Kuffler D P, Nicholls J, Drapeau P
J Comp Neurol. 1987 Feb 22;256(4):516-26. doi: 10.1002/cne.902560404.
An electron microscopic study has been made of chemical synapses that develop between identified nerve cells isolated from the CNS of the leech and maintained in culture. Structures resembling synapses were observed in pairs of Retzius cells and P sensory cells at which chemical transmission had been demonstrated by recording with microelectrodes. Vesicle recycling was shown by following the uptake of extracellular markers after stimulation. The membrane separation between the presynaptic Retzius cell (which is known to liberate serotonin) and the postsynaptic P cell was wider in synaptic than in extrasynaptic regions. The Retzius cell contained clusters of clear vesicles apposed to thickenings of the presynaptic membrane. These clear vesicle clusters were capped by a layer of dense core vesicles that did not contact the presynaptic membrane thickenings. Subsynaptic cisternae were found in the postsynaptic cell opposite the presynaptic membrane thickenings. Occasional slight postsynaptic membrane thickenings were seen. Extracellular material was observed within the synaptic cleft. Similar synaptic structures developed between pairs of Retzius cells in culture; even a single Retzius cell was able to form autapses upon itself. Structures resembling transmitter release sites were found in Retzius cells at a distance from any postsynaptic membranes. These are presumed to be locations for the diffuse release of transmitter. Presynaptic structures resembling release sites were never observed in P cells apposed to Retzius cells. Antibody to serotonin (5-HT) labelled with colloidal gold showed serotonin to be localized in the dense core vesicles in Retzius cells. Stimulation of pairs of Retzius and P cells by raised concentrations of K+ resulted in uptake of extracellular markers. Only Retzius cells became labelled. Ferritin was found in cisternae, in dense core vesicles, and in clear vesicles. HRP was found in cisternae and in clear vesicles. Colloidal gold was taken up by coated vesicles and was occasionally found in both clear and dense core vesicles. The uptake of extracellular markers following stimulation was blocked by high Mg++. These results show that structures develop between pairs of cells at which chemical transmission develops and that transmitter release leads to turnover of dense core and clear vesicles.
对从水蛭中枢神经系统分离并在培养中维持的已鉴定神经细胞之间形成的化学突触进行了电子显微镜研究。在Retzius细胞和P感觉细胞对中观察到了类似突触的结构，通过微电极记录已证明在这些细胞对中存在化学传递。通过追踪刺激后细胞外标记物的摄取，显示了囊泡循环。突触前Retzius细胞（已知释放5-羟色胺）与突触后P细胞之间的膜间距在突触区域比在突触外区域更宽。Retzius细胞含有与突触前膜增厚处相对的清亮囊泡簇。这些清亮囊泡簇被一层不与突触前膜增厚处接触的致密核心囊泡覆盖。在突触后细胞中，与突触前膜增厚处相对的位置发现了突触下池。偶尔可见轻微的突触后膜增厚。在突触间隙内观察到细胞外物质。培养中的Retzius细胞对之间也形成了类似的突触结构；甚至单个Retzius细胞也能自身形成自突触。在远离任何突触后膜的Retzius细胞中发现了类似递质释放位点的结构。推测这些是递质扩散释放的位置。在与Retzius细胞相对的P细胞中从未观察到类似释放位点的突触前结构。用胶体金标记的5-羟色胺抗体显示5-羟色胺定位于Retzius细胞的致密核心囊泡中。用升高的钾离子浓度刺激Retzius细胞和P细胞对导致细胞外标记物的摄取。只有Retzius细胞被标记。在池中、致密核心囊泡和清亮囊泡中发现了铁蛋白。在池中以及清亮囊泡中发现了辣根过氧化物酶。胶体金被有被小泡摄取，偶尔在清亮囊泡和致密核心囊泡中都能发现。刺激后细胞外标记物的摄取被高浓度的镁离子阻断。这些结果表明，在发生化学传递的细胞对之间形成了结构，并且递质释放导致致密核心囊泡和清亮囊泡的更新。
