Department of Neuroscience, University of Wisconsin - Madison, 1111 Highland Ave, Madison, WI 53705, United States.
Department of Neuroscience, University of Wisconsin - Madison, 1111 Highland Ave, Madison, WI 53705, United States.
Cell Calcium. 2022 Jul;105:102606. doi: 10.1016/j.ceca.2022.102606. Epub 2022 May 21.
Exocytosis operates through two distinct modes. Full-fusion leads to rapid expulsion of the entire content of a vesicle; kiss-and-run leads to slow and partial expulsion. These two modes have important biological consequences for endocrine regulation and synaptic transmission. Amperometry recordings of catecholamine release from chromaffin cells reveal single-vesicle fusion events corresponding to both of these modes, but classification is often difficult. This study introduces a new method of analyzing amperometry data to improve this classification. The ratio of the average amplitude to the peak amplitude differs between full-fusion and kiss-and-run, and the probability distribution of this ratio is well fitted by a double-Gaussian. Kiss-and-run events identified by this method have fusion pores with kinetic properties different from pores associated with full-fusion. They have slower transition rates and lifetime distributions indicative of irreversible transitions. The total-charge of an amperometric spike is expected to scale with vesicle volume during a full-fusion event. The cube root of this quantity should therefore scale with diameter, but the distribution of this quantity differs from the distribution of vesicle diameter seen in the electron microscope. Fusion pore lifetimes associated with full-fusion depend on vesicle size, and this makes the choice of mode size dependent. The fusion pore thus bifurcates after opening, and vesicle size influences this choice. The secretory vesicle protein synaptophysin influences the size dependence of fusion pore lifetime and the choice of release mode. Incorporating vesicle size into an analysis of release mode reconciled the kinetics of fusion pores, as well as the distributions of vesicle diameter and catecholamine content. Thus, the initial fusion pore emerges as a critical focus in endocrine regulation. By modulating the size-dependence of the mode of exocytosis, changes in the molecular makeup of the exocytotic apparatus can impact the shape and size of an amperometric event, and the speed and composition of secretion.
胞吐作用通过两种不同的模式进行。完全融合导致囊泡的整个内容物迅速排出；吻-跑模式导致缓慢和部分排出。这两种模式对内分泌调节和突触传递具有重要的生物学后果。儿茶酚胺从嗜铬细胞释放的安培记录显示出与这两种模式相对应的单个囊泡融合事件，但分类通常很困难。本研究介绍了一种新的分析安培数据的方法来改进这种分类。平均幅度与峰值幅度之间的比值在完全融合和吻跑之间有所不同，并且该比值的概率分布很好地符合双高斯分布。通过这种方法识别的吻跑事件具有与完全融合相关的融合孔不同的动力学特性。它们具有较慢的转变率和寿命分布，表明不可逆转变。在完全融合事件中，安培峰的总电荷量预计与囊泡体积成正比。因此，该数量的立方根应与直径成比例，但该数量的分布与电子显微镜中观察到的囊泡直径的分布不同。与完全融合相关的融合孔寿命取决于囊泡大小，因此模式大小的选择是相关的。融合孔在打开后分叉，囊泡大小影响这种选择。突触小体蛋白 synaptophysin 影响融合孔寿命的尺寸依赖性和释放模式的选择。将囊泡大小纳入释放模式分析中，协调了融合孔的动力学以及囊泡直径和儿茶酚胺含量的分布。因此，初始融合孔成为内分泌调节的关键焦点。通过调节胞吐作用模式的尺寸依赖性，外泌体装置的分子组成变化可以影响安培事件的形状和大小，以及分泌的速度和组成。
