Cerebellar Roles in Self-Timing for Sub- and Supra-Second Intervals

Abstract

Previous studies suggest that the cerebellum and basal ganglia are involved in sub-second and supra-second timing, respectively. To test this hypothesis at the cellular level, we examined the activity of single neurons in the cerebellar dentate nucleus in monkeys performing the oculomotor version of the self-timing task. Animals were trained to report the passage of time of 400, 600, 1200, or 2400 ms following a visual cue by making self-initiated memory-guided saccades. We found a sizeable preparatory neuronal activity before self-timed saccades across delay intervals, while the time course of activity correlated with the trial-by-trial variation of saccade latency in different ways depending on the length of the delay intervals. For the shorter delay intervals, the ramping up of neuronal firing rate started just after the visual cue and the rate of rise of neuronal activity correlated with saccade timing. In contrast, for the longest delay (2400 ms), the preparatory activity started late during the delay period, and its onset time correlated with self-timed saccade latency. Because electrical microstimulation applied to the recording sites during saccade preparation advanced self-timed but not reactive saccades, regardless of their directions, the signals in the cerebellum may have a causal role in self-timing. We suggest that the cerebellum may regulate timing in both sub-second and supra-second ranges, although its relative contribution might be greater for sub-second than for supra-second time intervals.SIGNIFICANCE STATEMENT How we decide the timing of self-initiated movement is a fundamental question. According to the prevailing hypothesis, the cerebellum plays a role in monitoring sub-second timing, whereas the basal ganglia are important for supra-second timing. To verify this, we explored neuronal signals in the monkey cerebellum while animals reported the passage of time in the range 400-2400 ms by making eye movements. Contrary to our expectations, we found that neurons in the cerebellar dentate nucleus exhibited a similar preparatory activity for both sub-second and supra-second intervals, and that electrical simulation advanced self-timed saccades in both conditions. We suggest that the cerebellum plays a causal role in the fine adjustment of self-timing in a larger time range than previously thought.

Keywords: cognitive function; extracellular recording; internal time; oculomotor; primate; time reproduction.

Figure 1.

Behavioral paradigms. A, B, Sequence of events on the display (A) and their time courses (B). In the conventional memory saccade task, monkeys made a saccade in response to the FP offset that occurred 400, 600, 1200, or 2400 ms following the cue offset. In the self-timed memory saccade task, animals were required to report the end of the mandatory delay interval (400, 600, 1200, or 2400 ms) following the cue by making a self-initiated saccade without any immediate trigger. The mandatory interval was constant in each block of 10 trials. Note that the color of the FP differed between the two tasks. In the self-timed task, an audiovisual feedback was presented 100 ms after the mandatory interval. The amount of reward was determined according to the difference in time between saccade and the mandatory delay interval (Δt). C, Trial order in a block. Forty self-timed saccade trials (10 trials for each mandatory delay interval, black dots) were followed by eight conventional memory-guided saccade trials (black Xs). Note that saccade direction (arrow) alternated every self-timed saccade trial, but not for the conventional memory saccade trials.

Figure 2.

Recording sites of monkey P. A, A photograph of a coronal section at the level 8 mm posterior to the interaural line (P8). The track of 30 gauge injection cannula, which was inserted after all recording sessions, is visible at P8L7 (arrowheads). Most neurons were recorded from the penetration at the track location. D, Dentate nucleus; AI, anterior interpositus nucleus; PI, posterior interpositus nucleus; F, fastigial nucleus. B, Locations of task-related neurons. Some data points (red dots) are horizontally jittered for presentation. The brackets indicate the lateral position relative to the midline (L7–L8).

Figure 3.

Behavioral data. A, Latency distribution of self-timed saccades for 62 recording sessions in two monkeys. Different colors represent the data for different mandatory intervals, which are indicated by inverted triangles. B, Distribution of saccade timing relative to the end of the mandatory delay interval (Δt) for each monkey. Timing of the audiovisual feedback is indicated by a solid vertical line (100 ms following the mandatory delay). The bottom panel plots the amount of reward relative to that in the conventional memory saccade task as a function of Δt.

Figure 4.

Neuronal activity of a representative neuron during the two saccade paradigms. Eye position traces and spike data during the self-timed task (top panels) and the conventional memory saccade task (bottom panels) are shown. Data are aligned with either the cue offset (left) or saccade initiation (right). Trials are sorted according to saccade latency for each condition. Times of the cue onset, cue offset, saccade initiation, and the end of mandatory delay interval are overlaid on the raster plot. Different colors indicate the data for different mandatory intervals. Trials with yellow dots were unrewarded and were excluded when computing the spike density. Note the absence of ramp-up of neuronal activity in the conventional memory saccade trials, which are presented in a block containing different delay intervals (see Materials and Methods; Fig. 1C).

Figure 5.

Time courses of the population activity. A, Population activity for 45 neurons that exhibited a ramp-up of firing rate during the self-timed task. B, Population activity for 12 neurons that exhibited a ramp-down of firing rate. Note that the ramp-up neurons showed no firing modulation around the time of saccades in the conventional memory saccade task (A), while the ramp-down neurons exhibited a transient activity (B).

Figure 6.

Effects of saccade direction and the mandatory delay interval on the magnitude of neuronal activity before self-timed saccades. A, Comparison of presaccadic activity (150–50 ms before saccade initiation, subtracted from the baseline activity) between contraversive and ipsiversive self-timed saccades. Sixty-three (68%) neurons exhibited a weak but significant directional modulation (unpaired t test, p < 0.05; filled symbols). There was no directional bias in the population (paired t test, p = 0.09; regression slope, 0.93; mean ± SD = 39 ± 66 spikes/s and 36 ± 64 spikes/s for ipsiversive and contraversive saccades, respectively). Note that some neurons (n = 12) exhibited decreased activity. Two outliers ([–107, –85], [–102, –95]) have been omitted from the scatter plot, but contribute to the histogram. B, Median and interquartile range of the population activity for different mandatory delay intervals. Neurons with ramping up of activity (n = 45) are plotted separately from those with ramping down of activity (n = 12).

Figure 7.

Time courses of population activity of ramp-up neurons for different saccade latencies. A, For each neuron and mandatory delay interval, the data were divided into six groups according to saccade latencies. Data were aligned to either the cue offset (left, end of horizontal black bar) or saccade initiation (right) and were then averaged for each of the six groups. For each panel, the right traces are shifted in time so that the times of data alignment are placed at the means of saccade latency relative to the cue offset. Vertical black lines indicate the end of the mandatory delay interval. Crosses in different colors indicate the means and SDs of saccade latency for different groups. Twenty-one of 66 neurons were examined for only three mandatory delay intervals. B, The time scale is normalized for the mandatory delay interval to visualize the different time courses of neuronal activity for short intervals.

Figure 8.

Time courses of population activity of ramp-down neurons for different self-timed saccade latencies. Conventions are the same as in Figure 7.

Figure 9.

Correlation between the time course of ramping activity and self-timing. A, Sample regression lines for three sets of six data points (dots) of the population activity of ramp-up neurons. Data for the 600 ms mandatory delay interval were divided into 12 groups according to saccade latency, but only the data for the 2nd (red), 6th (orange), and 12th (purple) groups are displayed. The onset of the preparatory activity (triangles) was measured as the intersection of the regression line with the baseline firing rate (black horizontal line). B, Correlation between the rate of rise of neuronal activity and saccade latency. The regression slopes measured as in A are plotted against the normalized saccade latency. Data points with black circles indicate the data shown in A. Different colors indicate the data for different mandatory intervals. C, Relationship between the times of activity onset and saccade latency. Note that the regression line for the 2400 ms delay (blue) is almost parallel to the black solid line with a unity slope, while that for the 600 ms delay (cyan) is nearly horizontal. D, Summary of correlation coefficients (Pearson's r) between the slope of ramp-up activity and saccade latency. Red circles indicate the data derived from the analysis shown in B. Filled symbols denote significant correlation (p < 0.05). Box–whisker plots show the median, quartiles, and range of the results of the bootstrap analysis. The horizontal thick lines indicate the median values that differed significantly from zero (p < 0.05). E, Regression slope measured between the onset of neuronal activity and saccade latency. Red circles show the data derived from the analysis in C. F, G, Correlation coefficients and regression slopes for ramp-down neurons. In G, the whiskers are truncated for the shorter delay intervals ([–3.7, 2.8], [–2.3, 2.0], and [–2.1, 2.1] for 400, 600, and 1200 ms, respectively).

Figure 10.

Effects of electrical microstimulation on saccade latency. A, Eye position traces in trials with and without electrical stimulation. Left, Vertical solid and dashed lines indicate the times of audiovisual feedback and the termination of the 600 ms mandatory delay interval, respectively. Right, The vertical dashed line indicates the FP offset in the conventional memory saccade task. Only ipsiversive saccade trials with a 600 ms mandatory delay interval are shown. Bars in different colors indicate the timing of electrical stimulation (100 μA, 200 ms in duration). Inverted triangles indicate the median latency. Filled triangles indicate the data with a significant stimulation effect (Wilcoxon rank sum test, p < 0.05). B, Distribution of ipsiversive saccade latency with and without electrical stimulation delivered to the same site as in A. For self-timed saccades (top), different colors represent different mandatory delay intervals. The timing of the feedback signal is indicated by vertical solid lines, while the end of the mandatory interval is shown by vertical dashed lines. For the conventional memory saccade trials (bottom), different colors indicate different stimulation timings, whereas the distributions for the two timings almost overlap. Vertical dashed lines denote the FP offset that occurred either 400, 600, 1200, or 2400 ms following the cue offset.

Figure 11.

Quantitative summary of the stimulation effects on saccade latency. A, Comparison of median saccade latencies relative to the mandatory delay interval between the trials with and without electrical stimulation. Different colors represent different delay intervals. Filled symbols indicate data with a significant difference (Wilcoxon rank sum test, p < 0.05). B, The means and 95% confidence intervals of changes in median latencies during stimulation of 19 sites. Note that the stimulation effects were greater for self-timed saccades (red) than for conventional memory saccades (blue and yellow).

Figure 12.

Possible role of the cerebellum in the fine adjustment of self-timing. A, Time courses of population activity for different mandatory delay intervals, replotted from Figure 5. B, Possible sequence of neuronal activity with time in the sub-second and supra-second ranges.