Attenuation of G protein‐mediated inhibition of N‐type calcium currents by expression of caveolins in mammalian NG108–15 cells

1 Caveolins are integral proteins of glycolipid/cholesterol‐rich plasmalemmal caveolae domains, where, they may function as a plasma membrane scaffold onto which many classes of signalling molecules, including receptors and heterotrimeric G proteins, can assemble. To ascertain whether caveolins influence G protein‐mediated signal transduction, we stably expressed caveolin‐1 and −3 isoforms in the neuroblastoma × glioma NG108–15 hybrid cell line, lacking endogenous caveolins. Subsequently, using whole‐cell voltage clamp methods, we examined whether the modulation of N‐type voltage‐gated Ca 2+ channels by G o protein‐coupled, δ‐type opioid receptors might be affected by recombinant caveolin expression. 2 In transfected NG108–15 cells, caveolins localized at the plasma membrane and, upon subcellular fractionation on sucrose density gradients, they co‐localized in Triton‐resistant, low buoyancy fractions, with endogenous G o protein α‐subunits. 3 The voltage‐dependent inhibition of ω‐conotoxin GVIA‐sensitive Ba 2+ currents following either activation of δ‐opioid receptors by the agonist [o‐pen 2 ,o‐pen 5 ]‐enkephalin (DPDPE), or direct stimulation of G proteins with guanosine 5′‐ O ‐(thiotriphosphate) (GTPγS) was significantly attenuated in caveolin‐expressing cells. The kinetics of Ca 2+ channel inhibition were also modified by caveolins. 4 Overall, these results suggest that caveolins may negatively affect G protein‐dependent regulation of voltage‐gated N‐type Ca 2+ channels, presumably by causing a reduction of the available pool of activated G proteins.