Roles of 3,4-methylenedioxymethamphetamine (MDMA)-induced alteration of connexin43 and intracellular Ca2+ oscillation in its cardiotoxicity

Abstract

Although it is well known that 3,4-methylenedioxymethamphetamine (MDMA) can cause various cardiovascular abnormalities and even sudden death from cardiac arrhythmia, whether it has any effect on myocardial gap junctions, which might be one of the targets mediating MDMA-induced cardiotoxicity, remains unclear. ObjectiveTo test the hypothesis that MDMA may affect the myocardial gap junction protein connexin43 (Cx43) and induce cardiac dysrhythmia. Method(1) In vivo study: adult rats were treated with a single dose MDMA administration (20mg/kg, i.p.). Electrocardiogram detection and immunohistochemical analysis were performed to evaluate cardiac function and expression of Cx43, respectively; (2) in vitro study: cultured ventricular myocytes of neonatal rats were treated with MDMA (10, 100, 1000μmol/L) for 1h. Western blotting and real-time quantitative polymerase chain reaction (RT-qPCR) were performed to investigate the total Cx43 mRNA expression. Immunofluorescent analysis was used to evaluate the amount of junctional Cx43. The phosphorylation status of Cx43 at site Ser368 and intracellular Ca2+ oscillation were also studied. ResultsObvious changes in electrocardiographic patterns were found in rats following MDMA administration. They were characterized by prolonged QRS duration associated with increased amplitude of QRS complex. The heart rates in treated rats were significantly decreased compared to the rats in the control group. The immunohistochemical findings revealed a significant decrease in Cx43 expression. The in vitro study also showed a marked decline in total Cx43 protein associated with reduction of Cx43 mRNA, whereas the phosphorylated Cx43 at Ser368 was increased. Decrease of junctional Cx43 was found correlated with reduction in N-cadherin induced by high concentration of MDMA. Additionally, confocal microscopy findings revealed alteration of intracellular calcium oscillation patterns characterized by high frequency and increasing influx Ca2+. ConclusionsMDMA reduces expression of cardiac gap junction protein Cx43. The increase of phosphorylation status of Cx43 at Ser368 induced by MDMA is attributed, at least in part, to the Ca2+-dependent regulation of protein kinase C (PKC) activity. Our findings provide first evidence of MDMA-mediated changes in those cardiac gap junctions that may underlie MDMA-induced cardiac arrhythmia.