Dana C, Vial M, Leonard K, Beauregard A, Kitabgi P, Vincent J P, Rostène W, Beaudet A
INSERM U-55, Hôpital St. Antoine, Paris, France.
J Neurosci. 1989 Jul;9(7):2247-57. doi: 10.1523/JNEUROSCI.09-07-02247.1989.
The distribution of specifically labeled neurotensin (NT) binding sites was examined by light and electron microscopic radioautography in the ventral tegmental area (VTA) and nucleus interfascicularis of the rat following incubation of lightly prefixed midbrain slices with the monoiodinated ligand, 125I-(Tyr3)-NT. Film radioautograms of whole 125I-NT-incubated slices exhibited intense NT displaceable binding throughout the VTA and interfascicular nucleus. In light microscopic radioautographs from 1-microns-thick sections taken from the surface of the slices, the label was found to be present both inside and outside neuronal perikarya. Probability circle analysis of silver grain distribution in electron microscopic radioautographs confirmed that a significant proportion (greater than 20%) of the specifically labeled binding sites was intraneuronal. The frequent association of these sites with profiles of rough endoplasmic reticulum or Golgi apparatus suggested that they corresponded in part to receptors undergoing synthesis and/or glycosylation. The remainder was associated with neuronal and/or glial plasma membranes, as attested by comparing the distribution of grains overlying apposed cellular elements with the distribution of hypothetical grains originating from randomly distributed membrane bound radioactive sources. Although the resolution of the technique did not make it possible to ascribe labeled membrane-bound receptors to either one of the apposed plasma membranes, their frequent association with interfaces involving the plasmalemma of perikarya and dendrites, together with the occurrence of silver grain alignments along the membrane of certain somata and dendrites suggested that a proportion of them was associated with the perikarya and dendrites of a subpopulation of ventral tegmental neurons. Interestingly, these perikaryal and dendritic receptors were not exclusively present on, or even concentrated opposite, abutting axon terminals but instead were more or less evenly distributed along the plasma membrane. Only an exceedingly small proportion was found to be associated with synaptic junctions. Such a low incidence makes it unlikely that only the synapse-linked binding sites correspond to functional receptors. On the contrary, the dispersion of labeled receptors seen here along the plasma membrane of presumptive dopamine neurons suggests that NT acts mainly in a paracrine or parasynaptic fashion in the ventral midbrain tegmentum.
在用单碘化配体125I-(Tyr3)-神经降压素（NT）孵育轻度固定的中脑切片后，通过光学和电子显微镜放射自显影术检查大鼠腹侧被盖区（VTA）和束间核中特异性标记的NT结合位点的分布。整个用125I-NT孵育的切片的胶片放射自显影片显示，在整个VTA和束间核中均存在强烈的NT可置换结合。在从切片表面切取的1微米厚切片的光学显微镜放射自显影片中，发现标记物存在于神经元胞体的内部和外部。电子显微镜放射自显影片中银颗粒分布的概率圆分析证实，相当一部分（超过20%）特异性标记的结合位点位于神经元内。这些位点经常与粗面内质网或高尔基体的轮廓相关联，表明它们部分对应于正在进行合成和/或糖基化的受体。其余的与神经元和/或神经胶质细胞膜相关，通过将覆盖相邻细胞成分的颗粒分布与源自随机分布的膜结合放射源的假设颗粒分布进行比较得到证实。尽管该技术的分辨率无法将标记的膜结合受体归属于相邻的任何一个质膜，但它们经常与涉及胞体和树突质膜的界面相关联，以及某些胞体和树突膜上银颗粒排列的出现表明，其中一部分与腹侧被盖神经元亚群的胞体和树突相关。有趣的是，这些胞体和树突受体并非仅存在于相邻轴突终末上，甚至也不是集中在其对面，而是或多或少均匀地分布在质膜上。仅发现极小比例的受体与突触连接相关。如此低的发生率使得不太可能只有与突触相关的结合位点对应于功能性受体。相反，此处所见的标记受体在假定的多巴胺能神经元质膜上的分散表明，NT主要以旁分泌或旁突触方式在腹侧中脑被盖区起作用。
