The HVCN1 voltage-gated proton channel contributes to pH regulation in canine ventricular myocytes

Abstract

Regulation of intracellular pH (pH_i ) in cardiomyocytes is crucial for cardiac function; however, currently known mechanisms for direct or indirect extrusion of acid from cardiomyocytes seem insufficient for energetically efficient extrusion of the massive H⁺ loads generated under in vivo conditions. In cardiomyocytes, voltage-sensitive H⁺ channel activity mediated by the HVCN1 proton channel would be a highly efficient means of disposing of H⁺ , while avoiding Na⁺ loading, as occurs during direct acid extrusion via Na⁺ /H⁺ exchange or indirect acid extrusion via Na⁺ -HCO₃^- cotransport. PCR and immunoblotting demonstrated expression of HVCN1 mRNA and protein in canine heart. Patch clamp analysis of canine ventricular myocytes revealed a voltage-gated H⁺ current that was highly H⁺ -selective. The current was blocked by external Zn²⁺ and the HVCN1 blocker 5-chloro-2-guanidinobenzimidazole. Both the gating and Zn²⁺ blockade of the current were strongly influenced by the pH gradient across the membrane. All characteristics of the observed current were consistent with the known hallmarks of HVCN1-mediated H⁺ current. Inhibition of HVCN1 and the NHE1 Na⁺ /H⁺ exchanger, singly and in combination, showed that either mechanism is largely sufficient to maintain pH_i in beating cardiomyocytes, but that inhibition of both activities causes rapid acidification. These results show that HVCN1 is expressed in canine ventricular myocytes and provides a major H⁺ extrusion activity, with a capacity similar to that of NHE1. In the beating heart in vivo, this activity would allow Na⁺ -independent extrusion of H⁺ during each action potential and, when functionally coupled with anion transport mechanisms, could facilitate transport-mediated CO₂ disposal. KEY POINTS: Intracellular pH (pH_i ) regulation is crucial for cardiac function, as acidification depresses contractility and causes arrhythmias. H⁺ ions are generated in cardiomyocytes from metabolic processes and particularly from CO₂ hydration, which has been shown to facilitate CO₂ venting from mitochondria. Currently, the NHE1 Na⁺ /H⁺ exchanger is viewed as the dominant H⁺ extrusion mechanism in cardiac muscle. We show that the HVCN1 voltage-gated proton channel is present and functional in canine ventricular myocytes, and that HVCN1 and NHE1 both contribute to pH_i regulation. HVCN1 provides an energetically efficient mechanism of H⁺ extrusion that would not cause Na⁺ loading, which can cause pathology, and that could contribute to transport-mediated CO₂ disposal. These results provide a major advance in our understanding of pH_i regulation in cardiac muscle.

Keywords: HVCN1; Hv1; SLC9A1; cardiac myocytes; intracellular pH; proton channel.