Calcineurin Increases Cardiac Transient Outward K+ Currents via Transcriptional Up-regulation of Kv4.2 Channel Subunits

Abstract

Fast transient outward potassium currents (I to,f ) are critical determinants of regional heterogeneity of cardiomyocyte repolarization as well as cardiomyocyte contractility. Additionally, I to,f densities are markedly down-regulated in cardiac hypertrophy and heart disease, conditions associated with activation of the serine/threonine phosphatase calcineurin (Cn). In this study, we investigated the regulation of I to,f expression by Cn in cultured neonatal rat ventricular myocytes (NRVMs) with and without ␣ 1 -adrenoreceptor stimulation with phenylephrine (PE). Overexpression of constitutively active Cn in NRVMs induced hypertrophy and caused profound increases in I to,f density as well as Kv4.2 mRNA and protein expression and promoter activity, without affecting Kv4.3 or KChIP2 levels. The effects of Cn on hypertrophy, I to,f , and Kv4.2 transcription were associated with NFAT activation and were abrogated by NFAT inhibition. Despite activating Cn and inducing hypertrophy in NRVMs, PE resulted in profound down-regulation of I to,f densities as well as Kv4.2, Kv4.3, and KChIP2 expression. Although hypertrophy and NFAT activation were inhibited by the Cn inhibitory peptide CAIN, I to,f and Kv4.2 expression were further reduced by CAIN, whereas Cn overexpression eliminated PEinduced reductions in I to,f and Kv4.2 expression without affecting Kv4.3 or KChIP2 levels. We conclude that Cn increases cardiac I to,f densities by positively regulating Kv4.2 gene transcription. Consistent with this conclusion, we found that I to,f was increased in myocytes isolated from young mice overexpressing Cn prior to the development of heart disease. This positive regulation of Kv4.2 transcription by Cn activation is expected to minimize the reductions in I to,f and Kv4.2 expression observed in hypertrophic cardiomyocytes.

Nuclear Factor κB Downregulates the Transient Outward Potassium Current I to,f Through Control of KChIP2 Expression

“…Here, we show that chronic ␣-AR activation (with PE/Pro), which occurs with cardiac hypertrophy and disease, 9,16 decreases I to,f in association with KChIP2 and Kv4.3 subunit mRNA reductions and Kv4.2 elevations. These I to,f reductions, despite increases in Kv4.2 expression, suggest that KChIP2 and/or Kv4.3 subunits are limiting factors for I to,f channel production when ␣-ARs are stimulated, consistent A, Representative I to,f current traces from NRVMs.…”

“…CAIN also decreased (PϽ0.05) Kv4.2 expression both in the presence and absence of PE/Pro, consistent with the conclusion that calcineurin is a positive regulator of Kv4.2 expression. 9 Kv4.3 expression was unchanged by CAIN. Despite these complex effects of CAIN, a 2-way ANOVA analysis showed no significant interaction (PϾ0.05) between PE/Pro and CAIN.…”

“…9 Adenoviral transfections and drug treatments were performed 24 hours after isolation with electrophysiological and mRNA measurements made 40 hours thereafter.…”

“…Previous studies have established that activation of receptors, such as angiotensin II and ␣ 1 -adrenergic (␣ 1 -AR), reduces I to,f and the expression of its molecular constituents. 1 Although I to,f changes induced by chronic ␣ 1 -AR stimulation have been linked to activation of calcineurin and its downstream transcriptional factor, NFAT (nuclear factor of activated T cells), 8 calcineurin-NFAT activation can either increase 9 or decrease 8 I to,f levels, suggesting that this pathway is not the primary regulator of I to,f . Thus, the search continues for the molecular regulators of I to,f .…”

Properties of WT and mutant hERG K+ channels expressed in neonatal mouse cardiomyocytes

“…Compared with adult mouse cardiomyocytes, neonatal mouse cardiomyocytes lack transverse tubules, which may have an advantage in voltage-clamp experiments (27). Neonatal mouse cardiomyocytes have been used as a model for ion channel expression, including for the Na ϩ channel Nav1.5 (39,44), Tand L-type Ca 2ϩ channels (17,21), the K ϩ channel Kv4.2 (18), and the pacemaker HCN channel (9). The study of WT and mutant ion channels in native cardiac myocyte models is important since ion channel mutations have been reported to function differently in native myocyte and HEK-293 cell expression systems (26).…”