V-107 | Studies on the mechanisms involved in H2O2 modulation of GABAAα1β2 receptors

Neurochemistry and Neuropharmacology
Author: Andrea Beltrán González | Email: andreabeltrangonzalez@gmail.com

Manuel I López Pazos^1°, Victoria Vitali^2°,Karina Alleva^2°, Mariana del Vas^3°, Andrea N Beltrán González^1°, Daniel J Calvo^1°

^1° IFIBYNE-CONICET-UBA
^2° IQUIFIB-CONICET-UBA
^3° IABIMO-INTA y CONICET

Reactive oxygen species are highly reactive molecules generated during cellular metabolism. We studied the effects of H2O2 on GABAA receptors using heterologous receptor expression in Xenopus laevis oocytes followed by two-electrode voltage clamp recording. Previous results indicated that GABAA receptors with αβ subunits were sensitive to H2O2 while those containing the γ2 subunit were highly resistant.
Now, we aimed to elucidate the mechanism involved in H2O2-induced enhancement of GABAAα1β2 receptor activity. The thiol-modifying agent N-ethylmaleimide (NEM) partially inhibited H2O2 potentiation of GABAAα1β2 responses (%Pot= 76.7 ± 5.3 vs. %PotNEM = 45.9 ± 2.6; n = 6; P < 0.001), suggesting participation of one or more cysteine residues. Additionally, we looked whether H2O2 acts via an intracellular mechanism, by developing a co-expression assay that uses aquaporin MtPIP2,2, capable to permeate both water and H2O2, to facilitate the entry of H2O2 into the cell. The presence of MtPIP2,2 did not significantly alter the effect of H2O2 on GABAAα1β2 responses (%Pα1β2 = 40.14 ± 7.4; %Pα1β2+AQP = 33.4 ± 5.2; n = 8; ns). Results suggest that facilitation of H2O2 influx to oocyte cytoplasm does not enhance receptor potentiation. Further experiments will be necessary to address whether the site of action of H2O2 is intracellular.