the CNS, has a major role in the pathophysiological process of concussion. The indiscriminate liberation of glutamate immediately after a concussion triggers an excitotoxic response that leads to cell death, neuronal damage, and the dysfunction of surviving neurons, largely by overactivation of N-methyl-d-aspartate (NMDA) glutamatergic receptors. The aim of the present study was to investigate the efficacy of prophylactic versus therapeutic administration of MK-801, a promising NMDA receptor antagonist, on the acute changes in amino acid extracellular concentrations involved in excitotoxicity resulting from a concussive trauma. METHODS: The immediate neurochemical response to a concussion cannot be characterized in humans. Therefore, the authors used their previously validated combination of a weight-drop concussion rat model and in vivo cerebral microdialysis. The microdialysis probe was inserted inside the hippocampus and left inserted at impact to allow uninterrupted sampling of amino acids of interest immediately after concussion. The primary outcome included amino acid concentrations and the secondary outcome included righting time. Samples were taken in 10-minute increments for 60 minutes before, during, and 60 minutes after impact, and analyzed for glutamate, gamma-aminobutyric acid, taurine, glycine, glutamine, and serine using high-performance liquid chromatography. Righting time was acquired as a neurological restoration indicator. Physiological saline or 10 mg/kg MK-801 was administrated intraperitoneally 60 minutes before or immediately following induction of sham injury or concussion. RESULTS: Following induction of concussion, glutamate, taurine, and glycine levels as well as righting times in cases from the MK-801 treatment group were comparable to those of vehicle-treated animals. In contrast, righting times and amino acid concentrations observed within the first 10 minutes after induction of concussion in cases assigned to the MK-801 prophylaxis group were comparable to those of sham-injured animals. CONCLUSIONS: These results suggest that presynaptic actions and peak availability of MK-801 following prophylactic administration significantly inhibit the immediate and indiscriminate release of glutamate, taurine, and glycine in extracellular fluid after a concussion.

Interhemispheric regulation of the medial prefrontal cortical glutamate stress response in rats.

While stressors are known to increase medial prefrontal cortex (PFC) glutamate (GLU) levels, the mechanism(s) subserving this response remain to be elucidated. We used microdialysis and local drug applications to investigate, in male Long-Evans rats, whether the PFC GLU stress response might reflect increased interhemispheric communication by callosal projection neurons. We report here that tail-pinch stress (20 min) elicited comparable increases in GLU in the left and right PFC that were sodium and calcium dependent and insensitive to local glial cystine-GLU exchanger blockade. Unilateral ibotenate-induced PFC lesions abolished the GLU stress response in the opposite hemisphere, as did contralateral mGlu(2/3) receptor activation. Local dopamine (DA) D(1) receptor blockade in the left PFC potently enhanced the right PFC GLU stress response, whereas the same treatment applied to the right PFC had a much weaker effect on the left PFC GLU response. Finally, the PFC GLU stress response was attenuated and potentiated, respectively, following alpha(1)-adrenoreceptor blockade and GABA(B) receptor activation in the opposite hemisphere. These findings indicate that the PFC GLU stress response reflects, at least in part, activation of callosal neurons located in the opposite hemisphere and that stress-induced activation of these neurons is regulated by GLU-, DA-, norepinephrine-, and GABA-sensitive mechanisms. In the case of DA, this control is asymmetrical, with a marked regulatory bias of the left PFC DA input over the right PFC GLU stress response. Together, these findings suggest that callosal neurons and their afferentation play an important role in the hemispheric specialization of PFC-mediated responses to stressors.

Mice with dopaminergic neuron-specific deletion of DTNBP-1 gene show blunted nucleus accumbens dopamine release and associated behaviors.

Reduced expression of a schizophrenia-associated gene Dystrobrevin Binding Protein 1 (DTNBP1) and its protein product dysbindin-1, has been reported in the brains of schizophrenia patients. DTNBP1-null mutant Sdy (Sandy) mice exhibit several behavioral features relevant to schizophrenia. Changes in dopaminergic as well as glutamatergic and GABAergic neurotransmission in cortico-limbic regions have been reported in Sdy mice. Since dysbindin-1 is expressed in multiple brain regions, it is not known whether dopamine (DA) changes observed in Sdy null mutants are due to dysbindin-1 deficiency in DAergic neurons specifically. Here, using a mouse line with conditional knockout (cKO) of DTNBP1 in DA neurons, we studied the effects of dysbindin-1 deficiency on DA release and DA-dependent behaviors. Spontaneous locomotor activity of cKO mice in novel environment was significantly reduced initially but was comparable at later time points with littermate controls. However, the locomotion-enhancing effect of a low dose of d-amphetamine (d-AMPH; 2.5 mg/kg, ip) was significantly attenuated in the cKO mice suggesting a dampened mesolimbic DA transmission. Similarly, the prepulse inhibition disrupting effect of d-AMPH was found to be significantly reduced in the mutant mice. No significant differences between the cKO and control mice were observed in tests of anxiety, spatial learning and memory and social interaction. In- vivo microdialysis in the nucleus accumbens (NAc) showed a decrease in d-AMPH-induced extracellular DA release in the cKO mice. No significant alterations in protein levels of DA transporter, phosphorylated CaM kinase-II or Akt308 in the NAc were observed in the cKO mice. Taken together, our data suggest an important role of dysbindin-1 in maintaining mesolimbic DA tone and call for further investigations identifying mechanisms linking dysbindin-1, DA and schizophrenia.

Lesch-Nyhan disease causes impaired energy metabolism and reduced developmental potential in midbrain dopaminergic cells.

Mutations in HPRT1, a gene encoding a rate-limiting enzyme for purine salvage, cause Lesch-Nyhan disease which is characterized by self-injury and motor impairments. We leveraged stem cell and genetic engineering technologies to model the disease in isogenic and patient-derived forebrain and midbrain cell types. Dopaminergic progenitor cells deficient in HPRT showed decreased intensity of all developmental cell-fate markers measured. Metabolic analyses revealed significant loss of all purine derivatives, except hypoxanthine, and impaired glycolysis and oxidative phosphorylation. real-time glucose tracing demonstrated increased shunting to the pentose phosphate pathway for de novo purine synthesis at the expense of ATP production. Purine depletion in dopaminergic progenitor cells resulted in loss of RHEB, impairing mTORC1 activation. These data demonstrate dopaminergic-specific effects of purine salvage deficiency and unexpectedly reveal that dopaminergic progenitor cells are programmed to a high-energy state prior to higher energy demands of terminally differentiated cells.

Rapid microelectrode measurements and the origin and regulation of extracellular glutamate in rat prefrontal cortex.

Glutamate in the prefrontal cortex (PFC) plays a significant role in several mental illnesses, including schizophrenia, addiction and anxiety. Previous studies on PFC glutamate-mediated function have used techniques that raise questions on the neuronal versus astrocytic origin of glutamate. The present studies used enzyme-based microelectrode arrays to monitor second-by-second resting glutamate levels in the PFC of awake rats. Locally applied drugs were employed in an attempt to discriminate between the neuronal or glial components of the resting glutamate signal. Local application of tetrodotoxin (sodium channel blocker), produced a significant (∼ 40%) decline in resting glutamate levels. In addition significant reductions in extracellular glutamate were seen with locally applied ω-conotoxin (MVIIC; ∼ 50%; calcium channel blocker), and the mGluR(2/3) agonist, LY379268 (∼ 20%), and a significant increase with the mGluR(2/3) antagonist LY341495 (∼ 40%), effects all consistent with a large neuronal contribution to the resting glutamate levels. Local administration of D,L-threo-ß-benzyloxyaspartate (glutamate transporter inhibitor) produced an ∼ 120% increase in extracellular glutamate levels, supporting that excitatory amino acid transporters, which are largely located on glia, modulate clearance of extracellular glutamate. Interestingly, local application of (S)-4-carboxyphenylglycine (cystine/glutamate antiporter inhibitor), produced small, non-significant bi-phasic changes in extracellular glutamate versus vehicle control. Finally, pre-administration of tetrodotoxin completely blocked the glutamate response to tail pinch stress. Taken together, these results support that PFC resting glutamate levels in rats as measured by the microelectrode array technology are at least 40-50% derived from neurons. Furthermore, these data support that the impulse flow-dependent glutamate release from a physiologically -evoked event is entirely neuronally derived.

Stimulation of L-type calcium channels increases tyrosine hydroxylase and dopamine in ventral midbrain cells induced from somatic cells.

Making high-quality dopamine (DA)-producing cells for basic biological or small molecule screening studies is critical for the development of novel therapeutics for disorders of the ventral midbrain. Currently, many ventral midbrain assays have low signal-to-noise ratio due to low levels of cellular DA and the rate-limiting enzyme of DA synthesis, tyrosine hydroxylase (TH), hampering discovery efforts. Using intensively characterized ventral midbrain cells derived from human skin, which demonstrate calcium pacemaking activity and classical electrophysiological properties, we show that an L-type calcium agonist can significantly increase TH protein levels and DA content and release. Live calcium imaging suggests that it is the immediate influx of calcium occurring simultaneously in all cells that drives this effect. Genome-wide expression profiling suggests that L-type calcium channel stimulation has a significant effect on specific genes related to DA synthesis and affects expression of L-type calcium receptor subunits from the CACNA1 and CACNA2D families. Together, our findings provide an advance in the ability to increase DA and TH levels to improve the accuracy of disease modeling and small molecule screening for disorders of the ventral midbrain, including Parkinson's disease.

Cholinergic dysfunction in the dorsal striatum promotes habit formation and maladaptive eating.

Dysregulation of habit formation has been recently proposed as pivotal to eating disorders. Here, we report that a subset of patients suffering from restrictive anorexia nervosa have enhanced habit formation compared with healthy controls. Habit formation is modulated by striatal cholinergic interneurons. These interneurons express vesicular transporters for acetylcholine (VAChT) and glutamate (VGLUT3) and use acetylcholine/glutamate cotransmission to regulate striatal functions. Using mice with genetically silenced VAChT (VAChT conditional KO, VAChTcKO) or VGLUT3 (VGLUT3cKO), we investigated the roles that acetylcholine and glutamate released by cholinergic interneurons play in habit formation and maladaptive eating. Silencing glutamate favored goal-directed behaviors and had no impact on eating behavior. In contrast, VAChTcKO mice were more prone to habits and maladaptive eating. Specific deletion of VAChT in the dorsomedial striatum of adult mice was sufficient to phenocopy maladaptive eating behaviors of VAChTcKO mice. Interestingly, VAChTcKO mice had reduced dopamine release in the dorsomedial striatum but not in the dorsolateral striatum. The dysfunctional eating behavior of VAChTcKO mice was alleviated by donepezil and by l-DOPA, confirming an acetylcholine/dopamine deficit. Our study reveals that loss of acetylcholine leads to a dopamine imbalance in striatal compartments, thereby promoting habits and vulnerability to maladaptive eating in mice.

A Novel and Translational Rat Model of Concussion Combining Force and Rotation with In Vivo Cerebral Microdialysis.

Persistent cognitive and motor symptoms are known consequences of concussions/mild traumatic brain injury (mTBIs) that can be partly attributable to altered neurotransmission. Indeed, cerebral microdialysis studies in rodents have demonstrated an excessive extracellular glutamate release in the hippocampus within the first 10 min following trauma. Microdialysis offers the clear advantage of in vivo neurotransmitter continuous sampling while not having to sacrifice the animal. In addition to the aforementioned technique, a closed head injury model that exerts rapid acceleration and deceleration of the head and torso is needed, as such a factor is not available in many other animal models. The Wayne State weight-drop model mimics this essential component of human craniocerebral trauma, allowing the induction of an impact on the head of an unrestrained rodent with a falling weight. Our novel and translational rat model combines cerebral microdialysis with the Wayne State weight-drop model to study, in lightly anesthetized and unrestrained adult rats, the acute changes in extracellular neurotransmitter levels following concussion. In this protocol, the microdialysis probe was inserted inside the hippocampus as region of interest, and was left inserted in the brain at impact. There is a high density of terminals and receptors in the hippocampus, making it a relevant region to document altered neurotransmission following concussion. When applied to adult Sprague-Dawley rats, our combined model induced increases in hippocampal extracellular glutamate concentrations within the first 10 min, consistent with the previously reported post-concussion symptomology. This combined weight-drop model provides a reliable tool for researchers to study early therapeutic responses to concussions in addition to repetitive brain injury, since this protocol induces a closed-head mild trauma.

Changes in nucleus accumbens dopamine transmission associated with fixed- and variable-time schedule-induced feeding.

We examined the changes in nucleus accumbens (NAcc) dopamine (DA) transmission associated with non-contingent meal presentations under conditions of high (fixed time-, FT-schedule) and low (variable time-, VT-schedule) predictability. Of interest were the changes in NAcc DA transmission associated with discrepancies between the time food is expected and when it is actually presented. We used in vivo voltammetry to monitor NAcc DA levels as rats received, on the first and second test days, 30-s meals of condensed milk on a VT-52 schedule (inter-meal intervals of 32, 35, 40, 45, 52, 64, and 95 s). On the third and subsequent days meals were presented first on a VT-52 s schedule and then on an FT-52 s schedule. On day 1, monotonic increases in NAcc DA signals were observed during both meal consumption and the intervals between VT meal presentations. By day 2, however, meal presentations on the VT schedule elicited biphasic DA signal fluctuations; DA signals increased prior to each meal presentation but then started to decline during the feeding bout that followed. Fixed-time meal presentations on day 3 disrupted this pattern, resulting in a concurrent escalation of DA signal fluctuations upon subsequent VT meal presentations. These findings provide further evidence that, in trained animals, NAcc DA transmission is activated by conditioned incentive cues rather than by primary rewards. They also suggest that the increases in NAcc DA transmission associated with reward expectancy are sensitive to temporal cues (e.g. interval timing) and to discrepancies between expected and actual outcomes.

Maternal high fat diet during the perinatal period alters mesocorticolimbic dopamine in the adult rat offspring: reduction in the behavioral responses to repeated amphetamine administration.

RATIONALE: Early environment can shape the development and function of the mesocorticolimbic dopamine (DA) system and represents a possible risk factor for adult pathologies. One critical variable in the early environment is nutrition, and exposure to high fat (HF) in adulthood is known to change this DA system. OBJECTIVES: We tested whether perinatal HF intake in rats could have long-term effects on DA function and behavior in adult offspring. MATERIALS AND METHODS: Rat dams were fed either a control (5% fat, CD) or high fat (30% fat, HF) diet during the last week of gestation and lactation, and adult offspring were tested (PND 56-90) after weaning on CD. Locomotor responses to acute and repeated doses of D: -amphetamine (AMP, 0.75 mg/kg bw) were determined as were indices of DA function in the ventral tegmental area (VTA), nucleus accumbens (NAc), and the prefrontal cortex (PFC). RESULTS: Adult HF offspring displayed increased tyrosine hydroxylase expression in the VTA and NAc and significant increases in DA and DOPAC content in the NAc, suggesting an elevated DA tone in this target field. In the NAc, there were no significant changes in D1, D2 receptors, or DA transporter (DAT) levels between diet groups. Perinatal HF feeding reduced AMP-induced locomotion and behavioral sensitization to AMP, suggesting that early diet might have caused long-lasting desensitization of postsynaptic receptor mechanisms in the NAc. CONCLUSIONS: Our results demonstrate that both synthetic activity in VTA neurons and the responsiveness of accumbens DA neurons is altered by maternal nutrition. These effects subside long after termination of exposure to the HF diet.

Increased Alcohol Seeking in Mice Lacking Gpr88 Involves Dysfunctional Mesocorticolimbic Networks.

BACKGOUND: Alcohol use disorder (AUD) is devastating and poorly treated, and innovative targets are actively sought for prevention and treatment. The orphan G protein-coupled receptor GPR88 is enriched in mesocorticolimbic pathways, and Gpr88 knockout mice show hyperactivity and risk-taking behavior, but a potential role for this receptor in drug abuse has not been examined. METHODS: We tested Gpr88 knockout mice for alcohol-drinking and -seeking behaviors. To gain system-level understanding of their alcohol endophenotype, we also analyzed whole-brain functional connectivity in naïve mice using resting-state functional magnetic resonance imaging. RESULTS: Gpr88 knockout mice showed increased voluntary alcohol drinking at both moderate and excessive levels, with intact alcohol sedation and metabolism. Mutant mice also showed increased operant responding and motivation for alcohol, while food and chocolate operant self-administration were unchanged. Alcohol place conditioning and alcohol-induced dopamine release in the nucleus accumbens were decreased, suggesting reduced alcohol reward in mutant mice that may partly explain enhanced alcohol drinking. Seed-based voxelwise functional connectivity analysis revealed significant remodeling of mesocorticolimbic centers, whose hallmark was predominant weakening of prefrontal cortex, ventral tegmental area, and amygdala connectional patterns. Also, effective connectivity from the ventral tegmental area to the nucleus accumbens and amygdala was reduced. CONCLUSIONS: Gpr88 deletion disrupts executive, reward, and emotional networks in a configuration that reduces alcohol reward and promotes alcohol seeking and drinking. The functional connectivity signature is reminiscent of alterations observed in individuals at risk for AUD. The Gpr88 gene, therefore, may represent a vulnerability/resilience factor for AUD, and a potential drug target for AUD treatment.

Medial prefrontal cortical alpha1 adrenoreceptor modulation of the nucleus accumbens dopamine response to stress in Long-Evans rats.

RATIONALE: The medial prefrontal cortex (PFC) receives stress-sensitive dopamine (DA) and noradrenergic (NE) projections from the ventral tegmental area and locus coeruleus, respectively, and evidence from various sources point to a complex functional interaction between these two systems. Stress will also stimulate DA transmission in the nucleus accumbens (NAcc), and our previous work has shown that this response is under the indirect inhibitory control of a DA-sensitive mechanism in PFC. OBJECTIVE: We examined the possibility that the NAcc DA stress response is also modulated by prefrontal cortical NE. MATERIALS AND METHODS: We used voltammetry to study in freely behaving rats the effects of local applications of alpha(1) (benoxathian 0.1, 1, 10 nmol), alpha(2) (SKF86466), and beta(1/2) (alprenolol) receptor selective antagonists into the PFC on the NAcc DA response to tail-pinch stress. RESULTS: The NAcc DA stress response was dose-dependently inhibited by local PFC blockade of alpha(1) receptors. Additional tests revealed, however, that the DA stress response in NAcc is unaffected after local alpha(1) receptor activation with cirazoline. Furthermore, at equivalent doses, neither alpha(2) nor beta(1/2) receptor blockade significantly affected the NAcc DA stress response. CONCLUSIONS: These data indicate that stress-induced activation of subcortical DA transmission is modulated by the NE input to PFC acting at alpha(1) receptors. They suggest that, under normal circumstances, this system exerts a facilitatory or enabling influence on the NAcc DA stress response.

Differential involvement of ventral tegmental GABA(A) and GABA(B) receptors in the regulation of the nucleus accumbens dopamine response to stress.

Evidence indicates that dopamine (DA) transmission in nucleus accumbens (NAcc) is modulated by glutamate (GLUT) projections from medial prefrontal cortex (PFC) to NAcc and the ventral tegmental area (VTA). Local NMDA receptor blockade in NAcc has previously been shown to enhance the DA stress response in this region as well as in the VTA. This raises the possibility that the NAcc DA stress response is regulated by GLUT acting at NMDA receptors located on NAcc GABA output neurons that project to the VTA where GABA is known to regulate DA cell activity. Thus, in the present study, we used voltammetry to examine the effects of intra-VTA administration of GABA(A) and GABA(B) agonists and antagonists on restraint stress-induced increases in NAcc DA. The results show that local VTA GABA(B) receptor activation with baclofen (0.01, 0.1 and 1.0 nmol) dose-dependently inhibited the NAcc DA stress response whereas GABA(B) receptor blockade with phaclofen had the opposite effect, resulting in a dose-dependent potentiation of the stress response. A similar potentiation of the NAcc DA stress response was observed following VTA GABA(A) receptor blockade with bicuculline, but only at the highest dose (1.0 nmol). Interestingly, intra-VTA injection of the GABA(A) receptor agonist, muscimol, at the lowest dose (0.01 nmol) but not at the higher doses (0.1 or 1.0 nmol) also potentiated the NAcc DA stress response, suggesting an action mediated primarily at GABA(A) receptors located on non-DA neurons. These results indicate that the NAcc DA stress response is regulated by GABA afferents to VTA DA cells and that this action is differentially mediated by GABA(A) and GABA(B) receptors. The data suggest that the relevant GABA(B) receptors are located on DA neurons whereas the GABA(A) receptors are located on GABA interneurons and perhaps also on DA cells. The present findings are also consistent with the idea that the corticofugal GLUT input to NAcc indirectly regulates stress-induced DA release in this region through the GABA feedback pathway to VTA.

Interhemispheric regulation of the rat medial prefrontal cortical glutamate stress response: role of local GABA- and dopamine-sensitive mechanisms.

RATIONALE: We previously reported that stressors increase medial prefrontal cortex (PFC) glutamate (GLU) levels as a result of activating callosal neurons located in the opposite hemisphere and that this PFC GLU stress response is regulated by GLU-, dopamine- (DA-), and GABA-sensitive mechanisms (Lupinsky et al. 2010). OBJECTIVES: Here, we examine the possibility that PFC DA regulates the stress responsivity of callosal neurons indirectly by acting at D1 and D2 receptors located on GABA interneurons. METHODS: Microdialysis combined with drug perfusion (reverse dialysis) or microinjections was used in adult male Long-Evans rats to characterize D1, D2, and GABAB receptor-mediated regulation of the PFC GABA response to tail-pinch (TP) stress. RESULTS: We report that TP stress reliably elicited comparable increases in extracellular GABA in the left and right PFCs. SCH23390 (D1 antagonist; 100 µM perfusate concentration) perfused by reverse microdialysis attenuated the local GABA stress responses equally in the left and right PFCs. Intra-PFC raclopride perfusion (D2 antagonist; 100 µM) had the opposite effect, not only potentiating the local GABA stress response but also causing a transient elevation in basal (pre-stress) GABA. Moreover, unilateral PFC raclopride microinjection (6 nmol) attenuated the GLU response to TP stress in the contralateral PFC. Finally, intra-PFC baclofen perfusion (GABAB agonist; 100 µM) inhibited the local GLU and GABA stress responses. CONCLUSIONS: Taken together, these findings implicate PFC GABA interneurons in processing stressful stimuli, showing that local D1, D2, and GABAB receptor-mediated changes in PFC GABA transmission play a crucial role in the interhemispheric regulation of GLU stress responsivity.