Zagon I S, Gibo D M, McLaughlin P J
Department of Anatomy, Pennsylvania State University, M.S. Hershey Medical Center, Hershey 17033.
Brain Res. 1991 Jun 14;551(1-2):28-35. doi: 10.1016/0006-8993(91)90909-f.
Endogenous opioids and opioid receptors (i.e. endogenous opioid systems) are expressed during neural ontogeny, and play a role in the development of the nervous system. Using [3H][Met5]-enkephalin, a potent ligand involved in neural growth, particularly cell proliferation, specific and saturable binding was detected in homogenates of 6-day-old rat cerebellum; the data were consistent with a single binding site. Scatchard analysis yielded a binding affinity (Kd) of 2.2 nM and a binding capacity (Bmax) of 22.3 fmol/mg protein. Binding was linear with protein concentration, dependent on time, temperature, and pH, and was sensitive to Na+, Mg2+, and guanyl nucleotides. Optimal binding required protease inhibitors, and pretreatment of the homogenates with trypsin markedly reduced binding, suggesting that the binding site was proteinaceous in character. The [Met5]-enkephalin binding site was an integral membrane protein located in the nuclear fraction. Competition experiments indicated that [Met5] enkephalin was the most potent displacer of [3H][Met5]-enkephalin, and that binding was stereospecific. In the adult rat cerebellum, non-opioid receptor binding of [3H][Met5]-enkephalin was recorded, mu and kappa receptors were also found in the developing rat cerebellum, while mu, delta, and kappa receptors were recorded in adult cerebellar tissue. The function, pharmacological and biochemical characteristics, subcellular distribution, and temporal expression of the [Met5]-enkephalin binding site suggest the presence of a unique opioid receptor, termed zeta (zeta), in the developing nervous system.
内源性阿片肽和阿片受体（即内源性阿片系统）在神经个体发生过程中表达，并在神经系统发育中发挥作用。使用[3H][Met5]-脑啡肽（一种参与神经生长，特别是细胞增殖的强效配体），在6日龄大鼠小脑匀浆中检测到特异性和可饱和结合；数据与单一结合位点一致。Scatchard分析得出结合亲和力（Kd）为2.2 nM，结合容量（Bmax）为22.3 fmol/mg蛋白质。结合与蛋白质浓度呈线性关系，依赖于时间、温度和pH，并且对Na+、Mg2+和鸟苷酸敏感。最佳结合需要蛋白酶抑制剂，用胰蛋白酶预处理匀浆可显著降低结合，这表明结合位点具有蛋白质性质。[Met5]-脑啡肽结合位点是位于核部分的整合膜蛋白。竞争实验表明，[Met5]脑啡肽是[3H][Met5]-脑啡肽最有效的置换剂，并且结合具有立体特异性。在成年大鼠小脑中，记录到[3H][Met5]-脑啡肽的非阿片受体结合，在发育中的大鼠小脑中也发现了μ和κ受体，而在成年小脑组织中记录到μ、δ和κ受体。[Met5]-脑啡肽结合位点的功能、药理学和生化特性、亚细胞分布以及时间表达表明，在发育中的神经系统中存在一种独特的阿片受体，称为ζ（zeta）。
