Guanine nucleotide-sensitive inhibition of L-type Ca2+ current by lysosphingolipids in RINm5F insulinoma cells.

The lysosphingolipids sphingosine-1-phosphate (SPP) and sphingosylphosphorylcholine (SPPC) reportedly increase free cytosolic Ca2+ concentration ([Ca2+]i) in a variety of cell types, apparently by activating G protein-coupled plasma membrane receptors. We investigated whether and how sphingolipids modulate Ca2+ homeostasis in the insulinoma cell line RINm5F. The addition of SPPC and glucopsychosine (GPS) did not affect basal [Ca2+]i but inhibited the KCl (30 mM)-induced increase in [Ca2+]i in a pertussis toxin-insensitive and concentration-dependent manner (EC50 approximately 5 micro M). Similar inhibitory effects were observed with dihydro-SPPC and psychosine, whereas SPP and various N-acylated sphingolipids (at 10 micro M each) had little or no effect on the KCl-induced [Ca2+]i increase. Because in RINm5F cells the primary pathway for depolarization-induced [Ca2+]i increase are L-type Ca2+ channels, we studied whether sphingolipids reduce L-type Ca2+ current (ICa.L). When added to the bath, GPS and SPPC, but not SPP (10 micro M each), rapidly reduced maximal ICa.L by approximately 35%, similar to the alpha2-adrenoceptor agonist clonidine (30 micro M). However, when applied internally, GPS had no effect on ICa. L. When the electrode solution contained the stable GDP analog guanosine-5'-O-(2-thio)diphosphate (1 and 10 mM), the inhibitory effect of GPS was abolished. In conclusion, a novel cellular action of lysosphingolipids is observed in RINm5F cells (i.e., a guanine nucleotide-sensitive inhibition of L-type Ca2+ currents). The pharmacological profile of this inhibition is unique and unlike any known lysosphingolipid receptor-mediated action.