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Inhibitory circuits in the auditory brainstem undergo multiple postnatal changes that are both activity-dependent and activity-independent. We tested to see if the shift from GABA- to glycinergic transmission, which occurs in the rat medial nucleus of the trapezoid body (MNTB) around the onset of hearing, depends on sound-evoked neuronal activity. We prevented the activity by bilateral cochlear ablations in early postnatal rats and studied ionotropic GABA and glycine receptors in MNTB neurons after hearing onset. The removal of the cochlea decreased responses of GABAA and glycine receptors to exogenous agonists as well as the amplitudes of inhibitory postsynaptic currents. The reduction was accompanied by a decrease in the number of glycine receptor- or vesicular GABA transporter-immunopositive puncta. Furthermore, the ablations markedly affected the switch in presynaptic GABAA to glycine receptors. The increase in the expression of postsynaptic glycine receptors and the shift in inhibitory transmitters were not prevented. The results suggest that inhibitory transmission in the MNTB is subject to multiple developmental signals and support the idea that auditory experience plays a role in the maturation of the brainstem glycinergic circuits.

Agomelatine is a new antidepressant drug acting as an antagonist of 5-hydroxytryptamine receptor 2C (5-HTR 2C ) and agonist of melatonergic receptors 1 and 2 (MT 1 and MT 2 ). Because of this dual action, it is an atypical antidepressant. The aim of this study was to investigate chronic anticonvulsant effects of agomelatine on penicillin-induced epilepsy model. Adult male Sprague–Dawley rats divided into four groups and were administered with tap water (vehicle), and agomelatine doses of 10 mg/kg, 50 mg/kg and 100 mg/kg for 14 days via oral gavage. After the last doses were given, epileptic seizures were induced by intracortical penicillin (500 IU/2.5 µl) application in rats under urethane (1.25 g/kg intraperitoneal) anesthesia. Electrocorticogram (ECoG) recordings were obtained from the somatomotor cortex through 90 min, and spike frequencies and amplitudes were analyzed. The spike frequency analyses revealed that only 50 mg/kg agomelatine administration decreased the spike frequencies of hypersynchronous discharge of neurons caused by penicillin (p < 0.05). No significant differences in amplitudes between experimental groups were observed. In addition, mRNA expressions of vesicular glutamate transporter 1 (VGLUT1) and vesicular gamma-aminobutyric acid transporter (VGAT) in response to the agomelatine active dose, 50 mg/kg, showed no significant effect of agomelatine on the mRNA expression. Our results indicate that chronic treatment with agomelatine may have potential anticonvulsant effects. Agomelatine may be a promising drug for epilepsy patients having depression due to its antiepileptic and antidepressant effects. © 2018 BAP141318006 This study was supported by Yeditepe University, Turkey and Necmettin Erbakan University Research Foundation (Project Number: BAP141318006), Turkey.

Chronic cerebral ischemia (CCI) is associated with cognitive decline in aging, vascular dementia and Alzheimer's disease. Substantial evidence has shown that chronic cerebral ischemia may cause cognitive impairment, but the underlying neurobiological mechanism is poorly understood so far. In the present study, we used a rat model of chronic cerebral ischemia by permanent bilateral common carotid artery occlusion (BCCAO) to investigate the alterations of glutamatergic and central cholinergic dysfunction, and their causal relationship with the cognitive deficits induced by chronic cerebral ischemia. We found that BCCAO rats exhibited spatial learning and memory impairments dysfunction 3 month after BCCAO. Meanwhile, vGluT levels as well as glutamatergic and central cholinergic positive neurons in the hippocampus CA1-3 field significantly decreased. The protection of glutamergic and cholinergic neurons or regulating glutamate and central cholinergic levels in hippocampal subregion may have beneficial effects on cognitive impairments associated with the possible mechanism in CCI-induced vascular dementia.

Alzheimer's disease (AD) is the most common cause of dementia in the elderly, and the underlying molecular mechanisms of this neurodegenerative disorder are still unclear. γ-Aminobutyric acid (GABA) neurons play an essential role in the excitatory/inhibitory (E/I) balance in the brain, and the GABAergic system may contribute to the pathogenesis of AD. We used human induced pluripotent stem cells (iPSCs) generated from sporadic AD (SAD) patients to analyze the phenotype and transcriptional profiles of SAD iPSC-derived neural cells. We observed reduced neurogenesis and increased astrogenesis in SAD neural differentiation. We discovered elevated levels of GABA, glutamate decarboxylase 67 (GAD67), and vesicular GABA transporter (vGAT) in SAD neurons that indicated increased GABAergic development. Gene expression profiling of SAD neural cultures showed upregulation of the GABAergic signaling pathway and downregulation of the neurogenesis pathway. We presumed that the GABAergic transmission system might be enhanced in SAD neurons, as an early pathological change of SAD, which provides a novel target and new direction for the development of more effective therapeutic strategies.

Diabetic retinopathy is a leading cause of vision loss and blindness. Increasing evidence has shown that the neuronal components of the retina are affected even before the detection of vascular lesions. Hyperglycemia is considered the main pathogenic factor for the development of diabetic complications. Nevertheless, other factors like neuroinflammation, might also contribute for neural changes. To clarify whether hyperglycemia can be the main trigger of synaptic changes, we evaluated whether prolonged elevated glucose per se, mimicking chronic hyperglycemia, is able to change the content and distribution of several exocytotic proteins and vesicular glutamate and GABA transporters in retinal neurons. Moreover, we also tested the hypothesis that an inflammatory stimulus (interleukin-1β) could exacerbate the effects induced by exposure to elevated glucose, contributing for changes in synaptic proteins in retinal neurons. Rat retinal neural cells were cultured for 9 days. Cells were exposed to elevated D-glucose (30 mM) or D-mannitol (osmotic control), for 7 days, or were exposed to interleukin-1β (10 ng/ml) or LPS (1 μg/ml) for 24 h. The protein content and distribution of SNARE proteins (SNAP-25, syntaxin-1, VAMP-2), synapsin-1, synaptotagmin-1, rabphilin 3a, VGluT-1 and VGAT, were evaluated by western blotting and immunocytochemistry. The protein content and immunoreactivity of syntaxin-1, synapsin-1, rabphilin 3a and VGAT increased in retinal neural cells exposed to high glucose. No changes were detected when cells were exposed to interleukin-1β, LPS or mannitol per se. Particularly, exposure to interleukin-1β for 24 h did not exacerbate the effect of high glucose on the content and immunoreactivity of exocytotic proteins, suggesting the primordial role of hyperglycemia for neuronal changes. In summary, prolonged exposure to elevated glucose alters the total content of several proteins involved in exocytosis, suggesting that hyperglycemia per se is a fundamental factor for neuronal changes caused by diabetes.

Valproic acid (VPA) has been used to treat epileptic patients because of its ability to potentiate GABA signaling in the brain. Despite its clinical significance, VPA administration during pregnancy increases the risk of congenital abnormalities, such as neural tube defects and neurodevelopmental disorders including autism. Furthermore, recent studies revealed that early postnatal administration of VPA also leads to neurodevelopmental deficits in rodents. Here, using cultured cortical neurons derived from postnatal day 1 rats, we examined whether exposure to VPA would affect synapse formation. When neurons were exposed to 1mM VPA during early development, expression of the vesicular GABA transporter (VGAT) was selectively reduced, whereas other synaptic markers, including the vesicular glutamate transporters 1 and 2 (VGLUT1 and 2), were not affected. This VPA effect was mediated through inhibition of histone deacetylases (HDACs), since the effects were mostly recapitulated by an HDAC inhibitor, trichostatin A, but not by a VPA derivative, valpromide, which lacks HDAC inhibitor activity. Immunocytochemical analysis demonstrated that VPA exposure resulted in a retardation of axonal growth specific to GABAergic neurons and a decrease in VGAT-positive synapses. Since disturbance of the excitatory and inhibitory (E-I) balance has been implicated as a potential cause of multiple psychiatric disorders, our results may account for one of the cellular mechanisms underlying the pathogenesis of VPA-induced neurodevelopmental impairments.

In the mature central nervous system, γ-amino butyric acid (GABA) is an inhibitory neurotransmitter, whereas during development, GABA induces depolarization. To examine the embryonic development of GABAergic transmission in the mouse spinal trigeminal nucleus interpolaris (SpVi), which receives sensory input from the face and is important in survival of rodents, we performed immunohistochemistry for three related molecules: glutamic acid decarboxylase (GAD), a marker of GABAergic neurons; vesicular GABA transporter (VGAT), a marker of GABAergic and glycinergic vesicles; and potassium chloride co-transporter 2 (KCC2), which shifts GABA action from excitatory to inhibitory. GAD-positive longitudinal projection fibers, where VGAT-positive dots were localized, were clearly discernible until embryonic day (E)17, and were markedly decreased in number on postnatal day 0. GAD-positive neurons were detected after E15, and GAD- and VGAT-positive axon varicosities were observed after E17. KCC2 immunolabeling was first localized in the dendrites and cell bodies of several neurons in the lateral part of the SpVi on E13 and throughout the nucleus on E17. These results suggest that the SpVi may first receive GABAergic projection fibers from extra-nuclear area before birth, and GABAergic interneurons may form synapses within the SpVi after E17. In addition, GABA action may gradually shift from excitatory to inhibitory between E13 and E17.

Gephyrin is a major postsynaptic scaffolding protein at GABAergic and glycinergic inhibitory synapses. Gephyrin-deficient (geph(-/-)) mice die after birth due to disinhibition of motor and sensory pathways resulting from a lack of postsynaptic glycine receptor and GABA(A) receptor clusters. Here, immunoelectron and confocal microscopy revealed that postsynaptic membrane specializations are formed in the absence of gephyrin. First, in brainstem sections obtained from newborn geph(-/-) mice inhibitory nerve terminals identified by immunogold labeling of either the vesicular inhibitory amino acid transporter (VIAAT) or GABA were found to be apposed to postsynaptic membrane areas decorated by electron-dense material. Second, neuroligin-2, a membrane protein of inhibitory postsynapses, was clustered beneath glutamate decarboxylase 65 (GAD-65) positive nerve terminals in geph(-/-) hippocampal cultures. These results indicate that proteins other than gephyrin define the ultrastructure of inhibitory postsynaptic membrane specializations.

Prompted by previous studies suggesting a regulatory role for the inhibitory amino acid γ -aminobutyric acid (GABA) within the mammalian pineal gland, we carried out a study of rat and gerbil pineal organs to elucidate whether there is evidence for a vesicular storage and release of GABA and/or glycine. Immunohistochemistry revealed the presence of the vesicular inhibitory amino acid transporter in pinealocytes. Moreover, we found that, in addition to glutamate and aspartate, cultured pinealocytes also released glycine upon stimulation by depolarizing concentrations of KCl, whereas the content of GABA in the culture medium did not exceed the detection limit either under control conditions or following KCl application. Therefore, we propose that glycine is a further component of the paracrine signaling system within the pineal organ which is based on the compartment of synaptic-like microvesicles (SLMVs) inside pinealocytes.

The neural cell recognition molecule NB-3, which is also referred to as contactin-6, is a member of the contactin subgroup molecules that are expressed prominently in the developing nervous system after birth. In mice, an NB-3 deficiency impairs motor coordination and reduces the synaptic density between parallel fibers and Purkinje cells in the cerebellum. Here, we studied the role of NB-3 in the formation of glutamatergic synapses in the hippocampal formation. At postnatal day 5, NB-3 immunoreactivity was detected in the subiculum, the stratum lacunosum-moleculare of the CA1 region and the hilus of the dentate gyrus. NB-3 expression in the strata radiatum and oriens was weak, and it was very weak in the granule cell layer of the dentate gyrus, the pyramidal cell layer of regions CA3 to CA1 and the stratum lucidum. NB-3-positive puncta partially overlapped with vesicular glutamate transporter 1 (VGLUT1) and 2 (VGLUT2), excitatory presynaptic markers, but not with vesicular GABA transporter (VGAT), an inhibitory presynaptic marker. The density of VGLUT1 and VGLUT2 puncta in the regions where NB-3 was strongly expressed in wild-type mice was reduced by approximately 20-30% in NB-3 knockout mice relative to wild-type mice, whereas that of VGAT puncta was not affected by NB-3 deficiency. Thus, NB-3 has key roles in the formation of glutamatergic, but not GABAergic, synapses during postnatal development of the hippocampal formation as well as the cerebellum.

Gamma-aminobutyric acid (GABA) is an inhibitory amino acid and acts as an intercellular transmitter in the central nervous system and peripheral tissues. In pineal glands, GABA is supposed to be a paracrine-like modulator of secretion of melatonin, although its mode of action, especially the sites of GABA signal appearance, is unknown. Vesicular inhibitory amino acid transporter (VIAAT) is a potential marker for the GABAergic phenotype. Here we presented evidence that VIAAT is expressed in GFAP-expressing astrocytes and a subpopulation of OX42-expressing microglia, but not in pinealocytes in cultured cells of rat pineal glands. The VIAAT-expressing cells also exhibit GABA immunoreactivity. Essentially the same results were obtained for pineal glands. These results suggest that GABA is stored and secreted from astrocytes and a subpopulation of microglia in pineal glands.