Relationship between spiking and 5K signal in vivo. Relationship between spiking and 5K signal in vivo. A, Visually evoked 5K response (top) and simultaneously recorded spikes (bottom) in a layer 2/3 pyramidal cell in V1. Arrow, Putative single spike-induced signal. B, 5K responses (top: gray, individual trials; purple, average of 5 trials) and corresponding spike raster (middle) and peristimulus time histogram (bottom) during the presentation of eight oriented grating stimuli. C, Peak GCaMP5K response during 2 s visual stimulation as a function of spike rate. D, Peak GCaMP5K response plotted against spike rate for nine cells. E, Average GCaMP5K response to 1, 2, and 3 APs within 200 ms search windows. Gray traces are mean ± SEM (n = 225, 81, and 21 for 1, 2, and 3 APs, respectively). F, Spike detection efficiency. G, Peak ΔF/F response to 1, 2, and 3 APs. H, Single exponential model (yellow trace) and nonlinear model (green trace) fit of the GCaMP5K signal (gray) from the simultaneously recorded spike response (black trace, bottom). Arrows, Underestimation of large events. Arrowheads, Overestimation of small events by the single exponential model. I, Trial-to-trial variability of GCaMP5K (coefficient of variation) during repeated presentation of preferred stimuli calculated using different measures. (**p = 0.0012; n.s., nonsignificant; n = 7 cells). J, Trial-to-trial variability of fluorescent responses at preferred orientation quantified for all visually responsive neurons, for all four calcium indicators. Jasper Akerboom et al. J. Neurosci. 2012;32:13819-13840 ©2012 by Society for Neuroscience