Characterization and subcellular targeting of GCaMP-type genetically-encoded calcium indicators.
Genetically-encoded calcium indicators (GECIs) hold the promise of monitoring [Ca(2+)] in selected populations of neurons and in specific cellular compartments. Relating GECI fluorescence to neuronal activity requires quantitative characterization. We have characterized a promising new genetically-encoded calcium indicator-GCaMP2-in mammalian pyramidal neurons. Fluorescence changes in response to single action potentials (17+/-10% DeltaF/F [mean+/-SD]) could be detected in some, but not all, neurons. Trains of high-frequency action potentials yielded robust responses (302+/-50% for trains of 40 action potentials at 83 Hz). Responses were similar in acute brain slices from in utero electroporated mice, indicating that long-term expression did not interfere with GCaMP2 function. Membrane-targeted versions of GCaMP2 did not yield larger signals than their non-targeted counterparts. We further targeted GCaMP2 to dendritic spines to monitor Ca(2+) accumulations evoked by activation of synaptic NMDA receptors. We observed robust DeltaF/F responses (range: 37%-264%) to single spine uncaging stimuli that were correlated with NMDA receptor currents measured through a somatic patch pipette. One major drawback of GCaMP2 was its low baseline fluorescence. Our results show that GCaMP2 is improved from the previous versions of GCaMP and may be suited to detect bursts of high-frequency action potentials and synaptic currents in vivo.
Mao
Tianyi
T
Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, United States of America.
O'Connor
Daniel H
DH
Scheuss
Volker
V
Nakai
Junichi
J
Svoboda
Karel
K
D000200:Action Potentials; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D006624:Hippocampus / Q000378:metabolism / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D051381:Rats; D013347:Subcellular Fractions / Q000378:metabolism*
D002118:Calcium