NEUROBIOLOGY OF ENDOMORPHINS IN SPINAL DORSAL HORN

脊髓中的胶状质(SG)神经元是初级传入的主要终末部位，其中许多支配伤害性感受器。内源性阿片剂或合成化合物被认为能产生抗伤害性感受器。内源性阿片剂或合成化合物被认为通过与SG神经元上的阿片受体亚型相互作用而产生抗伤害性感觉。最近分离的两个四肽内吗啡素(Endo)1和2被认为是阿片受体Mu亚型的内源性配体。初步结果显示阿片受体的内吗啡素样免疫反应(Endo-LI)亚型。初步结果表明，内吗啡样免疫反应阳性物质(Endo-LI)定位于大鼠背角浅层密集的神经纤维网络。因此，大鼠背角提供了一个独特的机会来检验这一假说，即Endo-1/Endo-2是内源性释放的，它可能调节SG神经元的活动。将解决两个主要问题。首先，利用放射性微探针技术研究麻醉大鼠体内内源性内吗啡的释放，以及体外大鼠脊髓内源性内源性内吗啡的释放。电刺激痛性刺激后爪的传入纤维将被用来评估在这些条件下内吗啡素的释放是否发生改变。其次，将利用全细胞膜片记录技术研究Endo对大鼠脊髓横断脑片单个SG神经元的细胞作用及其信号转导机制。我们的初步结果表明，Endo通过超极化和/或减弱突触传递来抑制SG神经元的活动。在这项提案中，将从电生理学和药理学的角度评估Endo的突触前和突触后的作用。K通道的亚型(S)可能是Endo超极化作用的基础，我们将对此进行研究。类似地，还将评估与突触前阿片受体偶联的钙通道和/或钾通道的亚型(S)，该通道可能介导内藤碱的突触抑制作用。这个项目的长期目标是提高我们目前对这类新的阿片肽在背角神经元上的作用部位和作用机制的了解，目的是开发一类具有治疗潜力的新型阿片类化合物。

Substantia gelatinosa (SG) neurons in the spinal cord are the principal site of termination of primary afferents, many of which innervate nociceptors. Endogenous opiates or synthetic compounds are thought to produce their anti-nociceptors. Endogenous opiates or synthetic compounds are thought to produce their anti-nociception by interacting with subtypes of opiate receptors on SG neurons. Two recently isolated tetrapeptides endomorphin (Endo) 1 and 2 are believed to be the endogenous ligand for the mu-subtype of opioid receptors. Preliminary results showed endomorphin-like immunoreactivity (Endo-LI) subtype of opioid receptors. Preliminary results showed that endomorphin-like immunoreactivity (Endo-LI) is localized to dense networks of nerve fibers in the superficial layers of the rat dorsal horn. Thus, the rat dorsal horn offers a unique opportunity to test the hypothesis that Endo-1/Endo-2 is released endogenously and that it may modulate the activity of SG neurons. Two major issues will be addressed. First, release of endogenous of endogenous endomorphins will be evaluated in anesthetized rats in vivo or isolated rat spinal cords in vitro by the radioactive microprobe techniques. Electrical stimulation of afferent fibers of painful stimulus to the hindpaw will be employed to evaluate whether or not endomorphin release is altered under these conditions. Second, whole-cell patch recording techniques will be used to study the cellular action and the signal transduction mechanism underlying the action of Endo on single SG neurons in rat transverse spinal cord slices. Our preliminary results show that Endo inhibits the activity of SG neurons by hyperpolarizing the membrane and/or attenuating synaptic transmission. In this proposal, the pre- and post-synaptic actions of Endo will be evaluated electrophysiologically and pharmacologically. The subtype(s) of K+ channels that may underlie the hyperpolarizing action of Endo will be examined. Similarly, the subtype(s) of Ca2+ and/or K+ channels coupled to the presynaptic opiate receptor that may mediate the synaptic depressant action of Endo will be evaluated. The long term goal of this project is to improve our current understanding of the site and mechanism of action of this new class of opioid peptides on dorsal horn neurons, with the aim toward developing a novel class of opiate compounds with therapeutic potentials.