Novel role and regulation of HDAC4 in cocaine-related behaviors.

Epigenetic mechanisms have been proposed to contribute to persistent aspects of addiction-related behaviors. One family of epigenetic molecules that may regulate maladaptive behavioral changes produced by cocaine use are the histone deacetylases (HDACs)-key regulators of chromatin and gene expression. In particular, the class IIa HDACs (HDAC4, HDAC5, HDAC7 and HDAC9) respond to changes in neuronal activity by modulating their distribution between the nucleus and cytoplasm-a process controlled in large part by changes in phosphorylation of conserved residues. Cocaine triggers a transient nuclear accumulation of HDAC5 that functions to limit the development of cocaine reward behavior. However, the role and regulation of the close family member, HDAC4, in cocaine behaviors remain largely unknown. In this study, we report that cocaine and cAMP signaling in striatum produced differential phosphorylation and subcellular localization of HDAC4 and HDAC5. Unlike HDAC5, cocaine exposure induced a modest hyperphosphorylation and nuclear export of HDAC4. Genetic deletion of HDAC4 in the nucleus accumbens reduced acute cocaine-produced locomotion, maximum locomotor sensitization and cocaine reward-related behavior. Interestingly, overexpression of an HDAC4 cytoplasm-concentrated mutant (S266E) increased cocaine reward behavior in the cocaine conditioned place preference assay, suggesting that cocaine-induced nuclear export of HDAC4 might function to facilitate the development of cocaine reward behaviors through a role in the cell cytoplasm. Together, our findings suggest that, despite high sequence homology, HDAC4 and HDAC5 are oppositely regulated by cocaine-induced signaling in vivo and have distinct roles in regulating cocaine behaviors.

Circadian genes Period 1 and Period 2 in the nucleus accumbens regulate anxiety-related behavior.

It has been suggested for some time that circadian rhythm abnormalities underlie the development of multiple psychiatric disorders. However, it is unclear how disruptions in individual circadian genes might regulate mood and anxiety. Here we found that mice lacking functional mPeriod 1 (mPer1) or mPeriod 2 (mPer2) individually did not have consistent behavioral abnormalities in measures of anxiety-related behavior. However, mice deficient in both mPer1 and mPer2 had an increase in levels of anxiety-like behavior in multiple measures. Moreover, we found that mPer1 and mPer2 expression was reduced in the nucleus accumbens (NAc) after exposure to chronic social defeat stress, a paradigm that led to increased anxiety-related behavior. Following social defeat, chronic treatment with fluoxetine normalized Per gene expression towards wild-type levels. Knockdown of both mPer1 and mPer2 expression via RNA interference specifically in the NAc led to a similar increase in anxiety-like behavior as seen in the mutant animals. Taken together, these results implicate the Per genes in the NAc in response to stress and the development of anxiety.

Differential effects of chronic social stress and fluoxetine on meal patterns in mice.

Both chronic stress and antidepressant medications have been associated with changes in body weight. In the current study, we investigate mechanisms by which stress and antidepressants interact to affect meal patterns. A group of mice was subjected to the chronic social defeat stress model of major depression followed by fluoxetine treatment and was subsequently analyzed for food intake using metabolic cages. We report that chronic social defeat stress increases food intake by specifically increasing meal size, an effect that is reversed by fluoxetine treatment. In an attempt to gain mechanistic insight into changes in meal patterning induced by stress and fluoxetine, fasting serum samples were collected every 4h over a 24-h period, and acyl-ghrelin, leptin, and corticosterone levels were measured. Chronic stress induces a peak in acyl-ghrelin levels just prior to the onset of the dark phase, which is shifted in mice treated with fluoxetine. Taken together, these results indicate that stress increases food intake by decreasing satiation, and that fluoxetine can reverse stress-induced changes in meal patterns.

NPAS4 in the medial prefrontal cortex mediates chronic social defeat stress-induced anhedonia-like behavior and reductions in excitatory synapses.

Chronic stress can produce reward system deficits (i.e., anhedonia) and other common symptoms associated with depressive disorders, as well as neural circuit hypofunction in the medial prefrontal cortex (mPFC). However, the molecular mechanisms by which chronic stress promotes depressive-like behavior and hypofrontality remain unclear. We show here that the neuronal activity-regulated transcription factor, NPAS4, in the mPFC is regulated by chronic social defeat stress (CSDS), and it is required in this brain region for CSDS-induced changes in sucrose preference and natural reward motivation in the mice. Interestingly, NPAS4 is not required for CSDS-induced social avoidance or anxiety-like behavior. We also find that mPFC NPAS4 is required for CSDS-induced reductions in pyramidal neuron dendritic spine density, excitatory synaptic transmission, and presynaptic function, revealing a relationship between perturbation in excitatory synaptic transmission and the expression of anhedonia-like behavior in the mice. Finally, analysis of the mice mPFC tissues revealed that NPAS4 regulates the expression of numerous genes linked to glutamatergic synapses and ribosomal function, the expression of upregulated genes in CSDS-susceptible animals, and differentially expressed genes in postmortem human brains of patients with common neuropsychiatric disorders, including depression. Together, our findings position NPAS4 as a key mediator of chronic stress-induced hypofrontal states and anhedonia-like behavior.

Activity-regulated cytoskeleton-associated protein (Arc/Arg3.1) regulates anxiety- and novelty-related behaviors.

The activity-regulated cytoskeleton-associated protein (Arc, also known as Arg3.1) regulates glutamatergic synapse plasticity and has been linked to neuropsychiatric illness; however, its role in behaviors associated with mood and anxiety disorders remains unclear. We find that stress upregulates Arc expression in the adult mouse nucleus accumbens (NAc)-a brain region implicated in mood and anxiety behaviors. Global Arc knockout mice have altered AMPAR-subunit surface levels in the adult NAc, and the Arc-deficient mice show reductions in anxiety-like behavior, deficits in social novelty preference, and antidepressive-like behavior. Viral-mediated expression of Arc in the adult NAc of male, global Arc KO mice restores normal levels of anxiety-like behavior in the elevated plus maze (EPM). Consistent with this finding, viral-mediated reduction of Arc in the adult NAc reduces anxiety-like behavior in male, but not female, mice in the EPM. NAc-specific reduction of Arc also produced significant deficits in both object and social novelty preference tasks. Together our findings indicate that Arc is essential for regulating normal mood- and anxiety-related behaviors and novelty discrimination, and that Arc's function within the adult NAc contributes to these behavioral effects.

HDAC5 and Its Target Gene, Npas4, Function in the Nucleus Accumbens to Regulate Cocaine-Conditioned Behaviors.

Individuals suffering from substance-use disorders develop strong associations between the drug's rewarding effects and environmental cues, creating powerful, enduring triggers for relapse. We found that dephosphorylated, nuclear histone deacetylase 5 (HDAC5) in the nucleus accumbens (NAc) reduced cocaine reward-context associations and relapse-like behaviors in a cocaine self-administration model. We also discovered that HDAC5 associates with an activity-sensitive enhancer of the Npas4 gene and negatively regulates NPAS4 expression. Exposure to cocaine and the test chamber induced rapid and transient NPAS4 expression in a small subpopulation of FOS-positive neurons in the NAc. Conditional deletion of Npas4 in the NAc significantly reduced cocaine conditioned place preference and delayed learning of the drug-reinforced action during cocaine self-administration, without affecting cue-induced reinstatement of drug seeking. These data suggest that HDAC5 and NPAS4 in the NAc are critically involved in reward-relevant learning and memory processes and that nuclear HDAC5 limits reinstatement of drug seeking independent of NPAS4.

MEF2C regulates cortical inhibitory and excitatory synapses and behaviors relevant to neurodevelopmental disorders.

Numerous genetic variants associated with MEF2C are linked to autism, intellectual disability (ID) and schizophrenia (SCZ) - a heterogeneous collection of neurodevelopmental disorders with unclear pathophysiology. MEF2C is highly expressed in developing cortical excitatory neurons, but its role in their development remains unclear. We show here that conditional embryonic deletion of Mef2c in cortical and hippocampal excitatory neurons (Emx1-lineage) produces a dramatic reduction in cortical network activity in vivo, due in part to a dramatic increase in inhibitory and a decrease in excitatory synaptic transmission. In addition, we find that MEF2C regulates E/I synapse density predominantly as a cell-autonomous, transcriptional repressor. Analysis of differential gene expression in Mef2c mutant cortex identified a significant overlap with numerous synapse- and autism-linked genes, and the Mef2c mutant mice displayed numerous behaviors reminiscent of autism, ID and SCZ, suggesting that perturbing MEF2C function in neocortex can produce autistic- and ID-like behaviors in mice.

Knockdown of Clock in the ventral tegmental area through RNA interference results in a mixed state of mania and depression-like behavior.

BACKGROUND: Circadian rhythm abnormalities are strongly associated with bipolar disorder; however the role of circadian genes in mood regulation is unclear. Previously, we reported that mice with a mutation in the Clock gene (ClockDelta19) display a behavioral profile that is strikingly similar to bipolar patients in the manic state. METHODS: Here, we used RNA interference and viral-mediated gene transfer to knock down Clock expression specifically in the ventral tegmental area (VTA) of mice. We then performed a variety of behavioral, molecular, and physiological measures. RESULTS: We found that knockdown of Clock, specifically in the VTA, results in hyperactivity and a reduction in anxiety-related behavior, which is similar to the phenotype of the ClockDelta19 mice. However, VTA-specific knockdown also results in a substantial increase in depression-like behavior, creating an overall mixed manic state. Surprisingly, VTA knockdown of Clock also altered circadian period and amplitude, suggesting a role for Clock in the VTA in the regulation of circadian rhythms. Furthermore, VTA dopaminergic neurons expressing the Clock short hairpin RNA have increased activity compared with control neurons, and this knockdown alters the expression of multiple ion channels and dopamine-related genes in the VTA that could be responsible for the physiological and behavioral changes in these mice. CONCLUSIONS: Taken together, these results suggest an important role for Clock in the VTA in the regulation of dopaminergic activity, manic and depressive-like behavior, and circadian rhythms.