Abstract
The cardiac slow delayed rectifier potassium channel (IKs), comprised of (KCNQ1) and beta (KCNE1) subunits, is regulated by sympathetic nervous stimulation, with activation of beta-adrenergic receptors PKA phosphorylating IKs channels. We examined the effects of 2-adrenergic receptors (beta2-AR) on IKs in cardiac ventricular myocytes from transgenic mice expressing fusion proteins of IKs subunits and hbeta2-ARs. KCNQ1 and beta2-ARs were localized to the same subcellular regions, sharing intimate localization within nanometers of each other. In IKs/B2-AR myocytes, IKs density was increased, and activation shifted in the hyperpolarizing direction; IKs was not further modulated by exposure to isoproterenol, and KCNQ1 was found to be PKA-phosphorylated. Conversely, beta2-AR overexpression did not affect L-type calcium channel current (ICaL) under basal conditions with ICaL remaining responsive to cAMP. These data indicate intimate association of KCNQ1 and beta2-ARs and that beta2-AR signaling can modulate the function of IKs channels under conditions of increased beta2-AR expression, even in the absence of exogenous beta-AR agonist.
Highlights
IKs, the slowly activating component of the human cardiac delayed rectifier Kϩ current, is a major contributor to repolarization of the cardiac action potential [1]
Localization of KCNQ1 and 2-AR in Murine Ventricular Myocytes—Using double labeling immunohistochemical techniques combined with laser scanning confocal microscopy KCNQ1 and 2-AR were found to be in similar subcellular localizations in ventricular myocytes
In cells isolated from the hKCNQ1-hKCNE1 (TGϩ) mice, KCNQ1 was localized to the intercalated disc (ICD) regions, the surface sarcolemmal membrane (SSM), and the transverse (T-) tubules
Summary
Transgenic mice expressing hKCNQ1-hKCNE1 fusion protein in the heart have been described in detail [9]. These mice express functional slow delayed rectifier potassium channel currents (IKs) normally absent from murine cardiac ventricular myocytes. Strains of TGϩ mice overexpressing 2-AR and hKCNQ1-hKCNE1 were crossed to produce double TGϩ (DTGϩ) mice overexpressing both 2-AR and expressing hKCNQ1-hKCNE1 Both transgene constructs were under the control of the ␣-myosin heavy chain promoter providing cardiomyocyte specific expression. All regions of the cell with fluorescence intensity equal to or greater than a threshold level were identified and marked in each image as a site of the protein as long as at least three contiguous pixels were at or above the threshold. Analysis was carried out using Velocity software by Improvision (Coventry, UK)
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