β2-adrenergic receptor overexpression in the developing mouse heart:: evidence for targeted modulation of ion channels

1. We studied the effect of overexpression of the beta(2)-adrenergic receptor (beta(2)-AR) in the heart on ion channel currents in single cells isolated from hearts of fetal and neonatal transgenic and wild-type mice. The beta(2)-AR transgene construct was under the control of the murine alpha-myosin heavy chain (alpha-MHC) promoter, and ion channel activity was measured at distinct developmental stages using whole-cell and perforated patch clamp techniques. 2. We found no change in L-type Ca2+ channel current (I-Ca) density in early embryonic stages (E11-13) of beta(2)-AR transgenic positive (TG+) mice, but significant increases in I-Ca density in intermediate (E14-16, 152%) and late (E17-19, 173.7%) fetal and neonatal (1 day post par turn, 161%) TG+ compared with transgenic negative (TG-) mice. This increase in I-Ca was accompanied by a negative shift in the peak of the current-voltage relationship in TG+ mice. 3. Transient (< 3 min) or prolonged (16-24 h) exposure of TG- neonatal stage myocytes to 8-Br-cAMP (300 mu M) increased I-Ca density and caused a shift in the current-voltage relationship to a similar extent to that seen in TG+ mice. In TG+ myocytes, 8-Br-cAMP had no effect. Exposure of TG+ cells to Rp-cAMPS reversed both the shift in voltage dependence and reduced the peak current density observed in these myocytes. We concluded from these results that the L-type Ca2+ channel is maximally modulated by cAMP-dependent protein kinase (PKA) in TG+ mice and that the alpha-MHC promoter is functional in the ventricle as early as embryonic day 14. 4. In contrast, we found that slow delayed rectifier K+ channels were not changed significantly at any of the developmental stages studied by the overexpression of beta(2)-ARs compared with TG- mice. The sensitivity of murine slow delayed rectifier K+ channels to cAMP was tested by both transient and prolonged exposure to 8-Br-cARMP (300 mu M), which increased the slow delayed rectifier K+ channel current (I-K,I-s) density to a similar extent in both TG- and TG+ neonatal myocytes. In addition, we found that there was no difference in the concentration dependence of the response of I-Ca and I-K,I-s to 8-Br-cAMP. 5. Thus, overexpression of the beta(2)-AR in the heart results in distinct modulation of I-Ca, but not I-K,I-s, and this is not due to differences in the 8-Br-cAMP sensitivity of the two channels. Instead, these results are consistent with both compartmentalization of beta(2)-AR-controlled cAMP and distinct localization of L-type Ca2+ and slow delayed rectifier R+ channels. This cAMP is targeted preferentially to the L-type Ca2+ channel and is not accessible to the slow delayed rectifier K+ channel.