Supplementary Figure 5: Distinct readouts of Purkinje cell activity for motor performance and motor learning

The effects of Purkinje cell activation on the induction of motor learning could be dissociated from its immediate effects on motor performance. (a) Examples illustrating the variation across mice in the effects of Purkinje cell stimulation on eye movement performance during training. In different mice, Purkinje cell stimulation during the contraversive phase of the vestibular stimulus could produce an immediate decrease (left, purple) or increase (right, orange) in eye movement amplitude, compared with the VOR response in the absence of Purkinje cell stimulation (grey). In both animals, 30-min of training with Purkinje cell activation during the contraversive phase of the vestibular stimulus induced a learned increase in the VOR, as measured in the absence of Purkinje cell stimulation (corresponding color symbols in panel b). (b) Within a given experiment, there was no significant correlation between the immediate effect of Purkinje cell stimulation on the eye movement performance during training and the learning it induced (R₍₂₅₎ = −0.17, P = 0.42, Pearson correlation; n = 25 mice). Each point represents data from an individual experiment in a different mouse. Learning was measured as the percent change in VOR amplitude at the end of training compared to pre-training, tested in the absence of Purkinje cell stimulation. The effect on performance was calculated as the percent change in eye movement amplitude observed immediately, when Purkinje cells were stimulated during the vestibular stimulus, as compared with responses made to the vestibular stimulus alone before training. The immediately evoked eye movements could increase (n = 9, right), decrease (n = 10, left), or have no significant effect (n = 6, points lying on vertical axis, including 4 mice with evoked vertical eye movements) on the amplitude of the on-going VOR during training. This variable immediate effect on VOR performance mirrors the heterogeneity of the Purkinje cell population within the flocculus, observed in single unit recordings. Variable expression levels of ChR2 and/or the placement of the optical fiber within the flocculus could differentially activate subpopulations of Purkinje cells driving ipsiversive or contraversive eye movements to yield an immediate increase or decrease in eye movement, or no change if the two populations were activated equally. Despite variable immediate effects on the on-going eye movement performance, there was a remarkably consistent effect of Purkinje cell activation on VOR-increase learning (points above solid horizontal axis, which shows mean for vestibular-alone control). Notably, the immediate effect of Purkinje cell stimulation on eye movement performance was similar at the end versus the beginning of training (1st 5-min block versus 6th 5-min block, t₍₂₄₎= 1.978, P = 0.06, paired t-test, n = 25; data not shown), suggesting that the effectiveness of optogenetic stimulation and Purkinje cell excitability were similar throughout training. (c) When Purkinje cell stimulation was paired with an ipsiversive vestibular stimulus, there was no significant effect on learning, and there was no significant correlation between the effect of Purkinje cell stimulation on the immediate eye movement performance and learning (R₍₂₄₎ = −0.26, P = 0.22, Pearson correlation; n = 25 mice). (d) Unsupported model: the results in panels b and c indicate that the effect of Purkinje cell activity on learning is not a secondary consequence of the eye movements present during training. (e) Our data support the model that there are independent read outs of the Purkinje cell activity for the control of movement on different time scales: immediate performance versus learning. Different subpopulations of Purkinje cells may make different contributions to eye movement performance, but their contribution to learning appears to be more uniform.