Cerebral artery Ca2+ signaling & subarachnoid hemorrhage

描述（由申请人提供）：动脉瘤破裂和蛛网膜下腔出血（SAH）后的脑血管痉挛是一种破坏性疾病，每年造成数千人残疾和死亡。目前，关于这种血液诱导的脑动脉狭窄的细胞机制知之甚少。该建议建立在我们的工作基础上，表明Ca2+信号和电压依赖性Ca2+通道（VDCC）特性的根本变化导致已建立的兔SAH模型中脑动脉收缩增强。我们已经获得了令人兴奋的初步数据，表明SAH通过以下途径导致脑动脉肌细胞Ca2+进入增强：1)由于r型VDCC （Cav 2.3）的出现而增强的VDCC电流，r型VDCC对传统的（“l型”）钙通道拮抗剂具有抗性；2)氧合血红蛋白，一种与血管痉挛相关的血液成分，可减少肌浆网Ca2+释放事件（Ca2+火花），通过膜电位去极化间接增加VDCC活性。本研究将采用一种综合的方法，通过采用创新和复杂的电生理学、细胞成像和分子生物学技术，阐明SAH对脑血流调节至关重要的小直径脑动脉的影响。特异性目的1将验证中心假设，即来自对照动物的脑动脉肌细胞含有单一类型的VDCCs （l型），而来自SAH动物的脑动脉肌细胞含有两种类型的VDCCs （l型和r型）。我们提出r型VDCCs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。特异性Aim 2将确定Ca2+火花在SAH后脑动脉直径调节中的作用。我们将研究SAH后肌浆网ryanodine受体（RyRs）和vdcs之间的通讯是否中断。SAH后脑动脉肌细胞中r型vdcs的出现为比较l型和r型vdcs与天然平滑肌中RyRs的耦合强度提供了独特的机会。特异性Aim 3将定义将氧合血红蛋白与VDCC表达增强和Ca2+火花抑制联系起来的细胞事件。这项研究将显著增强当前关于健康和疾病中Ca2+信号和脑动脉收缩的知识，并为脑血管痉挛的治疗提供可能的新疗法。

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.