Adenosine-Mediated Stimulation of Bone Cell Adenylate Cyclase Activity

Abstract

Adenosine rapidly stimulated adenylate cyclase activity but did not modify cyclic AMP degradation when added to a particulate fraction prepared from isolated bone cells. The effect of adenosine was one-half maximal at 5-10 micronM, and was not mimicked by 5' AMP, inosine, guanosine, uridine, adenine, or ribose. Basal and adenosine-stimulated adenylate cyclase activites were directly proportional to the concentration of particulate protein in the assay system. Theophylline decreased the degree to which adenosine stimulated adenylate cyclase activity, whereas another phosphodiesterase inhibitor, RO-20-1724, failed to iiinfluence the effect of adenosine. Adenosine itself, and not a metabolite of adenosine is the stimulator of adenylate cyclase, since it was neither phosphorylated nor deaminated appreciably by the particulate fraction. The particulate fraction did not convert substrate ATP to adenosine in amounts sufficient to enhance adenylate cyclase. The stimulatory effect of adenosine was maximal at 1.2 mM Mg2+, declined with increases in the Mg2+ concentration, and was replaced by inhibition at 20 mM Mg2+. At 2.4 mM Mg2+, basal adenylate cyclase activity peaked at 1.1 mM ATP, and was inhibited by higher ATP concentrations. The magnitude of adenosine stimulation was greater at inhibitory concentrations of ATP than at concentrations which yielded maximum activity. The results suggest that the previously demonstrated ability of adenosine to increase cyclic 3'5' AMP levels in intact bone cells stems from its effect on adenylate cyclase. Adenosine may act by modifying the regulatory nfluence of free Mg2+, uncomplexed ATP, (or both), on adenylate cyclase. Theophylline appears to interfere with the action of adenosine by a mechanism which is distinct from its capacity to inhibit cyclic AMP phosphodiesterase activity. (Endocrinology 99:901,1976)

Adenosine and the regulation of insulin secretion by isolated rat islets of Langerhans

“…These findings are consistent with the observations that adenosine, but not adenine, AMP or ADP, also affects the cyclic AMP content and adenylate cyclase activity of cerebral cortex (Sattin & Rall, 1970), adipocytes (Fain et al, 1972;Schwabe et al, 1973) and bone cells (Peck et al, 1976). Studies on the mode of action of adenosine in regulating adenylate cyclase activity have suggested that adenosine does not interact with a-and ,B-adrenergic receptors linked to adenylate cyclase in a variety of tissues (Clark et al, 1974;Zenser, 1975).…”

Purinergic Receptors

“…Low concentrations of adenosine stimulated platelet adenylate cyclase activity, but increasing nucleoside concentrations resulted in the appearance of an inhibitory adenosine effect. Similar biphasic curves have been observed with a variety of adenylate cyclase systems (Oark and Seney, 1976;Cooper and Londos, 1979;lakobs et al, 1979;Peck et al, 1976; .1 Effects of adenosine on hepatic adenylate cyclase. Adenylate cyclase activity was determined with dA TP as substrate, as described previously .…”

“…To circumvent these problems, two methods have been developed, as described below. It must also be pointed out that the use of methyl xanthines (theophylline, I-methyl-3-isobutylxanthine, and caffeine) as cyclic nucleotide phosphodiesterase inhibitors is precluded because these agents are antagonists of the adenosine receptor (Fain and Malbon, 1979;Marquardt et al, 1978;Dark et al, 1974;Green and Stanberry, 1977;Marone et al, 1978;Blume et al, 1973;Dark and Seney, 1976;Cooper and Londos, 1979;Fain, 1973;Fain and Wieser, 1975;Fredholm, 1977;Iizuka et al, 1976;Jakobs et al, 1979;Londos et al, 1979b;Maguire et al, 1975;Pecketal., 1974;Peck et al, 1976;Penitetal., 1976;Premont etal., 1977;Schwabe and Ebert, 1974;van Calker et al, 1978;Wilkening and Makman, 1975;Zenser, 1975;Wolberg et al, 1978;Premont et al, 1979b). However, other phosphodiesterase inhibitors, such as papaverine and d, 1-4-(3-butoxy-4-methoxy-benzyl)-2-imidazolidinone (RO 20-1724), do not interfere with the adenosine receptor (Fain and Malbon, 1979;Clark et al, 1974;Green and Stanberry, 1977;Marone et al, 1978;Blume et al, 1973;mume and Foster, 1975;Fain, 1973;Fredholm, 1977;lizuka et al, 1976;lakobs et ai., 1979;Peck et al, 1976;van Calker et al, 1978;…”

Two distinct adenosine-sensitive sites on adenylate cyclase.

“…Subsequently, isolated neuronal and other cells have been shown to respond similarly to adenosine (5)(6)(7)(8)(9)(10)(11). In a number of tissues it has been possible to demonstrate direct stimulation of adenylate cyclase by adenosine in isolated membrane preparations (12)(13)(14)(15)(16)(17).…”

“…Several adenylate cyclase systems exhibit biphasic responses to adenosine, with activation occurring at low concentrations and inhibition at higher concentrations of the nucleoside (12,14,15,(27)(28)(29). It seemed reasonable to ask whether the biphasic effects of these enzymes are mediated through different sites and whether the specificity of the sites mediating the opposite effects would conform to the specificities of the apparently different sites in the liver and Leydig cell adenylate cyclase (12) which is both stimulated and inhibited by adenosine and 2-chloroadenosine.…”