膜去极化和氨甲酰胆碱刺激完整神经末梢中的磷脂酰肌醇周转。
Membrane depolarization and carbamoylcholine stimulate phosphatidylinositol turnover in intact nerve terminals.
作者信息
Audigier S M, Wang J K, Greengard P
机构信息
Laboratory of Molecular and Cellular Neuroscience, Rockefeller University, New York, NY 10021.
出版信息
Proc Natl Acad Sci U S A. 1988 Apr;85(8):2859-63. doi: 10.1073/pnas.85.8.2859.
Synaptosomes, purified from rat cerebral cortex, were prelabeled with [3H]inositol to study phosphatidylinositol turnover in nerve terminals. Labeled synaptosomes were either depolarized with 40 mM K+ or exposed to carbamoylcholine (carbachol). K+ depolarization increased the level of inositol phosphates in a time-dependent manner. The inositol trisphosphate concentration increased rapidly and transiently, reaching maximum (250% of control) in less than 3 sec and returning to near basal levels by 30 sec. The inositol bisphosphate level also increased rapidly, but its elevated level (220% of control) was sustained during continued depolarization. The elevated level of inositol bisphosphate was reversed upon repolarization of the synaptosomes. The level of inositol monophosphate increased slowly to 120-130% of control. These effects of K+ depolarization depended on the presence of Ca2+ in the incubation medium. Carbachol stimulated the turnover of phosphatidylinositol in a dose- and time-dependent manner. The level of inositol trisphosphate increased only slightly (120-130% of control) during carbachol stimulation. The level of inositol bisphosphate increased to 210% of control, and this maximal response was seen from 15 to 60 min. Accumulation of inositol monophosphate (250% of control) was larger than that of inositol bisphosphate, but its time course was slower. Atropine and pirenzepine inhibited the carbachol effect with high affinities of 0.8 nM and 16 nM, respectively, indicating that the effect of carbachol was mediated by activation of a M1 muscarinic receptor. Incubation of synaptosomes in Ca2+-free buffer reduced the response to carbachol by 30%, and addition of EGTA abolished it. These data show that both Ca2+ influx and M1 muscarinic receptor activation stimulate phospholipase C activity in synaptosomes, suggesting that phosphatidylinositol turnover may be involved in regulating neurotransmitter release from nerve terminals.
从大鼠大脑皮层纯化得到的突触体，用[3H]肌醇进行预标记，以研究神经末梢中磷脂酰肌醇的周转情况。标记后的突触体要么用40 mM K+使其去极化，要么暴露于氨甲酰胆碱（卡巴胆碱）中。K+去极化以时间依赖的方式增加了肌醇磷酸的水平。肌醇三磷酸浓度迅速且短暂地升高，在不到3秒内达到最大值（对照的250%），并在30秒时恢复到接近基础水平。肌醇二磷酸水平也迅速升高，但其升高水平（对照的220%）在持续去极化过程中保持稳定。突触体复极化后，肌醇二磷酸升高的水平恢复正常。肌醇单磷酸水平缓慢升高至对照的120 - 130%。K+去极化的这些效应依赖于孵育介质中Ca2+的存在。卡巴胆碱以剂量和时间依赖的方式刺激磷脂酰肌醇的周转。在卡巴胆碱刺激期间，肌醇三磷酸水平仅略有升高（对照的120 - 130%）。肌醇二磷酸水平升高至对照的210%，并且在15至60分钟时出现这种最大反应。肌醇单磷酸的积累（对照的250%）大于肌醇二磷酸，但其时程较慢。阿托品和哌仑西平分别以0.8 nM和16 nM的高亲和力抑制卡巴胆碱的作用，表明卡巴胆碱的作用是通过M1毒蕈碱受体的激活介导的。在无Ca2+缓冲液中孵育突触体使对卡巴胆碱的反应降低30%，加入乙二醇双乙醚四乙酸（EGTA）则消除该反应。这些数据表明，Ca2+内流和M1毒蕈碱受体激活均刺激突触体中的磷脂酶C活性，提示磷脂酰肌醇周转可能参与调节神经末梢的神经递质释放。
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