Denis Hervé

Isolated dystonia can be caused by loss-of-function mutations in the GNAL gene (DYT-GNAL). This gene encodes the α _olf heterotrimeric G protein subunit, which, together with β ₂ γ ₇ subunits, mediates the stimulatory coupling of dopamine D1 and adenosine A2A receptors to adenylyl-cyclase. These receptors are expressed in distinct striatal projection neurons (SPNs) with complementary functions on motor behavior. To dissect the specific roles of Gα _olf in each subpopulation of SPNs, we generated and characterized mouse models in which Gnal was conditionally deleted in neurons expressing either D1 receptors (D1-SPNs) or A2A receptors (A2A-SPNs). Our results confirmed the critical role of Gα _olf in regulating adenylyl-cyclase 5 and its coupling with D1 and A2A receptors. Mice with a selective loss of Gα _olf in D1-SPNs showed nocturnal hyperactivity, deficits in motor performances, but no overt abnormal movements or generalized motor disability. Our experiments also revealed that Gα _olf in D1-SPNs is not systematically required for locomotor responses induced by D1 agonists or psychostimulants. Selective loss of Gα _olf in A2A-SPNs did not affect motor abilities nor learning. However, this loss strikingly increased spontaneous locomotor activity that was not further enhanced by psychostimulant drugs (cocaine, D-amphetamine, methylphenidate) or a selective A2 agonist, KW6002, and was paradoxically reduced by caffeine Our study identified specific roles of Gα _olf downstream of D1 and A2A receptors in the control of motor behavior and drug responses, highlighting their respective individual contribution in diseases associated with dysfunctional striatal signaling, including dystonia.

BACKGROUND Highly palatable food triggers behavioral alterations reminiscent of those induced by addictive drugs. These effects involve the reward system and dopamine neurons, which modulate neurons in the nucleus accumbens (NAc). The molecular mechanisms underlying the effects of highly palatable food on feeding behavior are poorly understood. METHODS We studied the effects of 2-week operant conditioning of mice with standard or isocaloric highly palatable food. We investigated the behavioral effects and dendritic spine modifications in the NAc. We compared the translating mRNA in NAc neurons identified by the type of dopamine receptors they express, depending on the type of food and training. We tested the consequences of invalidation of an abundant downregulated gene, Ncdn (Neurochondrin). RESULTS Operant conditioning for highly palatable food increases motivation for food even in well-fed mice. In control mice, free access to regular or highly palatable food results in increased weight as compared to regular food only. Highly palatable food increases spine density in the NAc. In animals trained for highly palatable food, translating mRNAs are modified in NAc dopamine D2-receptor-expressing neurons, mostly corresponding to striatal projection neurons, but not in those expressing D1-receptors. Knock-out of Ncdn, an abundant down-regulated gene, opposes the conditioning-induced changes in satiety-sensitive feeding behavior and apparent motivation for highly palatable food, suggesting down-regulation may be a compensatory mechanism. CONCLUSIONS Our results emphasize the importance of mRNA alterations D2 striatal projection neurons in the NAc in the behavioral consequences of highly palatable food conditioning and suggest a modulatory contribution of Ncdn downregulation.

Dystonia is often associated with functional alterations in the cerebello-thalamic pathways, which have been proposed to contribute to the disorder by propagating pathological firing patterns to the forebrain. Here, we examined the function of the cerebello-thalamic pathways in a model of DYT25 dystonia. DYT25 (Gnal^+/−) mice carry a heterozygous knockout mutation of the Gnal gene, which notably disrupts striatal function, and systemic or striatal administration of oxotremorine to these mice triggers dystonic symptoms. Our results reveal an increased cerebello-thalamic excitability in the presymptomatic state. Following the first dystonic episode, Gnal^+/- mice in the asymptomatic state exhibit a further increase of the cerebello-thalamo-cortical excitability, which is maintained after θ-burst stimulations of the cerebellum. When administered in the symptomatic state induced by a cholinergic activation, these stimulations decreased the cerebello-thalamic excitability and reduced dystonic symptoms. In agreement with dystonia being a multiregional circuit disorder, our results suggest that the increased cerebello-thalamic excitability constitutes an early endophenotype, and that the cerebellum is a gateway for corrective therapies via the depression of cerebello-thalamic pathways.

Dopamine D1 receptors play an important role in the effects of cocaine. Here, we investigated the role of neurons which express these receptors (D1‐neurons) in the acute locomotor effects of cocaine and the locomotor sensitization observed after a second injection of this drug, using the previously established two‐injection protocol of sensitization. We inhibited D1‐neurons using double transgenic mice conditionally expressing the inhibitory Gi‐coupled designer receptor exclusively activated by designer drugs (Gi‐DREADD) in D1‐neurons. Chemogenetic inhibition of D1‐neurons by a low dose of clozapine (0.1 mg/kg) decreased the cocaine‐induced expression of Fos in striatal neurons. It diminished the basal locomotor activity and acute hyper‐locomotion induced by cocaine (20 mg/kg). Clozapine 0.1 mg/kg had no effect by itself and did not alter cocaine effects in wild‐type mice. Inhibition of D1‐neurons during the first cocaine administration prevented the sensitization of the locomotor response in response to a second cocaine administration 10 days later. On Day 11, inhibition of D1‐neurons by clozapine stimulation of Gi‐DREADD blocked cocaine‐induced locomotion including in sensitized mice, whereas on Day 12, in the absence of clozapine and D1‐neurons inhibition, all mice displayed a sensitized response to cocaine. These results show that chemogenetic inhibition of D1‐neurons decreases spontaneous and cocaine‐induced locomotor activity. It prevents sensitization induction and blocks sensitized locomotion in a two‐injection protocol of sensitization but does not reverse established sensitization. Our study further supports the central role of D1‐neurons in mediating the acute locomotor effects of cocaine and its sensitization.

D1 dopamine receptors play an important role in the effects of cocaine. Here we investigated the role of neurons which express these receptors (D1-neurons) in the acute locomotor effects of cocaine and the locomotor sensitization observed after a second injection of this drug. We inhibited D1-neurons using double transgenic mice conditionally expressing the inhibitory Gi-coupled designer receptor exclusively activated by designer drugs (Gi-DREADD) in D1-neurons. Chemogenetic inhibition of D1-neurons by a low dose of clozapine (0.1 mg/kg) decreased the induction of Fos in striatal neurons. It diminished the basal locomotor activity and acute hyper-locomotion induced by cocaine (20 mg/kg). Clozapine 0.1 mg/kg had no effect by itself and did not alter cocaine effects in non-transgenic mice. Inhibition of D1-neurons during the first cocaine administration reduced the sensitization of the locomotor response in response to a second cocaine administration ten days later. At day 11, inhibition of D1-neurons by clozapine stimulation of Gi-DREADD, prevented the expression of the sensitized locomotor response, whereas at day 12, in the absence of clozapine and D1-neurons inhibition, all mice displayed a sensitized response to cocaine. These results show that chemogenetic inhibition of D1-neurons decreases spontaneous and cocaine-induced locomotor activity. It blunts the induction and prevents the expression of sensitization in a two-injection protocol of sensitization but does not alter established sensitization. Our study further supports the central role of D1-neurons in mediating the acute locomotor effects of cocaine and its sensitization.

Permanent tagging of neuronal ensembles activated in specific experimental situations is an important objective to study their properties and adaptations. In the context of learning and memory, these neurons are referred to as engram neurons. Here, we describe and characterize a novel mouse line, Egr1‐CreER^T2, which carries a transgene in which the promoter of the immediate early gene Egr1 drives the expression of the CreER^T2 recombinase that is only active in the presence of tamoxifen metabolite, 4‐hydroxy‐tamoxifen (4‐OHT). Egr1‐CreER^T2 mice were crossed with various reporter mice, Cre‐dependently expressing a fluorescent protein. Without tamoxifen or 4‐OHT, no or few tagged neurons were observed. Epileptic seizures induced by pilocarpine or pentylenetetrazol in the presence of tamoxifen or 4‐OHT elicited the persistent tagging of many neurons and some astrocytes in the dentate gyrus of hippocampus, where Egr1 is transiently induced by seizures. One week after cocaine and 4‐OHT administration, these mice displayed a higher number of tagged neurons in the dorsal striatum than saline/4‐OHT controls, with differences between reporter lines. Cocaine‐induced tagging required ERK activation and tagged neurons were more likely than others to exhibit ERK phosphorylation or Fos induction after a second injection. Interestingly neurons tagged in saline‐treated mice also had an increased propensity to express Fos, suggesting the existence of highly responsive striatal neurons susceptible to be re‐activated by cocaine repeated administration, which may contribute to the behavioral adaptations. Our report validates a novel transgenic mouse model for permanently tagging activated neurons and studying long‐term alterations of Egr1‐expressing cells.

Permanent tagging of neuronal ensembles activated in specific experimental situations is an important objective to study their properties and adaptations. In the context of learning and memory these neurons are referred to as engram neurons. Here we describe and characterize a novel mouse line, Egr1-CreER ^T2 , which carries a transgene in which the promoter of the immediate early gene Egr1 drives the expression of the CreER ^T2 recombinase that is only active in the presence of tamoxifen metabolite, 4-hydroxy-tamoxifen (4-OHT). Egr1-CreER ^T2 mice were crossed with various reporter mice, Cre-dependently expressing a fluorescent protein. Without tamoxifen or 4-OHT, no or few tagged neurons were observed. Epileptic seizures induced by pilocarpine or pentylenetetrazol in the presence of tamoxifen or 4-OHT, induced the persistent tagging of many neurons and some astrocytes in the dentate gyrus of hippocampus, where Egr1 is transiently induced by seizures. One week after cocaine and 4-OHT administration, these mice displayed a higher number of tagged neurons in the dorsal striatum than saline/4-OHT controls, with differences between reporter lines. Cocaine-induced tagging required ERK activation and tagged neurons were more likely than others to exhibit ERK phosphorylation or Fos induction after a second injection. Interestingly neurons tagged in saline-treated mice also had an increased propensity to express Fos, suggesting the existence of highly responsive striatal neurons susceptible to be re-activated by cocaine repeated administration, which may contribute to the behavioral adaptations. Our report validates a novel transgenic mouse model for permanently tagging activated neurons and studying long term alterations of Egr1 -expressing cells.

Forebrain dopaminoceptive neurons play a key role in movement, action selection, motivation, and working memory. Their activity is dysregulated in addiction, Parkinson’s disease and other conditions. To characterize the diverse dopamine target neuronal populations, we compare translating mRNAs in neurons of dorsal striatum and nucleus accumbens expressing D1 or D2 dopamine receptor and prefrontal cortex expressing D1 receptor. We identify D1/D2 and striatal dorso-ventral differences in the translational and splicing landscapes, which establish the characteristics of dopaminoceptive neurons. Expression differences and network analyses identify novel transcription factors with presumptive roles in these differences. Prostaglandin E2 appears as a candidate upstream regulator in the dorsal striatum, a hypothesis supported by converging functional evidence indicating its role in enhancing D2 dopamine receptor action. Our study provides powerful resources for characterizing dopamine target neurons, new information about striatal gene expression patterns, and reveals the unforeseen role of prostaglandin E2 in the dorsal striatum.