Ji Guangju, Feldman Morris E, Greene Kai Su, Sorrentino Vincenzo, Xin Hong-Bo, Kotlikoff Michael I
Department of Biomedical Sciences, Cornell University, Ithaca, NY 14853-6401, USA.
J Gen Physiol. 2004 Apr;123(4):377-86. doi: 10.1085/jgp.200308999. Epub 2004 Mar 15.
Calcium release through ryanodine receptors (RYR) activates calcium-dependent membrane conductances and plays an important role in excitation-contraction coupling in smooth muscle. The specific RYR isoforms associated with this release in smooth muscle, and the role of RYR-associated proteins such as FK506 binding proteins (FKBPs), has not been clearly established, however. FKBP12.6 proteins interact with RYR2 Ca(2+) release channels and the absence of these proteins predictably alters the amplitude and kinetics of RYR2 unitary Ca(2+) release events (Ca(2+) sparks). To evaluate the role of specific RYR2 and FBKP12.6 proteins in Ca(2+) release processes in smooth muscle, we compared spontaneous transient outward currents (STOCs), Ca(2+) sparks, Ca(2+)-induced Ca(2+) release, and Ca(2+) waves in smooth muscle cells freshly isolated from wild-type, FKBP12.6(-/-), and RYR3(-/-) mouse bladders. Consistent with a role of FKBP12.6 and RYR2 proteins in spontaneous Ca(2+) sparks, we show that the frequency, amplitude, and kinetics of spontaneous, transient outward currents (STOCs) and spontaneous Ca(2+) sparks are altered in FKBP12.6 deficient myocytes relative to wild-type and RYR3 null cells, which were not significantly different from each other. Ca(2+) -induced Ca(2+) release was similarly augmented in FKBP12.6(-/-), but not in RYR3 null cells relative to wild-type. Finally, Ca(2+) wave speed evoked by CICR was not different in RYR3 cells relative to control, indicating that these proteins are not necessary for normal Ca(2+) wave propagation. The effect of FKBP12.6 deletion on the frequency, amplitude, and kinetics of spontaneous and evoked Ca(2+) sparks in smooth muscle, and the finding of normal Ca(2+) sparks and CICR in RYR3 null mice, indicate that Ca(2+) release through RYR2 molecules contributes to the formation of spontaneous and evoked Ca(2+) sparks, and associated STOCs, in smooth muscle.
通过兰尼碱受体（RYR）释放钙可激活钙依赖性膜电导，并在平滑肌的兴奋-收缩偶联中发挥重要作用。然而，与平滑肌中这种钙释放相关的特定RYR亚型，以及RYR相关蛋白如FK506结合蛋白（FKBP）的作用尚未明确确立。FKBP12.6蛋白与RYR2钙释放通道相互作用，这些蛋白的缺失可预测地改变RYR2单一钙释放事件（钙火花）的幅度和动力学。为了评估特定的RYR2和FBKP12.6蛋白在平滑肌钙释放过程中的作用，我们比较了从野生型、FKBP12.6基因敲除（-/-）和RYR3基因敲除（-/-）小鼠膀胱中新鲜分离的平滑肌细胞中的自发瞬态外向电流（STOCs）、钙火花、钙诱导的钙释放和钙波。与FKBP12.6和RYR2蛋白在自发钙火花中的作用一致，我们发现，相对于野生型和RYR3基因敲除细胞，FKBP12.6缺陷型肌细胞中自发瞬态外向电流（STOCs）和自发钙火花的频率、幅度和动力学发生了改变，而野生型和RYR3基因敲除细胞之间没有显著差异。相对于野生型，FKBP12.6基因敲除（-/-）细胞中钙诱导的钙释放同样增强，但RYR3基因敲除细胞中没有。最后，RYR3细胞中由钙诱导钙释放（CICR）诱发的钙波速度与对照相比没有差异，表明这些蛋白对于正常的钙波传播不是必需的。FKBP12.6基因缺失对平滑肌中自发和诱发钙火花的频率、幅度和动力学的影响，以及在RYR3基因敲除小鼠中发现正常的钙火花和钙诱导钙释放（CICR），表明通过RYR2分子释放钙有助于平滑肌中自发和诱发钙火花以及相关STOCs的形成。
