Climbing fibers, the projections from the inferior olive to the cerebellar cortex, carry sensorimotor error and clock signals that trigger motor learning by controlling cerebellar Purkinje cell synaptic plasticity and discharge. Purkinje cells target the deep cerebellar nuclei, which are the output of the cerebellum and include an inhibitory GABAergic projection to the inferior olive. This pathway identifies a potential closed loop in the olivo-cortico-nuclear network. Therefore, sets of Purkinje cells may phasically control their own climbing fiber afferents. Here, using in vitro and in vivo recordings, we describe a genetically modified mouse model that allows the specific optogenetic control of Purkinje cell discharge. Tetrode recordings in the cerebellar nuclei demonstrate that focal stimulations of Purkinje cells strongly inhibit spatially restricted sets of cerebellar nuclear neurons. Strikingly, such stimulations trigger delayed climbing-fiber input signals in the stimulated Purkinje cells. Therefore, our results demonstrate that Purkinje cells phasically control the discharge of their own olivary afferents and thus might participate in the regulation of cerebellar motor learning.motor control | olivo-cerebellar loop | complex spikes T he cerebellar cortex is involved in a wealth of functions, from the control of posture to higher cognitive processes (1-3). Purkinje cells (PCs) are key processing units of the cerebellar cortex (4): each PC receives more than 175,000 parallel fiber synaptic inputs carrying information about the ongoing sensorymotor context. It also receives a single inferior olive afferent, the climbing fiber, which triggers a complex spike (CS), modulates PC firing (5), controls synaptic input plasticity, and has been proposed to carry error and clock signals to the cerebellum (2, 4-8). PCs are grouped in multiple parasagittal microzones, each receiving projections from separate areas of the inferior olive and projecting to subregions of the cerebellar nuclei (CN) (9-12). In the CN, PCs make inhibitory contacts on excitatory neurons that project to various premotor areas and propagate cerebellar computations to the motor system. Anatomical evidence indicates that PC terminals also contact CN inhibitory neurons that target inferior olive cells (13,14). This nucleoolivary pathway is topographically organized in multiple parallel projections to the inferior olive subnuclei (15), suggesting the existence of closed olivary-cortico-nuclear loops. Therefore, the discharge of a population of PCs in a microzone might not only shape the output of the cerebellum but also control its afferent climbing-fiber signal. Previous studies have shown that stimulation of the nucleo-olivary pathway significantly reduces olivary cell firing (16-18) and that pharmacological and genetic manipulations of PCs or olivary cell activity induce reciprocal modulations of the firing rate of PCs and climbing fibers (19, 20).These results indicate that PCs may tonically modulate the nucleo-olivary pathway. However, whether...

The compartmentalization of the cerebellum into modules is often used to discuss its function. What, exactly, can be considered a module, how do they operate, can they be subdivided and do they act individually or in concert are only some of the key questions discussed in this consensus paper. Experts studying cerebellar compartmentalization give their insights on the structure and function of cerebellar modules, with the aim of providing an up-to-date review of the extensive literature on this subject. Starting with an historical perspective indicating that the basis of the modular organization is formed by matching olivocorticonuclear connectivity, this is followed by consideration of anatomical and chemical modular boundaries, revealing a relation between anatomical, chemical, and physiological borders. In addition, the question is asked what the smallest operational unit of the cerebellum might be. Furthermore, it has become clear that chemical diversity of Purkinje cells also results in diversity of information processing between cerebellar modules. An additional important consideration is the relation between modular compartmentalization and the organization of the mossy fiber system, resulting in the concept of modular plasticity. Finally, examination of cerebellar output patterns suggesting cooperation between modules and recent work on modular aspects of emotional behavior are discussed. Despite the general consensus that the cerebellum has a modular organization, many questions remain. The authors hope that this joint review will inspire future cerebellar research so that we are better able to understand how this brain structure makes its vital contribution to behavior in its most general form.

The mossy fiber (MF)-granule cell (GC) pathway conveys multiple modalities of information to the cerebellar cortex, converging on Purkinje cells (PC), the sole output of the cerebellar cortex. Recent in vivo experiments have shown that activity in GCs varies from tonic firing at a few hertz to phasic bursts Ͼ500 Hz. However, the responses of parallel fiber (PF)-PC synapses to this wide range of input frequencies are unknown, and there is controversy regarding several frequency-related parameters of transmission at this synapse. We performed recordings of unitary synapses and combined variance-mean analysis with a carefully adapted extracellular stimulation method in young and adult rats. We show that, although the probability of release at individual sites is low at physiological calcium concentration, PF-PC synapses release one or more vesicles with a probability of 0.44 at 1.5 mM [Ca 2ϩ ] e . Paired-pulse facilitation was observed over a wide range of frequencies; it renders burst inputs particularly effective and reproducible. These properties are primarily independent of synaptic weight and age. Furthermore, we show that the PF-PC synapse is able to sustain transmission at very high frequencies for tens of stimuli, as a result of accelerated vesicle replenishment and an apparent recruitment of release site vesicles, which appears to be a central mechanism of paired-pulse facilitation at this synapse. These properties ensure that PF-PC synapses possess a dynamic range enabling the temporal code of MF inputs to be transmitted reliably to the PC.

Motor coordination is supported by an array of highly organized heterogeneous modules in the cerebellum. How incoming sensorimotor information is channeled and communicated between these anatomical modules is still poorly understood. In this study, we used transgenic mice expressing GFP in specific subsets of Purkinje cells that allowed us to target a given set of cerebellar modules. Combining in vitro recordings and photostimulation, we identified stereotyped patterns of functional synaptic organization between the granule cell layer and its main targets, the Purkinje cells, Golgi cells and molecular layer interneurons. Each type of connection displayed position-specific patterns of granule cell synaptic inputs that do not strictly match with anatomical boundaries but connect distant cortical modules. Although these patterns can be adjusted by activity-dependent processes, they were found to be consistent and predictable between animals. Our results highlight the operational rules underlying communication between modules in the cerebellar cortex.DOI: http://dx.doi.org/10.7554/eLife.09862.001

Two-photon microscopy is widely used to investigate brain function across multiple spatial scales. However, measurements of neural activity are compromised by brain movement in behaving animals. Brain motion-induced artefacts are typically corrected using post-hoc processing of 2D images, but this approach is slow and does not correct for axial movements. Moreover, the deleterious effects of brain movement on high speed imaging of small regions of interest and photostimulation cannot be corrected post-hoc . To address this problem, we combined random access 3D laser scanning using an acousto-optic lens and rapid closed-loop FPGA processing to track 3D brain movement and correct motion artifacts in real-time at up to 1 kHz. Our recordings from synapses, dendrites and large neuronal populations in behaving mice and zebrafish demonstrate real-time movement corrected 3D two-photon imaging with sub-micrometer precision.

The segregation of the readily releasable pool of synaptic vesicles (RRP) in sub-pools that are differentially poised for exocytosis shapes short-term plasticity. However, the frequency-dependent mobilization of these sub-pools is poorly understood. Using slice recordings and modeling of synaptic activity at cerebellar granule cell to Purkinje cell synapses of mice, we describe two sub-pools in the RRP that can be differentially recruited upon ultrafast changes in the stimulation frequency. We show that at low-frequency stimulations, a first sub-pool is gradually silenced, leading to full blockage of synaptic transmission. Conversely, a second pool of synaptic vesicles that cannot be released by a single stimulus is recruited within milliseconds by high-frequency stimulation and support an ultrafast recovery of neurotransmitter release after low-frequency depression. This frequency-dependent mobilization or silencing of sub-pools in the RRP in terminals of granule cells may play a role in the filtering of sensorimotor information in the cerebellum.

BACKGROUND AND PURPOSEThe tail of the ventral tegmental area (tVTA), also called the rostromedial tegmental nucleus, is a newly defined brain structure and a potential control centre for dopaminergic activity. It was identified by the induction of DeltaFosB following chronic cocaine exposure. In this work, we screened 20 drugs for their ability to induce FosB/DeltaFosB in the tVTA. EXPERIMENTAL APPROACHImmunohistochemistry following systemic drug administration was used to study FosB/DeltaFosB induction in the tVTA of adult rats. Double-staining was used to determine whether dopamine or GABA neurones are involved in this induction. KEY RESULTSThe acute injection of the psychostimulant drugs cocaine, D-amphetamine, (+/-)-3,4-methylenedioxymethamphetamine (MDMA), methylphenidate or caffeine, induced the expression of FosB/DeltaFosB in the tVTA GABAergic cells. No induction was observed following exposure to ethanol, diazepam, g-hydroxybutyric acid (GHB), morphine, ketamine, phencyclidine (PCP), D 9 -tetrahydrocannabinol (THC), sodium valproic acid or gabapentin. To evaluate the role of monoamine transporters in the psychostimulant-induced expression of FosB/DeltaFosB, we tested the antidepressant drugs reboxetine, nortriptyline, fluoxetine and venlafaxine (which target the noradrenaline and/or the 5-hydroxytryptamine transporters), the 5-hydroxytryptamine releasing agent dexfenfluramine, and the dopamine transporter inhibitor GBR12909. Only GBR12909 was able to induce FosB/DeltaFosB expression in the tVTA, showing that this induction is mediated by dopamine. CONCLUSIONS AND IMPLICATIONSNewly described brain structures may help to increase our knowledge of brain function, pathology and targets for treatments. FosB/DeltaFosB induction in the tVTA is a common feature of drugs sharing psychostimulant properties but not of drugs sharing risk of abuse. Abbreviations

The balance between excitation (E) and inhibition (I) in neuronal networks controls the firing rate of principal cells through simple network organization, such as feedforward inhibitory circuits. Here, we demonstrate in male mice, that at the granule cell (GrC)-molecular layer interneuron (MLI)-Purkinje cell (PC) pathway of the cerebellar cortex, E/I balance is dynamically controlled by short-term dynamics during bursts of stimuli, shaping cerebellar output. Using a combination of electrophysiological recordings, optogenetic stimulation, and modeling, we describe the wide range of bidirectional changes in PC discharge triggered by GrC bursts, from robust excitation to complete inhibition. At high frequency (200 Hz), increasing the number of pulses in a burst (from 3 to 7) can switch a net inhibition of PC to a net excitation. Measurements of EPSCs and IPSCs during bursts and modeling showed that this feature can be explained by the interplay between short-term dynamics of the GrC-MLI-PC pathway and E/I balance impinging on PC. Our findings demonstrate that PC firing rate is highly sensitive to the duration of GrC bursts, which may define a temporal-to-rate code transformation in the cerebellar cortex. Sensorimotor information processing in the cerebellar cortex leads to the occurrence of a sequence of synaptic excitation and inhibition in Purkinje cells. Granule cells convey direct excitatory inputs and indirect inhibitory inputs to the Purkinje cells, through molecular layer interneurons, forming a feedforward inhibitory pathway. Using electrophysiological recordings, optogenetic stimulation, and mathematical modeling, we found that presynaptic short-term dynamics affect the balance between synaptic excitation and inhibition on Purkinje cells during high-frequency bursts and can reverse the sign of granule cell influence on Purkinje cell discharge when burst duration increases. We conclude that short-term dynamics may play an important role in transforming the duration of sensory inputs arriving on cerebellar granule cells into cerebellar cortical output firing rate.