Osaka T, Joyner R W
Todd Franklin Cardiac Research Laboratory, Children's Heart Center, Department of Pediatrics, Emory University, Atlanta, Ga 30322.
Circ Res. 1992 Jan;70(1):104-15. doi: 10.1161/01.res.70.1.104.
We studied the developmental changes in the beta-adrenergic modulation of L-type calcium current (ICa) in enzymatically isolated adult (AD) and newborn (NB, 1-4-day-old) rabbit ventricular cells using the whole-cell patch-clamp method. ICa was measured as the peak inward current at a test potential of +15 mV by applying a 180-450-msec pulse from a holding potential of -40 mV with Cs(+)-rich pipettes and a K(+)-free bath solution at room temperature. In control, ICa density (obtained by normalizing ICa to the cell capacitance) was significantly higher in AD cells (5.5 +/- 0.2 [mean +/- SEM] pA/pF, n = 65) than in NB cells (2.6 +/- 0.1 pA/pF, n = 60). Isoproterenol (ISO, 1 nM-30 microM) increased ICa in a dose-dependent manner for both groups. The maximal effect (Emax) of ISO, expressed as percent increase in ICa over control levels, and the concentration for one half of the maximal effect (EC50) were 203% and 51 nM, respectively, for AD cells and 111% and 81 nM, respectively, for NB cells. The effect of ISO (1 microM) on ICa was decreased as the test potential was increased from -10 to +40 mV. However, the ratio of the percent increase in ICa for AD versus NB cells was almost constant (2.09-2.45) at each test potential. Dose-response curves of forskolin (FOR, 0.3-50 microM) gave Emax and EC50 of 268% and 0.74 microM, respectively, for AD cells and 380% and 1.15 microM, respectively, for NB cells. After stimulating ICa by 10 microM ISO, the addition of 10 microM FOR produced a further increase in ICa of only 12 +/- 2% in AD cells (n = 4) but a further increase of 140 +/- 41% in NB cells (n = 6). FOR (10 microM) did not produce any increase in ICa for AD and NB cells after stimulating ICa by intracellular application of 200 microM cAMP. ICa density stimulated by 10 microM ISO (17.8 +/- 1.1 pA/pF, n = 7), 10 microM FOR (21.0 +/- 1.3 pA/pF, n = 8), or 200 microM cAMP (18.0 +/- 1.3 pA/pF, n = 5) was equivalent in AD cells, whereas ICa density stimulated by 10 microM ISO (5.8 +/- 0.6 pA/pF, n = 9) was significantly lower than that stimulated by either 10 microM FOR (13.8 +/- 1.5 pA/pF, n = 7) or 200 microM cAMP (13.4 +/- 0.7 pA/pF, n = 7) in NB cells.(ABSTRACT TRUNCATED AT 400 WORDS)
我们采用全细胞膜片钳技术，研究了成年（AD）和新生（NB，1 - 4日龄）兔心室肌细胞中L型钙电流（ICa）的β - 肾上腺素能调节的发育变化。在室温下，使用富含Cs(+)的微电极并置于无K(+)的浴液中，从 - 40 mV的钳制电位施加180 - 450毫秒脉冲至 + 15 mV的测试电位，以此测量ICa，记录为内向电流峰值。在对照组中，AD细胞（5.5 ± 0.2 [均值 ± 标准误] pA/pF，n = 65）的ICa密度（通过将ICa除以细胞电容获得）显著高于NB细胞（2.6 ± 0.1 pA/pF，n = 60）。异丙肾上腺素（ISO，1 nM - 30 μM）使两组细胞的ICa呈剂量依赖性增加。以ICa相对于对照水平增加的百分比表示，ISO对AD细胞的最大效应（Emax）和产生最大效应一半时的浓度（EC50）分别为203%和51 nM，对NB细胞则分别为111%和81 nM。当测试电位从 - 10 mV增加到 + 40 mV时，ISO（1 μM）对ICa的效应减弱。然而，在每个测试电位下，AD细胞与NB细胞的ICa增加百分比之比几乎恒定（2.09 - 2.45）。福斯可林（FOR，0.3 - 50 μM）的剂量 - 反应曲线显示，AD细胞的Emax和EC50分别为268%和0.74 μM，NB细胞分别为380%和1.15 μM。在10 μM ISO刺激ICa后，添加10 μM FOR使AD细胞（n = 4）的ICa仅进一步增加12 ± 2%，而NB细胞（n = 6）则进一步增加140 ± 41%。在通过细胞内施加200 μM cAMP刺激ICa后，FOR（10 μM）对AD和NB细胞的ICa均未产生任何增加。在AD细胞中，10 μM ISO（17.8 ± 1.1 pA/pF，n = 7）、10 μM FOR（21.0 ± 1.3 pA/pF，n = 8）或200 μM cAMP（18.0 ± 1.3 pA/pF，n = 5）刺激后的ICa密度相当，而在NB细胞中，10 μM ISO（5.8 ± 0.6 pA/pF，n = 9）刺激后的ICa密度显著低于10 μM FOR（13.8 ± 1.5 pA/pF，n = 7）或200 μM cAMP（13.4 ± 0.7 pA/pF，n = 7）刺激后的密度。（摘要截短于400字）
