PLASMA-MEMBRANE CA2-MEDIATED CHANGES IN [CA2+](I) IN DDT1 MF-2 CELLS(PUMPING PLAYS A PROMINENT ROLE IN ADENOSINE A(1) RECEPTOR)

Adenosine A(1) receptor mediated formation of inositol 1,4,5-trisphosp hate (Ins(1,4,5)P-3) and accumulation of cytoplasmic Ca2+ ([Ca2+](i)) were investigated in DDT1 MF-2 smooth muscle cells. A strong reduction of the adenosine and N-6-cyclopentyladenosine (CPA) induced rise in [ Ca2+](i) was observed after blocking Ca2+ entry across the plasma memb rane with LaCl3. This effect of LaCl3 was not observed in the absence of extracellular Ca2+, it was not caused by reduced Ins(1,4,5)P-3 form ation or changed Ins(1,4,5)P-3 induced Ca2+ release, or influenced by temperature. The inhibition of the CPA induced increase in [Ca2+](i) b y LaCl3 was strongly counteracted in the presence of ortho-vanadate, a n inhibitor of plasma membrane Ca2+ ATPase. Ortho-vanadate might also reduce protein tyrosine-phosphate phosphatase activity involved in tyr osine kinase mediated phospholipase C (PLC) activation. However, ortho -vanadate and tyrphostin 25, a tyrosine kinase inhibitor, did not affe ct the CPA induced formation of Ins(1,4,5)P-3. Taken together, these r esults show a strong contribution of Ca2+ pumping across the plasma me mbrane to the regulation of [Ca2+](i) mediated by adenosine A(1) recep tors. Na+/Ca2+ exchange only played a minor role in the initial phase of CPA induced Ca2+ metabolism as measured in low Na+ containing solut ion. The mechanism by which adenosine A(1) receptors activate plasma m embrane Ca2+ ATPase pumps does not include direct stimulation of pumps , but most likely involves an indirect pathway activated by a rapid in crease in [Ca2+](i).