Extended Data Fig. 2: Varying intensity and duration of Purkinje cell optogenetic stimulation to dissociate stimulation onset, simple spike modulation, and evoked blinks.
From: Climbing fibers provide essential instructive signals for associative learning
a, Experimental scheme for pairing a visual CS with optogenetic Purkinje cell-US. b,c, Example electrophysiological traces (b) and histogram (c) from a Purkinje cell with identified SSpk (grey dots) and CSpk (red) in response to 300 ms medium intensity Pkj-ChR2 laser stimulation in the cerebellar cortex (Pkj-ChR2-Ctx-med; corresponds to Fig. 3e-h). d, SSpk rate (top) and CSpk rate (bottom) pre-, during- and post- laser epochs for the recordings in (b,c) and Fig. 2d (N = 3 units from 2 mice). SSpks: spont. vs during laser, P = 0. 4.8e-7***, paired t-test; spont. vs after laser, P = 0.0003***, paired t-test. CSpks: spont. vs during laser, P = 0.22n.s., paired t-test; spont. vs after laser, P = 0.87n.s., paired t-test. Each orange circle and line represents a unit, linked through conditions; the black solid circles and line represent the average. e, Average eyelid closures ±SEM (shadows) to 300 ms Pkj-ChR2-Ctx medium intensity laser stimulation (N = 2 mice, shading represents laser stimulation). f-j Higher intensity Pkj-ChR2-Ctx laser stimulation was used to evoke a pause in simple spikes and a short-latency evoked blink at stimulus onset (corresponds to Fig. 3i-l). This stimulation elicited electrophysiological signatures of complex spike-like events at laser onset48 and, with a longer and more variable delay, at laser offset (likely due to rebound from release of Purkinje cell inhibition via olivo-cerebellar loop. f,g, SSpk and CSpk traces and histograms (N = 2 units from 2 mice). h, Same as in d, but for high intensity Pkj-ChR2-Ctx laser stimulation. SSpks: spont. vs during laser, P = 9.9e-17***, paired t-test; spont. vs after laser, P = 9.86e-20***, paired t-test. CSpks: spont. vs during laser, P = 1.335e-15***, paired t-test; spont. vs after laser, P = 2.45e-14***, paired t-test. i, SSpk (grey) and spontaneous CSpk (red) waveforms. Yellow trace represents complex spike-like events at laser stimulation onset; note the correspondence to spontaneous CSpk waveforms (red). j, Pkj-ChR2 laser stimulation at higher intensities yields a blink at stimulus onset (N = 3 mice, shading represents laser stimulation, ±SEM in shadows).
