Faure M P, Nouel D, Beaudet A
Montreal Neurological Institute, McGill University, Quebec, Canada.
Neuroscience. 1995 Sep;68(2):519-29. doi: 10.1016/0306-4522(95)00145-9.
Previous studies have demonstrated that neurotensin is internalized and retrogradely transported in neurons of the substantia nigra following its intracerebral injection in the neostriatum. The aim of the present study was to compare the intracellular distribution of retrogradely transported material with that observed following internalization of the peptide at the somatodendritic level and to confirm that the internalization was confined to dopamine neurons. To document somatodendritic internalization, slices (350 microns) from the rat ventral midbrain were incubated in vitro with 20 mM fluoresceinylated neurotensin, a fluorescent derivative of neurotensin, and immunostained 5-60 min later for tyrosine hydroxylase. To document retrograde transport, rats were injected with the same compound into the neostriatum and the brains processed for tyrosine hydroxylase immunohistochemistry 4.5 and 8 h later. Confocal laser microscopic examination of superfused slices revealed that fluoresceinylated neurotensin was internalized at the level of the perikarya and processes of neurons in the substantia nigra, ventral tegmental area and interfascicular nucleus. At short time intervals, the label was detected in the form of small, intensely fluorescent particles distributed within the cytoplasm of both perikarya and dendrites. At longer time intervals, these fluorescent particles were larger, less numerous and confined to the perikarya where they eventually clustered against the nucleus. Following intrastriatal injection of fluoresceinylated neurotensin, retrogradely labeled cells were apparent throughout the substantia nigra, pars compacta, as well as in the lateral part of the ventral tegmental area. Here again, the label took the form of small fluorescent particles, comparable in size, shape and distribution to those detected following superfusion of midbrain slices. In both labeling conditions, fluoresceinylated neurotensin was almost exclusively confined to tyrosine hydroxylase-immunoreactive cells. These results indicate that neurotensin is internalized throughout the terminal and dendritic arborization of mesostriatal dopamine cells and that the internalized peptide is transported centripetally from both locations to the soma of the cells. The clustering of fluorescent particles in the perinuclear region of the cells further suggests that the internalized process may play a role in the long term transcellular signalling.
先前的研究表明，在新纹状体内脑内注射神经降压素后，它会在黑质神经元中被内化并逆行运输。本研究的目的是比较逆行运输物质的细胞内分布与在树突体水平肽内化后观察到的分布，并确认内化仅限于多巴胺能神经元。为了记录树突体的内化，将来自大鼠腹侧中脑的切片（350微米）与20 mM荧光素化神经降压素（神经降压素的一种荧光衍生物）在体外孵育，并在5 - 60分钟后用酪氨酸羟化酶进行免疫染色。为了记录逆行运输，将相同的化合物注射到大鼠的新纹状体内，并在4.5小时和8小时后对大脑进行酪氨酸羟化酶免疫组织化学处理。对灌流切片的共聚焦激光显微镜检查显示，荧光素化神经降压素在黑质、腹侧被盖区和束间核的神经元胞体和突起水平被内化。在短时间间隔内，标记物以小的、强烈荧光的颗粒形式被检测到，分布在胞体和树突的细胞质内。在较长时间间隔内，这些荧光颗粒更大、数量更少，并局限于胞体，最终聚集在细胞核周围。在纹状体内注射荧光素化神经降压素后，整个黑质致密部以及腹侧被盖区外侧的逆行标记细胞明显可见。同样，标记物呈小荧光颗粒形式，其大小、形状和分布与中脑切片灌流后检测到的相似。在两种标记条件下，荧光素化神经降压素几乎完全局限于酪氨酸羟化酶免疫反应阳性细胞。这些结果表明，神经降压素在中脑纹状体多巴胺细胞的终末和树突分支中被内化，并且内化的肽从这两个位置向细胞体进行向心性运输。细胞内核周区域荧光颗粒的聚集进一步表明，内化过程可能在长期跨细胞信号传导中起作用。
