We studied the effects of β₂-adrenergic receptors (β₂-AR) on the slow delayed rectifier current (I_Ks) in single ventricular myocytes isolated from hearts of transgenic (TG+) mice expressing an hKCNQ1-hKCNE1 fusion protein and overexpressing the hβ₂-AR. Both transgene constructs were under the control of the α-myosin heavy chain promoter (α-MHC). Strains of TG+ mice overexpressing β₂-AR (βTG+) and hKCNQ1-hKCNE1 (KTG+) were crossed to produce double TG+ (DTG+) mice overexpressing both β₂-AR and expressing hKCNQ1-hKCNE1.

Animals were humanely killed by intraperitoneal injection of pentobarbital sodium (50 mg kg^-1). KCNQ1 and β₂-AR were found to be in similar subcellular localisations of isolated ventricular myocytes by immunohistochemical techniques combined with laser scanning confocal microscopy. KCNQ1 was localised to the surface sarcolemmal membrane, the transverse tubules and intercalated disc regions (ICD). In contrast, β₂-AR were located in the T-tubules and ICD, but were absent from the surface sarcolemmal membrane. I_Ks density was increased (peak tail current after 60 mV pulses: KTG+ 4.0 ± 0.7 pA pF^-1 (mean ± S.E.M.), n = 8; DTG+ 9.2 ± 1.7 pA pF^-1, n = 6, P < 0.05, Student’s unpaired t test), and activation was shifted in the hyperpolarizing direction (24 mV) in myocytes from DTG+ mice compared to TG+ mice expressing the KCNQ1-KCNE1 transgene alone (KTG+), indicating overexpression of β₂-AR results in β-AR dependent stimulation of I_Ks, even in the absence of exogenous β-AR agonist. In fact, we found I_Ks current density in myocytes from DTG+ mice to have similar characteristics to that in myocytes from KTG+ mice plus β-AR stimulation by isoproterenol (ISO, 1 µM) (peak tail current after 60 mV pulses: KTG+ control 5.0 ± 1.7 pA pF^-1; KTG+ ISO 7.3 ± 2.1 pA pF^-1, n = 5, P < 0.05, Student’s paired t test, or 67 + 7 % increase). We found no further increase in I_Ks density or change in activation voltage dependence with β-AR stimulation in DTG+ mice indicating β₂-AR overexpression results in maximal activation of I_Ks. Back phosphorylation experiments revealed hyperphosphorylation of KCNQ1 in DTG+ mice overexpressing β₂-AR. Hyperphosphorylation of KCNQ1 was also detected in lysates from failing human hearts. Thus, overexpression of β₂-AR in the heart results in hyperphosphorylation of KCNQ1 and maximal activation of I_Ks. Hyperphosphorylation of KCNQ1 is also observed in myocytes from failing human hearts. As such, myocytes overexpressing β₂-AR may provide a good system to examine the effects on ion channel function of chronic activation of β-AR as seen in human heart failure.