CALRETININ-CONTAINING NEURONS AND EXCITOTOXIC INJURY

钙调素（CR）是一种钙结合蛋白， 主要存在于大脑神经元的亚群中， 黑质（SN）和腹侧被盖区（VTA）。 我们之前已经证明，一些SN/VTA多巴胺- 含有与CR共定位的细胞。 的进一步研究 完整的大鼠胚胎脑显示共定位 从E16开始 我们观察到许多CR免疫反应性（IR） 酪氨酸羟化酶（TH） 组织培养中的免疫反应细胞，但我们没有 发现CR + TH共定位于生长的胚胎细胞（E14）中 培养12天，似乎有些因素 存在于大脑中但不解离 cultures.我们还研究了兴奋性氨基的影响 EAA对SN细胞CR和TH表达的影响。 24小时后，CR和TH神经元受到不同程度的影响， 与红藻氨酸（KA）、NMDA或AMPA（100 - 500 总的来说，TH-IR神经元表现出更大的变化 EAA处理后CR-IR神经元比EAA处理后CR-IR神经元更少，并且两种类型的CR-IR神经元都比EAA处理后CR-IR神经元更少。 细胞受影响的严重程度降序：KA AMPA大于NMDA。 变性 神经元细胞和突起较少被观察到 KA（47%）和NMDA（22%）后CR-IR细胞中， TH-IR细胞（KA，57%; NMDA和AMPA，35%）。 两人都幸存下来 CR和TH细胞表现出增加的纤维分支， 但细胞的外观不同。 而存活的CR 细胞突起延伸很长的距离，TH细胞 出现了越来越多的轮辐状突起 直接来自细胞体，其中许多已经肿胀 静脉曲张 长分枝的戏剧性外观 EAA后的静脉曲张过程，尤其是KA， 我们的注意力集中在新纤维是否发芽 在EAA后24小时内，或在可能性， CR的周转和/或轴浆流增加 现有的纤维在控制下是不可见的 条件 定量CR的放射免疫测定显示， KA（42%）和AMPA（17%）后CR蛋白丢失，但无 在NMDA处理的细胞中的作用。 裂解核糖核酸酶 保护试验显示， KA治疗组CR和TH均为81%，TH为76%， AMPA治疗组的下降幅度较小（CR：62%，TH：43%）， NMDA处理的细胞（CR：58%TH：36%）。 这些数据不 明确地揭示了增加的 神经丛的纤维后EAA，作为CR-IR表明一个主要的 蛋白质从细胞体进入挪威啃食器或 在一些细胞中发芽，而在两个细胞中的减少， 蛋白质和mRNA的变化表明， 细胞 这是否反映了 SN A9或VTA（A10）细胞仍有待显示。 这种区别以前被发现， 用SN神经毒素MPTP治疗猴子， 破坏A9细胞但不破坏A10亚群。 以来 CR在培养的TH细胞内的共定位不是 但是，我们不能从这些实验中得出结论， 含有CR的多巴胺细胞被保护免受神经毒性 伤害EAA。

Calcretinin (CR) is a calcium binding protein that is mostly found in subpopulations of brain neurons such as the substantia nigra (SN) and the ventral tegmental area (VTA). We have previously shown that some of the SN/VTA dopamine- containing cells colocalize with CR. A further study of intact rat embryo brains revealed that colocalization begins at E16. We observed many CR immunoreactive (IR) neurons in addition to tyrosine hydroxylase (TH) immunoreactive cells in tissue culture, but we have not found CR+TH colocalization in embryonic cells (E14) grown for up to 12 days in culture, It appears that some factor is required which is present in brain but not dissociated cultures. We also studied the effects of excitatory amino acids (EAA) on the expression of CR and TH in SN cultures. CR and TH neurons were differentially affected by 24 hour incubation with kainic acid (KA), NMDA or AMPA (100-500 muM)> In general, TH-IR neurons exhibited greater change than CR-IR neurons after EAA treatment, and both types of cells were affected in a decreasing order of severity: KA greater than AMPA greater than NMDA. Degeneration of neuronal cells and processes were less frequently observed in CR-IR cells after KA (47%) and NMDA (22%) compared with TH-IR cells (KA, 57%; NMDA and AMPA, 35%). Both surviving CR and TH cells exhibited increased fiber ramifications, but the cells' appearances differed. Whereas surviving CR cell processes extended for very long distances, TH cells had increased numbers of spoke-like processes emerging directly from cell bodies, many of which had swollen varicosities. The dramatic appearance of long ramifying varicose processes following EAA, especially KA, focuses our attention on whether sprouting of new fibers occurs within 24 h following the EAA, or on the possibility that there is an increased turnover and/or axoplasmic flow of CR to existing fibers that were not visible under control conditions. A radioimmunoassay to quantify CR revealed a loss of CR protein after KA (42%), and AMPA (17%), but no effect in NMDA-treated cells. A lysate ribonuclease protection assay revealed a significant loss of mRNA for both CR and TH in the KA-treated group (CR:81%, TH:76%) and a lesser decrease in the AMPA-treated (CR:62%, TH:43%) and NMDA-treated cells (CR:58% TH:36%). These data do not definitively reveal the significance of the increased plexus of fibers following EAA, as CR-IR suggests a major movement of protein from cell bodies into norway gnawer or sprouting in some cells while a reduction in both the protein and mRNA suggests degeneration of a subset of cells. Whether this is a reflection of differences between SN compacta (A9) or VTA (A10) cells remains to be shown. Such a distinction was previously uncovered following treatment in monkeys with the SN neurotoxin, MPTP, which destroyed A9 cells but not the A10 subpopulation. Since colocalization of CR within TH cells in culture was not revealed, we cannot conclude from these experiments that CR-containing dopamine cells are protected from neurotoxic injury by EAA.