GABAA - Coupled Cl − / 3 HCO − - ATPase from Plasma Membrane of the Rat Brain : Role of 3 HCO − in the Enzyme Activation

This work examines the influence of Cl− (2.5 125 mM) and 3 HCO − (2 30 mM) on the Cl−/ 3 HCO − ATPase complex of the neuronal membrane and this enzyme is a Cl−-pump that is coupled to GABAA receptors. The greatest (44%) activating effect on the enzyme is found with 3 HCO − (20 30 mM), while the maximum activity occurs in the presence of a ratio of ~25 mM 3 HCO − /~5mM Cl−. Blockers of the GABAA receptor, namely bicuculline (10 50 μM) and picrotoxin (50 100 μM), inhibit this anion activation, whereas the 3 HCO − -ATPase activity is not sensitive to these ligands. Autoradiographic analysis of the spectrum of the partially purified enzyme phosphorylated with [γ-32P]ATP allowed us to distinguish three major 32P-labeled protein whose molecular weight are about 57, 53, and 48 kDa. In the presence of 5 mM Cl−/25mM 3 HCO − and 100 μM picrotoxin, the intensity of the phosphorylation of bands significantly decreased, thereby confirming the assumption about coupled of binding sites for anions and GABAA-ergic ligands. It was suggested scheme of Cl−-transport through the plasma membrane by utilizing neuronal Cl−/ 3 HCO − -ATPase in the low (5 mM) Cl− and high (25 mM) 3 HCO − concentrations. The data demonstrated for the first time that the GABAA-coupled Cl−/ 3 HCO − -ATPase from rat brain neuronal membranes is maximally activated at a Cl−/ 3 HCO − ratio of 1:5 and it remains stable at high concentrations of substrate and buffer. Corresponding author. S. A. Menzikov et al.