Response of nitrergic system in the brain of rat conditioned to intracranial self-stimulation.

The role of nitrergic system in modulating the action of psychostimulants on reward processing is well established. However, the relevant anatomical underpinnings and scope of the involved interactions with mesolimbic dopaminergic system have not been clarified. Using immunohistochemistry, we track the changes in neuronal nitric oxide synthase (nNOS) containing cell groups in the animals conditioned to intracranial self-stimulation (ICSS) via an electrode implanted in the lateral hypothalamus-medial forebrain bundle (LH-MFB) area. An increase in the nNOS immunoreactivity was noticed in the cells and fibers in the ventral tegmental area (VTA) and nucleus accumbens shell (AcbSh), the primary loci of the reward system. In addition, nNOS was up-regulated in the nucleus accumbens core (AcbC), vertical limb of diagonal band (VDB), locus coeruleus (LC), lateral hypothalamus (LH), superficial gray layer (SuG) of the superior colliculus, and periaqueductal gray (PAG). The brain tissue fragments drawn from these areas showed a change in nNOS mRNA expression, but in opposite direction. Intracerebroventricular (icv) administration of nNOS inhibitor, 7-nitroindazole (7-NI) showed decreased lever press activity in a dose-dependent manner in ICSS task. While an increase in the dopamine (DA) and 3, 4-dihydroxyphenylacetic acid (DOPAC) efflux was noted in the microdialysates collected from the AcbSh of ICSS rats, pre-administration of 7-NI (icv route) attenuated the response. The study identifies nitrergic centers that probably mediate sensory, cognitive, and motor components of the goal-directed behavior.

DNA Methylation-Mediated Mfn2 Gene Regulation in the Brain: A Role in Brain Trauma-Induced Mitochondrial Dysfunction and Memory Deficits.

Repeated mild traumatic brain injuries (rMTBI) affect mitochondrial homeostasis in the brain. However, mechanisms of long-lasting neurobehavioral effects of rMTBI are largely unknown. Mitofusin 2 (Mfn2) is a critical component of tethering complexes in mitochondria-associated membranes (MAMs) and thereby plays a pivotal role in mitochondrial functions. Herein, we investigated the implications of DNA methylation in the Mfn2 gene regulation, and its consequences on mitochondrial dysfunction in the hippocampus after rMTBI. rMTBI dramatically reduced the mitochondrial mass, which was concomitant with decrease in Mfn2 mRNA and protein levels. DNA hypermethylation at the Mfn2 gene promoter was observed post 30 days of rMTBI. The treatment of 5-Azacytidine, a pan DNA methyltransferase inhibitor, normalized DNA methylation levels at Mfn2 promoter, which further resulted into restoration of Mfn2 function. The normalization of Mfn2 function was well correlated with recovery in memory deficits in rMTBI-exposed rats. Since, glutamate excitotoxicity serves as a primary insult after TBI, we employed in vitro model of glutamate excitotoxicity in human neuronal cell line SH-SY5Y to investigate the causal epigenetic mechanisms of Mfn2 gene regulation. The glutamate excitotoxicity reduced Mfn2 levels via DNA hypermethylation at Mfn2 promoter. Loss of Mfn2 caused significant surge in cellular and mitochondrial ROS levels with lowered mitochondrial membrane potential in cultured SH-SY5Y cells. Like rMTBI, these consequences of glutamate excitotoxicity were also prevented by 5-AzaC pre-treatment. Therefore, DNA methylation serves as a vital epigenetic mechanism involved in Mfn2 expression in the brain; and this Mfn2 gene regulation may play a pivotal role in rMTBI-induced persistent cognitive deficits. Closed head weight drop injury method was employed to induce repeated mild traumatic brain (rMTBI) in jury in adult, male Wistar rats. rMTBI causes hyper DNA methylation at the Mfn2 promoter and lowers the Mfn2 expression triggering mitochondrial dysfunction. However, the treatment of 5-azacytidine normalizes DNA methylation at the Mfn2 promoter and restores mitochondrial function.

Role for Histone Deacetylation in Traumatic Brain Injury-Induced Deficits in Neuropeptide Y in Arcuate Nucleus: Possible Implications in Feeding Behavior.

INTRODUCTION: Repeated traumatic events result in long-lasting neuropsychiatric ailments, including neuroendocrine imbalances. Neuropeptide Y (NPY) in the arcuate nucleus (Arc) is an important orexigenic peptide. However, the molecular underpinnings of its dysregulation owing to traumatic brain injury remain unknown. METHODS: Rats were subjected to repeated mild traumatic brain injury (rMTBI) using the closed head weight-drop model. Feeding behavior and the regulatory epigenetic parameters of NPY expression were measured at 48 h and 30 days post-rMTBI. Further, sodium butyrate (SB), a pan-histone deacetylase (HDAC) inhibitor, was administered to examine whether histone deacetylation is involved in NPY expression post-rMTBI. RESULTS: The rMTBI attenuated food intake, which was coincident with a decrease in NPY mRNA and protein levels in the Arc post-rMTBI. Further, rMTBI also reduced the mRNA levels of the cAMP response element-binding protein (CREB) and CREB-binding protein (CBP) and altered the mRNA levels of the various isoforms of the HDACs. Concurrently, the acetylated histone 3-lysine 9 (H3-K9) levels and the binding of CBP at the NPY promoter in the Arc of the rMTBI-exposed rats were reduced. However, the treatment with SB corrected the rMTBI-induced deficits in the H3-K9 acetylation levels and CBP occupancy at the NPY promoter, restoring both NPY expression and food intake. CONCLUSIONS: These findings suggest that histone deacetylation at the NPY promoter persistently controls NPY function in the Arc after rMTBI. This study also demonstrates the efficacy of HDAC inhibitors in mitigating trauma-induced neuroendocrine maladaptations in the hypothalamus.

Protocatechuic Acid Prevents Early Hour Ischemic Reperfusion Brain Damage by Restoring Imbalance of Neuronal Cell Death and Survival Proteins.

OBJECTIVE: To investigate the neuroprotective effect of protocatechuic acid (PCA) on cell death/survival protein imbalance in a rat model of middle cerebral artery occlusion and reperfusion. METHODS: Focal ischemia was induced by middle cerebral artery occlusion in adult male Wistar rats and confirmed by measuring infarction of brain by 2,3,5-Triphenyltetrazolium chloride (TTC) staining. Rats were treated with vehicle or PCA at 10, 30 or 50 mg/kg dose intraperitoneally and subjected to neurological deficits or beam walk assessment at 24 h of reperfusion. Effective dose of PCA (50 mg/kg) was administered at 1, 2 and 3 h time point of post-ictus ischemia. Cellular damage and nuclear condensation was observed by haematoxylin and eosin (H and E) staining and Hoechst 33342 staining respectively. Additionally, immunohistochemical expression of caspase 3 and cAMP-response element binding protein (CREB) and their mRNA's were observed. RESULTS: PCA at 30 and 50 mg/kg significantly improved behavioural performance and reduced infarction. Maximum neuroprotective effect of PCA (50 mg/kg) was found at 1 h (early hours) post-ictus ischemia along with reduction in cellular damage and nuclear condensation. PCA increased CREB protein and it's mRNA, while suppressed caspase-3 protein and mRNA at 1 h of reperfusion injury. CONCLUSION: PCA exhibit the potential to prevent early hour (1h) reperfusion injury restoring balance of survival and death protein may offer a cost effective adjuvant therapy in stroke.

Essential Role of Histone Methyltransferase G9a in Rapid Tolerance to the Anxiolytic Effects of Ethanol.

BACKGROUND: Tolerance to ethanol-induced anxiolysis promotes alcohol intake, thus contributing to alcohol use disorder development. Recent studies implicate histone deacetylase-mediated histone H3K9 deacetylation in regulating neuropeptide Y expression during rapid ethanol tolerance to the anxiolytic effects of ethanol. Furthermore, the histone methyltransferase, G9a, and G9a-mediated H3K9 dimethylation (H3K9me2) have recently emerged as regulators of addiction and anxiety; however, their role in rapid ethanol tolerance is unknown. Therefore, we investigated the role of G9a-mediated H3K9me2 in neuropeptide Y expression during rapid ethanol tolerance. METHODS: Adult male rats were administered one injection of n-saline followed by single acute ethanol injection (1 g/kg) 24 hours later (ethanol group) or 2 injections (24 hours apart) of either n-saline (saline group) or ethanol (tolerance group). Anxiety-like behaviors and global and Npy-specific G9a and H3K9me2 levels in the amygdala were measured. Effects of G9a inhibitor (UNC0642) treatment on behavioral and epigenetic measures were also examined. RESULTS: Acute ethanol produced anxiolysis and decreased global H3K9me2 and G9a protein levels in the central and medial nucleus of the amygdala as well as decreased occupancy levels of H3K9me2 and G9a near a putative binding site for cAMP-response element binding protein on the Npy gene. Two identical doses of ethanol produced no behavioral or epigenetic changes relative to controls, indicating development of rapid ethanol tolerance. Interestingly, treatment with UNC0642, before the second ethanol dose reversed rapid ethanol tolerance, decreased global H3K9me2 and increased neuropeptide Y levels in the central and medial nucleus of the amygdala. CONCLUSIONS: These results implicate amygdaloid G9a-mediated H3K9me2 mechanisms in regulating rapid tolerance to the anxiolytic effects of ethanol via neuropeptide Y expression regulation.

Neuropeptide CART prevents memory loss attributed to withdrawal of nicotine following chronic treatment in mice.

Although chronic nicotine administration does not affect memory, its withdrawal causes massive cognitive deficits. The underlying mechanisms, however, have not been understood. We test the role of cocaine- and amphetamine-regulated transcript peptide (CART), a neuropeptide known for its procognitive properties, in this process. The mice on chronic nicotine treatment/withdrawal were subjected to novel object recognition task. The capability of the animal to discriminate between the novel and familiar objects was tested and represented as discrimination index (DI); reduction in the index suggested amnesia. Nicotine for 49 days had no effect on DI, but 8-hour withdrawal caused a significant reduction, followed by full recovery at 24-hour withdrawal timepoint. Bilateral CART infusion in dorsal hippocampus rescued deficits in DI at 8-hours, whereas CART-antibody infusion into the dorsal hippocampus attenuated the recovery at 24-hours. Commensurate changes were observed in the CART as well as CART mRNA profiles in the hippocampus. CART mRNA expression and the peptide immunoreactivity did not change significantly following chronic nicotine treatment. However, there was a significant reduction at 8-hour withdrawal, followed by a drastic increase in CART immunoreactivity as well as CART mRNA at 24-hour withdrawal, compared with 8-hour withdrawal. Distinct α7-nicotinic receptor immunoreactivity was detected on the hippocampal CART neurons, suggesting cholinergic inputs. An increase in the synaptophysin immunoreactive elements around CART cells in the dentate gyrus, cornu ammonis 3 and subiculum at 24-hour post-withdrawal timepoint suggested neuronal plasticity. CART circuit dynamics in the hippocampus seems to modulate short-term memory associated with nicotine withdrawal.

Minimal traumatic brain injury causes persistent changes in DNA methylation at BDNF gene promoters in rat amygdala: A possible role in anxiety-like behaviors.

Minimal traumatic brain injury (MTBI) often transforms into chronic neuropsychiatric conditions including anxiety, the underlying mechanisms of which are largely unknown. In the present study, we employed the closed-head injury paradigm to induce MTBI in rats and examined whether DNA methylation can explain long-term changes in the expression of the brain-derived neurotrophic factor (BDNF) in the amygdala as well as trauma-induced anxiety-like behaviors. The MTBI caused anxiety-like behaviors and altered the expression of DNA methyltransferase (DNMT) isoforms (DNMT1, DNMT3a, and DNMT3b) and factors involved in DNA demethylation such as the growth arrest and DNA damage 45 (GADD45a and GADD45b). After 30days of MTBI, the over-expression of DNMT3a and DNMT3b corresponded to heightened DNMT activity, whereas the mRNA levels of GADD45a and GADD45b were declined. The methylated cytosine levels at the BDNF promoters (Ip, IVp and IXp) were increased in the amygdala of the trauma-induced animals; these coincided negatively with the mRNA levels of exon IV and IXa, but not of exon I. Interestingly, treatment with 5-azacytidine, a pan DNMT inhibitor, normalized the MTBI-induced DNMT activity and DNA hypermethylation at exon IVp and IXp. Furthermore, 5-azacytidine also corrected the deficits in the expression of exons IV and IXa and reduced the anxiety-like behaviors. These results suggest that the DNMT-mediated DNA methylation at the BDNF IVp and IXp might be involved in the regulation of BDNF gene expression in the amygdala. Further, it could also be related to MTBI-induced anxiety-like behaviors via the regulation of synaptic plasticity.

Adolescent Alcohol Exposure-Induced Changes in Alpha-Melanocyte Stimulating Hormone and Neuropeptide Y Pathways via Histone Acetylation in the Brain During Adulthood.

Background: Adolescent intermittent ethanol exposure causes long-lasting alterations in brain epigenetic mechanisms. Melanocortin and neuropeptide Y signaling interact and are affected by ethanol exposure in the brain. Here, the persistent effects of adolescent intermittent ethanol on alpha-melanocyte stimulating hormone, melanocortin 4 receptor, and neuropeptide Y expression and their regulation by histone acetylation mechanisms were investigated in adulthood. Methods: Male rats were exposed to adolescent intermittent ethanol (2 g/kg, i.p.) or volume-matched adolescent intermittent saline from postnatal days 28 to 41 and allowed to grow to postnatal day 92. Anxiety-like behaviors were measured by the elevated plus-maze test. Brain regions from adult rats were used to examine changes in alpha-melanocyte stimulating hormone, melanocortin 4 receptor, and neuropeptide Y expression and the histone acetylation status of their promoters. Results: Adolescent intermittent ethanol-exposed adult rats displayed anxiety-like behaviors and showed increased pro-opiomelanocortin mRNA levels in the hypothalamus and increased melanocortin 4 receptor mRNA levels in both the amygdala and hypothalamus compared with adolescent intermittent saline-exposed adult rats. The alpha-Melanocyte stimulating hormone and melanocortin 4 receptor protein levels were increased in the central and medial nucleus of the amygdala, paraventricular nucleus, and arcuate nucleus of the hypothalamus in adolescent intermittent ethanol-exposed compared with adolescent intermittent saline-exposed adult rats. Neuropeptide Y protein levels were decreased in the central and medial nucleus of the amygdala of adolescent intermittent ethanol-exposed compared with adolescent intermittent saline-exposed adult rats. Histone H3K9/14 acetylation was decreased in the neuropeptide Y promoter in the amygdala but increased in the melanocortin 4 receptor gene promoter in the amygdala and the melanocortin 4 receptor and pro-opiomelanocortin promoters in the hypothalamus of adolescent intermittent ethanol-exposed adult rats compared with controls. Conclusions: Increased melanocortin and decreased neuropeptide Y activity due to changes in histone acetylation in emotional brain circuitry may play a role in adolescent intermittent ethanol-induced anxiety phenotypes in adulthood.

Adolescent alcohol exposure alters lysine demethylase 1 (LSD1) expression and histone methylation in the amygdala during adulthood.

Alcohol exposure in adolescence is an important risk factor for the development of alcoholism in adulthood. Epigenetic processes are implicated in the persistence of adolescent alcohol exposure-related changes, specifically in the amygdala. We investigated the role of histone methylation mechanisms in the persistent effects of adolescent intermittent ethanol (AIE) exposure in adulthood. Adolescent rats were exposed to 2 g/kg ethanol (2 days on/off) or intermittent n-saline (AIS) during postnatal days (PND) 28-41 and used for behavioral and epigenetic studies. We found that AIE exposure caused a long-lasting decrease in mRNA and protein levels of lysine demethylase 1(Lsd1) and mRNA levels of Lsd1 + 8a (a neuron-specific splice variant) in specific amygdaloid structures compared with AIS-exposed rats when measured at adulthood. Interestingly, AIE increased histone H3 lysine 9 dimethylation (H3K9me2) levels in the central nucleus of the amygdala (CeA) and medial nucleus of the amygdala (MeA) in adulthood without producing any change in H3K4me2 protein levels. Acute ethanol challenge (2 g/kg) in adulthood attenuated anxiety-like behaviors and the decrease in Lsd1 + 8a mRNA levels in the amygdala induced by AIE. AIE caused an increase in H3K9me2 occupancy at the brain-derived neurotrophic factor exon IV promoter in the amygdala that returned to baseline after acute ethanol challenge in adulthood. These results indicate that AIE specifically modulates epizymes involved in H3K9 dimethylation in the amygdala in adulthood, which are possibly responsible for AIE-induced chromatin remodeling and adult psychopathology such as anxiety.

Potential role of adolescent alcohol exposure-induced amygdaloid histone modifications in anxiety and alcohol intake during adulthood.

Binge drinking is common during adolescence and can lead to the development of psychiatric disorders, including alcoholism in adulthood. Here, the role and persistent effects of histone modifications during adolescent intermittent ethanol (AIE) exposure in the development of anxiety and alcoholism in adulthood were investigated. Rats received intermittent ethanol exposure during post-natal days 28-41, and anxiety-like behaviors were measured after 1 and 24 h of the last AIE. The effects of AIE on anxiety-like and alcohol-drinking behaviors in adulthood were measured with or without treatment with the histone deacetylase (HDAC) inhibitor, trichostatin A (TSA). Amygdaloid brain regions were collected to measure HDAC activity, global and gene-specific histone H3 acetylation, expression of brain-derived neurotrophic factor (BDNF) and activity-regulated cytoskeleton-associated (Arc) protein and dendritic spine density (DSD). Adolescent rats displayed anxiety-like behaviors after 24 h, but not 1 h, of last AIE with a concomitant increase in nuclear and cytosolic amygdaloid HDAC activity and HDAC2 and HDAC4 levels leading to deficits in histone (H3-K9) acetylation in the central (CeA) and medial (MeA), but not in basolateral nucleus of amygdala (BLA). Interestingly, some of AIE-induced epigenetic changes such as, increased nuclear HDAC activity, HDAC2 expression, and decreased global histone acetylation persisted in adulthood. In addition, AIE decreased BDNF exons I and IV and Arc promoter specific histone H3 acetylation that was associated with decreased BDNF, Arc expression and DSD in the CeA and MeA during adulthood. AIE also induced anxiety-like behaviors and enhanced ethanol intake in adulthood, which was attenuated by TSA treatment via normalization of deficits in histone H3 acetylation of BDNF and Arc genes. These novel results indicate that AIE induces long-lasting effects on histone modifications and deficits in synaptic events in the amygdala, which are associated with anxiety-like and alcohol drinking behaviors in adulthood.

Regulation of DNA methylation by ethanol induces tissue plasminogen activator expression in astrocytes.

Alcohol exposure affects neuronal plasticity in the adult and developing brain. Astrocytes play a major role in modulating neuronal plasticity and are a target of ethanol. Tissue plasminogen activator (tPA) is involved in modulating neuronal plasticity by degrading the extracellular matrix proteins including fibronectin and laminin and is up-regulated by ethanol in vivo. In this study we explored the hypothesis that ethanol affects DNA methylation in astrocytes thereby increasing expression and release of tPA. It was found that ethanol increased tPA mRNA levels, an effect mimicked by an inhibitor of DNA methyltransferase (DNMT) activity. Ethanol also increased tPA protein expression and release, and inhibited DNMT activity with a corresponding decrease in DNA methylation levels of the tPA promoter. Furthermore, it was observed that protein levels of DNMT3A, but not DNMT1, were reduced in astrocytes after ethanol exposure. These novel studies show that ethanol inhibits DNA methylation in astrocytes leading to increased tPA expression and release; this effect may be involved in astrocyte-mediated inhibition of neuronal plasticity by alcohol.

Reversal of deficits in dendritic spines, BDNF and Arc expression in the amygdala during alcohol dependence by HDAC inhibitor treatment.

Development of anxiety-like behaviours during ethanol withdrawal has been correlated with increased histone deacetylase (HDAC) activity and decreased brain-derived neurotrophic factor (BDNF) and activity-regulated cytoskeleton-associated protein (Arc) gene expression in the amygdala. Furthermore, HDAC-mediated histone modifications play a role in synaptic plasticity. In this study we used the HDAC inhibitor trichostatin A (TSA) to determine whether HDAC inhibition could prevent ethanol withdrawal-induced deficits in dendritic spine density (DSD), BDNF or Arc expression in the amygdala of rats. It was found that decreased BDNF and Arc expression in the central (CeA) and medial nucleus of amygdala (MeA), observed during withdrawal after chronic ethanol exposure, were normalized following acute TSA treatment. TSA treatment was also able to attenuate anxiety-like behaviours during ethanol withdrawal and correct the observed decrease in DSD in the CeA and MeA of ethanol-withdrawn rats. Taken together, these findings demonstrate that correcting the deficits in histone acetylation through TSA treatment also amends downstream synaptic plasticity-related deficits such as BDNF and Arc expression, and DSD in the CeA and MeA as well as attenuates anxiety-like behaviours in rats during withdrawal after chronic ethanol exposure.

Effects of histone deacetylase inhibitors on amygdaloid histone acetylation and neuropeptide Y expression: a role in anxiety-like and alcohol-drinking behaviours.

Recent studies have demonstrated the involvement of epigenetic mechanisms in psychiatric disorders, including alcoholism. Here, we investigated the effects of histone deacetylase (HDAC) inhibitor, trichostatin A (TSA) on amygdaloid HDAC-induced histone deacetylation and neuropeptide Y (NPY) expression and on anxiety-like and alcohol-drinking behaviours in alcohol-preferring (P) and -non-preferring (NP) rats. It was found that P rats displayed higher anxiety-like and alcohol-drinking behaviours, higher amygdaloid nuclear, but not cytosolic, HDAC activity, which was associated with increased HDAC2 protein levels and deficits in histone acetylation and NPY expression in the central (CeA) and medial nucleus of amygdala (MeA), as compared to NP rats. TSA treatment attenuated the anxiety-like and alcohol-drinking behaviours, with concomitant reductions in amygdaloid nuclear, but not cytosolic HDAC activity, and HDAC2, but not HDAC4, protein levels in the CeA and MeA of P rats, without effect in NP rats. TSA treatment also increased global histone acetylation (H3-K9 and H4-K8) and NPY expression in the CeA and MeA of P, but not in NP rats. Histone H3 acetylation within the NPY promoter was also innately lower in the amygdala of P rats compared with NP rats; which was normalized by TSA treatment. Voluntary ethanol intake in P, but not NP rats, produced anxiolytic effects and decreased the HDAC2 levels and increased histone acetylation in the CeA and MeA. These results suggest that higher HDAC2 expression-related deficits in histone acetylation may be involved in lower NPY expression in the amygdala of P rats, and operative in controlling anxiety-like and alcohol-drinking behaviours.

Effects of acute ethanol exposure on anxiety measures and epigenetic modifiers in the extended amygdala of adolescent rats.

Epigenetic mechanisms appear to play an important role in neurodevelopment. We investigated the effects of acute ethanol exposure on anxiety measures and function of histone deacetylases (HDAC) and DNA methyltransferases (DNMT) in the amygdala and bed nucleus of stria terminalis (BNST) of adolescent rats. One hour after ethanol exposure, rats were subjected to anxiety measures. A subset of adolescent rats was exposed to two doses (24 h apart) of ethanol (2 g/kg) to measure rapid ethanol tolerance to anxiolysis. The HDAC and DNMT activities and mRNA levels of DNMT isoforms were measured in the amygdala and BNST. The lower dose of ethanol (1 g/kg) produced neither anxiolysis, nor inhibited the HDAC and DNMT activities in the amygdala and BNST, except DNMT activity in BNST was attenuated. Anxiolysis by ethanol was observed at 2 and 2.25 g/kg, whereas higher doses (2.5 and 3 g/kg) were found to be sedative. DNMT activity in the amygdala and BNST, and nuclear HDAC activity in the amygdala, but not in the BNST were also inhibited by these doses of ethanol. A lack of tolerance was observed on ethanol-induced inhibition of DNMT activity in the amygdala and BNST, and nuclear HDAC activity in the amygdala, as well to anxiolysis produced by ethanol (2 g/kg). The DNMT1, DNMT3a, and DNMT3b mRNA expression in the amygdala was not affected by either 1or 2 doses of 2 g/kg. However, DNMT1 and DNMT3a expression in the BNST was increased, whereas DNMT3l mRNA was decreased in the amygdala, after 2 doses of 2 g/kg ethanol. These results suggest that reduced sensitivity to anxiolysis and the lack of rapid tolerance to the anxiolytic effects of ethanol and inhibition of HDAC and DNMT functions may play a role in engaging adolescents in binge drinking patterns.

Gut Microbiota Regulates Epigenetic Remodelling in the Amygdala: A Role in Repeated Mild Traumatic Brain Injury (rMTBI)-Induced Anxiety.

Gut microbiota serves in the development and maintenance of phenotype. However, the underlying mechanisms are still in its infancy. The current study shows epigenetic remodelling in the brain as a causal mechanism in the gut microbiota-brain axis. Like in trauma patients, gut dysbiosis and anxiety were comorbid in adult male Wistar rats subjected to repeated mild traumatic brain injuries (rMTBI). rMTBI caused epigenetic dysregulation of brain-derived neurotrophic factor (Bdnf) expression in the amygdala, owing to the formation of transcriptional co-repressor complex due to dynamic interaction between histone deacetylase and DNA methylation modification at the Bdnf gene promoter. The probiosis after faecal microbiota transplantation (FMT) from healthy naïve rats or by administration of single strain probiotic (SSP), Lactobacillus rhamnosus GG (LGG), recuperated rMTBI-induced anxiety. Concurrently, LGG infusion or naïve FMT also dislodged rMTBI-induced co-repressor complex resulting in the normalization of Bdnf expression and neuronal plasticity as measured by Golgi-Cox staining. Furthermore, sodium butyrate, a short-chain fatty acid, produced neurobehavioural effects similar to naïve FMT or LGG administration. Interestingly, the gut microbiota from rMTBI-exposed rats per se was able to provoke anxiety in naïve rats in parallel with BDNF deficits. Therefore, gut microbiota seems to be causally linked with the chromatin remodelling necessary for neuroadaptations via neuronal plasticity which drives experience-dependent behavioural manifestations.

Histone deacetylases (HDAC)-induced histone modifications in the amygdala: a role in rapid tolerance to the anxiolytic effects of ethanol.

BACKGROUND: Rapid tolerance to the anxiolytic effects of ethanol appears to be an important factor in the development of alcoholism. Here, we investigated the involvement of amygdaloid histone deacetylases (HDAC)-induced epigenetic changes in rapid ethanol tolerance (RET). METHODS: RET in rats was induced by 2 ethanol injections administered 24 hours apart. Both ethanol-tolerant and control rats were treated with the HDAC inhibitor, trichostatin A (TSA), and anxiety-like behaviors were measured. HDAC activity, histone (H3 and H4) acetylation, and neuropeptide Y (NPY) expression in the amygdala of these rats were also measured. RESULTS: A single ethanol exposure was able to produce an anxiolytic response, inhibit amygdaloid HDAC activity, and increase both histone acetylation and NPY expression (mRNA and protein levels) in the central nucleus of amygdala (CeA) and medial nucleus of amygdala (MeA) of rats. In contrast, 2 exposures of the same dose of ethanol (24 hours apart) neither elicited a similar anxiolytic response nor modulated HDAC activity, histone acetylation, or NPY expression in the amygdala. However, exposure to a higher dose of ethanol on the second day was able to produce an anxiolytic response and also inhibit amygdaloid HDAC activity. TSA treatment caused the reversal of RET by inhibiting HDAC activity, thereby increasing histone acetylation and NPY expression in the CeA and MeA. CONCLUSIONS: Cellular tolerance to the initial acute ethanol-induced inhibition of HDAC activity and the subsequent upregulation of histone acetylation and NPY expression in the amygdala may be involved in the mechanisms underlying rapid tolerance to the anxiolytic effects of ethanol.

Understanding the role of nACE2 in neurogenic hypertension among COVID-19 patients.

Currently, the third and fourth waves of the coronavirus disease -19 (COVID-19) pandemic are creating havoc in many parts of the world. Although vaccination programs have been launched in most countries, emerging new strains of the virus along with geographical variations are leading to varying success rates of the available vaccines. The presence of comorbidities such as diabetes, cardiovascular diseases and hypertension is responsible for increasing the severity of COVID-19 and, thus, the COVID-19 mortality rate. Angiotensin-converting enzyme 2 (ACE2), which is utilized by SARS-CoV-2 for entry into host cells, is widely expressed in the lungs, kidneys, testes, gut, adipose tissue, and brain. Infection within host cells mediates RAS overactivation, which leads to a decrease in the ACE2/ACE ratio, AT2R/AT1R ratio, and MasR/AT1R ratio. Such imbalances lead to the development of heightened inflammatory responses, such as cytokine storms, leading to post-COVID-19 complications and mortality. As the association of SARS-CoV-2 infection and hypertension remains unclear, this report provides an overview of the effects of SARS-CoV-2 infection on patients with hypertension. We discuss here the interaction of ACE2 with SARS-CoV-2, focusing on neuronal ACE2 (nACE2), and further shed light on the possible involvement of nACE2 in hypertension. SARS-CoV-2 enters the brain through neuronal ACE2 and spreads in various regions of the brain. The effect of viral binding to neuronal ACE2 in areas of the brain that regulate salt/water balance and blood pressure is also discussed in light of the neural regulation of hypertension in COVID-19.

TET1-induced DNA demethylation in dentate gyrus is important for reward conditioning and reinforcement.

Neuroadaptations in neurocircuitry of reward memories govern the persistent and compulsive behaviors. The study of the role of hippocampus in processing of reward memory and its retrieval is critical to our understanding of addiction and relapse. The aim of this study is to probe the epigenetic mechanisms underlying reward memory in the frame of dentate gyrus (DG). To that end, the rats conditioned to the food baited arm of a Y-maze and subjected to memory probe trial. The hippocampus of conditioned rats displayed higher mRNA levels of Ten-eleven translocase 1 (Tet1) and brain-derived neurotrophic factor (Bdnf) after memory probe trial. The DNA hydroxymethylation and TET1 occupancy at the Bdnf promoters showed concomitant increase. Stereotactic administration of Tet1 siRNA in the DG before and after conditioning inhibited reward memory formation and recall, respectively. Administration of Tet1 siRNA impaired the reward memory recall that was reinstated following administration of exogenous BDNF peptide or after wash-off period of 8 days. Infusion of a MEK/ERK inhibitor, U0126 in the DG inhibited reward memory retrieval. The TET1-induced DNA demethylation at the Bdnf promoters raised BDNF levels in the hippocampus, thereby setting the stage for reward memory retrieval. The study underscores the causative role of TET1 in the DG for reward memory formation and recall.

Reproductive phase related variations in the expression of neuropeptide, cocaine- and amphetamine- regulated transcript (CART) in the brain and pituitary gland of adult male Microhyla ornata.

Cocaine- and amphetamine-regulated transcript (CART) peptide is a multifaceted neuropeptide involved in several physiological functions including appetite and reproduction. While studies in mammals, aves and fishes suggest evolutionary conserved role of CART, the information in amphibian is scanty. We have investigated the reproductive phase related variations of CART in the brain of adult male Microhyla ornata. Seasonal changes in the expression of CART peptide were noticed in the brain and pituitary of M. ornata. Significant differences were observed in the nucleus infundibularis ventralis (NIV), epiphysis (E), anteroventral tegmental region (AV), raphe nucleus (Ra) of the brain and pars intermedia (PI), pars distalis (PD) of the pituitary. Compared to the pre-breeding and post-breeding seasons, increase in CART immunoreactivity was seen in E, NIV, AV, Ra of brain and PI, PD of pituitary gland of animals collected during breeding season. Similarly, highest mRNA levels of CART were also observed in the breeding season in the middle region of brain that includes hypothalamus and pituitary gland. Variation in the levels of CART peptide and mRNA in the brain of M. ornata suggests its conserved role in seasonal control of appetite and reproduction.

Neuroplastic Changes in the Superior Colliculus and Hippocampus in Self-rewarding Paradigm: Importance of Visual Cues.

Coincident excitation via different sensory modalities encoding objects of positive salience is known to facilitate learning and memory. With a view to dissect the contribution of visual cues in inducing adaptive neural changes, we monitored the lever press activity of a rat conditioned to self-administer sweet food pellets in the presence/absence of light cues. Application of light cues facilitated learning and consolidation of long-term memory. The superior colliculus (SC) of rats trained on light cue showed increased neuronal activity, dendritic branching, and brain-derived neurotrophic factor (BDNF) protein and mRNA expression. Concomitantly, the hippocampus showed augmented neurogenesis as well as BDNF protein and mRNA expression. While intra-SC administration of U0126 (inhibitor of ERK 1/2 and long-term memory) impaired memory formation, lidocaine (local anaesthetic) hindered memory recall. The light cue-dependent sweet food pellet self-administration was coupled with increased efflux of dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) in the nucleus accumbens shell (AcbSh). In conditioned rats, pharmacological inhibition of glutamatergic signalling in dentate gyrus (DG) reduced lever press activity, as well as DA and DOPAC secretion in the AcbSh. We suggest that the neuroplastic changes in the SC and hippocampus might represent memory engrams sculpted by visual cues encoding reward information.