N-type Calcium Channels in Nociceptive Neurons

描述（由申请人提供）：N型钙通道调节脊髓背角浅层初级感觉传入神经释放谷氨酸和P物质。脊髓背角突触前N型钙通道是治疗神经性和慢性疼痛综合征药物的主要靶点。然而，我们还不知道为什么N型钙通道阻滞剂对神经性疼痛如此有效，我们也不知道它们在介导阿片类药物的脊髓镇痛作用中有多重要。在我们的项目的第二阶段，我们将测试我们的假设，一个不同的亚型的N型钙通道是必不可少的脊髓水平的镇痛作用的一个子集的药物，包括阿片类药物在体内。在该基金的第一个资助期间，我们发现这种新的剪接异构体N型钙通道CaV2.2-e37 a对通过G蛋白偶联受体起作用的药物（包括吗啡）显着更敏感。这是重要的，因为i）我们先前表明CaV2.2-e37 a通道在伤害感受器中富集，而CaV2.2-e37 b通道在整个神经系统中发现，和ii）吗啡和其他药物的脊髓水平镇痛作用至少部分地通过抑制背角中的突触前N型通道来介导。我们现在准备测试CaV2.2-e37 a在正常和疾病状态下的体内功能意义。我们已经开发了四种新的小鼠品系，使用单外显子基因靶向。这些小鼠经遗传修饰以仅表达含有CaV2.2-e37 a或CaV2.2-e37 b但不同时含有两者的N型通道。我们已经开发了所有必要的技术专长，以评估这些小鼠在电生理学，生物化学，免疫组织化学和行为水平。我们将询问CaV2.2-e37 a通道是否介导吗啡的基础伤害感受和基础镇痛作用。我们还将询问CaV2.2-e37 a通道是否是神经性疼痛产生痛觉过敏所必需的，以及它们是否是神经性疼痛期间介导吗啡镇痛作用所必需的。我们的研究与寻找有效治疗神经性疼痛和更普遍的慢性疼痛的挑战高度相关。周围神经病变可在急性神经损伤后发展，并且与糖尿病、自身免疫性疾病、营养不良和感染密切相关。对于数百万神经性和慢性疼痛患者没有有效的治疗方法。

DESCRIPTION (provided by applicant): N-type calcium channels regulate release of glutamate and substance P from primary sensory afferents in the superficial dorsal horn of the spinal cord. Presynaptic N-type calcium channels in the spinal dorsal horn are major targets of drugs to treat neuropathic and chronic pain syndromes. However, we don't yet understand why N-type calcium channel blockade is so effective against neuropathic pain and we don't understand how important they are in mediating the spinal analgesic actions of opiates. In this second phase of our project we will test our hypothesis that a distinct isoform of N-type calcium channels is essential for spinal level analgesic actions of a sub-set of drugs including opiates in vivo. During the 1st funding period of this grant we discovered that this novel splice isoform the N-type calcium channel, CaV2.2-e37a, is significantly more sensitive to drugs that act through G protein coupled receptors, including morphine. This is important because i) we previously showed that CaV2.2-e37a channels are enriched in nociceptors whereas CaV2.2-e37b channels are found throughout the nervous system, and ii) spinal level analgesic actions of morphine and other drugs are, at least in part, mediated by inhibition of presynaptic N-type channels in the dorsal horn. We are now ready to test the functional significance of CaV2.2-e37a in vivo in normal and disease states. We have developed four novel mouse lines using single exon gene targeting. These mice are genetically modified to express exclusively N-type channels containing either CaV2.2-e37a or CaV2.2-e37b but not both. We have developed all the necessary technical expertise to assess these mice at electrophysiology, biochemical, immunohistochemical, and behavioral levels. We will ask if CaV2.2-e37a channels mediate basal nociception and basal analgesic effects of morphine. We will also ask if CaV2.2-e37a channels are required for hyperalgesia to develop with neuropathic pain, and if they are necessary to mediate the analgesic effects of morphine during neuropathic pain. Our studies are highly relevant to the challenge of finding effective treatment for neuropathic and more generally chronic pain. Peripheral neuropathies can develop following acute nerve injury, and are strongly associated with diabetes, autoimmune disorders, malnutrition, and infections. There are no effective treatments for the millions of neuropathic and chronic pain sufferers.