Department of Biomedical Engineering, University of Connecticut, Storrs, CT 06269, USA.
Department of Physiology and Neurobiology, University of Connecticut, Storrs, CT 06269, USA.
Function (Oxf). 2023 Apr 25;4(3):zqad018. doi: 10.1093/function/zqad018. eCollection 2023.
Small-conductance calcium-activated potassium (SK) channels show a ubiquitous distribution on neurons, in both somatodendritic and axonal regions. SK channels are associated with neuronal activity regulating action potential frequency, dendritic excitability, and synaptic plasticity. Although the physiology of SK channels and the mechanisms that control their surface expression levels have been investigated extensively, little is known about what controls SK channel diffusion in the neuronal plasma membrane. This aspect is important, as the diffusion of SK channels at the surface may control their localization and proximity to calcium channels, hence increasing the likelihood of SK channel activation by calcium. In this study, we successfully investigated the diffusion of SK channels labeled with quantum dots on human embryonic kidney cells and dissociated hippocampal neurons by combining a single-particle tracking method with total internal reflection fluorescence microscopy. We observed that actin filaments interfere with SK mobility, decreasing their diffusion coefficient. We also found that during neuronal maturation, SK channel diffusion was gradually inhibited in somatodendritic compartments. Importantly, we observed that axon barriers formed at approximately days 6 and restricted the diffusion of SK channels on the axon initial segment (AIS). However, after neuron maturation, SK channels on the AIS were strongly immobilized, even after disruption of the actin network, suggesting that crowding may cause this effect. Altogether, our work provides insight into how SK channels diffuse on the neuronal plasma membrane and how actin and membrane crowding impacts SK channel diffusion.
小电导钙激活钾 (SK) 通道在神经元中具有广泛的分布，存在于树突和轴突区域。SK 通道与神经元活动有关，可调节动作电位频率、树突兴奋性和突触可塑性。尽管 SK 通道的生理学及其表面表达水平的调控机制已被广泛研究，但对于控制 SK 通道在神经元质膜中扩散的因素知之甚少。这一方面很重要，因为 SK 通道在表面的扩散可能控制其定位和与钙通道的接近程度，从而增加钙激活 SK 通道的可能性。在这项研究中，我们通过将单粒子跟踪方法与全内反射荧光显微镜相结合，成功地研究了量子点标记的 SK 通道在人胚肾细胞和分离的海马神经元中的扩散。我们观察到肌动蛋白丝干扰 SK 运动性，降低其扩散系数。我们还发现，在神经元成熟过程中，SK 通道在树突和胞体区室中的扩散逐渐受到抑制。重要的是，我们观察到大约在第 6 天时形成的轴突屏障限制了 SK 通道在轴突起始段（AIS）上的扩散。然而，在神经元成熟后，即使破坏了肌动蛋白网络，AIS 上的 SK 通道也被强烈固定，这表明拥挤可能导致这种效应。总之，我们的工作深入了解了 SK 通道如何在神经元质膜上扩散，以及肌动蛋白和膜拥挤如何影响 SK 通道的扩散。
