Dolenšek Jurij, Pohorec Viljem, Skelin Klemen Maša, Gosak Marko, Stožer Andraž
Faculty of Medicine, University of Maribor, Maribor, Slovenia.
Faculty of Natural Sciences and Mathematics, University of Maribor, Maribor, Slovenia.
Acta Physiol (Oxf). 2025 Feb;241(2):e14261. doi: 10.1111/apha.14261.
The crucial steps in beta cell stimulus-secretion coupling upon stimulation with glucose are oscillatory changes in metabolism, membrane potential, intracellular calcium concentration, and exocytosis. The changes in membrane potential consist of bursts of spikes, with silent phases between them being dominated by membrane repolarization and absence of spikes. Assessing intra- and intercellular coupling at the multicellular level is possible with ever-increasing detail, but our current ability to simultaneously resolve spikes from many beta cells remains limited to double-impalement electrophysiological recordings.
Since multicellular calcium imaging of spikes would enable a better understanding of coupling between changes in membrane potential and calcium concentration in beta cell collectives, we set out to design an appropriate methodological approach.
Combining the acute tissue slice method with ultrafast calcium imaging, we were able to resolve and quantify individual spikes within bursts at a temporal resolution of >150 Hz over prolonged periods, as well as describe their glucose-dependent properties. In addition, by simultaneous patch-clamp recordings we were able to show that calcium spikes closely follow membrane potential changes. Both bursts and spikes coordinate across islets in the form of intercellular waves, with bursts typically displaying global and spikes more local patterns.
This method and the associated findings provide additional insight into the complex signaling within beta cell networks. Once extended to tissue from diabetic animals and human donors, this approach could help us better understand the mechanistic basis of diabetes and find new molecular targets.
在葡萄糖刺激下，β细胞刺激-分泌偶联的关键步骤包括代谢、膜电位、细胞内钙浓度和胞吐作用的振荡变化。膜电位的变化由一系列尖峰组成，其间的静息期以膜复极化和无尖峰为主。在多细胞水平上评估细胞内和细胞间的偶联可以越来越详细，但我们目前同时分辨多个β细胞尖峰的能力仍局限于双电极电生理记录。
由于对尖峰进行多细胞钙成像能够更好地理解β细胞群体中膜电位变化与钙浓度之间的偶联，我们着手设计一种合适的方法。
将急性组织切片法与超快钙成像相结合，我们能够在长时间内以大于150Hz的时间分辨率分辨和量化爆发中的单个尖峰，并描述其葡萄糖依赖性特性。此外，通过同时进行膜片钳记录，我们能够表明钙尖峰紧密跟随膜电位变化。爆发和尖峰都以细胞间波的形式在胰岛中协调，爆发通常表现出全局模式，而尖峰表现出更多的局部模式。
这种方法及相关发现为β细胞网络内的复杂信号传导提供了更多见解。一旦扩展到糖尿病动物和人类供体的组织，这种方法可以帮助我们更好地理解糖尿病的机制基础并找到新的分子靶点。
