Theta-patterned tactile stimulation modifies deep cerebellar nuclei neurons responsiveness in vivo
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1
University of Pavia, Dept. of Brain and Behavioral Sciences, Italy
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2
Brain Connectivity Center, C. Mondino National Neurological Institute, Italy
Introduction
Several forms of synaptic plasticity have been described in the cerebellar network in vitro, but how plasticity may be induced in vivo remains poorly explored. Sensory tactile stimuli organized in theta patterns have been reported to induce long-term changes in cerebellar granule cells (Roggeri et al., 2008), Purkinje cells and molecular layer interneurons (Ramakrishnan et al., 2016) in vivo. Deep cerebellar nuclei (DCN) neurons are known to respond to low frequency sensory stimulation through typical discharge patterns reflecting the inhibitory and excitatory inputs converging onto these nuclei, provided by Purkinje cells and mossy fibers respectively (Rowland and Jaeger, 2008). Nevertheless, whether and how DCN are able to modify their discharges following theta-patterned sensory stimulation remains unexplored. Herein, we addressed this issue performing single-unit recordings in vivo, from the medial nucleus of anesthetized mice. Our results provide the first evidence that DCN neurons are indeed able to modify their discharge properties following sensory stimulation in vivo, completing the picture of the theta sensory stimulation (TSS) impact on cerebellar neurons discharge in vivo.
Materials and Method
Multiple single-unit recordings were performed from the medial nucleus of C57BL/6 mice (31,4±0.5 days old; n=20) under urethane anesthesia. Recording electrodes were located over the vermis (-7.8 AP, +0,50 ML from bregma), ipsilateral to the stimulus source and lowered perpendicularly to the surface at a depth of 2002±68 µm (n=20). DCN neurons were identified by recording depth, spontaneous activity, stimulus-evoked responses, electrical lesion of the recording site and histological sections.
Tactile sensory stimulation was performed using air-puffs (100 pulses- 30 ms- 30-60 psi) delivered to the mouse lips or whisker pad to evoke the neuronal responses. DCN neurons responses were revealed in real-time by making peri-stimulus time histogram (PSTH) during the on-going experiment. Following 5 minutes of spontaneous activity recording, air-puffs were delivered at 0.5 Hz to reveal the stimulus-evoked responses. Once the responsive neuron was detected, control stimuli at 0.5 Hz were delivered for 20 min. Then, a theta sensory stimulation (TSS: 100 air-puffs at 4 Hz) was delivered, followed by post-induction control at 0.5 Hz for at least 40 minutes.
Recordings were analyzed offline using custom-written software in MATLAB and Excel. PSTHs constructed with 5 or 15 ms bin widths and raster plots of 300 trials were used for the analysis. Spike-related changes of the neuronal responses were evaluated from the corresponding PSTHs. Spike-related changes of the responses after TSS over than ± 10% were considered for potentiation or depression. Statistical comparisons were carried out using the unpaired Student’s t-test unless otherwise stated or one-way ANOVA in Origin software. Data in the text are reported as mean ± SEM.
Results
Tactile stimulation evoked 3 typical patterns of peaks and pauses in the peri-stimulus-time-histograms (PSTHs): (1) Pure inhibition (pause occurring at 20.23±4.50 ms; duration 28.82±11.42 ms; n=5); (2) excitation followed by inhibition (short-latency peak at 14.10±4.53 ms; duration 13.65±4.54 ms; n=7; Figure 1A); (3) excitation following inhibition (long-latency rebound peak at 88.34±24.26 ms, duration 17.28±3.42 ms; n=4). We have also observed pure excitation (only short-latency peak; n=2), in these cases the onset of the peak responses (13.10±2.15ms) was similar to that of the above mentioned cells with short-latency excitation followed by inhibition. Strikingly, when the inhibition followed the short-latency excitation, the onset of the pause was delayed, compared to the onset of the pause observed in pure inhibition responses (62.59±7.14 ms, duration 37.33±8.56ms, p˂0.00002). In some cells, showing short-latency excitation responses, we observed a recurrent long-latency rebound excitation that occurred every 205.6±48.5 ms for 1350.4±265.3 ms (n=2), or a second peak following the first one (second peak delay to the first peak: 16.4±4.2 ms; duration= 16.4±3.6ms; n=2). The TSS protocol induced significant amplitude changes in DCN neurons responses, though lasting for no longer than 30 minutes: increase of peak or pause amplitudes in 10 neurons (peak: 41.2±17.6%, n=5, p˂0.01; pause: 28.4±10.1%, n=5, p˂0.003; Figure 2A); decrease of peak or pause amplitudes in 8 neurons (peak: -18.5±5.2%, n=4, p˂0.02; pause: -20.3±4.6%, n=4, p˂0.005; Figure 2B). No significant amplitude changes were observed in 2 neurons. Interestingly, we did find a positive correlation between the variation of peak and pause amplitudes after TSS, in cells showing short-latency excitation followed by inhibition, with a tendency of the pauses amplitude to increase or decrease according with the corresponding variation of the peaks amplitude (and/or viceversa) (R2=0.60, p˂0.02, n=6).
Discussion and Conclusions
In in vivo urethane anesthetized mice, facial tactile stimulation evokes typical patterns of peaks and/or pauses responses in the PSTHs, which likely reflect the different combinations of excitatory and inhibitory synaptic inputs they may receive. The average latency of the inhibitory responses was well in line with the signal transmitted through granule cell-Purkinje cell pathway to DCN. The onset of the short-latency excitation well-matched with the delay of the signals known to be transmitted to DCN through the trigeminal pathway. The long-latency rebound excitation following the inhibition was likely due to disinhibition of DCN discharge caused by a pause in Purkinje cells activity, which is known to determine a rebound increase in DCN neurons firing (Dykstra et al., 2016). Sensory stimuli organized in theta-patterns (TSS) proved able to induce spike-related amplitude changes in DCN responses, showing transient but significant increases or decreases of their discharge. In particular, when both short-latency excitation (mossy fibers driven) and inhibition (Purkinje cells driven) coexisted in the same neuron, a positive correlation was revealed in these two changes of neuronal response. These results suggest an interplay between excitatory and inhibitory synaptic inputs onto DCN neurons that is likely to affect the direction of the TSS-induced changes. Taken together these results suggest that theta-pattern stimulation engages multiple discharge modifications distributed along Purkinje cell-DCN and mossy fiber-DCN pathways. Excitatory and inhibitory plasticity impinging on nuclear neurons could act synergistically to modify processing and integration of sensory information, playing a crucial role in shaping the cerebellar output.
Acknowledgements
This work was supported by: European Union grant Human Brain Project (HBP-604102) to ED. The authors declare no competing financial interests.
References
Dykstra, S., Engbers, J. D. T., Bartoletti, M., Turner, R.W. (2016). Determinants of rebound burst responses in rat cerebellar nuclear neurons to physiological stimuli. J Physiol. 594(4):985-1003. doi: 10.1113/JP271894.
Ramakrishnan, K. B., Voges, K., De Propris, L., De Zeeuw, C. I., D’Angelo, E. (2016). Tactile stimulation evokes long-lasting potentiation of Purkinje cell discharge in vivo. Front Cell Neurosci. 10:36. doi: 10.3389/fncel.2016.00036.
Roggeri, L., Rivieccio, B., Rossi, P., D’Angelo, E. (2008). Tactile stimulation evokes long-term synaptic plasticity in the granular layer of cerebellum. J Neurosci. 28(25):6354-9. doi: 10.1523/JNEUROSCI.5709-07.2008.
Rowland, N.C., Jaeger, D. (2008). Responses to tactile stimulation in deep cerebellar nucleus neurons result from recurrent activation in multiple pathways. J. Neurophysiol. 2, 704-717. doi: 10.1152/jn.01100.2007.
Keywords:
Deep cerebellar nuclei,
Cerebellum,
in vivo electrophysiology,
sensory stimulation,
in vivo plasticity
Conference:
The Cerebellum inside out: cells, circuits and functions
, ERICE (Trapani), Italy, 1 Dec - 5 Dec, 2016.
Presentation Type:
poster
Topic:
Cellular & Molecular Neuroscience
Citation:
Moscato
L,
Mapelli
L,
De Propris
L,
Tritto
S and
D‘Angelo
E
(2019). Theta-patterned tactile stimulation modifies deep cerebellar nuclei neurons responsiveness in vivo.
Conference Abstract:
The Cerebellum inside out: cells, circuits and functions
.
doi: 10.3389/conf.fncel.2017.37.000022
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Received:
30 Nov 2016;
Published Online:
25 Jan 2019.
*
Correspondence:
Miss. Letizia Moscato, University of Pavia, Dept. of Brain and Behavioral Sciences, Pavia, Italy, letizia.moscato01@universitadipavia.it
Dr. Lisa Mapelli, University of Pavia, Dept. of Brain and Behavioral Sciences, Pavia, Italy, lisa.mapelli@unipv.it
Prof. Egidio D‘Angelo, University of Pavia, Dept. of Brain and Behavioral Sciences, Pavia, Italy, egidiougo.dangelo@unipv.it