SPHINGOSINE 1-PHOSPHATE STIMULATES HYDROGEN-PEROXIDE GENERATION THROUGH ACTIVATION OF PHOSPHOLIPASE C-CA2-5 THYROID-CELLS - POSSIBLE INVOLVEMENT OF GUANOSINE TRIPHOSPHATE-BINDING PROTEINS IN THE LIPID SIGNALING( SYSTEM IN FRTL)

Exogenous sphingosine 1-phosphate (S1P) stimulated hydrogen peroxide ( H2O2) generation in association with an increase in intracellular Ca2 concentration in FRTL-5 thyroid cells. S1P also induced inositol phos phate production, reflecting activation of phospholipase C (PLC) in th e cells. These three S1P-induced elements were inhibited partially by pertussis toxin (PTX) and markedly by U73122, a PLC inhibitor, and wer e conversely potentiated by N-6-(L-2-phenylisopropyl)adenosine, an A(1 )-adenosine receptor agonist. In FRTL-5 cell membranes, S1P also activ ated PLC in the presence of guanosine 5'-O-(3-thiotriphosphate) (GTP g amma S), but not in its absence. Guanosine 5'-O-(2-thiodiphosphate) in hibited the S1P-induced GTP gamma S-dependent activation of the enzyme . To characterize the signaling pathways, especially receptors and G p roteins involved in the S1P-induced responses, cross-desensitization e xperiments were performed. Under the conditions where homologous desen sitization occurred in S1P-, lysophosphatidic acid (LPA)-, and bradyki nin-induced induction of Ca2+ mobilization, no detectable cross-desens itization of S1P and bradykinin was observed. This suggests that the p rimary action of S1P in its activation of the PLC-Ca2+ system was not the activation of G proteins common to S1P and bradykinin, but the act ivation of a putative S1P receptor. On the other hand, there was a sig nificant cross-desensitization of S1P and LPA; however, a still signif icant response to S1P (50-80% of the response in the nontreated contro l cells) was observed depending on the lipid dose employed after a pri or LPA challenge. S1P also inhibited cAMP accumulation in a PTX-sensit ive manner. We conclude that S1P stimulates H2O2 generation through a PLC-Ca2+ system and also inhibits adenylyl cyclase in FRTL-5 thyroid c ells. The S1P-induced responses may be mediated partly through a putat ive lipid receptor that is coupled to both PTX-sensitive and insensiti ve G proteins.