ADENOSINE-A1-RECEPTOR STIMULATION OF INOSITOL PHOSPHOLIPID HYDROLYSIS AND CALCIUM MOBILIZATION IN DDT1 MF-2 CELLS

1. The effect of adenosine receptor-stimulation on inositol phospholipid hydrolysis and calcium mobilization has been investigated in the hamster vas deferens smooth muscle cell line DDT1 MF-2. 2 Adenosine receptor stimulation increased the accumulation of total [H-3]-inositol phosphates in DDT1 MF-2 cells prelabelled with [H-3]-myo-inositol. The rank order of agonist potencies was N6-cyclopentyladenosine > 5'-N-ethylcarboxamidoadenosine > 2-chloroadenosine > adenosine. 3 The response to 2-chloroadenosine was antagonized by the antagonists 8-cyclopentyl-1,3-dipropylxanthine (K(D) 1.2 nM), PD 115,199 (K(D) 39 nM) and 8-phenyltheophylline (K(D) 31 nM). 4 The inositol phosphate response to 2-chloradenosine (10-mu-M) was not significantly altered when the extracellular Ca2+ ion concentration was reduced from 2.4 mM to 1.2 mM or 0.6 mM. Under calcium-free conditions, however, a reduced but still significant response to 2-chloroadenosine was evident (39 +/- 10% of the response in calcium-containing medium). 5 The 5-lipoxygenase inhibitor AA861 (10 and 100-mu-M) inhibited the inositol phosphate response to 2-chloroadenosine by 40 +/- 9% and 60 +/- 4% respectively. The cyclo-oxygenase inhibitor, indomethacin, however, was without significant effect at 1-mu-M. 6 2-Chloroadenosine stimulated an increase in intracellular free Ca2+ ion concentration in fura-2 loaded DDT1 MF-2 cells in calcium-free medium containing 0.1 mM EGTA, which could be inhibited by the adenosine A1-receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (0.1-mu-M). 7 These data suggest that adenosine A1-receptor stimulation results in inositol phospholipid hydrolysis and calcium mobilization from intracellular stores in DDT1 MF-2 cells.