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Activation of CaMKII in single dendritic spines during long-term potentiation

Abstract

Calcium/calmodulin-dependent kinase II (CaMKII) plays a central part in long-term potentiation (LTP), which underlies some forms of learning and memory. Here we monitored the spatiotemporal dynamics of CaMKII activation in individual dendritic spines during LTP using two-photon fluorescence lifetime imaging microscopy, in combination with two-photon glutamate uncaging. Induction of LTP and associated spine enlargement in single spines triggered transient (1 min) CaMKII activation restricted to the stimulated spines. CaMKII in spines was specifically activated by NMDA receptors and L-type voltage-sensitive calcium channels, presumably by nanodomain Ca2+ near the channels, in response to glutamate uncaging and depolarization, respectively. The high degree of compartmentalization and channel specificity of CaMKII signalling allow stimuli-specific spatiotemporal patterns of CaMKII signalling and may be important for synapse-specificity of synaptic plasticity.

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Figure 1: Simultaneous measurements of CaMKII activation and structural plasticity in single spines using two-photon fluorescence lifetime imaging microscopy combined with two-photon glutamate uncaging.
Figure 2: Measurements of parameters that determine spine-specificity: inactivation kinetics and mobility of CaMKII.
Figure 3: Imaging CaMKII activation during LTP induction by two-photon glutamate uncaging paired with postsynaptic depolarization.
Figure 4: Differential activation of CaMKII in spines and dendrites by postsynaptic depolarization.
Figure 5: Effects of Ca 2+ chelators on CaMKII activation.

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Acknowledgements

We thank H. Murakoshi for purified mEGFP and technical advice, Y. Hayashi for complementary DNAs and shRNA, A. Wang for cultured slices, T. Zimmerman and D. Kloetzer for laboratory management, and M. Ehlers, C. Harvey, J. Lisman, M. Patterson, S. Raghavachari, K. Svoboda, R. Weinberg and H. Zhong for comments on the manuscript. This study was funded by Burroughs Wellcome Fund, Dana Foundation, Alfred P. Sloan Foundation, Autism Speaks, National Alliance of Autism Research, Whitehall Foundation, Alzheimer’s Association, National Institute of Mental Health (R01MH08004), National Science Foundation (0642000), Duke University Undergraduate Research Support Grants (S.-J.R.L.), Howard Hughes Neuroscience Forum Fellowship (S.-J.R.L.), and Ruth K. Broad Biomedical Research Foundation (E.M.S.).

Author Contributions S.-J.R.L. and R.Y. designed the experiments. R.Y. built the microscope and developed the green-Camuiα construct. S.-J.R.L. performed most of the experiments. S.-J.R.L. and Y.E.-R. performed calcium imaging. E.M.S. performed immunostaining and pull-down assay. S.-J.R.L. and R.Y. analysed the data. R.Y. wrote the paper. All authors discussed and commented on the manuscript.

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Correspondence to Ryohei Yasuda.

Supplementary information

Supplementary Information

This file contains Supplementary Notes, Supplementary References and Supplementary Figures 1-11 with Legends. (PDF 639 kb)

Supplementary Movie 1

This movie shows CaMKII activation during structural plasticity induced by 2-photon glutamate uncaging in the absence of extracellular Mg2+. The white arrowhead indicates the location of 2-photon glutamate uncaging. (AVI 6124 kb)

Supplementary Movie 2

This movie shows CaMKII activation during LTP induced by 2-photon glutamate uncaging paired with postsynaptic depolarization. The white box at the left top corner indicates the time the neuron is depolarized, and the white arrowhead indicates the location of 2-photon uncaging. (AVI 5225 kb)

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Lee, SJ., Escobedo-Lozoya, Y., Szatmari, E. et al. Activation of CaMKII in single dendritic spines during long-term potentiation. Nature 458, 299–304 (2009). https://doi.org/10.1038/nature07842

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