Electrophysiological and mechanical activity of cells triggered and traced on the level of single receptor-ligand interactions

信号通路的复杂性需要几种方法和策略来推断所涉及的分子之间的相互联系和细胞的可测量反应。在大多数实验中，信号受体是由可溶性配体刺激的。但有时，即使是对单个感受器的刺激也会导致强大的细胞活动(例如，单个光子可以触发整个感光细胞)。涉及膜片钳的方法在电刺激方面是精确的，并成功地跟踪信号过程的动态，因为它们中的大多数包括离子通道活动。神经元受体通常直接耦合到离子通道。关注细胞机械活动的方法仍然不太受欢迎。在这里，我们希望利用原子力显微镜的纳米机械精度对细胞上的某些分子施加刺激，并通过同时膜片钳结合单通道记录来记录细胞对刺激的反应。这种独特的设置记录了细胞在高采样率和高分辨率下对给定刺激的电生理和机械反应。该方法首次针对单个血管紧张素II多肽与其G蛋白偶联受体途径相互作用以及单个谷氨酸分子与其离子型受体相互作用的相互作用。

The complexity of signaling pathways calls for several approaches and strategies to deduce the interconnections between the involved molecules and a measured reaction of the cell. In a majority of experiments the signaling receptors are stimulated by soluble ligands. But sometimes even a stimulus on a single receptor results in a strong cellular activity (e.g. a single photon can trigger a whole photoreceptor cell). Approaches that involve patch-clamp are precise in electrical stimulation and successful in tracing the dynamics of signaling processes since most of them include ion channel activity. Neuronal receptors often directly couple to ion channels. Approaches that focus on mechanical activity of cells still are less favored. Here we want to utilize the nano mechanical precision of an AFM to apply a stimulus to certain molecules on a cell and to record the cellular movement as a response to a stimulus combined with single channel recordings by simultaneous patch-clamp. This unique setup records electro-physiological and mechanical reactions of a cell to a given stimulus at high sampling rate and resolution. The interplay of mechanical and electrophysiological reactions of individual angiotensin II peptides interacting with its G-protein coupled receptor pathway and of individual glutamate molecules interacting with its ionotropic receptor are targeted in this approach for the first time.