CHARACTERIZATION OF NOVEL NEUROACTIVE COMPOUNDS FROM CNIDARIA VENOMS

历史证明，毒液是强大的神经活性剂的丰富来源。SNX-325是一种选择性B类（N型）钙通道阻滞剂，为临床治疗顽固性疼痛提供了新的药物。目前可用的钙通道药物，包括二氢吡啶类，靶通道在功能上和结构上与外周L型通道不同，并且由于目前缺乏特异性激动剂和拮抗剂，限制了对它们的治疗调节的探索。靶向这些通道的药物将是有用的，例如，在最大限度地减少缺血期间过量钙进入引起的神经元损伤。申请人实验室的最新发现表明，夏威夷水腔肠动物毒液中最强的一种，Carybdea阿拉塔毒液，含有至少三种不同的活性。提出的合作研究的总体目标是增强申请人使用离子通量测量筛选新型神经活性剂的能力，所述离子通量测量利用各种离子（包括Ca 2+和Na+）的成像技术。新型钙通道阻滞剂的潜在鉴定可能最终导致阐明治疗干预以实现神经保护，其可应用于治疗缺血和其他类型的脑损伤。研究提出测试工作假设，这种毒液，以及其他阻滞剂。为了测试这一点，将在各种模型制备物上测试分离的组分毒素的作用，包括小龙虾神经索（电生理学测量）和单个神经元细胞（利用荧光成像技术的离子通量和cAMP测量）。该项目将作为夏威夷大学和约翰霍普金斯大学的调查人员之间的综合和协调合作进行。申请人的项目组成部分将侧重于独特毒素成分的生化分离及其对靶组织的电生理和形态学影响，而合作组成部分将侧重于离子通量和第二信使系统影响的评价，特别强调钙和cAMP通量测量。在这两个组成部分进行测试的假设是密切相关的，因为是实验设计，并将允许调查人员的互补专业知识，以最大限度地提高生产力和互惠互利的方式应用。这项合作还将为夏威夷大学的学生和研究员提供充分的机会，使他们能够接受在进行这项研究工作中将要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.