GUANINE NUCLEOTIDE-SENSITIVE INHIBITION OF L-TYPE CA2+ CURRENT BY LYSOSPHINGOLIPIDS IN RINM5F INSULINOMA CELLS

The lysosphingolipids sphingosine-1-phosphate (SPP) and sphingosylphos phorylcholine (SPPC) reportedly increase free cytosolic Ca2+ concentra tion ([Ca2+](i)) in a variety of cell types, apparently by activating G protein-coupled plasma membrane receptors. We investigated whether a nd how sphingolipids modulate Ca2+ homeostasis in the insulinoma cell line RINm5F. The addition of SPPC and glucopsychosine (GPS) did not af fect basal [Ca2+](i) but inhibited the KCl (30 mM)-induced increase in [Ca2+](i) in a pertussis toxin-insensitive and concentration-dependen t manner (EC50 similar to 5 mu M). Similar inhibitory effects were obs erved with dihydro-SPPC and psychosine, whereas SPP and various N-acyl ated sphingolipids (at 10 mu M each) had little or no effect on the KC I-induced [Ca2+](i) increase. Because in RINm5F cells the primary path way for depolarization-induced [Ca2+](i) increase are L-type Ca2+ chan nels, we studied whether sphingolipids reduce L-type Ca2+ current (I-C aL). When added to the bath, GPS and SPPC, but not SPP (10 mu M each), rapidly reduced maximal I-CaL by similar to 35%, similar to the alpha (2)-adrenoceptor agonist clonidine (30 mu M). However, when applied in ternally, GPS had no effect on I-CaL. When the electrode solution cont ained the stable GDP analog guanosine-5'-O-(2-thio)diphosphate (1 and 10 mM), the inhibitory effect of GPS was abolished. In conclusion. a n ovel cellular action of lysosphingolipids is observed in RINm5F cells (i.e., a guanine nucleotide-sensitive inhibition of L-type Ca2+ curren ts). The pharmacological profile of this inhibition is unique and unli ke any known lysosphingolipid receptor-mediated action.