Expression Of Glutamic Acid Decarboxylase Research Articles

Sex differences in behavior and glutamic acid decarboxylase in Long Evans rats after prolonged social isolation beginning in adolescence.

Social isolation can have long-term effects on brain development and behavior and increases the risk of developing clinical conditions, including anxiety disorders. One modulator of the stress response is gamma-aminobutyric acid, an inhibitory neurotransmitter synthesized by glutamic acid decarboxylase (GAD). This study examined sex differences in behavior and GAD expression following prolonged social isolation beginning in adolescence in Long Evans rats. Males and females were equally divided into group-housed (GH) and socially isolated conditions on Postnatal Day 28 (n = 8 per group). Beginning 5 weeks later, tests were conducted for anxietylike behaviors (open-field test and elevated plus maze), social interactions (sociability test), and spatial memory (novel object location). Sex differences in behavior were observed, with GH females showing fewer anxietylike behaviors in the open-field test and elevated plus maze and spending more time with objects (sociability task) compared to GH males. Isolation had no effect on males but increased anxiety and reduced neophilic measures in females, removing sex differences. On the sociability task, all groups spent more time with novel rats compared to objects, suggesting social interest was retained after isolation. In the hippocampus, isolation reduced GAD in both sexes, and sex differences were seen (F > M). However, no group differences in behavior were observed in the hippocampal-dependent novel object location task. Our findings suggest that prolonged social isolation beginning in adolescence is anxiogenic for female Long Evans rats. Furthermore, sex and housing impact hippocampal GABA-ergic activity, which may have important implications in the treatment of anxiety disorders. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Perinatal exposure to bisphenol A impairs cognitive function via the gamma-aminobutyric acid signaling pathway in male rat offspring.

Bisphenol A (BPA) is a common synthetic endocrine disruptor that can be utilized in the fabrication of materials such as polycarbonates and epoxy resins. Numerous studies have linked BPA to learning and memory problems, although the precise mechanism remains unknown. Gamma-aminobutyric acid (GABA) is the most abundant inhibitory neurotransmitter in the vertebrate central nervous system, and it is intimately related to learning and memory. This study aims to evaluate whether altered cognitive behavior involves the GABA signaling pathway in male offspring of rats exposed to BPA during the prenatal and early postnatal periods. Pregnant rats were orally given BPA (0, 0.04, 0.4, and 4 mg/kg body weight (BW)/day) from the first day of pregnancy to the 21st day of breastfeeding. Three-week-old male rat offspring were selected for an open-field experiment and a new object recognition experiment to evaluate the effect of BPA exposure on cognitive behavior. Furthermore, the role of GABA signaling markers in the cognition affected by BPA was investigated at the molecular level using western blotting and real-time polymerase chain reaction (RT-PCR). The research demonstrated that BPA exposure impacted the behavior and memory of male rat offspring and elevated the expression of glutamic acid decarboxylase 67 (GAD67), GABA type A receptors subunit (GABAARα1), and GABA vesicle transporter (VGAT) in the hippocampus while decreasing the expression levels of GABA transaminase (GABA-T) and GABA transporter 1 (GAT-1). These findings indicate that the alteration in the expression of GABA signaling molecules may be one of the molecular mechanisms by which perinatal exposure to BPA leads to decreased learning and memory in male rat offspring.

Postnatal development of the hippocampal GABAergic system in rats genetically prone to audiogenic seizures.

Epileptogenesis can be associated with altered genetic control of the GABAergic system. Here we analyzed Krushinsky-Molodkina (KM) rats genetically prone to audiogenic epilepsy. KM rats express fully formed audiogenic seizures (AGSs) not early, then they reach 3months. At the age of 1-2months, KM rats either do not express AGS or demonstrate an incomplete pattern of seizure. Such long-term development of AGS susceptibility makes KM rats an especially convenient model to investigate the mechanisms and dynamics of the development of inherited epilepsy. The analysis of the GABAergic system of the hippocampus of KM rats was done during postnatal development at the 15th, 60th, and 120th postnatal days. Wistar rats of corresponding ages were used as a control. In the hippocampus of KM pups, we observed a decrease in the expression of glutamic acid decarboxylase 67 (GAD67) and parvalbumin (PV), which points to a decrease in the activity of GABAergic neurons. Analysis of the 2-month-old KM rats showed an increase in GAD67 and PV expression while synapsin I and vesicular GABA transporter (VGAT) were decreased. In adult KM rats, the expression of GAD67, PV, and synapsin I was upregulated. Altogether, the obtained data indicate significant alterations in GABAergic transmission in the hippocampus of audiogenic KM rats during the first postnatal months.

Age-related changes in glutamic acid decarboxylase 1 gene expression in the medial prefrontal cortex and ventral hippocampus of fear-potentiated rats subjected to isolation stress

Gamma-aminobutyric acid (GABA) plays a crucial role as a neurotransmitter in anxiety circuits, prominently in the hippocampus, amygdala, and prefrontal cortex. The synthesis of GABA in the central nervous system is primarily governed by glutamic acid decarboxylase 67 (GAD67). Aging is associated with emotional alterations, and isolation stress has been linked to increased anxiety. This study aimed to investigate the impact of aging on the gene expression of GAD67 (Gad1) in the medial prefrontal cortex (m PC) and ventral hippocampus (v Hip) of fear-potentiated rats subjected to isolation stress. To conduct the study, Wistar rats of different age groups 21-day-old (immature), 42-day-old (peri-adolescent), and 365-day-old (mature adult) were utilized. Each age level was categorized into four groups: 1) Control group - no pre-stressor, no maze, no drug, 2) Innate fear group (M) - no pre-stressor, maze, no drug, 3) Fear-potentiated group (IM) - isolation pre-stressor for 120 min, maze, no drug, and 4) Diazepam-treated group (IMD) - isolation pre-stressor for 120 min, maze, and diazepam administration. Following the tests, the (m PC) and (v Hip) regions were dissected, and Gad1 gene expression changes were assessed using Real-time PCR technique. The results revealed that, across all age groups, Gad1 expression in both the (m PC) and (v Hip) was significantly higher in the fear-potentiated groups (IM) compared to the control and innate fear (M) groups. Notably, in aged 365-day-old rats from the innate fear group (M), the expression of Gad1 in (v Hip) was also higher than that in the control group. Additionally, aged fear-potentiated rats exhibited elevated Gad1 gene expression in both structures compared to other age groups. These findings suggest that isolation stress before exposure to the elevated plus maze (EPM) can elevate Gad1 gene expression in both the (v Hip) and (m PC), and age may play a role in modulating its expression.

Zerumbone, a major sesquiterpene from Syringa pinnatifolia Hemsl., exerts the sedative effect by regulating GABAergic nervous system

Ethnopharmacological relevanceZerumbone (ZER) is a humulane sesquiterpenoid isolated from Syringa pinnatifolia Hemsl. (SP), its content accounts for 64.7% of volatile oil and 0.86% of total ethanol extract (TEE), representing one of characteristic ingredient of SP. As a representative Mongolian medicine with anti-“Khii”, anti-asthma, and clearing-heat effects, SP has been used for the treatment of cardiovascular diseases, upset, insomnia, and other symptoms. Aim of studyPrevious results showed that TEE has sedative effect, but the pharmacological substances and its sedative mechanism remains unclear. This study aims to determine whether ZER, as one of major and characteristic sesquiterpenoids of SP, contributes to the sedative effect of SP and its underlying mechanism. Materials and methodsLocomotor activity and threshold dose of pentobarbital sodium sleep experiments were used to evaluate the sedative effects in mice. ELISA assay was used to examine the level of GABA/Glu ratio in rats hippocampus, cortex and hypothalamus tissue. The binding ability of ZER with glutamic acid decarboxylase 67 (GAD67) and Gephyrin protein were predicted by molecular docking. Western blot and Immunohistochemistry assay were used to determine the expression of GABAergic nerve system related proteins (GAD67, Gephyrin) in rat’s hypothalamus. ZER was co-administrated with flumazenil and bicuculline (GABAA antagonist) to determine whether it acts on GABAA receptor. Furthermore, MQAE assay was used to test the effect of ZER on the chloride ion concentration in cerebellar granule cells. ResultsCurrent data demonstrated that ZER dose-dependently (5–20 mg/kg) reduces the locomotor activity and sleep latency of mice, and extend sleeping time of mice. The results of ELISA showed that ZER increases the level of GABA/Glu in rats brain tissue, in particular in hypothalamus. Molecular docking results revealed that ZER has a strong affinity to GAD67 and Gephyrin protein. The Western blot and Immunohistochemistry data indicated that ZER up-regulates the expression of GAD67 and Gephyrin protein in rat’s hypothalamus. Antagonism test results demonstrated that flumazenil and bicuculline reverse the effect of ZER on threshold dose of pentobarbital sodium sleep experiments. In addition, ZER also could dose-dependently (5–20 μM) increase the chloride ion concentration in cerebellar granule cell, suggesting that ZER induces the opening of chloride channel, exerts central inhibitory effect. ConclusionZER has a significant sedative effect in mice and rat, and the effect is associated with GABAergic nervous system. The present results suggest that ZER, as one of the major bioactive ingredients of SP, contributes to the sedative effect and provide substantial evidence for its traditional use of anti-“Khii” in clinic of Syringa pinnatifolia.

A GABA-producing probiotic for the protection of CNS demyelinating inflammation

Abstract The gut-microbiota-brain axis has emerged as a critical pathway in the regulation of neuroinflammation. The gut microbiome regulates the severity of many experimental models of autoimmune central nervous system (CNS) demyelinating inflammatory diseases. Our most recent findings demonstrate that the microbiota of mice from different sources affects the severity of CNS inflammatory demyelination. Neuroinflammation triggered in experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis, modified the gut microbiota composition. The disease progression resulted in a significant reduction in the relative abundances of members of lactic acid bacteria when compared to healthy control mice. Among the altered taxa, bacteria producing gamma-aminobutyric acid (GABA) were significantly reduced. We hypothesized that modifying the microbiota with a probiotic while increasing intestinal GABA levels would reduce EAE’s severity. We genetically engineered a Lactococcus lactis probiotic that produces excessed levels of GABA. Real-time quantitative PCR analysis demonstrated an elevated expression of glutamic acid decarboxylase (GAD). GABA-specific ELISA showed a significant increase in neurotransmitter production when exposed to increasing concentrations of glutamic acid and time. In vivo, five times/week oral gavages with 5 × 108 CFU/mouse of GAD L. lactis but not with empty-plasmid carrier L. lactis protects against EAE in mice compared with sham-treated mice and prevents weight loss of animals while modulating the microbiome’s composition. Our results show that the increase of GABA at the intestinal level with the oral treatment with a probiotic strain protects against neuroinflammation in the CNS. Supported by grant from NIH (R15 NS107743)

Body Weight-Supported Treadmill Training Ameliorates Motoneuronal Hyperexcitability by Increasing GAD-65/67 and KCC2 Expression via TrkB Signaling in Rats with Incomplete Spinal Cord Injury

Spasticity is a typical consequence after spinal cord injury (SCI). The critical reasons are reducing the synthesis of Gamma-Aminobutyric Acid (GABA), glycine and potassium chloride co-transporter 2 (KCC2) inside the distal spinal cord. The current work aimed to test whether exercise training could increase the expression of glutamic acid decarboxylase 65/67 (GAD-65/67, the key enzymes in GABA synthesis) and KCC2 in the distal spinal cord via tropomyosin-related kinase B (TrkB) signaling. The experimental rats were randomly assigned to the following five groups: Sham, SCI/phosphate-buffered saline (PBS), SCI-treadmill training (TT)/PBS, SCI/TrkB-IgG, and SCI-TT/TrkB-IgG. After that, the model of T10 contusion SCI was used, then TrkB-IgG was used to prevent TrkB activity at 7 days post-SCI. Body weight-supported treadmill training started on the 8th day post-SCI for four weeks. The Hmax/Mmax ratio and the rate-dependent depression of H-reflex were used to assess the excitability of spinal motoneuronal networks. Western blotting and Immunohistochemistry techniques were utilized for measuring the expression of GAD-65, GAD-67, and KCC2. The findings revealed that exercise training could reduce motoneuronal excitability and boost GAD-65, GAD-67, and KCC2 production in the distal region of the spinal cord after SCI. The effects of exercise training were decreased after the TrkB signaling was inhibited. The present exploration demonstrated that exercise training increases GAD-65, GAD-67, and KCC2 expression in the spinal cord via TrkB signaling and that this method could also improve rats with motoneuronal hyperexcitability and spasticity induced by incomplete SCI.

The GABAB receptor agonist STX209 reverses the autism‑like behaviour in an animal model of autism induced by prenatal exposure to valproic acid.

Autism spectrum disorder (ASD) is a lifelong neurodevelopmental condition characterized by impaired social interaction, compromised communication, and restrictive or stereotyped behaviours and interests. Due to the complex pathophysiology of ASD, there are currently no available medical therapies for improving the associated social deficits. Consequently, the present study investigated the effects of STX209, a selective γ-aminobutyric acid type B receptor (GABABR2) agonist, on an environmental rodent model of autism. The mouse model of autism induced by prenatal exposure to valproic acid (VPA) was used to assess the therapeutic potential of STX209 on autism-like behaviour in the present study. This study investigated the effects of STX209 on VPA model mice via behavioral testing and revealed a significant reversal of core/associated autism-like behavior, including sociability and preference for social novelty, novelty recognition, locomotion and exploration activity and marble-burying deficit. This may be associated with STX209 correcting dendritic arborization, spine density and GABABR2 expression in hippocampus of VPA model mice. However, expression of glutamic acid decarboxylase 65/67 in the hippocampus were not altered by STX209. The present results demonstrated that STX209 administration ameliorated autism-like symptoms in mice exposed to VPA prenatally, suggesting that autism-like symptoms in children with a history of prenatal VPA exposure may also benefit from treatment with the GABABR2 agonist STX209.

Specific Expression of KCC2 in the α Cells of Normal and Type 1 Diabetes Model Mouse Pancreatic Islets.

Gamma-aminobutyric acid (GABA) is an inhibitory neurotransmitter in the mature brain; however, it acts excitatory during development. This difference in action depends on the intracellular chloride ion concentration, primarily regulated by potassium chloride co-transporter2 (KCC2). Sufficient KCC2 expression results in its inhibitory action. GABA is also abundant in pancreatic islets, where it acts differentially on the islet cells, and is involved in carbohydrate metabolism. However, the mechanisms underlying the differential action remain unknown. We performed immunohistochemistry for glutamic acid decarboxylase (GAD), a synthetic enzyme for GABA, and KCC2 in normal adult islets. GAD was co-localized with insulin in β cells, whereas KCC2 was expressed in glucagon-positive α cells. These results are in line with previous observations that GABA decreases glucagon release but increases insulin release, and suggest that GABA and insulin may work together in reducing blood glucose levels under hyperglycemia. Next, we examined the streptozotocin-induced type1 diabetes mellitus mouse model. GAD and insulin expression levels were markedly decreased. KCC2 was expressed in glucagon-positive cells, whereas insulin- and somatostatin-positive cells were KCC2-negative. These findings suggest that in diabetes model, reduced GABA release may cause disinhibition of glucagon release, resulting in increased blood sugar levels and the maintenance of hyperglycemic state.

Expression of GABAergic and glutamatergic neurons after olfactory stimulation in the mouse piriform cortex during postnatal development

Introduction. The control of the survival and differentiation of immature neurons in the piriform cortex of rodents, which can transform into GABAergic and/or glutamatergic neurons under the influence of olfactory stimuli, is an important factor for prevention of neurological dysfunction.
 The aim of the study was to assess the expression of GABAergic and glutamatergic neurons after olfactory stimulation (OS) in the mouse piriform cortex during postnatal development.
 Materials and methods. The study was carried out on CD1 male mice aged 2 (n = 20; group Р2), 21 (n = 20; group Р21) and 60 (n = 20; group Р60) days. The mice were presented with olfactory stimuli, and brain tissue was collected for immunohistochemical analysis 2 hours, 24 hours and 7 days later, to assess glutamic acid decarboxylase 67 (GAD67) and vesicular glutamate transporter 1 (VGlut1) expression.
 Results. OS in the group P2 animals increased VGlut1 expression in the first 2 hours after OS, followed by a return to baseline level by day 7, while GAD67 expression showed no significant changes. The animals in group P21 showed increased expression of VGlut1 and GAD67 two hours after OS, followed by a significant decrease. Expression of both molecules demonstrated a statistically significant increase in the group P60 animals 24 hours after OS, and remained at the same level on day 7 (GAD67) or returned to baseline levels (VGlut1).
 Conclusion. OS increases the number of GABAergic (GAD67+) и glutamatergic (VGlut1+) neurons in the piriform cortex (P60). The predominance of glutamatergic effects is a possible mechanism for associative memory cell recruitment.

Fukutin regulates tau phosphorylation and synaptic function: Novel properties of fukutin in neurons.

Fukutin, a product of the causative gene of Fukuyama congenital muscular dystrophy (FCMD), is known to be responsible for basement membrane formation. Patients with FCMD exhibit not only muscular dystrophy but also central nervous system abnormalities, including polymicrogyria and neurofibrillary tangles (NFTs) in the cerebral cortex. The formation of NFTs cannot be explained by basement membrane disorganization. To determine the involvement of fukutin in the NFT formation, we performed molecular pathological investigations using autopsied human brains and cultured neurons of a cell line (SH‐SY5Y). In human brains, NFTs, identified with an antibody against phosphorylated tau (p‐tau), were observed in FCMD patients but not age‐matched control subjects and were localized in cortical neurons lacking somatic immunoreactivity for glutamic acid decarboxylase (GAD), a marker of inhibitory neurons. In FCMD brains, NFTs were mainly distributed in lesions of polymicrogyria. Immunofluorescence staining revealed the colocalization of immunoreactivities for p‐tau and phosphorylated glycogen synthase kinase‐3β (GSK‐3β), a potential tau kinase, in the somatic cytoplasm of SH‐SY5Y cells; both the immunoreactivities were increased by fukutin knockdown and reduced by fukutin overexpression. Western blot analysis using SH‐SY5Y cells revealed consistent results. Enzyme‐linked immunosorbent assay (ELISA) confirmed the binding affinity of fukutin to tau and GSK‐3β in SH‐SY5Y cells. In the human brains, the density of GAD‐immunoreactive neurons in the frontal cortex was significantly higher in the FCMD group than in the control group. GAD immunoreactivity on Western blots of SH‐SY5Y cells was significantly increased by fukutin knockdown. On immunofluorescence staining, immunoreactivities for fukutin and GAD were colocalized in the somatic cytoplasm of the human brains and SH‐SY5Y cells, whereas those for fukutin and synaptophysin were colocalized in the neuropil of the human brains and the cytoplasm of SH‐SY5Y cells. ELISA confirmed the binding affinity of fukutin to GAD and synaptophysin in SH‐SY5Y cells. The present results provide in vivo and in vitro evidence for novel properties of fukutin as follows: (i) there is an inverse relationship between fukutin expression and GSK‐3β/tau phosphorylation in neurons; (ii) fukutin binds to GSK‐3β and tau; (iii) tau phosphorylation occurs in non‐GAD‐immunoreactive neurons in FCMD brains; (iv) neuronal GAD expression is upregulated in the absence of fukutin; and (v) fukutin binds to GAD and synaptophysin in presynaptic vesicles of neurons.

Evaluation of suprachiasmatic nucleus in Alzheimer’s disease with non-invasive magnetic resonance methods

Evaluation of suprachiasmatic nucleus in Alzheimer’s disease with non-invasive magnetic resonance methods

Reduced GABAergic neuropil and interneuron profiles in schizophrenia: Complementary analysis of disease course-related differences

BackgroundGABAergic interneuron dysfunction has been implicated in the pathophysiology of schizophrenia. Expression of glutamic acid decarboxylase (GAD), a key enzyme in GABA synthesis, may also be altered. Here, we have simultaneously evaluated GAD-immunoreactive (GAD-ir) neuropil and cell profiles in schizophrenia-relevant brain regions, and analysed disease-course related differences. MethodsGAD65/67 immunoreactivity was quantified in specific brain regions for profiles of fibres and cell bodies of interneurons by automated digital image analysis in post-mortem brains of 16 schizophrenia patients from paranoid (n = 10) and residual (n = 6) diagnostic subgroups and 16 matched controls. Regions of interest were superior temporal gyrus (STG) layers III and V, mediodorsal (MD) and laterodorsal (LD) thalamus, and hippocampal CA1 and dentate gyrus (DG) regions. ResultsA reduction in GAD-ir neuropil profiles (p < 0.001), particularly in STG layer V (p = 0.012) and MD (p = 0.001), paralleled decreased GAD-ir cell profiles (p = 0.029) in schizophrenia patients compared to controls. Paranoid schizophrenia patients had lower GAD-ir neuron cell profiles in STG layers III (p = 0.007) and V (p = 0.001), MD (p = 0.002), CA1 (p = 0.001) and DG (p = 0.043) than residual patients. There was no difference in GAD-ir neuropil profiles between paranoid and residual subgroups (p = 0.369). ConclusionsThese results support the hypothesis of GABAergic dysfunction in schizophrenia. They show a more prominent reduction of GAD-ir interneurons in paranoid versus residual patients, suggestive of more pronounced GABAergic dysfunction in the former. Fully automated analyses of histological sections represent a step towards user-independent assessment of brain structure.

Prenatal androgenization causes expression changes of progesterone and androgen receptor mRNAs in the arcuate nucleus of female mice across development.

Prenatal exposure to excess androgens is associated with the development of polycystic ovary syndrome (PCOS). In prenatally androgenised (PNA) mice, a model of PCOS, progesterone receptor (PR) protein expression is reduced in arcuate nucleus (ARC) GABA neurons. This suggests a mechanism for PCOS-related impaired steroid hormone feedback and implicates androgen excess with respect to inducing transcriptional repression of the PR-encoding gene Pgr in the ARC. However, the androgen sensitivity of ARC neurons and the relative gene expression of PRs over development and following prenatal androgen exposure remain unknown. Here, we used a quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) of microdissected ARC to determine the relative androgen receptor (Ar) and progesterone receptor (Pgr) gene expression in PNA and control mice at five developmental timepoints. In a two-way analysis of variance, none of the genes examined showed expression changes with a statistically significant interaction between treatment and age, although PgrA showed a borderline interaction. For all genes, there was a statistically significant main effect of age on expression levels, reflecting a general increase in expression with increasing age, regardless of treatment. For PgrB and Ar, there was a statistically significant main effect of treatment, indicating a change in expression following PNA (increased for PgrB and decreased for Ar), regardless of age. For PgrA, there was a borderline main effect of treatment, suggesting a possible change in expression following PNA, regardless of age. PgrAB gene expression changes showed no significant main effect of treatment. We additionally examined androgen and progesterone responsiveness specifically in P60 ARC GABA neurons using RNAScope® (Advanced Cell Diagnostics, Inc.) in situ hybridization. This analysis revealed that Pgr and Ar were expressed in the majority of ARC GABA neurons in normal adult females. However, our RNAScope® analysis did not show significant changes in Pgr or Ar expression within ARC GABA neurons following PNA. Lastly, because GABA drive to gonadotropin-releasing hormone neurons is increased in PNA, we hypothesised that PNA mice would show increased expression of glutamic acid decarboxylase (GAD), the rate-limiting enzyme in GABA production. However, the RT-qPCR showed that the expression of GAD encoding genes (Gad1 and Gad2) was unchanged in adult PNA mice compared to controls. Our findings indicate that PNA treatment can impact Pgr and Ar mRNA expression in adulthood. This may reflect altered circulating steroid hormones in PNA mice or PNA-induced epigenetic changes in the regulation of Pgr and Ar gene expression in ARC neurons.

Effects of electroacupuncture on pain sensation in a rat model of hyperalgesia with nicotine dependence.

Tobacco smoking is considered to be one of the main risk factors in the development of chronic pain. Long-term chronic exposure to nicotine and other forms of tobacco have been shown to be associated with an increased incidence of pain. Studies have shown that acupuncture can help smokers to reduce their desire to smoke, reduce their withdrawal symptoms, and avoid a relapse after treatment. However, little has been reported about the effects of acupuncture on pain sensitivity caused by long-term smoking. Models of hyperalgesia were established in rats exposed to nicotine for 6 weeks. After 6 weeks of continuous nicotine exposure, electroacupuncture at bilateral acupoints Zusanli (ST36) and Taichong (LR3) was performed 20 minutes per day for 6 days at a continuous wave with a frequency of 2 Hz and a stimulus intensity of 1 mA. The results revealed that electroacupuncture treatment increased the mechanical response threshold of hind paw of nicotine-dependent rats with hyperalgesia and up-regulated the protein expression of pain-related factors μ-opioid receptor, β-endorphin and glutamic acid decarboxylase 65 in the spinal cord and midbrain periaqueductal gray and the protein expression of glutamic acid decarboxylase 67 in the spinal cord. These findings suggest that electroacupuncture treatment has positive analgesic effects on pain sensitivity caused by long-term chronic nicotine exposure. One possible mechanism for the improved analgesia is that electroacupuncture increases the expression of pain-related factors in the spinal cord and midbrain periaqueductal gray. This study was approved by Institutional Animal Care and Use Committee (IACUC) of the University of Miami (#18-167) on December 12, 2018.

Chemogenetic Activation of Cortical Parvalbumin-Positive Interneurons Reverses Noise-Induced Impairments in Gap Detection.

Exposure to loud noises not only leads to trauma and loss of output from the ear but also alters downstream central auditory circuits. A perceptual consequence of noise-induced central auditory disruption is impairment in gap-induced prepulse inhibition, also known as gap detection. Recent studies have implicated cortical parvalbumin (PV)-positive inhibitory interneurons in gap detection and prepulse inhibition. Here, we show that exposure to loud noises specifically reduces the density of cortical PV but not somatostatin (SOM)-positive interneurons in the primary auditory cortex in mice (C57BL/6) of both sexes. Optogenetic activation of PV neurons produced less cortical inhibition in noise-exposed than sham-exposed animals, indicative of reduced PV neuron function. Activation of SOM neurons resulted in similar levels of cortical inhibition in noise- and sham-exposed groups. Furthermore, chemogenetic activation of PV neurons with the hM3-based designer receptor exclusively activated by designer drugs completely reversed the impairments in gap detection for noise-exposed animals. These results support the notions that cortical PV neurons encode gap in sound and that PV neuron dysfunction contributes to noise-induced impairment in gap detection.SIGNIFICANCE STATEMENT Noise-induced hearing loss contributes to a range of central auditory processing deficits (CAPDs). The mechanisms underlying noise-induced CAPDs are still poorly understood. Here we show that exposure to loud noises results in dysfunction of PV-positive but not somatostatin-positive inhibitory interneurons in the primary auditory cortex. In addition, cortical PV inhibitory neurons in noise-exposed animals had reduced expression of glutamic acid decarboxylases and weakened inhibition on cortical activity. Noise exposure resulted in impaired gap detection, indicative of disrupted temporal sound processing and possibly tinnitus. We found that chemogenetic activation of cortical PV inhibitory interneurons alleviated the deficits in gap detection. These results implicate PV neuron dysfunction as a mechanism for noise-induced CAPDs.

Mechanisms for establishment of GABA signaling in adrenal medullary chromaffin cells.

γ-Aminobutyric acid (GABA) is thought to play a paracrine role in adrenal medullary chromaffin (AMC) cells. Comparative physiological and immunocytochemical approaches were used to address the issue of how the paracrine function of GABA in AMC cells is established. GABAA receptor Cl- channel activities in AMC cells of rats and mice, where corticosterone is the major glucocorticoid, were much smaller than those in AMC cells of guinea-pigs and cattle, where cortisol is the major. The extent of enhancement of GABAA receptor α3 subunit expression in rat pheochromocytoma (PC12) cells by cortisol was larger than that by corticosterone in parallel with their glucocorticoid activities. Thus, the species difference in GABAA receptor expression may be ascribed to a difference in glucocorticoid activity between corticosterone and cortisol. GABAA receptor Cl- channel activity in mouse AMC cells was enhanced by allopregnanolone, as noted with that in guinea-pig AMC cells, and the enzymes involved in allopregnanolone production were immunohistochemically detected in the zona fasciculata in both mice and guinea pigs. The expression of glutamic acid decarboxylase 67 (GAD67), one of the GABA synthesizing enzymes, increased after birth, whereas GABAA receptors already developed at birth. Stimulation of pituitary adenylate cyclase-activating polypeptide (PACAP) receptors, but not nicotinic or muscarinic receptors, in PC12 cells, resulted in an increase in GAD67 expression in a protein-kinase A-dependent manner. The results indicate that glucocorticoid and PACAP are mainly responsible for the expressions of GABAA receptors and GAD67 involved in GABA signaling in AMC cells, respectively.

Antinociceptive effect of intrathecal P7C3 via GABA in a rat model of inflammatory pain

The recently identified molecule P7C3 has been highlighted in the field of pain research. We examined the effect of intrathecal P7C3 in tissue injury pain evoked by formalin injection and determined the role of the GABA system in the activity of P7C3 at the spinal level. Male Sprague-Dawley rats with intrathecal catheters implanted for experimental drug delivery were studied. The effects of intrathecal P7C3 and nicotinamide phosphoribosyltransferase (NAMPT) administered 10 min before the formalin injection were examined. Animals were pretreated with bicuculline, a GABA-A receptor antagonist; saclofen, a GABA-B receptor antagonist; L-allylglycine, a glutamic acid decarboxylase (GAD) blocker; and CHS 828, an NAMPT inhibitor; to observe involvement in the effects of P7C3. The effects of P7C3 alone and the mixture of P7C3 with GABA receptor antagonists on KCl-induced calcium transients were examined in rat dorsal root ganglion (DRG) neurons. The expression of GAD and the concentration of GABA in the spinal cord were evaluated. Intrathecal P7C3 and NAMPT produced an antinociceptive effect in the formalin test. Intrathecal bicuculline, saclofen, L-allylglycine, and CHS 828 reversed the antinociception of P7C3 in both phases. P7C3 decreased the KCl-induced calcium transients in DRG neurons. Both bicuculline and saclofen reversed the blocking effect of P7C3. The levels of GAD expression and GABA concentration decreased after formalin injection and were increased by P7C3. These results suggest that P7C3 increases GAD activity and then increases the GABA concentration in the spinal cord, which in turn may act on GABA receptors causing the antinociceptive effect against pain evoked by formalin injection.

Paraventricular Nucleus Infusion of Oligomeric Proantho Cyanidins Improves Renovascular Hypertension.

Oxidative stress plays an important role in the pathogenesis of hypertension. Oligomeric proantho cyanidins (OPC) is the main polyphenol presents in grape seed and is known for its potent antioxidant and anti-inflammatory properties. In the present study, we hypothesize that OPC can attenuate oxidative stress in the paraventricular nucleus of hypothalamus (PVN), ameliorate neurotransmitter imbalance, decrease the blood pressure and sympathetic activity in renovascular hypertensive rats. After induction of renovascular hypertension by the two-kidney one-clip (2K-1C) method, male Sprague-Dawley rats received chronic bilateral PVN infusion of OPC (20 μg/h) or vehicle via osmotic minipump for 4 weeks. We found that hypertension induced by 2K-1C was associated with the production of reactive oxygen species (ROS) in the PVN. Infusion of OPC in the PVN significantly reduced the systolic blood pressure and norepinephrine in plasma of 2K-1C rats. In addition, PVN infusion of OPC decreased the level of ROS and the expression of stress-related nicotinamide adenine dinucleotide phosphate (NADPH) oxidases subunit NOX4, increased the levels of nuclear factor E2-related factor 2 (Nrf2) and antioxidant enzyme, balanced the content of cytokines, increased expression of glutamic acid decarboxylase and decreased the expression of tyrosine hydroxylase in the PVN of 2K-1C rats. Our findings provided strong evidence that PVN infusion of OPC inhibited the progression of renovascular hypertension through its potent anti-oxidative and anti-inflammatory function in the PVN.

Molecular mechanisms involved in the prevention and reversal of ketamine-induced schizophrenia-like behavior by rutin: the role of glutamic acid decarboxylase isoform-67, cholinergic, Nox-2-oxidative stress pathways in mice

Mounting evidences have shown that nicotinamide adenine dinucleotide phosphate oxidase-2 (Nox-2) pathway modifies glutamic-acid decarboxylase-67 (GAD67) (GABAergic enzyme) and cholinergic systems via oxidative-nitrergic mechanisms in schizophrenia pathology. Rutin, a neuroactive antioxidant compound, with proven neuroprotective property has been shown to reduce schizophrenic-like behavior in mice. This study sought to investigate the mechanisms of action of the psychopharmacological activity of rutin in the preventive and reversal effects of ketamine-induced schizophrenic-like behavior, oxidative-nitrergic stress, cholinergic and GABAergic derangements in mice. In the preventive treatment, male mice were given rutin (0.1, 0.2 and 0.4mg/kg) or risperidone (0.5mg/kg) orally for 14days prior to ketamine (20mg/kg, i.p.) treatment from the 8 to 14th day. However, in the reversal treatment, ketamine was given for 14days prior to rutin and risperidone. Behavioral (open-field, social-interaction and Y-maze tests), biochemical (oxidative/nitrergic stress markers, acetylcholinesterase activity), immunohistochemical (GAD67, Nox-2) and neuronal cell deaths in the striatum, prefrontalcortex, and hippocampus were evaluated. Ketamine-induced behavioral impairments were prevented and reversed by rutin. Exposure of mice to ketamine increased malondialdehyde, nitrite contents, acetylcholinesterase activity, neuronal cell death and Nox-2 expressions in the striatum, prefrontalcortex and hippocampus. Conversely, these derangements were prevented and reversed by rutin. The decreased glutathione levels due to ketamine were marked increased by rutin. Rutin only prevented ketamine-induced decrease in GAD67 expression in the striatal-hippocampal region. Altogether, the study showed that the prevention and reversaltreatments of mice with rutin attenuated ketamine-induced schizophrenic-like behaviors via reduction of Nox-2 expression, oxidative/nitrergic stresses, acetylcholinesterase activity, and increased GAD67 enzyme.