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ref: -2015 tags: PaRAC1 photoactivatable Rac1 synapse memory optogenetics 2p imaging mouse motor skill learning date: 10-30-2019 20:35 gmt revision:1 [0] [head]

PMID-26352471 Labelling and optical erasure of synaptic memory traces in the motor cortex

  • Idea: use Rac1, which has been shown to induce spine shrinkage, coupled to a light-activated domain to allow for optogenetic manipulation of active synapses.
  • PaRac1 was coupled to a deletion mutant of PSD95, PSD delta 1.2, which concentrates at the postsynaptic site, but cannot bind to postsynaptic proteins, thus minimizing the undesirable effects of PSD-95 overexpression.
    • PSD-95 is rapidly degraded by proteosomes
    • This gives spatial selectivity.
  • They then exploited the dendritic targeting element (DTE) of Arc mRNA which is selectively targeted and translated in activiated dendritic segments in response to synaptic activation in an an NMDA receptor dependent manner.
    • Thereby giving temporal selectivity.
  • Construct is then PSD-PaRac1-DTE; this was tested on hippocampal slice cultures.
  • Improved sparsity and labelling further by driving it with the Arc promoter.
  • Motor learning is impaired in Arc KO mice; hence inferred that the induction of AS-PaRac1 by the Arc promoter would enhance labeling during learning-induced potentiation.
  • Delivered construct via in-utero electroporation.
  • Observed rotarod-induced learning; the PaRac signal decayed after two days, but the spine volume persisted in spines that showed Arc / DTE hence PA labeled activity.
  • Now, since they had a good label, performed rotarod training followed by (at variable delay) light pulses to activate Rac, thereby suppressing recently-active synapses.
    • Observed both a depression of behavioral performance.
    • Controlled with a second task; could selectively impair performance on one of the tasks based on ordering/timing of light activation.
  • The localized probe also allowed them to image the synapse populations active for each task, which were largely non-overlapping.