β2-Adrenergic receptor overexpression in the developing mouse heart: evidence for targeted modulation of ion channels
Corresponding author R. S. Kass: Department of Pharmacology, College of Physicians and Surgeons of Columbia University, New York, NY 10032, USA. Email: email@example.com
- 1We studied the effect of overexpression of the β2-adrenergic receptor (β2-AR) in the heart on ion channel currents in single cells isolated from hearts of fetal and neonatal transgenic and wild-type mice. The β2-AR transgene construct was under the control of the murine α-myosin heavy chain (α-MHC) promoter, and ion channel activity was measured at distinct developmental stages using whole-cell and perforated patch clamp techniques.
- 2We found no change in L-type Ca2+ channel current (ICa) density in early embryonic stages (E11-13) of β2-AR transgenic positive (TG+) mice, but significant increases in ICa density in intermediate (E14-16, 152 %) and late (E17-19, 173.7 %) fetal and neonatal (1 day post partum, 161 %) TG+ compared with transgenic negative (TG-) mice. This increase in ICa was accompanied by a negative shift in the peak of the current-voltage relationship in TG+ mice.
- 3Transient (< 3 min) or prolonged (16-24 h) exposure of TG- neonatal stage myocytes to 8-Br-cAMP (300 μM) increased ICa 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 α-MHC promoter is functional in the ventricle as early as embryonic day 14.
- 4In contrast, we found that slow delayed rectifier K+ channels were not changed significantly at any of the developmental stages studied by the overexpression of β2-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-cAMP (300 μM), which increased the slow delayed rectifier K+ channel current (IK,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 ICa and IK,s to 8-Br-cAMP.
- 5Thus, overexpression of the β2-AR in the heart results in distinct modulation of ICa, but not IK,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 β2-AR-controlled cAMP and distinct localization of L-type Ca2+ and slow delayed rectifier K+ channels. This cAMP is targeted preferentially to the L-type Ca2+ channel and is not accessible to the slow delayed rectifier K+ channel.