期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2021
卷号：118
期号：7
页码：1
DOI：10.1073/pnas.2017937118
出版社：The National Academy of Sciences of the United States of America
摘要：The number and activity of Ca v 1.2 channels in the cardiomyocyte sarcolemma tunes the magnitude of Ca 2 -induced Ca 2 release and myocardial contraction. β-Adrenergic receptor ( βAR ) activation stimulates sarcolemmal insertion of Ca V 1.2. This supplements the preexisting sarcolemmal Ca V 1.2 population, forming large “superclusters” wherein neighboring channels undergo enhanced cooperative-gating behavior, amplifying Ca 2 influx and myocardial contractility. Here, we determine this stimulated insertion is fueled by an internal reserve of early and recycling endosome-localized, presynthesized Ca V 1.2 channels. βAR -activation decreased Ca V 1.2/endosome colocalization in ventricular myocytes, as it triggered “emptying” of endosomal Ca V 1.2 cargo into the t-tubule sarcolemma. We examined the rapid dynamics of this stimulated insertion process with live-myocyte imaging of channel trafficking, and discovered that Ca V 1.2 are often inserted into the sarcolemma as preformed, multichannel clusters. Similarly, entire clusters were removed from the sarcolemma during endocytosis, while in other cases, a more incremental process suggested removal of individual channels. The amplitude of the stimulated insertion response was doubled by coexpression of constitutively active Rab4a, halved by coexpression of dominant-negative Rab11a, and abolished by coexpression of dominant-negative mutant Rab4a. In ventricular myocytes, βAR -stimulated recycling of Ca V 1.2 was diminished by both nocodazole and latrunculin-A, suggesting an essential role of the cytoskeleton in this process. Functionally, cytoskeletal disruptors prevented βAR -activated Ca 2 current augmentation. Moreover, βAR -regulation of Ca V 1.2 was abolished when recycling was halted by coapplication of nocodazole and latrunculin-A. These findings reveal that βAR -stimulation triggers an on-demand boost in sarcolemmal Ca V 1.2 abundance via targeted Rab4a- and Rab11a-dependent insertion of channels that is essential for βAR -regulation of cardiac Ca V 1.2.
