Characterization of acetylcholinesterase in rabbit intrapulmonary arteries.

Acetylcholine (ACh) acts on the pulmonary vasculature to evoke vasodilation and, in some species, vasoconstriction. The actions of ACh are terminated by its rapid hydrolysis by cholinesterases. Aside from histochemical localization studies, there is little information on cholinesterase enzymes in pulmonary blood vessels. The present study addresses the hypothesis that pulmonary blood vessels contain sufficient cholinesterase activity to regulate the action of ACh in these tissues. Accordingly, studies were undertaken to characterize and quantify cholinesterase activities in pulmonary arteries and veins, quantify inhibition of enzyme activity, and investigate functional physiological consequences of cholinesterase inhibition. Cholinesterase activities in aorta and trachea also were examined for comparison. Kinetic studies showed that the lobar pulmonary arterial enzyme has a Michaelis constant of 55.3 +/- 17.0 microM and a maximum velocity of 8.6 +/- 2.7 nmol/min/mg protein similar to cholinesterases found in other peripheral tissues. Studies with selective inhibitors revealed that > 98% of total enzyme activity was attributable to acetylcholinesterase. Similar levels of enzyme activity were found in homogenates of lobar branch intrapulmonary arteries, intrapulmonary veins, and aorta. The majority of enzyme activity was localized to the membrane fraction, although a moderate amount was found in the cytosol. Studies in intact intrapulmonary lobar arteries showed that these vessels had cholinesterase activity comparable with that found in intact trachealis muscle and that neostigmine (10 nM to 10 microM) caused concentration-dependent inhibition of enzyme activity. In isolated intrapulmonary lobar arteries, functional studies showed that 1 and 10 microM neostigmine significantly potentiated ACh-induced contractions.(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacological characterization of muscarinic receptors mediating acetylcholine-induced contraction and relaxation in rabbit intrapulmonary arteries.

Rabbit pulmonary arteries exhibit a biphasic response to acetylcholine consisting of an endothelium-dependent contraction in tissues at resting tone and an endothelium-dependent relaxation in vessels with elevated tone. Each response was studied separately by treating the arteries with inhibitors of nitric oxide synthase to block the relaxant response or with inhibitors of cyclooxygenase to inhibit the contractile response. In the present study, experiments in isolated pulmonary arteries were undertaken to characterize the muscarinic receptor subtypes mediating the two separate responses by utilizing subtype-selective antagonists and determining pA2 values of the antagonists through Schild analysis. Both the relaxant and the contractile responses were inhibited most potently by atropine and by the M3-selective antagonist 4-diphenylacetoxy-N-methylpiperidine methiodide. The pA2 values for inhibition of the contractile and relaxant responses were 9.44 and 8.79 for atropine and 8.92 and 9.29 for 4-diphenylacetoxy-N-methylpiperidine methiodide, respectively. The M1-selective antagonist pirenzepine and the M2-selective antagonist (11-([2-[(diethylamino)methyl]-1-piperidinyl]- acetyl)-5, 11-dihydro-6H-pyrido[2,3-b][1,4]-benzodiazepine-6-one) displayed much lower affinities for the muscarinic receptors mediating these responses. The pA2 values for inhibition of the contractile and relaxant responses were 6.77 and 6.74 for pirenzepine and 6.06 and 5.70 for (11-([2-[(diethylamino)methyl]-1-piperidinyl] acetyl)-5, 11-dihydro-6H-pyrido[2,3-b][1,4]-benzodiazepine-6-one), respectively.