Modulation of cyclic AMP in purified rat mast cells. I. Responses to pharmacologic, metabolic, and physical stimuli

The cyclic adenosine 3',5' monophosphate (cAMP) content of isolated unstimulated mast cells and the changes induced by a variety of pharmacologic, metabolic, and physical stimuli were studied. A modified bovine serum albumin density gradient purification method consistently provided mast cell preparations which were 95% or more pure, without apparent damage, and a 73% recovery of the mast cells applied to the gradients. The measured cAMP in unstimulated mast cells was high, a mean of 16 picomoles per million cells. Moderate agitation or contact with glass increased cAMP content about 2 fold. When calcium was omitted from the medium mast cell cAMP was markedly elevated; incremental increases in added calcium ion concentration from 1 μM to 1 mM caused a linear decrease in cAMP content. Theophylline (3 to 20 mM) caused a dose related increase in mast cell cAMP content, approximately 2 fold at 20 mM theophylline. Epinephrine (0.01 to 1 mM) caused a modest, dose related increase in cAMP. In the presence of theophylline, epinephrine increased cAMP levels equal to or greater than the sum of the effects of the agents used individually. The increase in cAMP induced by epinephrine was completely inhibited by 100 μM propranolol and partially inhibited by 10 μM propranolol, thus suggesting that a β adrenergic receptor is involved. Prostaglandin E₁ (PGE₁) and histamine (in the presence of theophylline) also raised cAMP. Carbamylcholine (1 nM) lowered cAMP 38%. Atropine (0.1 mM) inhibited the decrease in cAMP induced by 1 nM carbamylcholine by 83% indicating that a muscarinic receptor is involved. In these homogeneous single cell suspensions, therefore, cholinergic and β adrenergic agents have opposing effects on cAMP levels. Diazoxide (10 μM) and adenine (1 μM) caused 37 and 32% decreases in cAMP, respectively. The availability of highly purified mast cells and the identification of agents which either decrease or increase cAMP content allows a direct examination of the role of cAMP in histamine release.