N‐Terminal α2 Na+ Pump Constructs are Dominant Negative for Native α2 Na+ Pumps in Smooth, but not Cardiac Muscles, and Increase Salt Sensitivity

Abstract

Plasmids that encode the N‐terminal 90–150 AAs of Na+ pumps with an α2 catalytic subunit are dominant negative (DN) for α2 expression in astrocytes and artery myocytes (Song et al., J Biol Chem 281:12929, 2006). To test the effect of reduced myocyte α2 on mean blood pressure (MBP), we generated α2SM/DN mice that express α2(1‐120)/flag under smooth muscle (SM) myosin heavy chain promoter control. Normal α2 Na+ pumps as well as Na/Ca exchanger‐1 (NCX1) were markedly downregulated in artery and urinary bladder SMs, but not brain. MBPs were recorded by telemetry during 12 hr light (low locomotor activity) periods of 12 hr dark/light cycles. Baseline MBP (0.5% salt diet) did not differ in α2SM/DN mice and wild type (WT) siblings (99 ± 2 vs 97 ± 2 mm Hg; n = 6 each), perhaps because of low NCX1 expression. Nevertheless, MBP increased in α2SM/DN (106 ± 4 vs baseline; P <0.03, ANOVA), but not WT (98 ± 3 vs baseline; NS), mice fed a 6% salt diet + 1% NaCl drinking water. Thus, reduced SM α2 expression increases salt sensitivity. Rat cardiomyocytes, which express α2 Na+ pumps, were transfected (viruses) with mCherry/α2(1‐150)/flag120 or α2(1‐130)/α1‐flag chimera. Cardiomyocytes expressed both constructs (in confocal, mCherry expressed in transverse and longitudinal tubules), but neither downregulated α2. Thus, unlike SMs and astrocytes, the cardiomyocyte α2 N‐terminus may be necessary, but is insufficient to displace normal α2 Na+ pumps.