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

• Published in: The Journal of physiology 2001-10, Vol.536 (2), p.361
• Main Authors: M Toselli, V Taglietti, V Parente, S Flati, A Pavan, F Guzzi, M Parenti
• Format: Article
• Language: English
• ISSN: 0022-3751; 1469-7793
• DOI: 10.1111/j.1469-7793.2001.0361c.xd

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. 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. 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. 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.