ATP and PIP2 dependence of the magnesium-inhibited, TRPM7-like cation channel in cardiac myocytes

The Mg2+-inhibited cation (MIC) current (I-MIC) in cardiac myocytes biophysically resembles currents of heterologously expressed transient receptor potential (TRP) channels, particularly TRPM6 and TRPM7, known to be important in Mg2+ homeostasis. To understand the regulation of MIC channels in cardiac cells, we used the whole cell voltage-clamp technique to investigate the role of intracellular ATP in pig, rat, and guinea pig isolated ventricular myocytes. IMIC, studied in the presence or absence of extracellular divalent cations, was sustained for >= 50 min after patch rupture in ATP-dialyzed cells, whereas in ATP-depleted cells IMIC exhibited complete rundown. Equimolar substitution of internal ATP by its nonhydrolyzable analog adenosine 5'-(beta, gamma-imido) triphosphate failed to prevent rundown. In ATP-depleted cells, inhibition of lipid phosphatases by fluoride + vanadate + pyrophosphate prevented IMIC rundown. In contrast, under similar conditions neither the inhibition of protein phosphatases 1, 2A, 2B or of protein tyrosine phosphatase nor the activation of protein kinase A (forskolin, 20 mu M) or protein kinase C (phorbol myristate acetate, 100 nM) could prevent rundown. In ATP-loaded cells, depletion of phosphatidylinositol 4,5-bisphosphate (PIP2) by prevention of its resynthesis (10 mu M wortmannin or 15 mu M phenylarsine oxide) induced rundown of IMIC. Finally, loading ATP-depleted cells with exogenous PIP2 (10 mu M) prevented rundown. These results suggest that PIP2, likely generated by ATP-utilizing lipid kinases, is necessary for maintaining cardiac MIC channel activity.