Synthesis, Storage and Drug-Induced Release of ATP-8- 3 H in the Perfused Bovine Adrenal Gland

The perfused bovine adrenal gland was able to take up and phosphorylate adenosine-8-3 H. Stimulation of the gland with carbachol resulted in a simultanious release of catecholamines, adenosine-8-3 H and labelled adenine nucleotides including a small quantity of ATP-3 H. A direct demonstration of the presence of ATP-3 H in isolated chromaffin granules was obrained by modified sucrose density gradient centrifugation. Glands were also perfused with a Ca++-Mg++-free-EDTA medium to inhibit adenosine triphosphatases. Using this medium, significant amounts of ATP-3 H and endogenous ATP were found in the effluent after drug stimulation by either Ba++ of aminophylline. Under Ca++-Mg++-free-EDTA conditions both Ba++ and aminophylline, but not carbachol, stimulated the release of catecholamines, ATP, ATP-3 H with no concomitant increase in total radioactivity. Using normal medium, Ba++ caused the simultaneous release of catecholamines and radioactivity while aminophylline only induced the release of catecholamines. Specific activity measurements of drug-released ATP indicated that the newly synthesized ATP-3 H was being released preferentially during spontaneous activity while the endogenous ATP was preferentially released after either Ba++ or aminophylline-induced secretion.