Mulligan S J, Davison I, Delaney K R
Department of Biological Sciences, Simon Fraser University, B.C., V5A 1S6, Burnaby, Canada.
Neuroscience. 2001;104(1):137-51. doi: 10.1016/s0306-4522(01)00057-4.
Dextran-conjugated Ca(2+) indicators were injected into the accessory olfactory bulb of frogs in vivo to selectively fill presynaptic terminals of mitral cells at their termination in the ipsilateral amygdala. After one to three days of uptake and transport, the forebrain hemisphere anterior to the tectum was removed and maintained in vitro for simultaneous electrophysiological and optical measurements. Ca(2+) influx into these terminals was compared to synaptic transmission between mitral cells and amygdala neurons under conditions of reduced Ca(2+) influx resulting from reduced extracellular [Ca(2+)], blockade of N- and P/Q-type channels, and application of the cholinergic agonist carbachol. Reducing extracellular [Ca(2+)] had a non-linear effect on release; release was proportional to Ca(2+) influx raised to the power of approximately 3.6, as observed at numerous other synapses. The N-type Ca(2+) channel blocker, omega-conotoxin-GVIA (1 microM), blocked 77% of Ca(2+) influx and 88% of the postsynaptic field potential. The P/Q-type Ca(2+) channel blocker, omega-agatoxin-IVA (200 nM), blocked 19% of Ca(2+) influx and 25% of the postsynaptic field, while the two toxins combined to block 92% of Ca(2+) influx and 97% of the postsynaptic field. The relationship between toxin blockade of Ca(2+) influx and synaptic transmission was therefore only slightly non-linear; release was proportional to Ca(2+) influx raised to the power approximately 1.4. Carbachol (100 microM) acting via muscarinic receptors had no effect on the afferent volley, but rapidly and reversibly reduced Ca(2+) influx through both N- and P/Q-type channels by 51% and postsynaptic responses by 78%, i.e. release was proportional to Ca(2+) raised to the power approximately 2.5. The weak dependence of release on changes in Ca(2+) when channel toxins block channels suggests little overlap between Ca(2+) microdomains from channels supporting release or substantial segregation of channel subtypes between terminals. The proportionately greater reduction of transmission by muscarinic receptors compared to Ca(2+) channel toxins suggests that they directly affect the release machinery in addition to reducing Ca(2+) influx.
将葡聚糖结合的钙离子指示剂注入青蛙体内的副嗅球，以选择性地填充同侧杏仁核中二尖瓣细胞终末的突触前终末。在摄取和运输一到三天后，切除中脑前方的前脑半球并在体外维持，用于同时进行电生理和光学测量。在细胞外钙离子浓度降低、N型和P/Q型通道被阻断以及应用胆碱能激动剂卡巴胆碱导致钙离子内流减少的情况下，比较这些终末的钙离子内流与二尖瓣细胞和杏仁核神经元之间的突触传递。降低细胞外钙离子浓度对释放有非线性影响；释放与钙离子内流的约3.6次方成正比，这在许多其他突触中也有观察到。N型钙离子通道阻滞剂ω-芋螺毒素-GVIA（1微摩尔）阻断了77%的钙离子内流和88%的突触后场电位。P/Q型钙离子通道阻滞剂ω-阿加毒素-IVA（200纳摩尔）阻断了19%的钙离子内流和25%的突触后场，而两种毒素联合阻断了92%的钙离子内流和97%的突触后场。因此，毒素对钙离子内流的阻断与突触传递之间的关系只是略微非线性；释放与钙离子内流的约1.4次方成正比。通过毒蕈碱受体起作用的卡巴胆碱（100微摩尔）对传入冲动没有影响，但迅速且可逆地使通过N型和P/Q型通道的钙离子内流减少51%，使突触后反应减少78%，即释放与钙离子的约2.5次方成正比。当通道毒素阻断通道时，释放对钙离子变化的弱依赖性表明，支持释放的通道的钙离子微区之间几乎没有重叠，或者终末之间通道亚型存在大量分离。与钙离子通道毒素相比，毒蕈碱受体对传递的抑制作用更大，这表明它们除了减少钙离子内流外，还直接影响释放机制。
