Baker R R, Dowdall M J, Whittaker V P
Brain Res. 1975 Dec 26;100(3):629-44. doi: 10.1016/0006-8993(75)90162-6.
(1) Crude synaptosomal fractions (P2) derived from guinea-pig cerebral cortex were incubated in the presence of 50 mM KCl in a Krebs-glucose medium. Torpedo marmorata electric organs were stimulated electrically in vivo at 5 pulses/sec for 30 min by electrodes placed on the electric lobe. Synaptic vesicles were isolated from each source and the phospholipid compositions analysed and compared with vesicles from unstimulated controls. (2) Lysophosphatidylcholine was the only lysophosphoglyceride demonstrable in the synaptic vesicles from either source and its low levels did not increase as a result of chemical or electircal stimulation. In each case there was a close similarity of the phospholipid distributions in the vesicles taken from control and stimulated samples. (3) Control experiments indicated extensive decreases in the acetylcholine content of the vesicles from the stimulated electric organ and smaller decreases in the acetylcholine content of the synaptic vesicles from stimulated crude synaptosomal fractions. These fractions were found to respire linearly in the presence of 10 mM glucose and the vesicle fractions were shown to have low levels of contaiminating membranes as judged by marker enzyme analyses. (4) Crude synaptosomal fractions from guinea-pig cerebral cortex were incubated in a Krebs-glucose medium with labelled fatty acids and [3H]glucose in the presence or absence of 50 mM KCl. Subsynaptosomal fractionation was carried out and specific radioactivities of phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine and phosphatidylinositol were determined in fractions D (synaptic vesicles), E (microsomes) and H (disrupted synaptosomes). The release of neurotransmitter did not significantly enhance the labelling of phospholipids in any of the fractions studied as compared with phospholipids from unstimulated fractions. This was found after two incubation times and using [14C]oleate, [14C]arachidonate, [3H]palmitate and [3H]glucose.
(1) 将豚鼠大脑皮层来源的粗制突触体组分(P2)在含有50 mM KCl的Krebs - 葡萄糖培养基中孵育。用电极置于电叶上,以每秒5次脉冲的频率对电鳐的电器官进行体内刺激30分钟。从每个来源分离突触小泡,并分析其磷脂组成,并与未刺激对照的小泡进行比较。(2) 溶血磷脂酰胆碱是两种来源的突触小泡中唯一可检测到的溶血甘油磷脂,其低水平不会因化学或电刺激而增加。在每种情况下,对照和刺激样品的小泡中磷脂分布都非常相似。(3) 对照实验表明,刺激的电器官的小泡中乙酰胆碱含量大幅降低,而刺激的粗制突触体组分的突触小泡中乙酰胆碱含量降低幅度较小。发现这些组分在10 mM葡萄糖存在下呈线性呼吸,通过标记酶分析判断,小泡组分的污染膜水平较低。(4) 将豚鼠大脑皮层的粗制突触体组分在含有标记脂肪酸和[3H]葡萄糖的Krebs - 葡萄糖培养基中,在有或无50 mM KCl的情况下孵育。进行亚突触体分级分离,并测定D组分(突触小泡)、E组分(微粒体)和H组分(破碎的突触体)中磷脂酰胆碱、磷脂酰乙醇胺、磷脂酰丝氨酸和磷脂酰肌醇的比放射性。与未刺激组分的磷脂相比,神经递质的释放并未显著增强所研究的任何组分中磷脂的标记。这是在两个孵育时间后,使用[14C]油酸、[14C]花生四烯酸、[3H]棕榈酸和[3H]葡萄糖后发现的。