Endothelium-mediated negative dromotropic effects of intravascular acetylcholine

Acetylcholine acting through specific muscarinic membrane receptors causes a negative dromotropic effect and, in blood vessels, causes a vasodilation which results from its action on the endothelial cells via release of nitri c oxide (NO). We decided to study this effect in isolated Krebs-Henseleit r etrogradely perfused guinea pig hearts. A pair of stimulating electrodes wa s placed in the right atrium and to record the auricular-ventricular interv al (A-V delay) one recording electrode was placed on the left atrium and th e other on the tip of the ventricle. Hearts were paced at a rate of 3.8 +/- 0.1 Hz and perfused at a coronary flow rate of 9 +/- 0.25 ml/min. To obtai n dose-response curves, single doses (as boluses) of acetylcholine were inf used and the maximal A-V delay induced by each dose was determined. Perfusi on of agents that inhibit NO accumulation (L-Arginine methyl ester (L-NAME) (0.5 mM)) or oxyhemoglobin (6 mu M) caused displacement of the acetylcholi ne dose-response curve downward and to the right. Perfusion of NO-sparing a gents like superoxide dismutase and dithiothreitol caused an upward and lef tward displacement. Infusion of NO solutions or a NO donor (diethylamine-ni tric oxide [DEA-NO]) caused a dose-dependent negative dromotropic effect. I n contrast, inhibition of the prostaglandin metabolic pathway by Indomethac in (0.01 mM) caused potentiation of acetylcholine effects which were revers ed when it was co-perfused with L-NAME. When endothelial intravascular musc arinic receptors were selectively blocked by perfusion of a non-permeable m acromolecule: dextran (> 2000 kDa) covalently complexed to the receptor blo cker (3-(2'-aminobenzhydryloxy) tropane), the negative dromotropic effect o f intravascular acetylcholine was diminished in a concentration-dependent m anner up to complete blockade. Our data indicate that the dromotropic effec t caused by intracoronary administration of acetylcholine is the result sol ely of activation of intravascular endothelial muscarinic receptors, that n itric oxide and prostaglandins are non-synergistic endothelial mediators of this effect and that there may be an interaction between NO and prostaglan din metabolic pathways. (C) 1998 Elsevier Science B.V. All rights reserved.