Heterogeneity of hypoxic Ca2+ release in pulmonary and *

描述（由申请人提供）： 缺氧可引起肺动脉平滑肌细胞[Ca ~（2+）]I升高及相关的血管收缩，但对体动脉平滑肌细胞无明显影响。 肺动脉平滑肌细胞对缺氧、缺氧Ca ~（2+）和收缩反应的抵抗力明显高于导管平滑肌细胞。 然而，对这些功能差异背后的细胞和分子机制知之甚少。 我们和其他研究者一致表明，Ryanodine受体（RyR）Ca 2+释放是PASMCs缺氧[Ca 2 +]I增加和相关血管收缩的关键。 我们的初步研究，加上以前的研究结果，表明特定的RyR亚型（RyR 1，RyR 2和RyR 3）在血管中的差异表达。 这种异质性的表达可能有助于观察到的兴奋-收缩偶联和缺氧Ca 2+和收缩反应在各种血管中的差异。 此外，内源性调节分子FKBP 12.6（RyR抑制剂）和cADPR（RyR激活剂）与不同RyR亚型的特异性相互作用也可能在生理性和缺氧性Ca 2+释放和血管收缩的异质性中发挥重要作用。 为了解决这些假设，本提案旨在研究以下三个问题（具体目的）：RyR亚型在阻力和管道肺动脉和肠系膜动脉SMC中表达不均匀吗？ 2)RyR Ca 2+释放在阻力和管道肺动脉和肠系膜动脉SMC中是不均匀的吗？ 3)RyR亚型表达的异质性以及RyR与FKBP12.6和cADPR的偶联是否解释了阻力性和管道性肺动脉和肠系膜动脉SMC之间缺氧Ca 2+释放的差异？ 这些目标将通过使用最先进的生物物理学（共聚焦显微镜，膜片钳等），分子和遗传学方法（基因敲除和过表达）来实现。 因此，从这个建议的结果将扩展我们的理解的细胞和分子机制，有助于生理和缺氧的Ca 2+和收缩反应的差异和异质性，并可能确定新的治疗肺动脉高压的目标。 所提出的研究也是基本的生理意义，在其他类型的细胞中的Ca 2+释放的调节。

DESCRIPTION (provided by applicant): Hypoxia induces [Ca2+]I increase and associated vasoconstriction in pulmonary artery smooth muscle cells (PASMCs), but not in systemic artery myocytes. Futhermore, hypoxic Ca2+ and contractile responses are much greater in resistance than conduit PASMCs. However, very little is known about the cellular and molecular mechanisms underlying these functional differences. We and other investigators have consistently demonstrated that ryanodine receptor (RyR) Ca2+ release is critical for hypoxic [Ca2+]I increase and associated vasoconstriction in PASMCs. Our preliminary studies, together with previous findings, suggest that specific RyR subtypes (RyR1, RyR2 and RyR3) are differentially expressed in the vasculature. This heterogeneous nature of expression may contribute to observed differences in excitation-contraction coupling and hypoxic Ca2+ and contractile responses in various blood vessels. In addition, specific interactions of endogenous regulatory molecules FKBP12.6 (RyR inhibitor) and cADPR (RyR activator with distinct RyR subtypes may also play an important role in the heterogeneity of physiological and hypoxic Ca2+ release and vasoconstriction. To address these hypotheses, this proposal seeks to examine the following three questions (Specific Aims): Are RyR subtypes heterogeneously expressed in resistance and conduit pulmonary and mesenteric artery SMCs? 2) Is RyR Ca2+ release heterogeneous in resistance and conduit pulmonary and mesenteric artery SMCs? 3) Do the heterogeneities of RyR subtypes expression and RyR coupling to FKBP12.6 and cADPR explain differences in hypoxic Ca2+ release between resistance and conduit pulmonary and mesenteric artery SMCs? These aims will be pursued by using the state-of-the-art biophysical (confocal microscopy, patch clamp, etc), molecular and genetic approaches (gene knockout and over-expression). Thus, the findings from this proposal will extend our understanding of the cellular and molecular mechanisms that contribute to the differences and heterogeneity of physiological and hypoxic Ca2+ and contractile responses, and may identify novel therapeutic targets for pulmonary hypertension. The proposed studies are also of fundamental physiological significance with respect to the regulation of Ca2+ release in other cell types.