Shiosaka S, Yamamoto T, Hertzberg E L, Nagy J I
Department of Physiology, Faculty of Medicine, University of Manitoba, Winnipeg, Canada.
J Comp Neurol. 1989 Mar 8;281(2):282-97. doi: 10.1002/cne.902810210.
Immunohistochemical techniques and an affinity-purified antibody directed against the 27-kD gap-junctional protein (GJP) from rat liver were used to determine the ultrastructural localization of GJP in the rat hippocampus. At the light microscope level, dense GJP immunoreactivity having a stringlike appearance was seen in a very small percentage of medium-sized neuronal somata located in the stratum pyramidale, and diffuse immunostaining was seen in many small cell bodies in the stratum pyramidale, stratum oriens, and the alveus. Abundant GJP-immunoreactive (GJP-IR) varicose fibers were observed in the strata pyramidale, radiatum, and oriens but were less concentrated in the alveus. Numerous punctate GJP-IR elements were observed in all hippocampal layers. Upon EM analysis, GJP-IR neuronal somata in the stratum pyramidale were found to be, without exception, nonpyramidal neurons as judged by such distinguishing features as their fusiform perikarya, indented nucleus, and well-developed rough endoplasmic reticulum (RER). Immunostaining within these cells was largely localized to the Golgi apparatus and associated vesicular components. Small, diffusely GJP-IR cells were identified ultrastructurally as protoplasmic and fibrous astrocytes. Immunostaining within these cells was localized to the Golgi apparatus, RER, and small, ribosomelike bodies 15-25 nm in diameter. Among neuronal processes GJP immunoreactivity was found within dendrites, axons, and axonal terminals. The latter structures contained numerous GJP-IR vesicles having an average diameter of about 40 nm. A frequent observation indicating some degree of specificity of the anti-GJP antibody employed here was immunostaining of typical gap junctions between dendrites and, more commonly, between processes of glial cells. Occasionally, however, GJP-IR dendrodendritic, axodendritic, and axoaxonic contacts were found that could be considered, at best, as being gap-junction-like (gj-L). In these cases, asymmetric immunostaining of adjacent plasma membranes forming gj-L structures was not uncommon. These results confirm the existence of gap junctions between dendrites in the rat hippocampus and demonstrate that GJP immunoreactivity on cytoplasmic membranes is restricted either to typical neuronal and glial gap junctions or to gj-L structures at circumscribed sites of contact between various types of neuronal elements where GJP may contribute to a novel mechanism of neural communication.
采用免疫组织化学技术和一种针对大鼠肝脏27-kD缝隙连接蛋白(GJP)的亲和纯化抗体,来确定GJP在大鼠海马体中的超微结构定位。在光学显微镜水平,位于锥体层的极少数中型神经元胞体中可见呈串状外观的密集GJP免疫反应性,在锥体层、原层和海马槽的许多小细胞体中可见弥漫性免疫染色。在锥体层、辐射层和原层观察到大量GJP免疫反应性(GJP-IR)曲张纤维,但在海马槽中浓度较低。在所有海马层均观察到大量点状GJP-IR元件。通过电子显微镜分析,根据其梭形胞体、凹陷核和发达的粗面内质网(RER)等特征判断,锥体层中GJP-IR神经元胞体无一例外均为非锥体神经元。这些细胞内的免疫染色主要定位于高尔基体和相关的囊泡成分。超微结构鉴定,小的、弥漫性GJP-IR细胞为原浆性和纤维性星形胶质细胞。这些细胞内的免疫染色定位于高尔基体、RER和直径为15-25nm的小核糖体样体。在神经元突起中,在树突、轴突和轴突终末内发现了GJP免疫反应性。后者结构包含许多平均直径约40nm的GJP-IR囊泡。一个常见的观察结果表明这里使用的抗GJP抗体具有一定程度的特异性,即树突之间以及更常见的胶质细胞突起之间的典型缝隙连接出现免疫染色。然而,偶尔也会发现GJP-IR树突-树突、轴突-树突和轴突-轴突接触,充其量可认为是缝隙连接样(gj-L)。在这些情况下,形成gj-L结构的相邻质膜不对称免疫染色并不罕见。这些结果证实了大鼠海马体中树突之间存在缝隙连接,并表明细胞质膜上的GJP免疫反应性要么局限于典型的神经元和胶质缝隙连接,要么局限于不同类型神经元元件之间特定接触部位的gj-L结构,其中GJP可能有助于一种新的神经通讯机制。