Kemmel Véronique, Miehe Monique, Roussel Guy, Taleb Omar, Nail-Boucherie Karine, Marchand Christiane, Stutz Christine, Andriamampandry Christian, Aunis Dominique, Maitre Michel
Institut de Chimie Biologique and INSERM U-575, Faculty of Medicine, 67085 Strasbourg, France.
J Comp Neurol. 2006 Oct 1;498(4):508-24. doi: 10.1002/cne.21072.
Gamma-hydroxybutyrate (GHB) is a substance derived from the metabolism of GABA and is heterogeneously distributed in various regions of the brain. This compound possesses a neuromodulatory role on several types of synapses, particularly those using GABA as a neurotransmitter. At physiological concentrations, this effect of GHB is mediated via specific receptors that induce neuronal hyperpolarization and bind radioactive GHB with a specific distribution, ontogenesis, kinetics, and pharmacology. A membrane protein that possesses six to seven transmembrane domains and which binds and is activated by micromolar amounts of GHB was recently cloned from rat brain hippocampus. In order to study the regional and cellular distribution of this receptor in rat brain, we selected several specific peptides belonging to the extracellular domains of the receptor to be used as specific immunogens to raise polyclonal antibodies in the rabbit. Among the antisera obtained, one of them gave particularly good results in terms of specificity and reactivity at high dilution. Immunohistochemical analyses, both at the confocal and electron microscopic level, showed receptor protein distribution closely resembling the distribution of GHB high-affinity binding sites, except for cerebellum, where GHB receptor(s) of lower affinity exist(s). In all regions studied the GHB receptor-like protein labeling appears to be distributed specifically in neurons and not in glial cells. At the cellular level the antibody specifically labels dendrites, and no immunoreactivity was detected in presynaptic endings or in axons. Accordingly, electron microscopy reveals strong labeling of postsynaptic densities and of neuronal cytosol.
γ-羟基丁酸（GHB）是一种由γ-氨基丁酸（GABA）代谢产生的物质，在大脑的各个区域呈不均匀分布。这种化合物对几种类型的突触具有神经调节作用，特别是那些以GABA作为神经递质的突触。在生理浓度下，GHB的这种作用是通过特定受体介导的，这些受体可诱导神经元超极化，并以特定的分布、个体发生、动力学和药理学特性结合放射性GHB。最近从大鼠脑海马体中克隆出一种膜蛋白，该蛋白具有六到七个跨膜结构域，能与微摩尔量的GHB结合并被其激活。为了研究该受体在大鼠脑中的区域和细胞分布，我们选择了属于该受体细胞外结构域的几种特定肽段作为特异性免疫原，用于在兔体内产生多克隆抗体。在所获得的抗血清中，其中一种在高稀释度下的特异性和反应性方面表现特别出色。共聚焦显微镜和电子显微镜水平的免疫组织化学分析表明，受体蛋白的分布与GHB高亲和力结合位点的分布非常相似，但小脑除外，小脑中存在低亲和力的GHB受体。在所研究的所有区域中，类GHB受体蛋白标记似乎特异性地分布在神经元中，而不是神经胶质细胞中。在细胞水平上，该抗体特异性标记树突，在突触前末梢或轴突中未检测到免疫反应性。因此，电子显微镜显示突触后致密物和神经元胞质溶胶有强烈的标记。
