Division of Physiology, Medical University Innsbruck, Fritz-Pregl-Str. 3, 6020 Innsbruck, Austria.
Neuroscience. 2013 Dec 3;253:330-40. doi: 10.1016/j.neuroscience.2013.08.052. Epub 2013 Sep 5.
Neurotransmitter release probability is related by high power to the local concentration of calcium in presynaptic terminals, which in turn is controlled by voltage-gated calcium channels. P/Q- and N-type channels trigger synaptic transmission in the majority of neurons of the central nervous system. However, whether and under which conditions both channel types act cooperatively or independently is still insufficiently understood. Previous studies suggested either a dominance of N- or P/Q-type channels, or a synergistic action of both channels, depending on the experimental paradigms. Thus, to provide insight into the properties of neurotransmitter release in cultured mouse hippocampal neurons, we used quantitative analysis of FM dye release from presynaptic boutons induced by high potassium membrane depolarization. Increasing extracellular potassium concentrations revealed a sigmoid dependence of FM dye release to the stimulation strength. Individual and combined application of the P/Q- and N-type channel-specific blockers ω-agatoxin-IVA and ω-conotoxin-GVIA, respectively, allowed us to specifically isolate the contribution of both channel types to release triggered with 40 mM KCl. Analysis of the release kinetics and the fractional release amplitude demonstrate that, whereas in only 15% of the synapses release depended exclusively on P/Q-type channels, the majority of synapses (85%) contained both N- and P/Q-type channels. Nevertheless, the kinetics of FM dye release in synapses containing both channel types was determined by the P/Q-type channels. Together, our data suggest a more direct coupling of P/Q-type channels to synaptic release compared to N-type channels, which may explain the high prevalence of neurological P/Q-type channelopathies.
神经递质释放概率与突触前末梢中钙离子的局部浓度密切相关，而钙离子的浓度又受电压门控钙通道的控制。P/Q 型和 N 型通道在中枢神经系统的大多数神经元中触发突触传递。然而，这两种通道类型是否以及在何种条件下协同或独立发挥作用，仍了解不足。以前的研究表明，这两种通道类型要么是 N 型通道占主导地位，要么是两种通道协同作用，这取决于实验方案。因此，为了深入了解培养的小鼠海马神经元中神经递质释放的特性，我们使用定量分析方法，研究了由高钾膜去极化诱导的 FM 染料从前突末梢中的释放。增加细胞外钾浓度显示 FM 染料释放对刺激强度呈 S 形依赖性。单独和联合应用 P/Q 型和 N 型通道特异性阻断剂 ω-芋螺毒素-GVIA 和 ω-蟾毒蛙毒素-GVIA，分别允许我们专门分离两种通道类型对 40mM KCl 引发的释放的贡献。释放动力学和分数释放幅度的分析表明，虽然在只有 15%的突触中释放仅依赖于 P/Q 型通道，但大多数突触（85%）包含 N 型和 P/Q 型通道。然而，包含两种通道类型的突触中 FM 染料释放的动力学由 P/Q 型通道决定。总的来说，我们的数据表明 P/Q 型通道与突触释放的直接耦合程度高于 N 型通道，这可能解释了神经 P/Q 型通道病的高发率。
