Cerebral artery Ca2+ signaling & subarachnoid hemorrhage

描述(申请人提供)：动脉瘤破裂和蛛网膜下腔出血(SAH)后的脑血管痉挛(SAH)是一种破坏性的疾病，每年导致数千人残疾和死亡。目前，关于这种血液引起的大脑动脉狭窄的细胞机制还知之甚少。这一建议建立在我们的工作基础上，表明在已建立的兔SAH模型中，钙信号和电压依赖性钙通道(VDCC)特性的根本变化导致脑动脉收缩增强。我们已经获得了令人兴奋的初步数据，表明SAH通过：1)由于R型VDCC(Cav2.3)的出现而增加VDCC电流，这种电流对传统的(“L”型)钙通道拮抗剂具有抵抗力；以及2)氧合血红蛋白，一种与血管痉挛有关的血液成分，它通过减少肌浆网钙释放事件(Ca~(2+)火花)通过膜电位去极化间接增加VDCC的活性，从而导致大脑动脉肌细胞钙内流增加。这项拟议的工作将采用一种综合的方法，通过创新和复杂的电生理学、细胞成像和分子生物学技术来阐明蛛网膜下腔出血对小直径脑动脉的影响，这些动脉对脑血流调节至关重要。特定目的1将验证中央假设，即对照组动物的大脑动脉肌细胞含有单一类型的VDCC(L型)，而蛛网膜下腔出血动物的脑动脉肌细胞含有两种类型的VDCC(L型和R型)。我们建议R型VDCC作为治疗SAIL的新靶点，它们的表达表明脑血管痉挛的可能性增加。具体目标2将确定钙离子火花在SAH后脑动脉内径调节中所起的作用。我们将检查SAH后肌浆网兰诺定受体(RyRs)和VDCC之间的通讯是否中断。蛛网膜下腔出血后脑动脉肌细胞中R型VDCC的出现为比较L型和R型VDCC与天然平滑肌RyR的偶联强度提供了独特的机会。具体目标3将定义将氧合血红蛋白与VDCC表达增强和钙火花抑制联系起来的细胞事件。这项研究将极大地增强健康和疾病中关于钙信号和脑动脉收缩的现有知识，并为治疗脑血管痉挛提供可能的新疗法。

DESCRIPTION (provided by applicant): Cerebral vasospasm following aneurysm rupture and subarachnoid hemorrhage (SAH) is a devastating disorder that inflicts disability and death upon thousands of individuals each year. Currently, little is known regarding the cellular mechanisms of this blood-induced cerebral artery narrowing. This proposal builds upon our work indicating that fundamental changes in Ca2+ signaling and voltage-dependent Ca2+ channel (VDCC) properties lead to enhanced constriction of cerebral arteries in an established rabbit SAH model. We have obtained exciting preliminary data indicating that SAH leads to enhanced Ca2+ entry in cerebral artery myocytes through: 1) Enhanced VDCC currents due to the emergence of R-type VDCCs (Cav 2.3), which are resistant to conventional ("L-type") calcium channel antagonists; and 2) Oxyhemoglobin, a blood component linked to vasospasm, which decreases sarcoplasmic reticulum Ca2+ release events (Ca2+ sparks) to indirectly increase VDCC activity via membrane potential depolarization. The proposed work will take an integrative approach to elucidate the impact of SAH on small diameter cerebral arteries critical to cerebral blood flow regulation by employing innovative and sophisticated electrophysiological, cell imaging and molecular biology techniques. Specific Aim 1 will test the central hypothesis that cerebral artery myocytes from control animals contain a single type of VDCCs (L-type), while cerebral artery myocytes from SAH animals contain two types of VDCCs (L-type and R-type). We propose R-type VDCCs as novel targets to treat SAIl, and that their expression indicates increased likelihood of cerebral vasospasm. Specific Aim 2 will determine the role Ca2+ sparks play in diameter regulation of cerebral arteries following SAH. We will examine whether the communication between sarcoplasmic reticulum ryanodine receptors (RyRs) and VDCCs is disrupted following SAH. The emergence of R-type VDCCs in cerebral artery myocytes following SAH provides the unique opportunity to compare the coupling strength of L-type and R-type VDCCs to RyRs in native smooth muscle. Specific Aim 3 will define the cellular events linking oxyhemoglobin to both enhanced VDCC expression and inhibition of Ca2+sparks. This study should significantly enhance current knowledge with respect to Ca2+signaling and constriction of cerebral arteries in health and disease and provide insight into possible new therapies for the treatment of cerebral vasospasm.