Increased intracellular magnesium attenuates β-adrenergic stimulation of the cardiac Cav1.2 channel

Abstract

Increases in intracellular Mg2+ (Mg2+i), as observed in transient cardiac ischemia, decrease L-type Ca2+ current of mammalian ventricular myocytes (VMs). However, cardiac ischemia is associated with an increase in sympathetic tone, which could stimulate L-type Ca2+ current. Therefore, the effect of Mg2+i on L-type Ca2+ current in the context of increased sympathetic tone was unclear. We tested the impact of increased Mg2+i on the β-adrenergic stimulation of L-type Ca2+ current. Exposure of acutely dissociated adult VMs to higher Mg2+i concentrations decreased isoproterenol stimulation of the L-type Ca2+ current from 75 ± 13% with 0.8 mM Mg2+i to 20 ± 8% with 2.4 mM 2+i. We activated this signaling cascade at different steps to determine the site or sites of 2+i action. Exposure of VMs to increased Mg2+i attenuated the stimulation of L-type Ca2+ current induced by activation of adenylyl cyclase with forskolin, inhibition of cyclic nucleotide phosphodiesterases with isobutylmethylxanthine, and inhibition of phosphoprotein phosphatases I and IIA with calyculin A. These experiments ruled out significant effects of Mg2+i on these upstream steps in the signaling cascade and suggested that 2+i acts directly on Cav1.2 channels. One possible site of action is the EF-hand in the proximal C-terminal domain, just downstream in the signaling cascade from the site of regulation of Cav1.2 channels by protein phosphorylation on the C terminus. Consistent with this hypothesis, Mg2+i had no effect on enhancement of Cav1.2 channel activity by the dihydropyridine agonist (S)-BayK8644, which activates Cav1.2 channels by binding to a site formed by the transmembrane domains of the channel. Collectively, our results suggest that, in transient ischemia, increased Mg2+i reduces stimulation of L-type Ca2+i current by theβ-adrenergic receptor by directly acting on Cav1.2 channels in a cell-autonomous manner, effectively decreasing the metabolic stress imposed on VMs until blood flow can be reestablished. © 2013 Brunet et al.