Bixel M, Shimomura Y, Hutson S, Hamprecht B
Physiologisch-chemisches Institut der Universität, Tübingen, Germany.
J Histochem Cytochem. 2001 Mar;49(3):407-18. doi: 10.1177/002215540104900314.
Transamination of branched-chain amino acids (BCAAs) catalyzed by the branched chain aminotransferase isoenzymes (BCATs) is believed to play an important role in nitrogen shuttling and excitatory neurotransmitter glutamate metabolism in brain. Recently, we have shown that the mitochondrial isoenzyme (BCATm) is the predominant form found in cultured astrocytes. In this study we used immunocytochemistry to examine the distribution of BCAT isoenzymes in cultured rat neurons and microglial cells. The cytoplasm of neurons displayed intense staining for the cytosolic isoenzyme (BCATc), whereas BCATm staining was not detectable in neurons. In contrast, microglial cells expressed BCATm in high concentration. BCATc appeared to be absent in this cell type. The second and committed step in the BCAA catabolic pathway is oxidative decarboxylation of the alpha-keto acid products of BCAT catalyzed by the branched-chain alpha-keto acid dehydrogenase (BCKD) enzyme complex. Because the presence of BCKD should provide an index of the ability of a cell to oxidize BCAA, we have also immunocytochemically localized BCKD in neuron and glial cell cultures from rat brain. Our results suggest ubiquitous expression of this BCKD enzyme complex in cultured brain cells. BCKD immunoreactivity was detected in neurons and in astroglial and microglial cells. Therefore, the expression of BCAT isoenzymes shows cell-specific localization, which is consistent with the operation of an intercellular nitrogen shuttle between neurons and astroglia. On the other hand, the ubiquitous expression of BCKD suggests that BCAA oxidation can probably take place in all types of brain cells and is most likely regulated by the activity state of BCKD rather than by its cell-specific localization.
支链氨基酸转氨酶同工酶（BCATs）催化的支链氨基酸（BCAAs）转氨作用被认为在大脑中的氮穿梭和兴奋性神经递质谷氨酸代谢中起重要作用。最近，我们已经表明线粒体同工酶（BCATm）是培养的星形胶质细胞中发现的主要形式。在本研究中，我们使用免疫细胞化学方法检测了BCAT同工酶在培养的大鼠神经元和小胶质细胞中的分布。神经元的细胞质对胞质同工酶（BCATc）显示出强烈染色，而在神经元中未检测到BCATm染色。相反，小胶质细胞高浓度表达BCATm。在这种细胞类型中似乎不存在BCATc。BCAA分解代谢途径的第二步也是关键步骤是由支链α-酮酸脱氢酶（BCKD）酶复合物催化的BCAT的α-酮酸产物的氧化脱羧。因为BCKD的存在应该提供细胞氧化BCAA能力的指标，我们还通过免疫细胞化学方法在大鼠脑的神经元和胶质细胞培养物中定位了BCKD。我们的结果表明这种BCKD酶复合物在培养的脑细胞中普遍表达。在神经元以及星形胶质细胞和小胶质细胞中检测到BCKD免疫反应性。因此，BCAT同工酶的表达显示出细胞特异性定位，这与神经元和星形胶质细胞之间细胞间氮穿梭的运作一致。另一方面，BCKD的普遍表达表明BCAA氧化可能发生在所有类型的脑细胞中，并且最有可能由BCKD的活性状态而不是其细胞特异性定位来调节。
