Transient receptor potential canonical 1 (TRPC1) protein is abundantly expressed in cardiomyocytes. While TRPC1 is supposed to be critically involved in cardiac hypertrophy, its physiological role in cardiomyocytes is poorly understood. We investigated the subcellular location of TRPC1 and its contribution to Ca2+ signaling in mammalian ventricular myocytes. Immunolabeling, three-dimensional scanning confocal microscopy and quantitative colocalization analysis revealed an abundant intracellular location of TRPC1 in neonatal rat ventricular myocytes (NRVMs) and adult rabbit ventricular myocytes. TRPC1 was colocalized with intracellular proteins including sarco/endoplasmic reticulum Ca2+ ATPase 2 in the sarcoplasmic reticulum (SR). Colocalization with wheat germ agglutinin, which labels the glycocalyx and thus marks the sarcolemma including the transverse tubular system, was low. Super-resolution and immunoelectron microscopy supported the intracellular location of TRPC1. We investigated Ca2+ signaling in NRVMs after adenoviral TRPC1 overexpression or silencing. In NRVMs bathed in Na+ and Ca2+ free solution, TRPC1 overexpression and silencing was associated with a decreased and increased SR Ca2+ content, respectively. In isolated rabbit cardiomyocytes bathed in Na+ and Ca2+ free solution, we found an increased decay of the cytosolic Ca2+ concentration [Ca2+]i and increased SR Ca2+ content in the presence of the TRPC channel blocker SKF-96365. In a computational model of rabbit ventricular myocytes at physiological pacing rates, Ca2+ leak through SR TRPC channels increased the systolic and diastolic [Ca2+]i with only minor effects on the action potential and SR Ca2+ content. Our studies suggest that TRPC1 channels are localized in the SR, and not present in the sarcolemma of ventricular myocytes. The studies provide evidence for a role of TRPC1 as a contributor to SR Ca2+ leak in cardiomyocytes, which was previously explained by ryanodine receptors only. We propose that the findings will guide us to an understanding of TRPC1 channels as modulators of [Ca2+]i and contractility in cardiomyocytes.

中文翻译：

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心室肌细胞中瞬时受体电位经典通道1的位置和功能。

瞬态受体电位经典1（TRPC1）蛋白在心肌细胞中大量表达。虽然应该认为TRPC1严重参与了心肌肥大，但人们对其在心肌细胞中的生理作用知之甚少。我们调查了TRPC1的亚细胞位置及其在哺乳动物心室肌细胞中对Ca2 +信号传导的贡献。免疫标记，三维扫描共聚焦显微镜和定量共定位分析显示，在新生大鼠心室肌细胞（NRVMs）和成年兔心室肌细胞中TRPC1的大量细胞内位置。TRPC1与肌浆网（SR）中包括肌浆/内质网Ca2 + ATPase 2的细胞内蛋白共定位。与小麦胚芽凝集素共定位，标记糖萼并因此标记包括横管系统在内的肌膜低。超分辨率和免疫电子显微镜支持TRPC1在细胞内的位置。我们研究了腺病毒TRPC1过表达或沉默后NRVM中的Ca2 +信号传导。在浸入无Na +和Ca2 +的溶液中的NRVM中，TRPC1过表达和沉默分别与SR Ca2 +含量的减少和增加有关。在游离于Na +和不含Ca2 +的溶液中的离体兔心肌细胞中，我们发现在存在TRPC通道阻滞剂SKF-96365的情况下，胞质Ca2 +浓度[Ca2 +] i的衰减增加，SR Ca2 +含量增加。在生理起搏速率下兔心室肌细胞的计算模型中，Ca2 +通过SR TRPC通道的泄漏增加了收缩期和舒张期[Ca2 +] i，而对动作电位和SR Ca2 +含量的影响很小。我们的研究表明，TRPC1通道位于SR中，而不存在于心室肌细胞的肌膜中。这项研究提供了证据，证明TRPC1是导致心肌细胞SR Ca2 +泄漏的原因之一，以前仅由ryanodine受体解释过。我们建议，这些发现将指导我们了解作为PC细胞[Ca2 +] i和心肌收缩力调节剂的TRPC1通道。以前仅由ryanodine受体解释过。我们建议，这些发现将指导我们对TRPC1通道作为[Ca2 +] i和心肌细胞收缩性调节剂的理解。以前仅由ryanodine受体解释过。我们建议，这些发现将指导我们了解作为PC细胞[Ca2 +] i和心肌收缩力调节剂的TRPC1通道。