Spontaneous activity during dual recordings of Purkinje cells and ventral roots.

(A) Left, dorsal view of larval zebrafish head illustrating the location of the cerebellum (Cb, arrow). In this image, cerebellar Purkinje cells are fluorescently labeled by Archaerhodopsin-3 (white). Dotted line, outline of brain. ot: optic tectum. L, lateral; C, caudal. Middle, a Purkinje cell filled with Alexa Fluor 488. Right, schematic of the preparation. (B) Sample ventral root (vr) recording showing episodic bouts of fictive swimming (compressed time base, black) and cyclical motor bursts (expanded time base, grey). (C) Schematic of the zebrafish cerebellum. Pkj: Purkinje cell. EC: eurydendroid cell; GC: granule cell; cf: climbing fiber; mf: mossy fibers; pf: parallel fibers; Cb: cerebellum. (D) Sample voltage-clamp recording (holding potential = −60 mV, all voltage-clamp records) of cfEPSC and pfEPSCs (inset). (E) Sample voltage-clamp recording (top) and simultaneous vr recording (bottom). Box, episode of spontaneous swimming, expanded in (H). (F) Sample current-clamp recording of complex and simple spikes (top) and pfEPSPs (bottom). (G) Sample current-clamp recording of Purkinje cell spikes and simultaneous vr recording. Box, episode of spontaneous swimming, expanded in (I). Recordings in (E) and (G) are from the same cell. (H) Higher gain voltage-clamp and vr recording from (E) of clustered parallel fiber EPSCs during spontaneous swimming. (I) Higher gain current-clamp and vr recording from (G) of a parallel fiber driven long-lasting depolarization and simple spikes during spontaneous swimming. Dotted line, inter-spike potential (−56 mV) to illustrate depolarization. (J) Distribution of conductances associated with all complex spikes (cxs), simple spikes (ss), and parallel fiber EPSPs (pfEPSPs) included in the study. Absolute conductances (bin width, cxs and ss = 0.02 nS; pfEPSP = 0.01 nS). (K) As in (J) but conductances normalized to the mean cxs conductance in each cell (bin width = 0.005).

Cerebellar associative learning.

(A) Sample ventral root recordings during training, illustrating the emergence and persistence of conditional responses (CRs) over time, as well as unconditional responses (URs). Trial numbers as indicated. Blue step, conditional stimulus (CS); purple step, unconditional stimulus (US). (B) Percentage of trials with a CR over all 70 trials. Bars are means; markers are data from individual fish; ns, not significant. F_(2,49)=0.97. (C) Percentage of trials with a CR per 10-trial block for learner and non-learner groups from fish in (B) trained with 2 s CS. F_(6,18)=4.92. (D) Left side view of exposed zebrafish brain before (top) and after (bottom) ablation of the cerebellum (arrow). V, ventral; C, caudal. Right, sample ventral root recordings of swimming evoked by a high-contrast visual stimulus (top) or tactile stimulus (bottom) after cerebellar ablation. (E) Changes in CR properties from first 10 CR trials in learner fish. UR data plotted for comparison. Left, swimming latency (relative to CS or US onset), CR: F_(9,81)=3.80, UR: F_(3.98,81) = 0.78, p=0.5. Middle, number of bursts per swim response, CR: F_(9,81)=2.8, UR: F_(9,81)=1.60, p=0.13. Right, frequency of bursts, CR: F_(9,81)=2.84, UR: F_(3.47,81) = 1.82, p=0.16.