Methodologic aspects of acetylcholine-evoked relaxation of rabbit aorta

The acetylcholine-evoked relaxation of rabbit isolated thoracic aorta preco ntracted by phenylephrine was studied. Phenylephrine caused a steady contra ction that was maintained for 6 h, In the presence of calcium disodium ethy lenediaminetetraacetate (EDTA) and ascorbic acid the contraction decreased with time. N-G-Nitro-L-arginine abolished the inhibitory effect of EDTA and ascorbic acid. Acetylcholine evoked a rapid concentration-dependent relaxa tion that recovered spontaneously and slowly, but fully, with time. Relaxat ion evoked by equieffective concentrations of carbachol and acetylcholine h ad the same time course. Cumulative addition of acetylcholine (10(-7) - 3 x 10(-5) M) caused a marked relaxation that was reverted slightly at high co ncentrations. The relaxation was the same with rings derived from the upper , middle, and lower part of the thoracic aorta. Two consecutive concentrati on-response curves for acetylcholine obtained at a 2-h interval demonstrate d a slight development of tachyphylaxis. The relaxation was inversely relat ed to precontractile tension evoked by phenylephrine when expressed as a pe rcentage, but independent when expressed as g tension. Storage of aorta in cold salt solution for 24 h did not alter the relaxation. EDTA and ascorbic acid did not alter the relaxation. It is concluded that (1) EDTA and ascor bic: acid can not be used with impunity to stabilize catecholamines used as preconstriction agents, (2) the reversal of the acetylcholine-evoked relax ation is not due to hydrolysis of acetylcholine; (3) the relaxation is unif orm in all segments of thoracic aorta; (4) cold storage of aorta does not a lter the relaxation; and (5) acetylcholine releases the same amount of rela xing factor, irrespective of the precontractile tension. (C) 2000 Elsevier Science Inc.