Rat diaphragm cyclic nucleotide-phosphodiesterase: Influence of drugs affecting skeletal muscle contractility

Summary In the present study a phosphodiesterase was partly purified from rat diaphragm and its properties as well as the effects of some drugs known to affect neuromuscular transmission were examined.The enzyme preparation had a pH optimum of 7.0–8.0 As for phosphodiesterase of other organs, the activity was dependent on Mg²⁺ and mainly located in the 100 000×g supernatant. It showed two apparent K _m values (6.4 and 390 M) for the cyclic adenosine-3,5-monophosphate hydrolysis. Various drugs inhibited diaphragm phosphodiesterase non-competitively in the following order of potency: eupaverine papaverine > 1-hexyl-3,7-dimethylxanthine > Ro 7-2956 > theophylline > d-tubocurarine > hydrochlorothiazide. Succinylcholine was ineffective.Of the cyclic nucleotides tested here only cyclic guanosine-3,5-monophosphate elicited an inhibiton at low concentrations (K _i=7M), while cyclic inosine-3,5-monophosphate and cyclic N₆-2-O-dibutyryl-adenosine-3,5-monophosphate inhibited only in high concentrations. Cyclic uridine-3,5-monophosphate did not inhibit phosphodiesterase. The type of inhibition was apparently competitive for cyclic N₆-2-O-dibutyryl-adenosine-3,5-monophosphate and cyclic guanosine-3,5-monophosphate, and non-competitive for cyclic inosine-3,5-monophosphate.The present findings on phosphodiesterase inhibitors agree well with our earlier results on the ability of these drugs (except d-tubocurarine) to increase muscular contractility. It is suggested that their mode of action might be facilitation of the release of acetylcholine from motor nerve endings via the accumulation of cyclic adenosine-3,5-monophosphate.