Piezo1 是一种 Ca2 渗透性机械激活离子通道，参与各种生理过程和细胞对机械刺激的反应。 Piezo1生物物理特性的研究对于理解其功能和调控机制具有重要意义。拉伸激活是一种用于刺激质膜中 Piezo1 活性的常规方法，具有许多显着的局限性，使精确的单通道分析变得复杂。在这里，我们的目的是确定天然 Piezo1 的孔特性，特别是使用 Piezo1 特异性化学激动剂 Yoda1 检查人髓性白血病 K562 细胞中生理相关信号二价离子（钙和镁）的渗透。通过结合响应 Yoda1 的低噪声单电流膜片钳记录 Piezo1 活性，我们确定了天然 Piezo1 在各种离子条件下的单通道特性。全细胞测定使我们能够在细胞外溶液中没有其他可渗透阳离子的情况下直接测量 Ca2 或 Mg2 离子携带的 Piezo1 单电流；在不同浓度的 Mg2 下估计的单一电导值显示出强烈的饱和效应。膜片钳数据与荧光成像相结合，清楚地证明了人白血病 K562 细胞中 Ca2 和 Mg2 通过天然 Piezo1 通道进入。在准生理条件下检测到通过 Piezo1 的 Mg2 流入，从而表明 Piezo1 通道可能为 Mg2 离子运输提供生理相关途径，并有助于调节 Mg2 依赖性细胞内信号传导。© 2024 Wiley periodicals LLC。

Piezo1 is a Ca2+-permeable mechanically activated ion channel that is involved in various physiological processes and cellular responses to mechanical stimuli. The study of biophysical characteristics of Piezo1 is important for understanding the mechanisms of its function and regulation. Stretch activation, a routine approach that is applied to stimulate Piezo1 activity in the plasma membrane, has a number of significant limitations that complicate precise single-channel analysis. Here, we aimed to determine pore properties of native Piezo1, specifically to examine permeation for physiologically relevant signaling divalent ions (calcium and magnesium) in human myeloid leukemia K562 cells using Piezo1-specific chemical agonist, Yoda1. Using a combination of low-noise single-current patch-clamp recordings of Piezo1 activity in response to Yoda1, we have determined single-channel characteristics of native Piezo1 under various ionic conditions. Whole-cell assay allowed us to directly measure Piezo1 single currents carried by Ca2+ or Mg2+ ions in the absence of other permeable cations in the extracellular solutions; unitary conductance values estimated at various concentrations of Mg2+ revealed strong saturation effect. Patch clamp data complemented with fluorescent imaging clearly evidenced Ca2+ and Mg2+ entry via native Piezo1 channel in human leukemia K562 cells. Mg2+ influx via Piezo1 was detected under quasi-physiological conditions, thus showing that Piezo1 channels could potentially provide the physiological relevant pathway for Mg2+ ion transport and contribute to the regulation of Mg2+-dependent intracellular signaling.© 2024 Wiley Periodicals LLC.