High-Speed SICM for the Visualization of Nanoscale Dynamic Structural Changes in Hippocampal Neurons

Abstract

Dynamic reassembly of the cytoskeleton and structural changes represented by dendritic spines, cargo transport, and synapse formation are closely related to memory. However, the visualization of the nanoscale topography is challenging because of the diffraction limit of optical microscopy. Scanning ion conductance microscopy (SICM) is an effective tool for visualizing the nanoscale topography changes of the cell surface without labeling. The temporal resolution of SICM is a critical issue of live-cell time-lapse imaging. Here, we developed a new scanning method, automation region of interest (AR)-mode SICM, to select the next imaging region by predicting the location of a cell, thus improving the scanning speed of time-lapse imaging. The newly developed algorithm reduced the scanning time by half. The time-lapse images provided not only novel information about nanoscale structural changes but also quantitative information on the dendritic spine and synaptic bouton volume changes and formation process of the neural network that are closely related to memory. Furthermore, translocation of plasmalemmal precursor vesicles (ppvs), for which fluorescent labeling has not been established, were also visualized along with the rearrangement of the cytoskeleton at the growth cone.

Topography and Permeability Analyses of Vasculature‐on‐a‐Chip Using Scanning Probe Microscopies

“…SICM with nanopipette overcomes the diffraction limit (≈200 nm) and provides nanometer-scale topographical imaging, [31] although the resolutions in this study were relatively low: 2.3 μm (64 pixel × 64 pixel for 150 μm × 150 μm images) or 1.2 μm (128 pixel × 128 pixel for 150 μm × 150 μm images) owing to the acquisition pixel sizes. Because the SECM and SICM measurements themselves did not damage the cell integrity, the analytical system has the potential to monitor spatiotemporal changes in the cell function in the same chip.…”

Ion Conductance-Based Perfusability Assay of Vascular Vessel Models in Microfluidic Devices

“…The detection mechanism is based on the increase in the electrical resistance of the pore in response to the excluded volume resulting from the introduced objects when applying electrical voltages. In scanning ion conductance microscopy, the ion conductance at a nanocapillary is used to measure the height of a sample [19,20]. In the present study, ion currents between channels separated by endothelial monolayers were monitored to evaluate their perfusion during vessel formation.…”

Micropipet-Based Navigation in a Microvascular Model for Imaging Endothelial Cell Topography Using Scanning Ion Conductance Microscopy

“…To widen the range of SICM applications, the hardware and scanning algorithm for SICM have been developed. For example, researchers have strived to develop SICM systems that are capable of speedy observations. − Others have measured different parameters, such as pH, , surface charge, , ion transport, and specific molecular, and cell-cell junctions, , which can be obtained by combining SICM with scanning electrochemical microscopy, − potentiometric-SICM, or a multibarrel probe format . As noted above, the development of high-speed and multiobservation SICM systems has been progressing steadily.…”