An N-ethylmaleimide-sensitive G-protein modulates Aplysia Ca2+ channels

Abstract

The sulfhydryl alkylating agent N-ethylmaleimide (NEM) was used to probe the possible modulation of calcium current (ICa) by G-proteins in identified neurons of Aplysia californica. ICa recorded with conventional two-electrode voltage clamp was irreversibly suppressed by bath applied NEM in a concentration-dependent manner. This effect was fully blocked by addition of dithiothreitol or intracellular pressure injection of GTP gamma S but was unaffected by pre-treatment with pertussis toxin. These findings suggest that NEM inhibits ICa by causing persistent activation of an inhibitory G-protein in the absence of applied agonist. It appears that alkylation of key cysteine residues involved in G-protein deactivation underlie this effect.