Non-Visual Opsins & Vasoregulation: Implications for Vascular Therapy

非视觉视蛋白

基本信息

批准号：
8965151
负责人：
DAN E BERKOWITZ
金额：
$ 40.97万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-07-01 至 2019-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8965151
关键词：
ADRBK1 gene Adenoviruses Adrenergic Agents Arrestins Arteries Beta-Adrenergic Receptor Kinase 1 Blood Blood Circulation Blood Vessels Blood flow Cells Cephalic Circadian Rhythms Complement Coronary Arteriosclerosis Cutaneous Cyclic GMP-Dependent Protein Kinases Data Disease Dominant-Negative Mutation Dose Electrodes Endothelium Erectile dysfunction Exhibits Fingers Functional disorder G protein coupled receptor kinase G-Protein-Coupled Receptors G-substrate GTP-Binding Proteins Grant Health Human Hypertension Immunohistochemistry Injury Invertebrate Photoreceptors Invertebrates Lasers Light Measurement Mediating Membrane Potentials Microdialysis Molecular Mus Myography Nitric Oxide Synthase Opsin Pain Paroxetine Pathway interactions Patients Phosphotransferases Physiological Potassium Channel Process Protein Isoforms Proteins Publications Rattus Raynaud Disease Recording of previous events Regulation Relaxation Retinal Ganglion Cells Reverse Transcriptase Polymerase Chain Reaction Role Scleroderma Signal Pathway Signal Transduction Signal Transduction Pathway Skin Sleep Smooth Muscle Myocytes Soluble Guanylate Cyclase Stimulus Stroke Tail Techniques Testing Tissues Universities Vascular Diseases Vascular Smooth Muscle Vasodilation Vasomotor Vertebrates Visual adrenergic base constriction desensitization in vivo inhibitor/antagonist insight light intensity melanopsin mouse model mutant new therapeutic target novel overexpression phosphodiesterase 6 prevent public health relevance receptor research study response small hairpin RNA small molecule targeted treatment vascular bed vasoconstriction

项目摘要

DESCRIPTION (provided by applicant): Impaired vasomotor function mediated by pertubations in neurohumoral signaling and/or impaired endothelial function is the proximate cause of vascular diseases such as hypertension, coronary artery disease, stroke, erectile dysfunction, and specific vasculopathies such as Raynaud's phenomenon (RP). The identification of novel pathways that regulate vascular tone provides new targets for treatment of vascular disorders. We have recently identified a novel mechanism that modulates vascular tone. Melanospin (Opn4) are classically found in the retinal ganglion cells where they regulate circadian rhythm and sleep. We have identified a non-visual light (opsin) receptor, Opn4, in blood vessels from a number of mammalian species, and from number of vascular beds. These receptors mediated intensity-dependent vasorelaxation to light of a specific wavelength (blue 430-460nM). Preliminary data suggest that signal transduction mechanism(s) involve soluble guanylate cyclase and phosphodiesterase 6 but not protein kinase G. Light activation leads to vascular hyperpolarization a process that involves K+ channels. The receptor is likely regulated as a classic G-protein coupled receptor in that inhibition of G-protein receptor kinase prevents stimulus-dependent desensitization and significantly decrease the light intensity needed to produce a relaxation response. Finally, in vivo, blue light is able evoke a physiologic response; significantly increasing blood flow in the mouse tail artery. We hypothesize that 1) Opn4 is an important regulator of vasodilatory function and mediates a physiologic function in vivo; 2) the signal transduction mechanism in vessels mimics visual opsins of vertebrates or "simple" photoreceptors of invertebrates; 3) desensitization/regulation occurs by a GRK2/arrestin mechanism; and 4) this pathway is upregulated in diseases in which NO signaling (vasodilation) is impaired and vasocontriction is enhanced such as RP. In this proposal, we plan to: 1) Further characterize the potential physiologic role of Opn 4 in vasoregulation utilizing Opn4-/- mice and newly discovered Opn4 inhibitors, opsinamides, using myography in isolated vessels and laser doppler in cranial windows as endpoints in vivo; 2) Determine signal transduction mechanism/s from receptor and G protein to channel with sharp electrode measurement of membrane potential in isolated vessels using specific inhibitors and shRNA knockdown of pathway proteins; 3) Understand receptor regulation using novel pharmacologic inhibitor of GRK2, paroxetine, vascular smooth muscle-selective GRK2-/-, and arrestin-/- mice, as well as GRK2 and arrestin shRNA adenovirus knockdown in isolated vessels. 4) Determine the role of Opn4 receptors as a target in mouse models of RP and explore mechanisms mediating photorelaxation in vivo in the cutaneous circulation of healthy humans and those with primary RP using laser flow doppler and skin microdialysis. In this way we hope to gain insight into the function and dysfunction of this novel pathway in health and disease, and determine its potential as a novel therapeutic target.

描述（由申请人提供）：由神经体液信号传导紊乱介导的血管舒缩功能受损和/或内皮功能受损是血管疾病（例如高血压、冠状动脉疾病、中风、勃起功能障碍）和特定血管病（例如雷诺现象）的直接原因。调节血管张力的新途径的鉴定为治疗血管疾病提供了新的靶标。 Melanospin (Opn4) 通常存在于视网膜神经节细胞中，调节昼夜节律和睡眠。我们在许多哺乳动物的血管中发现了一种非视觉光（视蛋白）受体 Opn4。初步数据表明，这些受体介导特定波长光（蓝色 430-460nM）的强度依赖性血管舒张作用。信号转导机制涉及可溶性鸟苷酸环化酶和磷酸二酯酶 6，但不涉及蛋白激酶 G。光激活导致血管超极化，这是一个涉及 K+ 通道的过程，该受体可能作为经典的 G 蛋白偶联受体进行调节。 G 蛋白受体激酶可防止刺激依赖性脱敏，并显着降低产生松弛反应所需的光强度。最后，在体内，蓝光能够显着增加小鼠的血流量。我们研究发现：1) Opn4 是血管舒张功能的重要调节剂，介导体内的生理功能；2) 血管中的信号转导机制模仿脊椎动物的视觉视蛋白或无脊椎动物的“简单”光感受器；3) 脱敏/调节。通过 GRK2/arrestin 机制发生；4) 在 NO 信号传导（血管舒张）受损的疾病中，该途径上调；在本提案中，我们计划： 1) 利用 Opn4-/- 小鼠和新发现的 Opn4 抑制剂、opsinamides，使用离体血管肌造影和激光多普勒进一步表征 Opn 4 在血管调节中的潜在逻辑生理作用。颅窗作为体内终点；2) 使用特定的电极测量分离血管中的膜电位，确定从受体和 G 蛋白到通道的信号转导机制。抑制剂和通路蛋白的 shRNA 敲除；3) 使用 GRK2、帕罗西汀、血管平滑肌选择性 GRK2-/- 和抑制蛋白 -/- 小鼠的新型药理学抑制剂，以及分离的 GRK2 和抑制蛋白 shRNA 腺病毒敲除来了解受体调节4) 确定 Opn4 受体作为 RP 小鼠模型中靶标的作用，并探索健康人皮肤循环体内介导光松弛的机制。通过这种方式，我们希望深入了解这一新途径在健康和疾病中的功能和功能障碍，并确定其作为新治疗靶点的潜力。