乙醇给药在大鼠脑中产生 GABA 能神经活性甾体免疫组织化学的不同变化。
Ethanol administration produces divergent changes in GABAergic neuroactive steroid immunohistochemistry in the rat brain.
机构信息
Departments of Psychiatry and Pharmacology , Bowles Center for Alcohol Studies, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
出版信息
Alcohol Clin Exp Res. 2014 Jan;38(1):90-9. doi: 10.1111/acer.12223. Epub 2013 Jul 26.
BACKGROUND
The 5α-reduced pregnane neuroactive steroid (3α,5α)-3-hydroxypregnan-20-one (3α,5α-THP or allopregnanolone) is a potent positive modulator of GABAA receptors capable of modulating neuronal activity. In rats, systemic ethanol (EtOH) administration increases cerebral cortical and hippocampal levels of 3α,5α-THP, but the effects of EtOH on 3α,5α-THP levels in other brain regions are unknown. There is a large body of evidence suggesting that 3α,5α-THP enhances EtOH sensitivity, contributes to some behavioral effects of EtOH, and modulates EtOH reinforcement and motivation to drink. In this study, we used immunohistochemistry (IHC) to determine EtOH-induced changes in cellular 3α,5α-THP expression in brain regions associated with EtOH actions and responses.
METHODS
Male Wistar rats were administered EtOH (2 g/kg) or saline intraperitoneally and after 60 minutes transcardially perfused. IHC was performed on free-floating sections (3 to 4 sections/animal/brain region) using an affinity purified anti-3α,5α-THP primary antibody, and immunoreactivity was visualized with 3,3'-diaminobenzidine.
RESULTS
EtOH significantly increased 3α,5α-THP immunoreactivity by 24 ± 6% in the medial prefrontal cortex, 32 ± 12% in the hippocampal Cornu Ammonis area 1 (CA1) pyramidal cell layer, 52 ± 5% in the polymorph cell layer of the dentate gyrus (DG), 44 ± 15% in the bed nucleus of the stria terminalis, and 36 ± 6% in the paraventricular nucleus of the hypothalamus. In contrast, EtOH administration significantly reduced 3α,5α-THP immunoreactivity by 25 ± 5% in the nucleus accumbens "shore" and 21 ± 3% in the central nucleus of the amygdala. No changes were observed in the ventral tegmental area, dorsomedial striatum, granule cell layer of the DG, or the lateral and basolateral amygdala.
CONCLUSIONS
The results suggest acute EtOH (2 g/kg) produces divergent, brain region specific, effects on cellular 3α,5α-THP levels. Regional differences in the effects of EtOH suggest there may be regional brain synthesis of 3α,5α-THP independent of the adrenal glands and novel mechanisms that reduce cellular 3α,5α-THP. Regional differences in EtOH-induced changes in 3α,5α-THP levels likely contribute to EtOH effects on neuronal function in brain.
背景
5α-还原孕烷神经活性甾体（3α,5α）-3-羟孕烷-20-酮（3α,5α-THP 或别孕烯醇酮）是一种能够调节神经元活性的 GABA A 受体的有效正向调节剂。在大鼠中，全身给予乙醇（EtOH）可增加大脑皮质和海马体中的 3α,5α-THP 水平，但 EtOH 对其他脑区 3α,5α-THP 水平的影响尚不清楚。有大量证据表明，3α,5α-THP 增强了 EtOH 的敏感性，有助于 EtOH 的一些行为效应，并调节 EtOH 的强化和饮酒动机。在这项研究中，我们使用免疫组织化学（IHC）来确定与 EtOH 作用和反应相关的脑区中 EtOH 诱导的细胞 3α,5α-THP 表达变化。
方法
雄性 Wistar 大鼠经腹腔内给予 EtOH（2 g/kg）或生理盐水，60 分钟后经心内灌流。使用亲和纯化的抗 3α,5α-THP 抗体进行游离漂浮切片（每个动物/脑区 3-4 个切片）的免疫组织化学染色，并使用 3,3'-二氨基联苯胺进行免疫反应可视化。
结果
EtOH 使内侧前额叶皮质的 3α,5α-THP 免疫反应性增加了 24±6%，海马体 CA1 锥体细胞层增加了 32±12%，齿状回多形细胞层增加了 52±5%，终纹床核增加了 44±15%，下丘脑室旁核增加了 36±6%。相比之下，EtOH 给药使伏隔核“岸区”的 3α,5α-THP 免疫反应性降低了 25±5%，杏仁核中央核的 3α,5α-THP 免疫反应性降低了 21±3%。腹侧被盖区、背内侧纹状体、齿状回颗粒细胞层以及外侧和基底外侧杏仁核未观察到变化。
结论
结果表明，急性 EtOH（2 g/kg）对细胞 3α,5α-THP 水平产生了不同的、特定脑区的影响。EtOH 作用的区域差异表明，可能存在与肾上腺无关的脑内 3α,5α-THP 合成的区域差异，以及降低细胞 3α,5α-THP 的新机制。EtOH 诱导的 3α,5α-THP 水平变化的区域差异可能有助于 EtOH 对脑内神经元功能的影响。
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