Bestrophin1-mediated tonic GABA release from reactive astrocytes prevents the development of seizure-prone network in kainate-injected hippocampi.

Tonic extrasynaptic GABAA receptor (GABAA R) activation is under the tight control of tonic GABA release from astrocytes to maintain the brain's excitation/inhibition (E/I) balance; any slight E/I balance disturbance can cause serious pathological conditions including epileptic seizures. However, the pathophysiological role of tonic GABA release from astrocytes has not been tested in epileptic seizures. Here, we report that pharmacological or genetic intervention of the GABA-permeable Bestrophin-1 (Best1) channel prevented the generation of tonic GABA inhibition, disinhibiting CA1 pyramidal neuronal firing and augmenting seizure susceptibility in kainic acid (KA)-induced epileptic mice. Astrocyte-specific Best1 over-expression in KA-injected Best1 knockout mice fully restored the generation of tonic GABA inhibition and effectively suppressed seizure susceptibility. We demonstrate for the first time that tonic GABA from reactive astrocytes strongly contributes to the compensatory shift of E/I balance in epileptic hippocampi, serving as a good therapeutic target against altered E/I balance in epileptic seizures.

Developmental changes in GABAA tonic inhibition are compromised by multiple mechanisms in preadolescent dentate gyrus granule cells.

The sustained tonic currents (Itonic) generated by γ-aminobutyric acid A receptors (GABAARs) are implicated in diverse age-dependent brain functions. While various mechanisms regulating Itonic in the hippocampus are known, their combined role in Itonic regulation is not well understood in different age groups. In this study, we demonstrated that a developmental increase in GABA transporter (GAT) expression, combined with gradual decrease in GABAAR α5 subunit, resulted in various Itonic in the dentate gyrus granule cells (DGGCs) of preadolescent rats. Both GAT-1 and GAT-3 expression gradually increased at infantile (P6-8 and P13-15) and juvenile (P20-22 and P27-29) stages, with stabilization observed thereafter in adolescents (P34-36) and young adults (P41-43). Itonic facilitation of a selective GAT-1 blocker (NO-711) was significantly less at P6-8 than after P13-15. The facilitation of Itonic by SNAP-5114, a GAT-3 inhibitor, was negligible in the absence of exogenous GABA at all tested ages. In contrast, Itonic in the presence of a nonselective GAT blocker (nipecotic acid, NPA) gradually decreased with age during the preadolescent period, which was mimicked by Itonic changes in the presence of exogenous GABA. Itonic sensitivity to L-655,708, a GABAAR α5 subunit inverse agonist, gradually decreased during the preadolescent period in the presence of NPA or exogenous GABA. Finally, Western blot analysis showed that the expression of the GABAAR α5 subunit in the dentate gyrus gradually decreased with age. Collectively, our results suggested that the Itonic regulation of altered GATs is under the final tune of GABAAR α5 subunit activation in DGGCs at different ages.

Facilitation of AMPA receptor-mediated steady-state current by extrasynaptic NMDA receptors in supraoptic magnocellular neurosecretory cells.

In addition to classical synaptic transmission, information is transmitted between cells via the activation of extrasynaptic receptors that generate persistent tonic current in the brain. While growing evidence supports the presence of tonic NMDA current (INMDA) generated by extrasynaptic NMDA receptors (eNMDARs), the functional significance of tonic INMDA in various brain regions remains poorly understood. Here, we demonstrate that activation of eNMDARs that generate INMDA facilitates the α-amino-3-hydroxy-5-methylisoxazole-4-proprionate receptor (AMPAR)-mediated steady-state current in supraoptic nucleus (SON) magnocellular neurosecretory cells (MNCs). In low-Mg(2+) artificial cerebrospinal fluid (aCSF), glutamate induced an inward shift in Iho lding (IGLU) at a holding potential (Vholding) of -70 mV which was partly blocked by an AMPAR antagonist, NBQX. NBQX-sensitive IGLU was observed even in normal aCSF at Vholding of -40 mV or -20 mV. IGLU was completely abolished by pretreatment with an NMDAR blocker, AP5, under all tested conditions. AMPA induced a reproducible inward shift in Iholding (IAMPA) in SON MNCs. Pretreatment with AP5 attenuated IAMPA amplitudes to ~60% of the control levels in low-Mg(2+) aCSF, but not in normal aCSF at Vholding of -70 mV. IAMPA attenuation by AP5 was also prominent in normal aCSF at depolarized holding potentials. Memantine, an eNMDAR blocker, mimicked the AP5-induced IAMPA attenuation in SON MNCs. Finally, chronic dehydration did not affect IAMPA attenuation by AP5 in the neurons. These results suggest that tonic INMDA, mediated by eNMDAR, facilitates AMPAR function, changing the postsynaptic response to its agonists in normal and osmotically challenged SON MNCs.

Antiallodynic effects of KDS2010, a novel MAO-B inhibitor, via ROS-GABA inhibitory transmission in a paclitaxel-induced tactile hypersensitivity model.

Monoamine oxidase (MAO) inhibitors have been investigated for the treatment of neuropathic pain. Here, we assessed the antiallodynic effects of a novel MAO-B inhibitor, KDS2010, on paclitaxel (PTX)-induced mechanical hypersensitivity. Oral administration of KDS2010 effectively relieved PTX-induced mechanical hypersensitivity in a dose-dependent manner. KDS2010 (25 mg/Kg) significantly prevented and suppressed PTX-induced pain responses with minimal effects on the body weight, motor activity, and working memory. KDS2010 significantly reduced reactive astrocytosis and reactive oxygen species (ROS) level in the L4-L6 spinal cord of PTX-treated mice. Furthermore, KDS2010 reversed the attenuation of GABAergic spontaneous inhibitory postsynaptic current (sIPSC) frequency in spinal dorsal horn neurons, although it failed to restore the reduced tonic GABAA inhibition nor the increased GABA transporter 1 (GAT1) expression in PTX-treated mice. In addition, bath application of a reactive oxygen species (ROS) scavenger (PBN) restored the sIPSC frequency in PTX-treated mice but not in control and PTX + KDS2010-treated mice. These results indicated that the antiallodynic effect of KDS2010 is not due to a MAO-B-dependent GABA production. Finally, PBN alone also exerted a similar analgesic effect as KDS2010, but a co-treatment of PBN with KDS2010 showed no additive effect, suggesting that inhibition of MAO-B-dependent ROS production is responsible for the analgesic effect by KDS2010 on PTX-induced allodynia. Overall, KDS2010 attenuated PTX-induced pain behaviors by restoring the altered ROS level and GABAergic inhibitory signaling in the spinal cord, suggesting that KDS2010 is a promising therapeutic strategy for chemotherapy-induced peripheral neuropathy.

Korean red ginseng excitation of paraventricular nucleus neurons via non-N-methyl-D-aspartate glutamate receptor activation in mice.

It has been reported that Korean red ginseng (KRG), a valuable and important traditional medicine, has varied effects on the central nervous system, suggesting its activities are complicated. The paraventricular nucleus (PVN) neurons of the hypothalamus has a critical role in stress responses and hormone secretions. Although the action mechanisms of KRG on various cells and systems have been reported, the direct membrane effects of KRG on PVN neurons have not been fully described. In this study, the direct membrane effects of KRG on PVN neuronal activity were investigated by using a perforated patch-clamp in ICR mice. In gramicidin perforated patch-clamp mode, KRG extract (KRGE) induced repeatable depolarization followed by hyperpolarization of PVN neurons. The KRGE-induced responses were concentration- dependent and persisted in the presence of tetrodotoxin, a voltage sensitive Na+ channel blocker. The KRGE-induced responses were suppressed by 6-cyano-7-nitroquinoxaline-2,3-dione (10 µM), a non-N-methyl-D-aspartate (NMDA) glutamate receptor antagonist, but not by picrotoxin, a type A gamma-aminobutyric acid receptor antagonist. The results indicate that KRG activates non-NMDA glutamate receptors of PVN neurons in mice, suggesting that KRG may be a candidate for use in regulation of stress responses by controlling autonomic nervous system and hormone secretion.

Molecular and electrophysiological characterization of nucleotide-sensitive chloride current-inducing protein of Fasciola hepatica.

Nucleotide-sensitive chloride current regulating proteins (ICln's) of the chloride channels have been characterized from man and animals. An ICln of Fasciola hepatica (ICln-Fh) consisting of 231 amino acids revealed high similarities to both consensus domain of ICln's and two acidic residue-abundant patches in its C-terminus. Native ICln-Fh protein was confirmed present in F. hepatica soluble extract by immunoblotting. The recombinant ICln-Fh protein expressed in collagenase-defolliculated Xenopus oocytes induced fast rising and outward rectifying Cl- currents (I(Cln-Fh)). The recombinant ICln-Fh protein, however, did not trigger cell swelling-induced Cl- currents (I(Cl-swell)). The I(Cln-Fh) currents were significantly reduced by substituting external Cl- with gluconic acid and by externally adding cAMP. Collectively, these results suggest that ICln-Fh protein is an inducer of Cl- currents in F. hepatica.