Properties of adenylate cyclase and cyclic nucleotide phosphodiesterase in hamster isolated capillary preparations

Abstract

Capillaries isolated by collagenase digestion of hamster epididymal fat pads were used to examine the properties of endothelial adenylate cyclase and cyclic nucleotide phosphodiesterase. Adenylate cyclase activity in capillary homogenates was increased by 10 μM GTP or 100 μM isoproterenol. Lower concentrations of the catecholamine and 5.7 μM prostagladin E 1 did not stimulate endothelial adenylate cyclase activity unless GTP was included in the assay system. The effects of isoproterenol on capillary adenylate cyclase activity were blocked by propranolol, but were not affected by phentolamine. Phosphodiesterase activity in endothelial homogenates showed anomalous kinetic behavior with either cyclic AMP or cyclic GMP as the enzyme substrate. At substrate concentrations below 1 μM, capillary phosphodiesterase activity hydrolyzed cyclic GMP 2–6 times faster than cyclic AMP. However, at high substrate levels, e.g., 100 μM, cyclic AMP and cyclic GMP were degraded at similar rates. Hydrolysis of 1 μM cyclic AMP by capillary homogenates was stimulated by 0.1 and 1 μM cyclic GMP. Caffeine, 1-methyl-3-isobutylxanthine, papaverine and dipyridamole SQ 20009 were effective inhibitors of capillary phosphodiesterase activity. In contrast, imidazole enhanced the activity of the enzyme. The presence of adenylate cyclase and phosphodiesterase activities in enzyme. The presence of adenylate cyclase and phosphodiesterase activities in hamster isolated capillaries is consistent with a role for cyclic AMP in the regulation of endothelial function. Moreover, the experiments described here indicate that hamster isolated capillaries are useful model systems for studying the metabolism of vascular endothelium.