Molecular pharmacological study for characterization of new identified iropioid receptor subclass

利用克隆的MOR-1、MOR-1A、MOR-1C和MOR-1E细胞，研究了这些μ-阿片受体剪接变异体的药理学特性。DAMGO是典型的μ-阿片受体激动剂，在这些剪接变异体中均表现出高亲和力和内在活性。此外，选择性μ-阿片受体激动剂氨基- tapa可以诱导内源性x-阿片肽的释放，在所有这些剪接变异体中也显示出较高的内在活性。这些证据清楚地表明，这4种剪接变体并不是导致内源性κ-阿片肽释放的μ-阿片受体剪接变体。为了鉴定导致内源性κ-阿片肽释放的剪接变异，我们决定首先利用μ-阿片受体基因的外显子特异性反义寡脱氧核苷酸，在30多个剪接变异中鉴定氨基- tapa敏感的damgo不敏感的剪接变异。结果表明，mir - 1j、mir - 1k和mir - 1l是对tapa敏感的damgo不敏感的剪接变体。此外，利用内源性κ-阿片肽、dynorphin A、dynorphin B或α-neoendorphin抗血清进行的实验发现，mir - 1j和mir - 1l是导致dynorphin A释放的剪接变体，而mir - 1k是导致dynorphin B和α-neoendorphin释放的剪接变体。基于以上证据，我们克隆出了MOR-1J、MOR-1K和MOR-1L，但在克隆出MOR-1K和MOR-1L的细胞中没有观察到氨基- tapa的内在活性。由于这两种剪接变体不含跨膜结构，如果没有其他含有7-跨膜结构的剪接变体，这些剪接变体可能没有任何功能。为了鉴定其表达细胞中mir - 1j、mir - 1k和mir - 1l的药理学特征，并对这些剪接变体的异源二聚体进行功能表征。Μ-opioid可能需要含有7-跨膜结构的受体剪接变异体。本项目所克隆的MOR-1J、MOR-1K和MOR-1L的cDNA拟用于新立项。少

With the cells expressed cloned MOR-1, MOR-1A, MOR-1C or MOR-1E, the pharmacological character of these μ-opioid receptor splice variants were investigated. DAMGO, typical μ-opioid receptor agonist, showed high affinity and intrinsic activity in all of these splice variants. Moreover, amidino-TAPA, selective μ-opioid receptor agonist can lead the release of endogenous x-opioid peptides, also showed high intrinsic activity in all of these splice variants. These evidences clearly suggest that these 4 splice variants are not the μ-opioid receptor splice variants lead the release of endogenous κ-opioid peptides. To identify the splice varinats lead the release of endogenous κ-opioid peptides, we decide to identify first the amidino-TAPA-sensitive DAMGO-insensitive splice variants among more than 30 splice variants in behavioral experiment using the exon-specific antisense oligodeoxynucleotides for μ-opioid receptor gene. As the results, MOR-1J, MOR-1K and MOR-1L were identified as the amid … More ino-TAPA-sensitive DAMGO-insensitive splice variants. Moreover, additional experiments using the antisera against endogenous κ-opioid peptides, dynorphin A, dynorphin B or α-neoendorphin, identified that MOR-1J and MOR-1L are splice variants lead the release of dynorphin A, whereas MOR-1K is a splice variant lead the release of dynorphin B and α-neoendorphin. With above evidence, MOR-1J, MOR-1K and MOR-1L were cloned, but no intrinsic activity of amidino-TAPA was observed in the cells expressed cloned MOR-1K and MOR-1L. Since these 2 splice variants do not contain transmembrane structure, these splice variants may not have any function without other splice variants contain 7-transmembrane structure. To identify the pharmacological character of MOR-1J, MOR-1K and MOR-1L in its expressed cell, the functional characterization of heterodimer of these splice variants with other. Μ-opioid receptor splice variants, which contain 7-transmembrane structure, may be required. The cDNA of MOR-1J, MOR-1K and MOR-1L, cloned in the present project, are scheduled to be used in the new project accepted. Less