MUSCARINIC CHOLINERGIC REGULATION OF CARDIAC MYOCYTE ICA-L IS ABSENT IN MICE WITH TARGETED DISRUPTION OF ENDOTHELIAL NITRIC-OXIDE SYNTHASE

Cardiac myocytes have been shown to express constitutively endothelial nitric oxide synthase (eNOS) (nitric oxide synthase 3), the activatio n of which has been implicated in the regulation of myocyte L-type vol tage-sensitive calcium channel current (ICa-L) and myocyte contractile responsiveness to parasympathetic nervous system signaling, although this implication remains controversial. Therefore, we examined the eff ect of the muscarinic cholinergic agonist carbachol (CCh) on ICa-L and contractile amplitude in isoproterenol (ISO)-prestimulated ventricula r myocytes isolated from adult mice, designated eNOS(null) mice, with targeted disruption of the eNOS gene. Although both eNOS(null) and wil d type (WT) ventricular myocytes exhibited similar increases in ICa-L in response to ISO, there was no measurable suppression of ICa-L by CC h in cells from eNOS(null) mice, in contrast to cells from WT mice. Th ese results were reflected in the absence of an effect of CCh on the p ositive inotropic effect of ISO in eNOS(null) myocytes. Also, unlike m yocytes from WT animals, eNOS(null) myocytes failed to exhibit an incr ease in cGMP content in response to CCh. Nevertheless, the pharmacolog ic nitric oxide donors 3-morpholino-sydnonimine and S-nitroso acetyl-c ystein increased cGMP generation and suppressed ISO-augmented ICa-L in eNOS(null) cells, suggesting that the signal transduction pathway(s) downstream of eNOS remained intact. Of importance, activation of the a cetylcholine-activated K+ channel by CCh was unaffected in atrial and ventricular eNOS(null) myocytes. These results confirm the obligatory role of eNOS in coupling muscarinic receptor activation to cGMP-depend ent control of ICa-L in cardiac myocytes.