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Comparative Study
. 2020 Jan 2;40(1):159-170.
doi: 10.1523/JNEUROSCI.0806-19.2019. Epub 2019 Nov 6.

Differential Coding Strategies in Glutamatergic and GABAergic Neurons in the Medial Cerebellar Nucleus

Orçun Orkan Özcan  1 , Xiaolu Wang  2 , Francesca Binda  1 , Kevin Dorgans  1 , Chris I De Zeeuw  2   3 , Zhenyu Gao  2 , Ad Aertsen  4   5 , Arvind Kumar  5   6 , Philippe Isope  7
Affiliations
Comparative Study

Differential Coding Strategies in Glutamatergic and GABAergic Neurons in the Medial Cerebellar Nucleus

Orçun Orkan Özcan et al. J Neurosci. .

Abstract

The cerebellum drives motor coordination and sequencing of actions at the millisecond timescale through adaptive control of cerebellar nuclear output. Cerebellar nuclei integrate high-frequency information from both the cerebellar cortex and the two main excitatory inputs of the cerebellum: the mossy fibers and the climbing fiber collaterals. However, how nuclear cells process rate and timing of inputs carried by these inputs is still debated. Here, we investigate the influence of the cerebellar cortical output, the Purkinje cells, on identified cerebellar nuclei neurons in vivo in male mice. Using transgenic mice expressing Channelrhodopsin2 specifically in Purkinje cells and tetrode recordings in the medial nucleus, we identified two main groups of neurons based on the waveform of their action potentials. These two groups of neurons coincide with glutamatergic and GABAergic neurons identified by optotagging after Chrimson expression in VGLUT2-cre and GAD-cre mice, respectively. The glutamatergic-like neurons fire at high rate and respond to both rate and timing of Purkinje cell population inputs, whereas GABAergic-like neurons only respond to the mean population firing rate of Purkinje cells at high frequencies. Moreover, synchronous activation of Purkinje cells can entrain the glutamatergic-like, but not the GABAergic-like, cells over a wide range of frequencies. Our results suggest that the downstream effect of synchronous and rhythmic Purkinje cell discharges depends on the type of cerebellar nuclei neurons targeted.SIGNIFICANCE STATEMENT Motor coordination and skilled movements are driven by the permanent discharge of neurons from the cerebellar nuclei that communicate cerebellar computation to other brain areas. Here, we set out to study how specific subtypes of cerebellar nuclear neurons of the medial nucleus are controlled by Purkinje cells, the sole output of the cerebellar cortex. We could isolate different subtypes of nuclear cell that differentially encode Purkinje cell inhibition. Purkinje cell stimulation entrains glutamatergic projection cells at their firing frequency, whereas GABAergic neurons are only inhibited. These differential coding strategies may favor temporal precision of cerebellar excitatory outputs associated with specific features of movement control while setting the global level of cerebellar activity through inhibition via rate coding mechanisms.

Keywords: Purkinje cells; cerebellar nuclei; cerebellum; electrophysiology in vivo; temporal coding.

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Figures

Figure 1.
Figure 1.
Optogenetic stimulation of PCs inhibits CN neurons in the medial nucleus. A, Diagram of the olivo-cerebellar network and experimental protocol for tetrode recordings. Recordings (dark blue) are simultaneously performed in the PC layer and the medial nucleus while an optic fiber (light blue) is positioned above the cerebellar cortex. MLI, Molecular interneurons; GoC, Golgi cells; GrC, granule cells. B, Tetrode position was reconstructed after the experiment. Left, Tetrode track monitored by DiI fluorescence (red) in a cerebellar section stained with DAPI. Right, 3D scatter plot of the coordinates of the recording sites. C, Example of multiunit recordings in the PC layer and medial CN. D, Example of interstimulus spike histogram (ISI) of PC discharge under light stimulation at 40, 65, and 115 Hz. E, Boxplot of p values for each cell from Rayleigh test calculated from the histograms of the spike latency from stimulation onset at 40, 65, and 115 Hz. p < 0.01 indicates nonuniform distribution (dotted red line). White line indicates median. Box limit indicates first (Q1) and third (Q3) quartile. Whiskers represent 1.5 x (Q3-Q1) from the edge of the box.
Figure 2.
Figure 2.
Tetrode recordings identify two distinct groups of CN neurons. A, Example of unit isolation: raw signal on the 4 recording sites of a single tetrode. Orange and blue dots represent spikes from the 2 units isolated by the PCA (Materials and Methods). B, Projection of the spikes from the 2 units against the first 2 principal components. C, Averaged spike waveforms of the 2 units on the 4 recording sites. D, Averaged waveforms for all the recorded units were classified using PCA followed by a hierarchical clustering on principal components (HCPC). Description of the measured parameters for the PCA and projection on the first two principal components (top right). Onset time, Time point where the spike waveform reached 5% of its positive peak; P. Rise slope, positive 10%–90% rise time; P. Half-width, spike duration at 50% of the positive peak; N. Peak amplitude, negative peak; Negative Peak time, time of negative peak; N. fall slope, 10%–90% fall time; N. Half-width, spike duration at 50% of the negative peak; N. Decay time, spike duration from negative peak to 10% of the negative peak; End time, time point at 5% of its negative peak. E, Ascending hierarchical tree highlighting 2 groups of action potential waveforms. F, Superimposed averaged spike waveforms of all units normalized to the maximum amplitude (left); mean of the averaged spike waveforms for the 2 groups (Group 1, red, n = 22; Group 2, blue, n = 17) (right) identified. Shaded area represents SD. G, Boxplot of firing rates, CV and CV2 for the 2 groups. Wilcoxon rank test, Firing rate: ***p = 1.4 × 10−7; CV: p = 0.37; CV2: p = 0.36.
Figure 3.
Figure 3.
Juxtacellular recordings in transgenic mice expressing Chrimson in glutamatergic or GABAergic neurons identify the same two groups of CN neurons. A, Diagram of the experimental protocol for glass pipette recordings. GLUT and GABA neurons are identified by the activation of Chrimson in neurons recorded either in VGLUT2-cre or in GAD2-cre mice, respectively, after injection of an AAV9-hSyn-FLEX-ChrimsonR-tdTomato. Boxplots of the delay of the first spike from light onset (bottom right). B, GABA and GLUT neuron identification. Red represents Chrimson-tdTomato. Green represents Gad65/67*. Blue represents DAPI. Left, GABAergic neurons can be identified as smaller cells with both Chrimson-tdTomato and Gad65/67* expression. Right, Glutamatergic neurons are Chrimson-tdTomato. C, Mean of the averaged spike waveforms for GLUT and GABA neurons recorded using glass pipettes juxtacellular recordings. Note the similarity with waveforms recorded using tetrodes. D, Boxplots of firing rates, CV and CV2, for the 2 groups: GLUT (n = 6) versus GABA (n = 11). Wilcoxon rank test firing rate: *p = 0.012; CV: p = 0.28; CV2: p = 0.42.
Figure 4.
Figure 4.
GLUT-like and GABA-like neuron firing rates are similarly affected by Purkinje cells. A, Raster plots of one example GLUT-like cell inhibited by trains (5 ms pulse length) of PC illumination at 40, 65, and 115 Hz. B, GLUT-like and GABA-like are gradually inhibited by PC illumination. Left, Mean firing rate of individual units during illumination. Right, Averaged normalized firing rate of GLUT-like and GABA-like neuron population. Error bars indicate mean ± SEM.
Figure 5.
Figure 5.
Differential temporal coding by GLUT-like and GABA-like neurons. Nuclear neuron discharges during trains of PC illumination at 40, 65, and 115 Hz. A–D, Example of GLUT-like neuron. A, Raw trace from a GLUT-like unit during illumination (blue bars) at 65 Hz. B, Histogram of the spike latency from stimulation onset. Gray bar represents the position of the subtracted stimulation artifact. C, ISI histograms at 40, 65, and 115 Hz. D, Power spectra of the unit autocorrelogram at the same frequencies. E–H, Same analyses for a GABA-like neuron. I, Boxplot of averaged power density for GLUT-like and GABA-like neurons at 40, 65, and 115 Hz. White line indicates median. Box limit indicates first (Q1) and third (Q3) quartiles. Whiskers represent 10th and 90th percentiles. Wilcoxon rank tests: ***p = 2.6 × 10−6, ***p = 3.7 × 10−5, ***p = 0.00076.
Figure 6.
Figure 6.
GLUT-like neurons can follow a wide range of frequencies. A, Example of raster plots for one GLUT-like neuron and one GABA-like neuron during train of irregular PC illuminations (5 ms pulse length; illumination times drawn according to a homogeneous Poisson process with average rate of 40 Hz). B, Histogram of the normalized IFR for the GLUT-like neuron (left) and the GABA-like neuron (right) during baseline (top, black) and during the trains of PC illuminations (bottom, green) superimposed to the IFR histogram of the stimulation (blue cityscape). Note the overlap of the IFR histogram for the GLUT-like neuron with the IFR histogram of the stimulation. KS test p, p value of the Kolmogorov–Smirnov test. C, Boxplot of the p value of the Kolmogorov–Smirnov test between IFR histogram of cerebellar neurons (for GLUT-like and GABA-like neurons) during PC stimulation and IFR histogram of the stimulation.
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References

    1. Abbasi S, Hudson AE, Maran SK, Cao Y, Abbasi A, Heck DH, Jaeger D (2017) Robust transmission of rate coding in the inhibitory Purkinje cell to cerebellar nuclei pathway in awake mice. PLoS Comput Biol 13:e1005578. 10.1371/journal.pcbi.1005578 - DOI - PMC - PubMed
    1. Aizenman CD, Linden DJ (1999) Regulation of the rebound depolarization and spontaneous firing patterns of deep nuclear neurons in slices of rat cerebellum. J Neurophysiol 82:1697–1709. 10.1152/jn.1999.82.4.1697 - DOI - PubMed
    1. Allen GI, Tsukahara N (1974) Cerebrocerebellar communication systems. Physiol Rev 54:957–1006. 10.1152/physrev.1974.54.4.957 - DOI - PubMed
    1. Alviña K, Walter JT, Kohn A, Ellis-Davies G, Khodakhah K (2008) Questioning the role of rebound firing in the cerebellum. Nat Neurosci 11:1256–1258. 10.1038/nn.2195 - DOI - PMC - PubMed
    1. Alviña K, Ellis-Davies G, Khodakhah K (2009) T-type calcium channels mediate rebound firing in intact deep cerebellar neurons. Neuroscience 158:635–641. 10.1016/j.neuroscience.2008.09.052 - DOI - PMC - PubMed

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