Gamma-aminobutyric acid (GABA) immunoreactivity in the mouse adrenal gland

Gamma-aminobutyric acid (GABA) immunoreactivity was revealed by immunocytochemistry in the mouse adrenal gland at the light and electron microscopic levels. Groups of weakly or faintly GABA immunoreactive chromaffin cells were often seen in the adrenal medulla. By means of immunohistochemistry combined with fluorescent microscopy, these GABA immunoreactive chromaffin cells showed noradrenaline fluorescence. The immunoreaction product was seen mainly in the granular cores of these noradrenaline cells. These results suggest the co-existence of GABA and noradrenaline within the chromaffin granules. Sometimes thick or thin bundles of GABA immunoreactive nerve fibers with or without varicosities were found running through the cortex directly into the medulla. In the medulla, GABA immunoreactive varicose nerve fibers were numerous and were often in close contact with small adrenaline cells and large ganglion cells; a few, however, surrounded clusters of the noradrenaline cells, where membrane specializations were formed. Single GABA immunoreactive nerve fibers, and thin or thick bundles of the immunoreactive varicose nerve fibers ran along the blood vessels in the medulla. The immunoreaction deposits were observed diffusely in the axoplasm and in small agranular vesicles of the GABA immunoreactive nerve fibers. Since no ganglion cells with GABA immunoreactivity were found in the adrenal gland, the GABA immunoreactive nerve fibers are regarded as extrinsic in origin.     

Gamma-aminobutyric acid (GABA) increases in vitro germ-tube formation and phospholipase B1 mRNA expression in Candida albicans

Candida albicans is a commensal yeast in humans that disseminates in immunocompromised persons. Its spreading is modulated by melanin, hormones, or some neurotransmitters, among other factors. The neurotransmitter gamma-aminobutyric acid (GABA) is used by bacteria, plants, and fungi as a carbon and nitrogen source. In this article, the in vitro effect of different doses of GABA on germ-tube formation and expression of phospholipase B1 (PLB1) mRNA in two Candida albicans strains was investigated. Results demonstrated that GABA increases both germ-tube formation and PLB1 mRNA expression in the two Candida strains in a dose-dependent manner, which suggests that GABA promotes the growth of C. albicans.     

Gamma-aminobutyric acid (GABA) in the rat brain: re-evaluation of sampling procedures and the post-mortem increase

—GABA levels determined after the brain was removed, then frozen, were found to be generally compatible to levels after the brain was frozen in situ provided removal and freezing were effected in less than 60 s. Optimal GABA values were realized when the brain was removed and frozen in liquid nitrogen within 30 s of death. Beginning at 60 s post-mortem, GABA levels increased until the 4th min with the greatest rate occurring between 60 and 120 s at 30 μg/g/min. When frozen brains were dissected into regions by a partial thawing technique, post-mortem increases were not found to occur. Microwave irradiation, investigated as a potential sampling technique for GABA assay, showed considerable variability between samples and was rejected as a practical alternative at this time.     

On the formation of gamma-aminobutyric acid from putrescine in brain.

Gamma-aminobutyric acid is not formed in significant amounts from putrescine by incubation with rat brain homogenates. However, it is formed if acetyl-CoA is added to the incubation medium. This is taken as further evidence for the existence of a metabolic pathway in mammalian brain which comprises acetylation of putrescine to monoacetyl putrescine and oxidative deamination of monoacetyl putrescine by MAO. Nerve cells and glia cells have comparable capacities for putrescine degradation along this pathway.     

Gamma-aminobutyric acid (GABA) and pentobarbital induce different conformational rearrangements in the GABA A receptor alpha1 and beta2 pre-M1 regions

Gamma-aminobutyric acid (GABA) binding to GABA(A) receptors (GABA(A)Rs) triggers conformational movements in the alpha(1) and beta(2) pre-M1 regions that are associated with channel gating. At high concentrations, the barbiturate pentobarbital opens GABA(A)R channels with similar conductances as GABA, suggesting that their open state structures are alike. Little, however, is known about the structural rearrangements induced by barbiturates. Here, we examined whether pentobarbital activation triggers movements in the GABA(A)R pre-M1 regions. Alpha(1)beta(2) GABA(A)Rs containing cysteine substitutions in the pre-M1 alpha(1) (K219C, K221C) and beta(2) (K213C, K215C) subunits were expressed in Xenopus oocytes and analyzed using two-electrode voltage clamp. The cysteine substitutions had little to no effect on GABA and pentobarbital EC(50) values. Tethering chemically diverse thiol-reactive methanethiosulfonate reagents onto alpha(1)K219C and alpha(1)K221C affected GABA- and pentobarbital-activated currents differently, suggesting that the pre-M1 structural elements important for GABA and pentobarbital current activation are distinct. Moreover, pentobarbital altered the rates of cysteine modification by methanethiosulfonate reagents differently than GABA. For alpha(1)K221Cbeta(2) receptors, pentobarbital decreased the rate of cysteine modification whereas GABA had no effect. For alpha(1)beta(2)K215C receptors, pentobarbital had no effect whereas GABA increased the modification rate. The competitive GABA antagonist SR-95531 and a low, non-activating concentration of pentobarbital did not alter their modification rates, suggesting that the GABA- and pentobarbital-mediated changes in rates reflect gating movements. Overall, the data indicate that the pre-M1 region is involved in both GABA- and pentobarbital-mediated gating transitions. Pentobarbital, however, triggers different movements in this region than GABA, suggesting their activation mechanisms differ.     

Gamma-aminobutyric acid (GABA) receptor mediates suanzaorentang, a traditional Chinese herb remedy, -induced sleep alteration

Summary The sedative-hypnotic medications, including benzodiazepines and non-benzodiazepines, are the most common treatments for insomnia. However, concerns regarding patterns of inappropriate use, dependence and adverse effects have led to caution in prescribing those sedative-hypnotic medications. On the other hand, a traditional Chinese herb remedy, suanzaorentang, has been efficiently and widely used in clinic for insomnia relief without severe side effects in Asia. Although suanzaorentang has been reported to improve sleep disruption in insomniac patients, its mechanism is still unclear. The present study was designed to elucidate the effects of oral administration of suanzaorentang on physiological sleep-wake architectures and its underlying mechanism in rats. We found that oral administration of suanzaorentang at the beginning of the dark onset dose-dependently increased non-rapid eye movement sleep (NREMS) during the dark period, but had no significant effect on rapid eye movement sleep (REMS). Our results also indicated that intracerebroventricular (ICV) administration of γ-aminobutyric acid (GABA) receptor type A antagonist, bicuculline, significantly blocked suanzaorentang-induced enhancement in NREMS during the dark period, but GABA_B receptor antagonist, 2-hydroxysaclofen had no effect. These results implicated that this traditional Chinese herb remedy, suanzaorentang increases spontaneous sleep activity and its effects may be mediated through the GABA_A receptors, but not GABA_B receptors.     

GABA from putrescine is bound in macromolecular form in keratinocytes

We report for the first time that the neurotransmitter γ-aminobutyric acid (GABA) exists in macromolecular form in keratinocytes. GABA derived from putrescine (Pu) has been identified as a component of acid-precipitable material of cultured human keratinocytes. Confluent, stratified cultures of human foreskin keratinocytes exposed to [³H]-Pu for 4 hours took up about 14% of the radioactivity from the medium and 1% of the total cell-associated radioactivity was precipitable by trichloroacetic acid (TCA). Both attached and shed cells were examined by HPLC for Pu and its radioactive metabolites in TCA-insoluble and TCA-soluble fractions. GABA accounted for the major portion (54%) of the radioactivity derived from Pu in the TCA-precipitable material of attached keratinocytes. Pu and spermidine represented lesser amounts, 35% and 9% respectively, of the total TCA-precipitable radioactivity. In addition, a large portion of acid soluble radioactivity derived from Pu (63%) was GABA, whereas Pu and spermidine represented 29% and 6% respectively of the total TCA-soluble radioactivity. The exact origin of GABA in acid-precipitable material, as well as its form of attachment, is currently under investigation.     

Gamma-aminobutyric acid type B (GABA(B)) receptor internalization is regulated by the R2 subunit

γ-Aminobutyric acid type B (GABA(B)) receptors are important for slow synaptic inhibition in the CNS. The efficacy of inhibition is directly related to the stability of cell surface receptors. For GABA(B) receptors, heterodimerization between R1 and R2 subunits is critical for cell surface expression and signaling, but how this determines the rate and extent of receptor internalization is unknown. Here, we insert a high affinity α-bungarotoxin binding site into the N terminus of the R2 subunit and reveal its dominant role in regulating the internalization of GABA(B) receptors in live cells. To simultaneously study R1a and R2 trafficking, a new α-bungarotoxin binding site-labeling technique was used, allowing α-bungarotoxin conjugated to different fluorophores to selectively label R1a and R2 subunits. This approach demonstrated that R1a and R2 are internalized as dimers. In heterologous expression systems and neurons, the rates and extents of internalization for R1aR2 heteromers and R2 homomers are similar, suggesting a regulatory role for R2 in determining cell surface receptor stability. The fast internalization rate of R1a, which has been engineered to exit the endoplasmic reticulum, was slowed to that of R2 by truncating the R1a C-terminal tail or by removing a dileucine motif in its coiled-coil domain. Slowing the rate of internalization by co-assembly with R2 represents a novel role for GPCR heterodimerization whereby R2 subunits, via their C terminus coiled-coil domain, mask a dileucine motif on R1a subunits to determine the surface stability of the GABA(B) receptor.     

Possible mechanisms of synapse formation during ontogenesis

Electron microscopic investigation of synaptogenesis in the sensomotor cortex and in the caudate nucleus has been performed in the prenatal ontogenesis (16-22 days) and in newborn rats. The first immature synapses are demonstrated to appear beginning on the 16th day of embryogenesis. At the end of the prenatal development and especially in newborn animals desmosome-like, asymmetric and symmetric, mixed and complex forms of the synaptic contacts are revealed. As a result of the analysis performed on the ultrastructural organization of the contacts, a hypothesis explaining mechanisms of development of various elements of the synapses has been suggested. A part of the synaptic contacts of the asymmetric and symmetric types is supposed to be genetically programmed and membrane specialization of these contacts is formed earlier than synaptic vesicles appear. Other part of the synapses undergoes certain stages of differentiation before the functionally mature contact is formed. The initial stage in the synapses formation in formation of the desmosome-like junction. The second stage is appearance of synaptic vesicles in the area of this contact. The third stage includes development of pre- and postsynaptic membranous specialization and owing to this the contact acquires either asymmetric or symmetric appearance. For the ontogenetic periods investigated establishment of complex forms of the intercellular junctions (tangent, reciprocal, etc.) is specific; this evidently demonstrates certain plastic rearrangements in the synapses during the process of development.     

Excitatory action of GABA in ontogenesis

GABA (gamma-aminobutyric acid) is the main inhibitory mediator in central nervous system. However, at early stages of ontogenesis, GABA has an excitatory effect on immature neurons. This review surveys modern concepts of the mechanisms of GABAergic excitation and physiological role of excitatory GABA in generation of patterns of network activity in developing brain.     

γ—Aminobutyric acid transporter （GAT1） overexpression in mouse affects the testicular morphology

γ-Aminobutyric acid and GABAergic receptors were previously reported to be distributed in reproductive systems besides CNS and predicted to participate in the modulation of testicular function.γ-Aminobutyric acid transporter was implicated to be involved in this process.However,the potential role of γ-aminobutyric transporter in testis has not been explored.In this study,we investigated the existence of mouse γ-aminobutyric acid transporter subtype I (mGAT1) in testis.Wild-type and transgenic mice,which overexpressing mGAT1 in a variety of tissues,especially in testis,were primarily studied to approach the profile of mGAT1 in testis.Mice with overexpressed mGAT1 develop normally but with reduced mass and size of testis as compared with wild-type.Testicular morphology of transgenic mice exhibited overt abnormalities including focal damage of the spermatogenic epithelium accompanied by capillaries proliferation and increased diameter of seminiferous tubules lumen.Reduced number of spermatids was also found in some seminiferous tubules.Our results clearly demonstrate the presence of GAT1 in mouse testis and imply that GAT1 is possibly involved in testicular function.     

Impaired reproduction in transgenic mice overexpressing γ-aminobutyric acid transporter I （GAT1）

It is well documented that 7-aminobutyric acid (GABA) system existed in reproductive organs. Recent researches showed that GABAA and GABAB receptors were present in testis and sperm, and might mediate the acrosome reaction induced by GABA and progesterone. GABA transporter I (GAT1) also existed in testis and sperm, but its physiological function was unknown. In the present study, we used GAT1 overexpressing mice to explore GAT1 function in male reproductive system. We found that the expression level of GAT1 continuously increased in wild-type mouse testis from 1 month to 2 months after birth. GAT1 overexpression in mouse affected testis development, which embodied reduced testis mass and slowed spermatogenesis in transgenic mice. Moreover, transgenic mice showed increase of the percentage of broken sperm. The further study revealed that the reproductive capacity was impaired in GAT1 overexpressing mice. In addition, testosterone level was significantly low in transgenic mice compared with that in wild-type mice. Our findings provided the first evidence that abnormal expression of GAT1 could result in dysgenesis,and indicated that GAT1 might be therapeutically targeted for contraception or dysgenesis treatment.