Bettendorff L, Hennuy B, Wins P, Schoffeniels E
Laboratory of General and Comparative Biochemistry, University of Liège, Belgium.
Neuroscience. 1993 Feb;52(4):1009-17. doi: 10.1016/0306-4522(93)90547-s.
Several membrane fractions were prepared from rat brain by differential and sucrose density gradient centrifugation. Most fractions took up 36Cl- rapidly at a rate linear with time during the first 30-60 s, then the rate progressively slowed down. The lowest rate of uptake was found in the mitochondrial fraction. Oxythiamin partially inhibited 36Cl- uptake in all fractions. In P2 (crude synaptosomal fraction), oxythiamin decreased the initial rate of uptake by 32%, the apparent Ki being 1.5 mM. Thiamin and amprolium were less effective as inhibitors. 4,4'-Diisothiocyanostilbene-2,2'-disulfonic acid (0.1-1 mM) inhibited 36Cl- uptake by 40-50%. In the presence of this compound at a concentration > or = 5 x 10(-4) M, oxythiamin became ineffective. 36Cl- uptake was increased by GABA (0.1 mM) and this effect was antagonized by picrotoxin as expected, but not by oxythiamin. The rate of 36Cl- uptake did not appreciably depend on the external chloride concentration and was unaffected by bumetanide or by replacement of external Na+ by choline. Taken together, these data suggest that the oxythiamin-sensitive 36Cl- influx is essentially diffusional and is not related to the GABA receptor or the Na:K:2Cl co-transport. Partial replacement of external Na+ by K+ or treatment with 0.1 mM veratridine (which should both result in membrane depolarization) increased 36Cl- uptake by 50 and 30% respectively; the inhibitory effect of oxythiamin was enhanced to the same proportion.(ABSTRACT TRUNCATED AT 250 WORDS)
通过差速离心和蔗糖密度梯度离心从大鼠脑中制备了几种膜组分。大多数组分在前30 - 60秒内以与时间呈线性的速率快速摄取³⁶Cl⁻，然后速率逐渐减慢。线粒体组分的摄取速率最低。氧硫胺部分抑制了所有组分对³⁶Cl⁻的摄取。在P2（粗突触体组分）中，氧硫胺使初始摄取速率降低了32%，表观Ki为1.5 mM。硫胺和氨丙啉作为抑制剂的效果较差。4,4'-二异硫氰基芪-2,2'-二磺酸（0.1 - 1 mM）抑制³⁶Cl⁻摄取40 - 50%。当该化合物浓度≥5×10⁻⁴ M时，氧硫胺变得无效。γ-氨基丁酸（0.1 mM）增加了³⁶Cl⁻摄取，并且如预期的那样，该效应被苦味毒拮抗，但未被氧硫胺拮抗。³⁶Cl⁻摄取速率在很大程度上不依赖于外部氯离子浓度，并且不受布美他尼的影响，也不受用胆碱替代外部钠离子的影响。综上所述，这些数据表明，对氧硫胺敏感的³⁶Cl⁻内流基本上是扩散性的，与γ-氨基丁酸受体或钠钾氯同向转运无关。用钾部分替代外部钠离子或用0.1 mM藜芦碱处理（两者均应导致膜去极化）分别使³⁶Cl⁻摄取增加50%和30%；氧硫胺的抑制作用也相应增强。（摘要截断于250字）
