Functional inhibition of native volume-sensitive outwardly rectifying anion channels in muscle cells and Xenopus oocytes by anti-ClC-3 antibody

1. Intracellular dialysis of NIH/3T3 cells with a commercially available an ti-ClC-3 polyclonal antibody (Ab) for similar to 30 min completely inhibite d expressed guinea-pig ClC-3 currents (IgpClC-3), while intracellular dialy sis with antigen-preabsorbed anti-ClC-5 Ab failed to affect IgpClC-3. 2. Anti-ClC-3 Ab was used as a selective probe to examine the relationship between endogenous ClC-3 expression and native volume-sensitive outwardly r ectifying anion channels (VSOACs) in guinea-pig cardiac cells, canine pulmo nary arterial smooth muscle cells (PASMCs) and Xenopus laevis oocytes. Intr acellular dialysis or injection of anti-ClC-3 Ab abolished native VSOAC: fu nction in cardiac cells and PASMCs and significantly reduced VSOACs in oocy tes. In contrast, native VSOAC function was unaltered by antigen-preabsorbe d anti-ClC-3 Ab. 3. It is suggested that endogenous ClC-3 represents a major molecular entit y responsible for native VSOACs in cardiac and smooth muscle cells and Xeno pus oocytes. Anti-ClC-3 Ab should be a useful experimental tool to directly test the relationship between endogenous ClC-3 expression and native VSOAC function, and help resolve existing controversies related to the regulatio n and physiological role of native VSOACs in a wide variety of different ce lls.