Calcium and osmotic regulation of the NA⁺/H⁺ exchanger in neonatal ventricular myocytes

Intracellular pH regulation in primary cultures of neonatal cardiac myocytes has been characterized. Myocytes were exposed to hyperosmolar solutions to examine the effects on pH regulation by the Na⁺/H⁺ exchanger. Exposure to 100 mM NaCl, sorbitol, N-methyl-D-glucamine, or choline chloride all caused significant increases in steady state pH(i) in myocytes. Omission of extracellular calcium or administration of calmodulin antagonists reduced the osmotic activation of the exchanger. The myosin light-chain inhibitor ML-7 completely blocked osmotic activation of the exchanger suggesting that myosin light-chain kinase is involved in osmotic activation of the exchanger in the myocardium. The calmodulin-dependent protein kinase II inhibitor KN-93 inhibited the rate of recovery from an acute acid load as did trifluoperazine (TFP) and the calmodulin blocker W7, [N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide]. Addition of the calcium ionophore ionomycin caused a large increase in resting pH(i) in isolated myocytes. However, this effect was largely resistant to HMA (5-(N,N-hexamethylene)-amiloride) indicating that an alternative mechanism of pH(i) regulation is responsible. The results demonstrate that the Na⁺/H⁺ exchanger of the neonatal myocardium is responsive to calcium and osmotically responsive pathways and that myosin light-chain kinase is a key protein involved in mediating the osmotic response. (C) 2000 Academic Press.