The biochemical activation of T-type Ca2+ channels in HEK293 cells stably expressing alpha(1G) and Kir2.1 subunits

Abstract

In order to investigate the currently unknown Cellular signaling pathways of T-type Ca2+ channels, we decided to construct a new cell line which would stably express alpha(1G) and Kir2.1 subunits in HEK293 cells (HEK293/alpha(1G)/Kir2.1). Compared to cells which only expressed alpha(1G) (HEK293/alpha(1G)) HEK293/alpha(1G)/Kir2.1 cells produced an enormous inward rectifying current which was blocked by external Ba2+ and Cs+ in a concentration-dependent manner. The expression of Kir-2.1 channels contributed significantly to the shift of membrane potential from -12.2 +/- 2.8 to -57.3 +/- 3.7 mV. However.. biophysical and pharmacological properties of alpha(1G)-mediated Ca2+ channels remained unaffected by the expression of Kir2.1 subunits, except for the enlarging of the window current region. Biochemical activation of alpha(1G) channels using 150 mM KCl brought about an increase in [Ca2+](i), which was blocked by mibefradil, the T-type Ca2+ channel blocker. These data suggest that the HEK293/alpha(1G)/Kir2.1 cell line would have potential uses in the study of T-type Ca2+ channel-mediated signaling pathways and possibly useful in the development of new therapeutic drugs associated with T-type Ca2+ channels. (C) 2004 Elsevier Inc. All rights reserved.