Stevens B R, Vo C B
Department of Physiology, University of Florida College of Medicine, Gainesville 32610-0274, USA.
J Neurochem. 1998 Aug;71(2):564-70. doi: 10.1046/j.1471-4159.1998.71020564.x.
The potential induction of cationic and zwitterionic amino acid transport systems and mRNA transcripts was investigated in primary neuronal cultures from rat hypothalamus/brainstem. Cultures exposed to bacterial lipopolysaccharide (LPS) plus interferon-gamma (IFNgamma) were assessed with respect to northern blot analyses, L-leucine/L-arginine cross-inhibition uptake profiles in the presence and absence of Na+, and initial rate sodium-independent L-arginine transport kinetics. L-Arginine uptake activity was constitutively expressed along with uninduced steady-state levels of CAT1 and 4F2hc transcripts. However, neither the high-affinity nor the low-affinity alternatively spliced inducible isoforms of CAT2 or CAT2a transcripts (encoding system y+ in control astrocytes, lymphocytes, or liver) nor the rBAT transcripts (encoding system b(o,+) in control intestinal epithelial cells) were detected by northern analysis of neuronal mRNA. Cross-inhibition profiles were consistent with physiologic system y+ activity, but not system b(o,+) or system y+ L. Transport kinetics gave a single component with Vmax = 113 +/- 7 pmol/min/mg of protein and Km = 47 +/- 8 microM L-arginine; these kinetic parameters were not influenced by addition of LPS/IFNgamma at concentrations that up-regulated CAT2 mRNA and system y+ activity in control astroglia from the same area of the brain. The data are consistent with L-arginine membrane uptake occurring via only system y+ encoded by constitutive CAT1, with possible physiologic contribution by constitutive 4F2hc transcripts in primary neuronal cultures.
研究了大鼠下丘脑/脑干原代神经元培养物中阳离子和两性离子氨基酸转运系统及mRNA转录本的潜在诱导情况。对暴露于细菌脂多糖（LPS）加干扰素-γ（IFNγ）的培养物进行了Northern印迹分析、在有无Na⁺存在下的L-亮氨酸/L-精氨酸交叉抑制摄取曲线以及初始速率的非钠依赖性L-精氨酸转运动力学评估。L-精氨酸摄取活性与CAT1和4F2hc转录本的未诱导稳态水平一起组成性表达。然而，通过对神经元mRNA的Northern分析，未检测到CAT2或CAT2a转录本（在对照星形胶质细胞、淋巴细胞或肝脏中编码系统y⁺）的高亲和力或低亲和力可变剪接诱导异构体，也未检测到rBAT转录本（在对照肠上皮细胞中编码系统b(o,+)）。交叉抑制曲线与生理系统y⁺活性一致，但与系统b(o,+)或系统y⁺L不一致。转运动力学给出了一个单一成分，Vmax = 113±7 pmol/分钟/毫克蛋白质，Km = 47±8 μM L-精氨酸；这些动力学参数不受在相同脑区对照星形胶质细胞中上调CAT2 mRNA和系统y⁺活性的浓度的LPS/IFNγ添加的影响。数据表明L-精氨酸膜摄取仅通过组成性CAT1编码的系统y⁺发生，组成性4F2hc转录本在原代神经元培养物中可能有生理贡献。
