所長招聘セミナー

In the central nervous system, most excitatory synapses terminate on dendritic spines, tiny (~0.1 femtoliter) protrusions emanating from the dendritic surface. Ca2+ influx into spines activates signaling networks composed of tens of species of molecules to induce diverse forms of synaptic plasticity, which is thought to underlie learning and memory. To further our understanding of signaling mechanisms underlying synaptic plasticity, we have developed a technique to monitor signaling activity in single dendritic spines in slices using 2-photon fluorescence lifetime imaging (2pFLIM) in combination with new FRET sensors extensively optimized for 2pFLIM. Using this technique, we succeeded in monitoring activity of many signaling proteins, including small GTPase proteins Ras, Rac, RhoA, Cdc42 and several Rab proteins and kinases PKC, ERK and CaMKII, during long-term potentiation (LTP). These signaling proteins have distinct spatiotemporal patterns: CaMKII, Cdc42 and Rab activations are restricted to the stimulated spines. In contrary, Ras, Rac, RhoA, PKC and ERK activations spread out of the stimulated spine and diffuse along the parent dendrite over 5-50 µm. Further, we found that Ca2+ elevation in the spine, which lasts only for ~0.1 s, is relayed in several different stages. First, Ca2+ signals activates CaMKII, of which activity decays over ~10 s. Then, downstream small GTPase proteins relay this transient CaMKII signal into signals lasting 10-30 min. This rich spatiotemporal regulation must play an essential role in coordinating cellular events occurring within spines and the dendritic shaft to induce synapse-specific plasticity.