Abstract
Optogenetic approaches for studying neuronal functions have proven their utility in the neurosciences. However, optogenetic tools capable of inducing synaptic plasticity at the level of single synapses have been lacking. Here, we engineered a photoactivatable (pa)CaMKII by fusing a light-sensitive domain, LOV2, to CaMKIIα. Blue light or two-photon excitation reversibly activated paCaMKII. Activation in single spines was sufficient to induce structural long-term potentiation (sLTP) in vitro and in vivo. paCaMKII activation was also sufficient for the recruitment of AMPA receptors and functional LTP in single spines. By combining paCaMKII with protein activity imaging by 2-photon FLIM-FRET, we demonstrate that paCaMKII activation in clustered spines induces robust sLTP via a mechanism that involves the actin-regulatory small GTPase, Cdc42. This optogenetic tool for dissecting the function of CaMKII activation (i.e., the sufficiency of CaMKII rather than necessity) and for manipulating synaptic plasticity will find many applications in neuroscience and other fields.
Highlights
Optogenetic approaches for studying neuronal functions have proven their utility in the neurosciences
We developed a photoactivatable calmodulin-dependent protein kinase II (CaMKII) that fuses CaMKII and the light-oxygen-voltage domain 2 (LOV2)-Jα of the plant photoreceptor, phototropin 127. By expressing this optogenetic paCaMKII in hippocampal and cortical neurons, we demonstrate that paCaMKII activation is sufficient to induce structural long-term potentiation (sLTP), recruit AMPARs into dendritic spines, and induce functional Long-term potentiation (LTP) at the single-spine level
By combining paCaMKII with twophoton fluorescence lifetime imaging microscopy-based Förster resonance energy transfer (2pFLIM-FRET) to image protein activity, we show that paCaMKII activation in a single spine robustly activates the actin-regulatory small GTPase Cdc[42] rather than RhoA. paCaMKII activation in clustered spines enhances Cdc[42] activity, and subsequent sLTP likely contributes to the molecular mechanism of clustered synaptic plasticity[28]
Summary
Optogenetic approaches for studying neuronal functions have proven their utility in the neurosciences. By combining paCaMKII with protein activity imaging by 2-photon FLIM-FRET, we demonstrate that paCaMKII activation in clustered spines induces robust sLTP via a mechanism that involves the actin-regulatory small GTPase, Cdc[42] This optogenetic tool for dissecting the function of CaMKII activation (i.e., the sufficiency of CaMKII rather than necessity) and for manipulating synaptic plasticity will find many applications in neuroscience and other fields. We developed a photoactivatable CaMKII (paCaMKII) that fuses CaMKII and the light-oxygen-voltage domain 2 (LOV2)-Jα of the plant photoreceptor, phototropin 127 By expressing this optogenetic paCaMKII in hippocampal and cortical neurons, we demonstrate that paCaMKII activation is sufficient to induce sLTP, recruit AMPARs into dendritic spines, and induce functional LTP at the single-spine level. PaCaMKII is a significant addition to the current optogenetic toolbox, and allows manipulation of synaptic plasticity and neuronal cell signaling
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