Run-Down of L-Type Ca 2+Channels Occurs on the α 1Subunit

Abstract

Run-down of L-type Ca 2+channels in CHO cells stably expressing α 1c, α 1cβ 1a, or α 1cβ 1aα 2δγ subunits was studied using the patch-clamp technique (single channel recording). The channel activity ( NPo) of α 1cchannels was increased 4- and 8-fold by coexpression with β 1aand β 1aα 2δγ, respectively. When membranes containing channels composed of different subunits were excised into basic internal solution, the channel activity exhibited run-down, the time-course of which was independent of the subunit composition. The run-down was restored by the application of calpastatin (or calpastatin contained in cytoplasmic P-fraction) + H-fraction (a high molecular mass fraction of bovine cardiac cytoplasm) + 3 mM ATP, which has been shown to reverse the run-down in native Ca 2+channels in the guinea-pig heart. The restoration level was 64.7, 63.5, and 66.4% for channels composed of α 1c, α 1cβ 1a, and α 1cβ 1aα 2δγ, respectively, and was thus also independent of the subunit composition. We conclude that run-down of L-type Ca 2+channels occurs via the α 1subunit and that the cytoplasmic factors maintaining Ca 2+channel activity act on the α 1subunit.