Chen-Izu Y, Xiao R P, Izu L T, Cheng H, Kuschel M, Spurgeon H, Lakatta E G
Laboratory of Cardiovascular Sciences, Gerontology Research Center, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21224-6823, USA.
Biophys J. 2000 Nov;79(5):2547-56. doi: 10.1016/S0006-3495(00)76495-2.
A plausible determinant of the specificity of receptor signaling is the cellular compartment over which the signal is broadcast. In rat heart, stimulation of beta(1)-adrenergic receptor (beta(1)-AR), coupled to G(s)-protein, or beta(2)-AR, coupled to G(s)- and G(i)-proteins, both increase L-type Ca(2+) current, causing enhanced contractile strength. But only beta(1)-AR stimulation increases the phosphorylation of phospholamban, troponin-I, and C-protein, causing accelerated muscle relaxation and reduced myofilament sensitivity to Ca(2+). beta(2)-AR stimulation does not affect any of these intracellular proteins. We hypothesized that beta(2)-AR signaling might be localized to the cell membrane. Thus we examined the spatial range and characteristics of beta(1)-AR and beta(2)-AR signaling on their common effector, L-type Ca(2+) channels. Using the cell-attached patch-clamp technique, we show that stimulation of beta(1)-AR or beta(2)-AR in the patch membrane, by adding agonist into patch pipette, both activated the channels in the patch. But when the agonist was applied to the membrane outside the patch pipette, only beta(1)-AR stimulation activated the channels. Thus, beta(1)-AR signaling to the channels is diffusive through cytosol, whereas beta(2)-AR signaling is localized to the cell membrane. Furthermore, activation of G(i) is essential to the localization of beta(2)-AR signaling because in pertussis toxin-treated cells, beta(2)-AR signaling becomes diffusive. Our results suggest that the dual coupling of beta(2)-AR to both G(s)- and G(i)-proteins leads to a highly localized beta(2)-AR signaling pathway to modulate sarcolemmal L-type Ca(2+) channels in rat ventricular myocytes.
受体信号特异性的一个合理决定因素是信号传播的细胞区室。在大鼠心脏中，与G(s)蛋白偶联的β(1)-肾上腺素能受体（β(1)-AR）或与G(s)和G(i)蛋白偶联的β(2)-AR受到刺激时，都会增加L型Ca(2+)电流，从而增强收缩强度。但只有β(1)-AR刺激会增加受磷蛋白、肌钙蛋白-I和C蛋白的磷酸化，导致肌肉松弛加速和肌丝对Ca(2+)的敏感性降低。β(2)-AR刺激不会影响这些细胞内蛋白中的任何一种。我们推测β(2)-AR信号可能定位于细胞膜。因此，我们研究了β(1)-AR和β(2)-AR信号在其共同效应器L型Ca(2+)通道上的空间范围和特征。使用细胞贴附式膜片钳技术，我们发现通过向膜片吸管中添加激动剂来刺激膜片中的β(1)-AR或β(2)-AR，都会激活膜片中的通道。但是当激动剂应用于膜片吸管外的膜时，只有β(1)-AR刺激能激活通道。因此，β(1)-AR向通道的信号是通过胞质溶胶扩散的，而β(2)-AR信号定位于细胞膜。此外，G(i)的激活对于β(2)-AR信号的定位至关重要，因为在百日咳毒素处理的细胞中，β(2)-AR信号变得具有扩散性。我们的结果表明，β(2)-AR与G(s)和G(i)蛋白的双重偶联导致了一条高度局限的β(2)-AR信号通路，以调节大鼠心室肌细胞肌膜上的L型Ca(2+)通道。
