Regulation of transmitter release from retinal neurons with synaptic ribbons

双极细胞是视网膜上的二级神经元，对光刺激的反应具有梯度电位。在它们的轴突末端有突触带，突触囊泡附着在突触带上。为了阐明递质（谷氨酸）释放的调控机制，我们应用膜片钳技术对从金鱼视网膜中酶解分离的Mb1双极细胞轴突末端进行了观察。我们同时测量了与突触囊泡融合相关的Ca电流和膜电容的变化。同时，利用谷氨酸受体丰富的神经元对释放的谷氨酸进行电生理监测。得到以下结论：1)谷氨酸在Ca电流开始后以最小延迟-1 ms释放。2) Ca电流终止后300ms内发射机释放停止。3)至少存在两种突触囊泡池；立即释放池和备用池。4)双极细胞中蛋白激酶C的激活通过增加储备池的大小来增强递质释放。5)立即释放池和蓄水池可能分别对应于突触带底排和上排的突触囊泡。此外，在蝾螈视网膜切片制备中，我们通过同时电压箝制一对连接的双极细胞及其突触后神经元（无突细胞或神经节细胞）来研究谷氨酸能突触传递的特性。得到以下结论：6)只有非NMDA受体可以在双相细胞释放位点下方的突触后区域表达，而NMDA受体则稍微远离释放位点。7)非NMDA和NMDA受体共同合作，将双极细胞的分级光反应按比例传递给突触后神经元。

Bipolar cells, the second order neurons in the retina, respond to light stimulation with graded potentials. In their axon terminals there are synaptic ribbons, to which synaptic vesicles are attached. To elucidate the regulatory mechanisms of transmitter (glutamate) release, the patch-clamp technique was applied to the axon terminals of Mb1 bipolar cells that were enzymatically isolated from the goldfish retina. We measured simultaneously the Ca current and membrane capacitance changes associated with fusion of synaptic vesicles. At the same time, the released glutamate was monitored electrophysiologically with glutamate receptor-rich neurons. The following conclusions were obtained. 1) Glutamate is released with a minimal delay of-1 ms after the onset of the Ca current. 2) Transmitter release is stopped within 300 ms after the termination of the Ca current. 3) There are at least two kinds of synaptic vesicle pools ; an immediately releasable pool and a reserve pool. 4) Activation of protein kinase C in bipolar cells potentiates transmitter release by increasing the size of the reserve pool. 5) The immediately releasable pool and the reservoir pool may correspond to synaptic vesicles attached to the bottom row of the synaptic ribbons and those to the upper rows, respectively. Furthermore, we investigated properties of glutamatergic synaptic transmission by simultaneously voltage-clamping a pair of connected bipolar cells and their post-synaptic neurons (amacrine or ganglion cells) in the newt retinal slice preparation. The following conclusions were obtained. 6) Only non-NMDA receptors may be expressed in the postsynaptic regions immediately beneath the release sites of bipolar cells, whereas NMDA receptors slightly away from the release sites. 7) Both non-NMDA and NMDA receptors cooperate to transfer the graded photoresponses of bipolar cells proportionally to post-synaptic neurons.