@article{13202,
  abstract     = {Phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) plays an essential role in neuronal activities through interaction with various proteins involved in signaling at membranes. However, the distribution pattern of PI(4,5)P2 and the association with these proteins on the neuronal cell membranes remain elusive. In this study, we established a method for visualizing PI(4,5)P2 by SDS-digested freeze-fracture replica labeling (SDS-FRL) to investigate the quantitative nanoscale distribution of PI(4,5)P2 in cryo-fixed brain. We demonstrate that PI(4,5)P2 forms tiny clusters with a mean size of ∼1000 nm2 rather than randomly distributed in cerebellar neuronal membranes in male C57BL/6J mice. These clusters show preferential accumulation in specific membrane compartments of different cell types, in particular, in Purkinje cell (PC) spines and granule cell (GC) presynaptic active zones. Furthermore, we revealed extensive association of PI(4,5)P2 with CaV2.1 and GIRK3 across different membrane compartments, whereas its association with mGluR1α was compartment specific. These results suggest that our SDS-FRL method provides valuable insights into the physiological functions of PI(4,5)P2 in neurons.},
  author       = {Eguchi, Kohgaku and Le Monnier, Elodie and Shigemoto, Ryuichi},
  issn         = {1529-2401},
  journal      = {The Journal of Neuroscience},
  number       = {23},
  pages        = {4197--4216},
  publisher    = {Society for Neuroscience},
  title        = {{Nanoscale phosphoinositide distribution on cell membranes of mouse cerebellar neurons}},
  doi          = {10.1523/JNEUROSCI.1514-22.2023},
  volume       = {43},
  year         = {2023},
}

@article{7665,
  abstract     = {Acute brain slice preparation is a powerful experimental model for investigating the characteristics of synaptic function in the brain. Although brain tissue is usually cut at ice-cold temperature (CT) to facilitate slicing and avoid neuronal damage, exposure to CT causes molecular and architectural changes of synapses. To address these issues, we investigated ultrastructural and electrophysiological features of synapses in mouse acute cerebellar slices prepared at ice-cold and physiological temperature (PT). In the slices prepared at CT, we found significant spine loss and reconstruction, synaptic vesicle rearrangement and decrease in synaptic proteins, all of which were not detected in slices prepared at PT. Consistent with these structural findings, slices prepared at PT showed higher release probability. Furthermore, preparation at PT allows electrophysiological recording immediately after slicing resulting in higher detectability of long-term depression (LTD) after motor learning compared with that at CT. These results indicate substantial advantages of the slice preparation at PT for investigating synaptic functions in different physiological conditions.},
  author       = {Eguchi, Kohgaku and Velicky, Philipp and Hollergschwandtner, Elena and Itakura, Makoto and Fukazawa, Yugo and Danzl, Johann G and Shigemoto, Ryuichi},
  issn         = {16625102},
  journal      = {Frontiers in Cellular Neuroscience},
  publisher    = {Frontiers Media},
  title        = {{Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions}},
  doi          = {10.3389/fncel.2020.00063},
  volume       = {14},
  year         = {2020},
}