CHARACTERIZATION OF NOVEL NEUROACTIVE COMPOUNDS FROM CNIDARIA VENOMS

历史证明，毒液是强大的神经活性物质的丰富来源。最近发现的SNX-325是一种B类(N型)钙通道的选择性阻滞剂，为临床医生提供了一种治疗顽固性疼痛的新药物。目前已有的钙通道药物，包括二氢吡啶类，靶向通道在功能和药理上都有别于外周L类通道，目前缺乏特异性的激动剂和拮抗剂，限制了对其治疗调节的探索。例如，针对这些通道的药物将有助于将缺血期间过量钙内流造成的神经元损伤降至最低。申请人实验室的最新研究结果表明，夏威夷水产腔肠动物毒液中最强的一种--翼状毒液--至少含有三种不同的活性。提出的合作研究的总体目标是增强申请者利用包括钙离子和钠离子在内的各种离子的成像技术的离子通量测量来筛选新的神经活性物质的能力。新型钙通道阻滞剂的潜在鉴定可能最终导致阐明实现神经保护的治疗干预措施，这可能应用于脑缺血和其他类型的脑损伤的治疗。有人建议进行研究，以检验这种毒液以及其他阻滞剂的工作假说。为了测试这一点，分离的成分毒素的影响将在各种模型准备上进行测试，包括小龙虾神经弦(电生理测量)和单个神经元细胞(使用荧光成像技术测量离子通量和cAMP)。该项目将在夏威夷大学和约翰·霍普金斯大学的研究人员之间进行综合和协调的合作。申请人的项目部分将侧重于独特成分毒素的生化分离及其对目标组织的电生理和形态影响，而合作部分将侧重于评估离子通量和次级信使系统的影响，特别强调钙和cAMP通量的测量。将在这两个部分中检验的假设与实验设计密切相关，并将使研究人员的互补专业知识以最大限度地产生效益和互惠互利的方式得到应用。合作还将为夏威夷大学的学生和研究员提供充分的机会，以获得在开展这项研究工作中将采用的基本神经科学技术方面的培训。

Venoms have historically proven to be a rich source of powerful neuroactive agents. Recent discovery of SNX-325, a selective blocker of class B (N-type) calcium channels, has provided clinicians with a novel agent for the treatment of intractable pain. Currently available calcium channel drugs, including the dihydropyridines, target channels are functionally and pharmacologically distinct from the peripheral L-type channels and the exploration of their therapeutic modulation is limited by the present lack of specific agonists and antagonists. Drugs targeted toward these channels would be useful, for example, in minimizing neuronal damage caused by excess calcium entry during periods of ischemia. Recent findings from the applicant's laboratory demonstrate that the most powerful of the Hawaiian water coelenterate venoms, Carybdea alata venom, contains at least three distinct activities. The general goal of the collaborative research proposed is to augment the ability of the applicant to screen for novel neuroactive agents using ion flux measurements utilizing imaging techniques for various ions including Ca2+ and Na+. Potential identification of novel calcium channel blockers may ultimately result in elucidation of therapeutic interventions to accomplish neuroprotection which may be applied in the treatment of ischemia and other types of brain injury. Studies are proposed to test the working hypothesis that this venom, as well as other blockers. To test this, the effects of isolated constituent toxins will be tested on various model preparations including crayfish nerve chord (electrophysiological measurement) and single neuronal cells (ion flux and cAMP measurements utilizing fluorescence imaging techniques). The project will be conducted as an integrated and coordinated collaboration between investigators at the University of Hawaii and Johns Hopkins University. The applicant's component of the project will focus on biochemical separation of unique constituent toxins and their respective electrophysiological and morphological effects upon target tissues, while the collaborating component will focus on the evaluation of ion flux and secondary messenger system effects, with particular emphasis on calcium and cAMP flux measurements. The hypotheses to be tested in both components are closely interrelated as are the experimental designs, and will allow the complementary expertise of the investigators to be applied in a maximally productive and mutually beneficial way. The collaboration will also provide ample opportunities for students and fellows at the University of Hawaii to obtain training in the basic neuroscientific techniques to be implemented in the conduct of this research effort.