MOR1--MU OPIOID RECEPTORS AND THEIR ENDOGENOUS LIGANDS

克隆每一种已知的阿片类药物 受体（即，delta，mu和kappa）已经极大地改变了 在这个不匹配的问题中， 阿片受体和它们的内源性配体之间的联系两个问题 出现的问题是：“克隆的受体是否能解释所有的阿片类药物， 神经系统中的受体“和“确定的内源性 配体表示可用于作用的配体的完整集合， 阿片受体？" 随着mu阿片样物质敲除小鼠的成功产生， 一个新的神经肽家族的分离和表征（ 内吗啡肽）在μ受体中起作用，这些问题可以 以新颖的方式解决。正是考虑到这一点，我们建议 实验将有助于实现以下目标 具体目标： 1)确定克隆的mu受体MOR1是否是mu受体 负责突触前抑制去甲肾上腺素的释放， 蓝斑神经元的轴突。 2)确定负责合成的生物合成前体 内吗啡肽，以及表达内吗啡肽的细胞的区域分布。 内吗啡肽及其生物活性产物的转录物。 3)确定内吗啡肽和内吗啡肽之间的空间关系。 MOR1。 4)确定缺乏内吗啡肽的小鼠中内吗啡肽的表达是否改变 表观同源受体（MOR1）。 这些目标将通过几种不同类型的实验来实现。 野生型和mu基因敲除小鼠前脑突触体制备 小鼠将有助于阐明MOR1在多大程度上负责 去甲肾上腺素释放的突触前抑制此外，建议 表达克隆实验，以分离生物合成前体 内吗啡肽，随后进行原位杂交研究。最后我们 建议开发针对内吗啡肽的特异性抗血清， 随后在野生型中进行双色免疫荧光实验， 基因敲除小鼠，以确定克隆小鼠之间的匹配程度， 受体和这个新的内源性配体家族。

The cloning of each of the pharmacologically identified types of opioid receptors (i.e., delta, mu and kappa) has dramatically altered the nature of the questions that can be addressed in the problem of the mismatch between opioid receptors and their endogenous ligands. Two questions that arise are: "Do the cloned receptors account for all of the opioid receptors in the nervous system?" and "Do the identified endogenous ligands represent the complete set of ligands available for action at opioid receptors?" With the successful production of a mu opioid knockout mouse and the isolation and characterization of a new family of neuropeptides (the endomorphins) that are active in mu receptors, these questions can be addressed in novel ways. It is with this in mind that we propose experiments that will contribute to the realization of the following specific aims: 1) Determine if the cloned mu receptor, MOR1, is the mu receptor responsible for presynaptic inhibition of norepinephrine release from axons of locus coeruleus neurons. 2) Determine the biosynthetic precursor responsible for synthesis of endomorphin, and the regional distribution of cells that express the transcript for endomorphin and its biologically active products. 3) Determine the spatial relationship between the endomorphin peptides and MOR1. 4) Determine if the expression of endomorphin is altered in mice that lack the apparent cognate receptor (MOR1). These aims will be accomplished by several different types of experiments. Synaptosomal preparations of the forebrain of wild type and mu knockout mice will help to elucidate the extend to which MOR1 is responsible for the presynaptic inhibition of norepinephrine release. Also, proposed are expression cloning experiments to isolate the biosynthetic precursor of the endomorphins, followed by in situ hybridization studies. Finally, we propose the development of specific antisera to the endomorphins and subsequent two-color immunofluorescent experiments in wild type and knockout mice to determine the extent of match between the cloned mu receptor and this new family of endogenous ligands.