Cardiac Na/K-ATPase and Na/Ca Exchange Function is Altered in Ankyrin B Heterozygous Mice

Abstract

Ankyrin-B (AnkB) is a multivalent “adaptor” protein that targets select membrane proteins to the cytoskeleton. Loss-of-function mutations in AnkB may cause ventricular arrhythmias and sudden cardiac death in humans. Direct interaction with AnkB is required for the membrane targeting and stability of Na/Ca exchanger (NCX) and Na/K-ATPase (NKA), key regulators of cardiac contractility and arrhythmogenesis. However, it is currently unknown how AnkB modulates NCX and NKA function. To investigate this, we used AnkB heterozygous mice (AnkB+/-) and their wild-type (WT) littermates. Cardiac myocytes from AnkB+/- mice show reduced expression (by ∼20%) and altered localization of both NCX and NKA. In agreement with the lower protein level, we found slower decay of the caffeine-induced Ca transient (τ=7.4±0.8 sec vs. 5.2±0.6 sec) and reduced maximum rate of NKA-mediated Na extrusion (5.0±0.5 vs. 6.4±0.4 mM/min) in intact myocytes from AnkB+/- mice vs. WT. Thus, NCX and NKA transport function are reduced in AnkB+/-vs. WT mice. We also measured the voltage-dependence of the currents carried by NCX (INCX) and NKA (Ipump) using whole-cell voltage-clamp. INCX and Ipump were recorded during descending voltage ramps, as Cd-sensitive and K-activated currents, respectively. INCX reflected the lower NCX expression in AnkB+/- myocytes, with no difference in the voltage-dependence vs. WT. In contrast, Ipump had a significantly (p<0.001) steeper voltage-dependence in AnkB+/-vs. WT myocytes. Thus, at −80 mV, close to the resting membrane potential, Ipump was reduced by ∼35% in AnkB+/- mice, whereas at +30 mV, close to the peak of the action potential, AnkB+/- Ipump was elevated by ∼18% vs. WT. Thus, in addition to reducing NKA protein expression, AnkB also directly modulates NKA function in cardiac myocytes, by reducing the voltage-dependent Ipump inactivation. This could significantly affect myocyte [Na]i and [Ca]i regulation.