Laser Diode for Ion Channel Stimulation

该第二阶段SBIR项目的目标是开发和验证激光二极管刺激器，以评估 用于体外和体内研究的热敏瞬时受体电位(Trp)离子通道，使用 膜片钳记录和钙成像及复合动作电位记录。 这种激光器将对新药的发现和评估非常有用，因为目前还没有 具有类似功能的商用工具。 色氨酸家族蛋白的评估可用于描述可能的止痛药的潜在效用 毒品。有选择地(通过温度阈值)、可靠且可重复地激活TRP的能力 也可以在理解拮抗剂作用(结合)和 提供有关不同色氨酸蛋白在细胞和细胞内相互作用机制的更多信息 神经组织。还有一个潜在的用途是能够通过以下方式对疼痛进行不同的评估 TRPV1或TRPV2蛋白在疼痛患者诊断和病情监测中的应用因此，它是 我们的总体目标是建立与激光刺激器一起使用的刺激方案，用于体外和体内 热门离子通道的激活。激光刺激用于选择性激活C和A 人类和大鼠的Delta纤维(在我们的平行项目中)，允许可重复的协议来测试 所谓的止痛药，从细胞培养到人类。 因此，我们假设激光离子通道刺激器将被证明是安全和可靠的，并将 符合FDA对激光设备的要求。为此，我们将开发一种额外的电子产品 允许通过显微镜阻挡激光发射的电路，保护眼睛免受任何 损坏。我们还将提交激光器及其部件进行电磁认证 干扰标准，并进行必要的设备更改以满足要求。 此外，我们还将开发一种直接测量激光诱导的温度的方案 温度和修改设备，以允许可调的加热速度。我们还将为 激光与钙成像的兼容性，并将继续使用TRPV1 DRG测试激光 和HEK293细胞，以研究致敏/脱敏过程。要做到这一点，我们将应用 短时间和长时间的激光脉冲和卡萨西平。我们将测试与TRP相关的概念 激光直接激活神经纤维激活C和A三角纤维及其应用 TRPV1和TRPV2拮抗剂为卡萨西平和钌红。

The goal of this Phase II SBIR project is to develop and validate a laser diode stimulator to assess heat-sensitive Transient Receptor Potential (TRP) ion channels for in vitro and in vivo studies, using patch clamp recording and calcium imaging and compound action potential recording, respectively. This laser will be very useful for new drug discovery and evaluation, as there is currently no commercially available tool of similar capabilities. Evaluation of TRP family proteins can be useful in describing the potential utility of putative analgesic drugs. The ability to selectively (by temperature thresholds), reliably and reproducibly activate TRPs may also provide unique utility in understanding the mechanisms of antagonist action (binding) and provide additional information about mechanisms of interaction of different TRP proteins in cells and nerve tissue. There is also a potential utility of being able to differentially evaluate pain mediated by TRPV1 or TRPV2 proteins in the diagnosis and monitoring of progression of pain patients. Thus, it is our overall goal to establish stimulation protocols for use with a laser stimulator for in vitro and in vivo activation of heat gated ion channels. Laser stimulation is used for selective activation of C and A delta fibers in humans and rats (in our parallel projects), allowing reproducible protocols to test putative analgesic, from cell culture to humans. Thus, we hypothesize that the laser ion channel stimulator will prove safe and reliable action and will meet the FDA requirements for a laser device. To do this, we will develop an additional electronic circuit that allows the blockade of laser emission through the microscope, protecting eyes from any damage. We will also submit the laser and its components for certification of electrical and magnetic interference standards, and make any device changes that are necessary to meet the requirements. In addition, we will develop a protocol of direct temperature measurement of laser-induced temperature and modify the device to allow a tunable rate of heating. We will also insure the compatibility of the laser with calcium imaging and will continue to test the laser with TRPV1 DRG and HEK293 cells in order to study sensitization/desensitization processes. To do this we will apply brief and prolonged laser pulses and capsazepine. We will test the concept of TRP-associated activation of C and A delta fibers by direct laser activation of nerve fibers and application of the TRPV1 and TRPV2 antagonists capsazepine and ruthenium red.