The Role of CGMP Dependent Nitric Oxide Signalling on Cardiac Repolarisation in Adult Guinea Pig Ventricular Myocytes

Abstract

Recently, roles for nitric oxide (NO) signalling pathways have been linked with abnormalities in cardiac repolarisation and cardiac arrhythmias. Studies from isolated heart preparations have demonstrated an antifibrillatory effect following vagus nerve stimulation, which is NO dependent. The cellular and molecular basis for this protective effect is unknown. In this study, cGMP dependent regulation of repolarisation was investigated using BAY 60-2770, a novel NO and haem independent soluble guanylyl cyclase (sGC) activator. Action potentials and currents were recorded from isolated guinea pig ventricular myocytes using the perforated patch clamp technique. Cellular cGMP was quantified by radioimmunoassay and expressed as a fold change relative to basal levels. 1 μM BAY 60-2770 increased cellular cGMP by 2.2 fold and by 7.1 fold in the presence of 100 μM IBMX (3-Isobutyl-1-methylxanthine), a non-selective phosphodiesterase (PDE) inhibitor. In vitro assays on purified sGC have shown that ODQ (1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one) potentiates the action of BAY 60-2770. This was corroborated in our experiments, BAY 60-2770 + 10 μM ODQ increased cGMP by 27.9 fold. Despite substantial increases in cellular cGMP, changes in times to 90% repolarisation (APD90) were modest. BAY 60-2770 shortened APD90 by 11.3 ms and adding ODQ to BAY 60-2770 caused no further APD90 shortening. In contrast, when cGMP hydrolysis was blocked by IBMX, BAY 60-2770 lengthened APD90 by 15.7 ms. Preliminary findings indicate that delayed rectifier potassium currents are modulated by these pathways. Overall, these findings demonstrate the complex interplay between cGMP and cAMP mediated effects on the ion channels regulating cardiac repolarisation. The modest effect of BAY 60-2770 + ODQ on APD90, despite a substantial increase in cellular cGMP, highlights that PDEs limit cGMP accumulation close to ion channels in the sarcolemma, thus compartmentalising cGMP signalling.