Objective: To study the intracellular pathways which mediate the inhibitory actions of adenosine on isoprenaline-stimulated calcium current (ICa) in atrioventricular (AV) nodal myocytes. Methods: The whole-cell patch-clamp technique was used to record ICa from rabbit AV nodal cells, isolated by enzymatic and mechanical dispersion. Results: Isoprenaline, 0.1 μM, increased peak ICa from 0.58±0.08 to 1.23±0.1 nA, and this increase was reversibly inhibited by adenosine, 10 μM (83±6%), which we have previously shown to be mediated by nitric oxide (NO) production. A membrane-permeable analogue of cyclic GMP, 8-Br-cGMP (300 μM), an inhibitor of cGMP-stimulated phosphodiesterase, prevented the effect of adenosine on ICa. Methylene blue (10 μM), an inhibitor of NO-sensitive guanylyl cyclase and a generator of superoxide (·O2−), did not prevent, but increased, the inhibiting action of adenosine (49.5±6.6%, P<0.01). Methylene blue (50 μM) caused a reduction of ICa, with further inhibition when combined with adenosine. A ·O2−-generating system, xanthine oxidase (0.02 U/ml) and purine (2.3 mM), also increased the inhibitory action of adenosine on ICa. Inhibition of ICa by adenosine in the presence of xanthine oxidase was not prevented by 8-Br-cGMP (300 μM) and was not influenced by pre-incubation of cells with a NO synthase inhibitor, l-NAME (0.5 mM). Conclusions: The inhibitory effect of adenosine on ICa in rabbit AV nodal myocytes can be mediated by two mechanisms—stimulation of cGMP-stimulated phosphodiesterase by NO-induced cGMP, and a mechanism which involves interaction with ·O2− production.
Keywords: Adenosine; Rabbit, heart; Atrioventricular nodal cells; Nitric oxide; cGMP; Superoxide; Calcium current
Journal Article. 4515 words. Illustrated.
Subjects: Cardiovascular Medicine
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