title: Nitric oxide directly activates GABA(A) receptor function through a cGMP/protein kinase-independent pathway in frog pituitary melanotrophs.
abstract: The direct effects of nitric oxide (NO) donors and sulfhydryl-modifying agents on the GABA(A) receptor function were examined by perforated patch, whole-cell and single channel recordings in cultured frog melanotrophs. In amphotericin B-perforated cells incubated with the soluble guanylyl cyclase inhibitors LY 83583 and ODQ (10-4 M each), the NO donor sodium nitroprusside (SNP) (10(-3) M) reversibly increased the current evoked by GABA (5 x 10(-6) M). In the whole-cell configuration, internal application of the oxidizing agent H2O2 (0.05%) potentiated the GABA-evoked current while the reducing agent 2-mercaptoethanol (5 x 10(-3) M) slightly decreased the current amplitude. In inside-out patches, GABA (2 x 10(-7) M) triggered single channel bursts of openings. Incubation with the NO donors SNP or DEA/NO (10(-4) M each) enhanced the open probability of the GABA(A) receptor channel but did not modify the chloride reversal potential and did not affect the conductance states. The oxidizing agents H2O2 (0.05%) or DTNB (10-4 M) mimicked the stimulatory effect of the NO donors on the open probability while the reducing compounds 2-mercaptoethanol (5 x 10(-3) M) or DTT (10(-4) M) markedly attenuated the channel activity. Potentiation of the GABA-induced single channel activity by SNP or H2O2 was blocked by 2-mercaptoethanol. Similarly, the potentiating effect produced by DEA/NO or DTNB on the open probability was reversed by DTT. In outside-out patches, incubation with SNP also significantly enhanced the open probability of single channels activated by GABA (10(-6) M). These data indicate that, in frog pituitary melanotrophs, NO potentiates the GABA-evoked current independently of the cGMP/protein kinase pathway. The effect of NO can be accounted for by S-nitrosylation/oxidation of thiol groups either directly on the GABA(A) receptor subunits or on a regulatory protein tightly associated with the GABA(A) receptor.
authors.affiliation: European Institute for Peptide Research (IFRMP 23), Laboratory of Cellular and Molecular Neuroendocrinology, INSERM U413, UA CNRS, University of Rouen, Mont-Saint-Aignan, France.
authors.forename: H
authors.initials: H
authors.lastname: Castel
authors.forename: H
authors.initials: H
authors.lastname: Vaudry
mesh_terms: D000634:Aminoquinolines / Q000494:pharmacology; D000818:Animals; D006152:Cyclic GMP / Q000378:metabolism; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006834:Hydrazines / Q000494:pharmacology; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D008297:Male; D008544:Melanocytes / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008623:Mercaptoethanol / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009589:Nitrogen Oxides; D009599:Nitroprusside / Q000494:pharmacology; D010069:Oxadiazoles / Q000494:pharmacology; D010084:Oxidation-Reduction; D018408:Patch-Clamp Techniques; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D011810:Quinoxalines / Q000494:pharmacology; D011895:Rana ridibunda; D011963:Receptors, GABA-A / Q000378:metabolism*; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
chemical_list: C095284:1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one; D000634:Aminoquinolines; D004791:Enzyme Inhibitors; D006834:Hydrazines; D020030:Nitric Oxide Donors; D009589:Nitrogen Oxides; D010069:Oxadiazoles; D011810:Quinoxalines; D011963:Receptors, GABA-A; D013439:Sulfhydryl Reagents; D009599:Nitroprusside; D009569:Nitric Oxide; D005680:gamma-Aminobutyric Acid; D008623:Mercaptoethanol; C084012:1,1-diethyl-2-hydroxy-2-nitrosohydrazine; C041715:6-anilino-5,8-quinolinedione; D004228:Dithionitrobenzoic Acid; D017869:Cyclic GMP-Dependent Protein Kinases; D006152:Cyclic GMP