Abstract
In order to understand learning and memory at the molecular level, we must uncover the mechanisms underlying synaptic plasticity. Rem2, a Ras-like GTPase from the RGK (Rad/Rem/Rem2/Gem/Kir) protein family, is an activity-regulated protein that mediates several plastic changes within hippocampal neurons, such as dendritic complexity and excitatory synapse formation, though a role for Rem2 in long-term potentiation (LTP) has not yet been reported. Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a well-studied serine-threonine kinase that has an established function in LTP induction at the Schaffer Collateral-CA1 (SC-CA1) synapse, and this contributes to learning and memory processes such as spatial learning. While the structure and function of CaMKII is well-understood, and several downstream signaling pathways during LTP have been proposed, there is little known about the negative regulation of CaMKII. Our previous work has shown that Rem2 potently inhibits CaMKII catalytic activity in an in vitro kinase assay. We also demonstrated that mutation of two key amino acid residues in the N-terminus (R79/R80) are required for this inhibition. In the following work, we sought to determine if Rem2 plays a role in CaMKII-dependent SC-CA1 LTP in the hippocampus. To investigate this question, we recorded field excitatory postsynaptic potentials (fEPSPs) in the hippocampal CA1 area, and induced LTP using tetanus stimulation. We observed enhanced LTP at the SC-CA1 synapse in the absence of Rem2; this effect is dependent on Rem2 inhibition of CaMKII. Our findings suggest that Rem2 normally functions to restrict LTP, indicating that Rem2 and CaMKII might interact in the postsynaptic density (PSD) of the synapse. To test this hypothesis, we performed 2-photon fluorescence lifetime imaging (2pFLIM) using mCherry-Rem2 and Clover-CaMKII transfected into organotypic hippocampal slices. We observe that Rem2 and CaMKII colocalize in dendritic spines during structural plasticity, establishing that Rem2 can interact with CaMKII during LTP in the hippocampus. We also undertook a series of experiments to determine the acute role for Rem2 inhibition of CaMKII using a Rem2-derived inhibitory peptide in field electrophysiology recordings. Taken together, our data lead us to propose that Rem2 normally serves as a brake on runaway synaptic potentiation via inhibition of CaMKII activity, and is recruited to the PSD during synaptic plasticity.