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This paper presents a model of learning by the cerebellar circuit. In the traditional and dominant learning model, training teaches finely graded parallel fibre synaptic weights which modify transmission to Purkinje cells and to interneurons that inhibit Purkinje cells. Following training, input in a learned pattern drives a training-modified response. The function is that the naive response to input rates is displaced by a learned one, trained under external supervision. In the proposed model, there is no weight-controlled graduated balance of excitation and inhibition of Purkinje cells. Instead, the balance has two functional states—a switch—at synaptic, whole cell and microzone level. The paper is in two parts. The first is a detailed physiological argument for the synaptic learning function. The second uses the function in a computational simulation of pattern memory. Against expectation, this generates a predictable outcome from input chaos (real-world variables). Training always forces synaptic weights away from the middle and towards the limits of the range, causing them to polarise, so that transmission is either robust or blocked. All conditions teach the same outcome, such that all learned patterns receive the same, rather than a bespoke, effect on transmission. In this model, the function of learning is gating—that is, to select patterns that trigger output merely, and not to modify output. The outcome is memory-operated gate activation which operates a two-state balance of weight-controlled transmission. Group activity of parallel fibres also simultaneously contains a second code contained in collective rates, which varies independently of the pattern code. A two-state response to the pattern code allows faithful, and graduated, control of Purkinje cell firing by the rate code, at gated times.

The cerebellum is largely conserved in its circuitry, but varies greatly in size and shape across species. The extent to which differences in cerebellar morphology is driven by changes in neuron numbers, neuron sizes or both, remains largely unknown. To determine how species variation in cerebellum size and shape is reflective of neuron sizes and numbers requires the development of a suitable comparative data set and one that can effectively separate different neuronal populations. Here, we generated the largest comparative dataset to date on neuron numbers, sizes, and volumes of cortical layers and surface area of the cerebellum across 54 bird species. Across different cerebellar sizes, the cortical layers maintained relatively constant proportions to one another and variation in cerebellum size was largely due to neuron numbers rather than neuron sizes. However, the rate at which neuron numbers increased with cerebellum size varied across Purkinje cells, granule cells, and cerebellar nuclei neurons. We also examined the relationship among neuron numbers, cerebellar surface area and cerebellar folding. Our estimate of cerebellar folding, the midsagittal foliation index, was a poor predictor of surface area and number of Purkinje cells, but surface area was the best predictor of Purkinje cell numbers. Overall, this represents the first comprehensive, quantitative analysis of cerebellar anatomy in a comparative context of any vertebrate. The extent to which these relationships occur in other vertebrates requires a similar approach and would determine whether the same scaling principles apply throughout the evolution of the cerebellum.

In classical theories of cerebellar cortex, high-dimensional sensorimotor representations are used to separate neuronal activity patterns, improving associative learning and motor performance. Recent experimental studies suggest that cerebellar granule cell (GrC) population activity is low-dimensional. To examine sensorimotor representations from the point of view of downstream Purkinje cell ‘decoders’, we used three-dimensional acousto-optic lens two-photon microscopy to record from hundreds of GrC axons. Here we show that GrC axon population activity is high dimensional and distributed with little fine-scale spatial structure during spontaneous behaviors. Moreover, distinct behavioral states are represented along orthogonal dimensions in neuronal activity space. These results suggest that the cerebellar cortex supports high-dimensional representations and segregates behavioral state-dependent computations into orthogonal subspaces, as reported in the neocortex. Our findings match the predictions of cerebellar pattern separation theories and suggest that the cerebellum and neocortex use population codes with common features, despite their vastly different circuit structures. By recording from hundreds of cerebellar granule cell axons with three-dimensional two-photon calcium imaging, Lanore et al. show that population activity is high-dimensional and that quiet wakeful and active states are orthogonally arranged in neuronal activity space.

Vigabatrin is the drug of choice in resistant epilepsy and infantile spasms. Ataxia, tremors, and abnormal gait have been frequently reported following its use indicating cerebellar involvement. This study aimed, for the first time, to investigate the involvement of necroptosis and apoptosis in the VG-induced cerebellar cell loss and the possible protective role of combined omega-3 and vitamin B12 supplementation. Fifty Sprague-Dawley adult male rats (160-200 g) were divided into equal five groups: the control group received normal saline, VG200 and VG400 groups received VG (200 mg or 400 mg/kg, respectively), VG200 + OB and VG400 + OB groups received combined VG (200 mg or 400 mg/kg, respectively), vitamin B12 (1 mg/kg), and omega-3 (1 g/kg). All medications were given daily by gavage for four weeks. Histopathological changes were examined in H&E and luxol fast blue (LFB) stained sections. Immunohistochemical staining for caspase-3 and receptor-interacting serine/threonine-protein kinase-1 (RIPK1) as well as quantitative real-time polymerase chain reaction (qRT-PCR) for myelin basic protein (MBP), caspase-3, and receptor-interacting serine/threonine-protein kinase-3 (RIPK3) genes were performed. VG caused a decrease in the granular layer thickness and Purkinje cell number, vacuolations, demyelination, suppression of MBP gene expression, and induction of caspases-3, RIPK1, and RIPK3 in a dose-related manner. Combined supplementation with B12 and omega-3 improved the cerebellar histology, increased MBP, and decreased apoptotic and necroptotic markers. In conclusion, VG-induced neuronal cell loss is dose-dependent and related to both apoptosis and necroptosis. This could either be ameliorated (in low-dose VG) or reduced (in high-dose VG) by combined supplementation with B12 and omega-3.

Spinocerebellar ataxia type 1 (SCA1) is a fatal neurodegenerative disease caused by an abnormal expansion of CAG repeats in the Ataxin1 (ATXN1) gene. SCA1 is characterized by motor deficits, cerebellar neurodegeneration, and gliosis and gene expression changes. Expression of brain-derived neurotrophic factor (BDNF), growth factor important for the survival and function of cerebellar neurons, is decreased in ATXN1[82Q] mice, the Purkinje neuron specific transgenic mouse model of SCA1. As this decrease in BDNF expression may contribute to cerebellar neurodegeneration, we tested whether delivery of extrinsic human BDNF via osmotic ALZET pumps has a beneficial effect on disease severity in this mouse model of SCA1. Additionally, to test the effects of BDNF on established and progressing cerebellar pathogenesis and motor deficits, we delivered BDNF post-symptomatically. We have found that post-symptomatic delivery of extrinsic BDNF ameliorated motor deficits and cerebellar pathology (i.e., dendritic atrophy of Purkinje cells, and astrogliosis) indicating therapeutic potential of BDNF even after the onset of symptoms in SCA1. However, BDNF did not alter Purkinje cell gene expression changes indicating that certain aspects of disease pathogenesis cannot be ameliorated/slowed down with BDNF and that combinational therapies may be needed.

Thiamine deficiency is associated with cerebellar dysfunction; however, the consequences of thiamine deficiency on the electrophysiological properties of cerebellar Purkinje cells are poorly understood. Here, we evaluated these parameters in brain slices containing cerebellar vermis. Adult mice were maintained for 12-13 days on a thiamine-free diet coupled with daily injections of pyrithiamine, an inhibitor of thiamine phosphorylation. Morphological analysis revealed a 20% reduction in Purkinje cell and nuclear volume in thiamine-deficient animals compared to feeding-matched controls, with no reduction in cell count. Under whole-cell current clamp, thiamine-deficient Purkinje cells required significantly less current injection to fire an action potential. This reduction in rheobase was not due to a change in voltage threshold. Rather, thiamine-deficient neurons presented significantly higher input resistance specifically in the voltage range just below threshold, which increases their sensitivity to current at these critical membrane potentials. In addition, thiamine deficiency caused a significant decrease in the amplitude of the action potential afterhyperpolarization, broadened the action potential, and decreased the current threshold for depolarization block. When thiamine-deficient animals were allowed to recover for 1 week on a normal diet, rheobase, threshold, action potential half-width, and depolarization block threshold were no longer different from controls. We conclude that thiamine deficiency causes significant but reversible changes to the electrophysiology properties of Purkinje cells prior to pathological morphological alterations or cell loss. Thus, the data obtained in the present study indicate that increased excitability of Purkinje cells may represent a leading indicator of cerebellar dysfunction caused by lack of thiamine.

We review advances in understanding Purkinje cell (PC) complex spike (CS) physiology that suggest increased CS synchrony underlies syndromic essential tremor (ET). We searched PubMed for papers describing factors that affect CS synchrony or cerebellar circuits potentially related to tremor. Inferior olivary (IO) neurons are electrically coupled, with the degree of coupling controlled by excitatory and GABAergic inputs. Clusters of coupled IO neurons synchronize CSs within parasagittal bands via climbing fibers (Cfs). When motor cortex is stimulated in rats at varying frequencies, whisker movement occurs at ~10 Hz, correlated with synchronous CSs, indicating that the IO/CS oscillatory rhythm gates movement frequency. Intra-IO injection of the GABAA receptor antagonist picrotoxin increases CS synchrony, increases whisker movement amplitude, and induces tremor. Harmaline and 5-HT2a receptor activation also increase IO coupling and CS synchrony and induce tremor. The hotfoot17 mouse displays features found in ET brains, including cerebellar GluRδ2 deficiency and abnormal PC Cf innervation, with IO- and PC-dependent cerebellar oscillations and tremor likely due to enhanced CS synchrony. Heightened coupling within the IO oscillator leads, through its dynamic control of CS synchrony, to increased movement amplitude and, when sufficiently intense, action tremor. Increased CS synchrony secondary to aberrant Cf innervation of multiple PCs likely also underlies hotfoot17 tremor. Deep cerebellar nucleus (DCN) hypersynchrony may occur secondary to increased CS synchrony but might also occur from PC axonal terminal sprouting during partial PC loss. Through these combined mechanisms, increased CS/DCN synchrony may plausibly underlie syndromic ET.

This commentary highlights a “cerebellar classic” by Heinrich Obersteiner (1847–1922), the founder of Vienna’s Neurological Institute. Obersteiner had a long-standing interest in the cerebellar cortex, its development, and pathology, having provided one of the early accurate descriptions of the external germinal layer (sometimes called the “marginal zone of Obersteiner” or “Obersteiner layer”). In his communication before the 81st meeting of the Society of German Natural Scientists and Physicians in Salzburg in September 1909, Obersteiner placed special emphasis on the histophysiology of the granule cell layer of the cerebellum and covered most of the fundamental elements of the cerebellar circuitry, on the basis of Ramon y Cajal’s neuronism. Those elements are discussed in a historic and a modern perspective, including some recent ideas about the role of granule cells, beyond the mere relay of sensorimotor information from mossy fibers to the Purkinje cells, in learning and cognition.

Although Krüppel-associated box domain-containing zinc-finger proteins (K-ZNFs) may be associated with sophisticated gene regulation in higher organisms, the physiological functions of most K-ZNFs remain unknown. The Zfp212 protein was highly conserved in mammals and abundant in the brain; it was mainly expressed in the cerebellum (Cb). Zfp212 (mouse homolog of human ZNF212) knockout (Zfp212-KO) mice showed a reduction in survival rate compared to wild-type mice after 20 months of age. GABAergic Purkinje cell degeneration in the Cb and aberrant locomotion were observed in adult Zfp212-KO mice. To identify genes related to the ataxia-like phenotype of Zfp212-KO mice, 39 ataxia-associated genes in the Cb were monitored. Substantial alterations in the expression of ataxin 10, protein phosphatase 2 regulatory subunit beta, protein kinase C gamma, and phospholipase D3 (Pld3) were observed. Among them, Pld3 alone was tightly regulated by Flag-tagged ZNF212 overexpression or Zfp212 knockdown in the HT22 cell line. The Cyclic Amplification and Selection of Targets assay identified the TATTTC sequence as a recognition motif of ZNF212, and these motifs occurred in both human and mouse PLD3 gene promoters. Adeno-associated virus-mediated introduction of human ZNF212 into the Cb of 3-week-old Zfp212-KO mice prevented Purkinje cell death and motor behavioral deficits. We confirmed the reduction of Zfp212 and Pld3 in the Cb of an alcohol-induced cerebellar degeneration mouse model, suggesting that the ZNF212–PLD3 relationship is important for Purkinje cell survival.

In studies on cardiac arrest (CA)/resuscitation (R) injury, Purkinje cell degeneration was described, however, with inconsistent data concerning severity and time point of manifestation. Moreover, CA/R studies paid only limited attention to inhibitory stellate interneurons. To this aim, the hypothesis that cerebellar could be relatively resilient toward CA/R because of diverse cellular defense mechanisms including interaction with stellate cells was tested. We examined rats with survival times of 6, 24, and 48 h, and 7 and 21 days in comparison with sham- and nonoperated animals. Thereby, we focused on the immunohistochemical expression of cfos, MnSOD, Bcl2, caspase 3, parvalbumin, calbindin D28 k, MAP2, IBA1, and GFAP, especially in the particular sensitivity to CA/R cerebellar lobule IX. Hippocampal CA1 degeneration was demonstrated by expression patterns of MAP2 and NeuN in combination with IBA1 and GFAP. Comparative analysis of hippocampal CA1 pyramidal cells and cerebellar Purkinje cells confirmed a relative resil-ience of Purkinje cells to CA/R. We found only a notable degeneration of Purkinje cell neuronal fiber network, which, however, not necessarily led to neuronal cell death. To induce significant Purkinje cell loss, a stronger ischemic trigger seems to be needed. As possible Purkinje cell-protecting mechanisms, we would propose: (1) activation of inhibitory stellate cells, shown by cfos, MnSOD, and Bcl2 expression, balancing out ischemia-induced excitation and inhibition of Purkinje cells; (2) translocation of the calcium-buffering system, shown by parvalbumin and calbindin D28 k expression, protecting Purkinje cells from detrimental calcium overload; (3) activation of the neuron–astrocyte cross talk, protecting Purkinje cells from over-excitation by removing potassium and neurotransmitters from the extracellular space; (4) activation of the effective and long-lasting MnSOD defense system; and (5) of the anti-apoptotic protein Bcl2 in Purkinje cells itself. Moreover, the results emphasize the limited comparability of animal CA/R studies because of the heterogeneity of the used experimental regimes.

Neurofibromatosis type 2 (NF2) patients are prone to develop glial-derived tumors in the peripheral and central nervous system (CNS). The Nf2 gene product -Merlin is not only expressed in glia, but also in neurons of the CNS, where its function still remains elusive. Here, we show that cerebellar Purkinje cells (PCs) of isoform-specific Merlin-deficient mice were innervated by smaller vGluT2-positive clusters at presynaptic terminals than those of wild-type mice. This was paralleled by a reduction in frequency and amplitude of miniature excitatory postsynaptic currents (mEPSC). On the contrary, in conditional transgenic mice in which Merlin expression was specifically ablated in PCs (L7Cre;Nf2fl/fl), we found enlarged vGluT2-positive clusters in their presynaptic buttons together with increased amplitudes of miniature postsynaptic currents. The presynaptic terminals of these PCs innervating neurons of the deep cerebellar nuclei were also enlarged. When exploring mice with Merlin-deficient granule cells (GCs) (Math1Cre;Nf2fl/fl), we found cerebellar extracts to contain higher amounts of vGluT1 present in parallel fiber terminals. In parallel, mEPSC frequency was increased in Math1Cre;Nf2fl/fl mice. On the contrary, VGluT2 clusters in cerebellar glomeruli composed of NF2-deficient presynaptic Mossy fiber terminals and NF2-deficient postsynaptic GC were reduced in size as shown for isoform-specific knockout mice. These changes in Math1Cre;Nf2fl/fl-deficient mice were paralleled by an increased activation of Rac1-Cofilin signaling which is known to impact on cytoskeletal reorganization and synapse formation. Consistent with the observed synaptic alterations in these transgenic mice, we observed altered ultrasonic vocalization, which is known to rely on proper cerebellar function. No gross morphological changes or motor coordination deficits were observed in any of these transgenic mice. We therefore conclude that Merlin does not regulate overall cerebellar development, but impacts on pre- and post-synaptic terminal organization.

The ATP/GTP-Binding Protein 1 (AGTPBP1) gene (OMIM *606830) catalyzes deglutamylation of polyglutamylated proteins, and its deficiency manifests by cerebellar ataxia and peripheral neuropathy in mice and lower motor neuron-like disease in sheep. In the mutant mice, cerebellar atrophy due to Purkinje cell degeneration is observed, likely due to increased tubulin polyglutamylation in affected brain areas. We report two unrelated individuals who presented with early onset cerebellar atrophy, developmental arrest with progressive muscle weakness, and feeding and respiratory difficulties, accompanied by severe motor neuronopathy. Whole exome sequencing followed by segregation analysis in the families and cDNA studies revealed deleterious biallelic variants in the AGTPBP1 gene. We conclude that complete loss-of-function of AGTPBP1 in humans, just like in mice and sheep, is associated with cerebellar and motor neuron disease, reminiscent of Pontocerebellar Hypoplasia Type 1 (PCH1).

One of the major impacts of spinal cord injury (SCI) is the cerebellar neurological malfunction and deformation of its sub-layers. This could be due to the enormous innervation of the spinocerebellar tract from the posterior gray horn in the spinal cord to the ipsilateral cerebellum. Although the neuroprotective role of estradiol in spinal cord (SC) injuries, as well as its ability to delay secondary cell death changes, is well-known, its effect on cerebellar layers is not fully investigated. In this study, a SCI model was achieved by injection of Kainic acid into SC of adult Male Wistar rats in order to assess the effects of SCI on the cerebellum. The animals were classified into SCI group (animals with SCI), estradiol-treated group (animals with SCI and received estradiol), control groups, and sham control group. The microscopical examination 24 h after induction of SCI revealed that KA induced the most characteristics of neurodegeneration including astrocytic propagation and microglial activation. The estradiol was injected intraperitoneally 20 min after induction of SCI, and the samples were collected at 1, 3, 7, 14, and 30 days. Histologically, the estradiol reduced the inflammatory response, enhanced the recovery of molecular, granular, and Purkinje cell layers, and therefore aided in the restoration of layer organization. These findings were also confirmed by immunohistochemical staining and gene expression profiling.

We here provide neural evidence that the cerebellar circuit can predict future inputs from present outputs, a hallmark of an internal forward model. Recent computational studies hypothesize that the cerebellum performs state prediction known as a forward model. To test the forward-model hypothesis, we analyzed activities of 94 mossy fibers (inputs to the cerebellar cortex), 83 Purkinje cells (output from the cerebellar cortex to dentate nucleus), and 73 dentate nucleus cells (cerebellar output) in the cerebro-cerebellum, all recorded from a monkey performing step-tracking movements of the right wrist. We found that the firing rates of one population could be reconstructed as a weighted linear sum of those of preceding populations. We then went on to investigate if the current outputs of the cerebellum (dentate cells) could predict the future inputs of the cerebellum (mossy fibers). The firing rates of mossy fibers at time t + t_1 could be well reconstructed from as a weighted sum of firing rates of dentate cells at time t, thereby proving that the dentate activities contained predictive information about the future inputs. The average goodness-of-fit (R^2) decreased moderately from 0.89 to 0.86 when t_1 was increased from 20 to 100 ms, hence indicating that the prediction is able to compensate the latency of sensory feedback. The linear equations derived from the firing rates resembled those of a predictor known as Kalman filter composed of prediction and filtering steps. In summary, our analysis of cerebellar activities supports the forward-model hypothesis of the cerebellum.

The current work was undertaken to evaluate the toxicologic effect of manganese chloride (MnCl2) and potential therapeutic effect of curcumin on male albino rats. Ninety-six rats were divided into three groups. MnCl2 toxicity group (I) included 32 rats were administered (MnCl2) at a dose of 5 mg/ml daily in drinking water orally for 8 weeks. Curcumin-treated group (II) included 32 rats were given (MnCl2) in the same dose and route as the first group. Each rat of this group was treated by oral administration of curcumin in a daily dose 100 mg/kg b.w. The control group (III) included 32 rats. Eight rats were randomly selected from groups I, II and III and sacrificed at (2, 4, 6, and 8 weeks) of experiment. Tissue samples from cerebellum and midbrain of all groups were collected for histopathological examination, and neuromorphometrical analysis as well as blood samples were taken for complete blood picture. The histopathological examination in group I showed Purkinje cell loss in the cerebellum with degeneration of motor neurons in midbrain. In group II, curcumin caused significant increase in number of healthy Purkinje cells in cerebellum with prevalence of healthy motor neurons and minimal vascular alterations in midbrain. The hematological results revealed that animals in group I showed a significant increase in WBCs and platelets compared to other groups, while RBCs count significantly decreased. The number of poikilocyts increased in group I compared to group II and group III. In conclusion, curcumin therapy has a powerful neuroprotective effect in Mncl2 toxicity.

The signals in cerebellar Purkinje cells sufficient to instruct motor learning have not been systematically determined. Therefore, we applied optogenetics in mice to autonomously excite Purkinje cells and measured the effect of this activity on plasticity induction and adaptive behavior. Ex vivo, excitation of channelrhodopsin-2-expressing Purkinje cells elicits dendritic Ca2+ transients with high-intensity stimuli initiating dendritic spiking that additionally contributes to the Ca2+ response. Channelrhodopsin-2-evoked Ca2+ transients potentiate co-active parallel fiber synapses; depression occurs when Ca2+ responses were enhanced by dendritic spiking. In vivo, optogenetic Purkinje cell activation drives an adaptive decrease in vestibulo-ocular reflex gain when vestibular stimuli are paired with relatively small-magnitude Purkinje cell Ca2+ responses. In contrast, pairing with large-magnitude Ca2+ responses increases vestibulo-ocular reflex gain. Optogenetically induced plasticity and motor adaptation are dependent on endocannabinoid signaling, indicating engagement of this pathway downstream of Purkinje cell Ca2+ elevation. Our results establish a causal relationship among Purkinje cell Ca2+ signal size, opposite-polarity plasticity induction, and bidirectional motor learning., Plastic reweighting of parallel fiber synaptic strength is a mechanism for the acquisition of cerebellum-dependent motor learning. Here, the authors found that optogenetic activation of PCs generates dendritic Ca2+ signals that induce plasticity in vitro and instruct learned changes to coincident eye movements in vivo.

Spinocerebellar ataxia type 1 (SCA1) is a fatal neurodegenerative disease caused by abnormal expansion of glutamine-encoding CAG repeats in the Ataxin-1 (ATXN1) gene. SCA1 is characterized by progressive motor deficits, cognitive decline, and mood changes including anxiety and depression, with longer number of repeats correlating with worse disease outcomes. While mouse models have been very useful in understanding etiology of ataxia and cognitive decline, our understanding of mood symptoms in SCA1 has lagged. It remains unclear whether anxiety or depression stem from an underlying brain pathology or as a consequence of living with an untreatable and lethal disease. To increase our understanding of the etiology of SCA1 mood alterations, we used the elevated-plus maze, sucrose preference and forced swim tests to assess mood in four different mouse lines. We found that SCA1 knock-in mice exhibit increased anxiety that correlated with the length of CAG repeats, supporting the idea that underlying brain pathology contributes to SCA1-like anxiety. Additionally, our results support the concept that increased anxiety is caused by non-cerebellar pathology, as Purkinje cell specific SCA1 transgenic mice exhibit decreased anxiety-like behavior. Regarding the molecular mechanism, partial loss of ATXN1 may play a role in anxiety, based on our results for Atxn1 haploinsufficient and null mice.

We hypothesized that early-life gut microbiota support the functional organization of neural circuitry in the brain via regulation of synaptic gene expression and modulation of microglial functionality. Germ-free mice were colonized as neonates with either a simplified human infant microbiota consortium consisting of four Bifidobacterium species, or with a complex, conventional murine microbiota. We examined the cerebellum, cortex, and hippocampus of both groups of colonized mice in addition to germ-free control mice. At postnatal day 4 (P4), conventionalized mice and Bifidobacterium-colonized mice exhibited decreased expression of synapse-promoting genes and increased markers indicative of reactive microglia in the cerebellum, cortex and hippocampus relative to germ-free mice. By P20, both conventional and Bifidobacterium-treated mice exhibited normal synaptic density and neuronal activity as measured by density of VGLUT2+ puncta and Purkinje cell firing rate respectively, in contrast to the increased synaptic density and decreased firing rate observed in germ-free mice. The conclusions from this study further reveal how bifidobacteria participate in establishing functional neural circuits. Collectively, these data indicate that neonatal microbial colonization of the gut elicits concomitant effects on the host CNS, which promote the homeostatic developmental balance of neural connections during the postnatal time period.

The cerebellum coordinates voluntary movements for balanced motor activity in a normal gravity condition. It remains unknown how hypergravity is associated with cerebellum-dependent motor behaviors and Purkinje cell’s activities. In order to investigate the relationship between gravity and cerebellar physiology, we measured AMPA-mediated fast currents and mGluR1-mediated slow currents of cerebellar Purkinje cells along with cerebellum-dependent behaviors such as the footprint and irregular ladder under a hypergravity condition. We found abnormal animal behaviors in the footprint and irregular ladder tests under hypergravity. They are correlated with decreased AMPA and mGluR1-mediated synaptic currents of Purkinje cells. These results indicate that gravity regulates the activity of Purkinje cells, thereby modulating cerebellum-dependent motor outputs.

The cerebellum is a region of the brain that plays an important role in motor control. It is classified phylogenetically into archicerebellum, paleocerebellum and neocerebellum. The Purkinje cells are lined in a row called Purkinje cell layer and it has a unique dendritic branches with many spines.The previous study reported that there is a difference of synapse density according to the lobules based on large two-dimensional data. However, recent study with high voltage electron microscopy showed there was no differences in dendritic spine density of the Purkinje cell according to its phylogenetic lobule. We analyzed Purkinje cell density in the II, VI and X lobules by stereological modules and synaptic density was estimated by double disector based on Purkinje cell density in the molecular layer of each lobule.The results showed that there was significant difference in the Purkinje cell density and synapse number according to their phylogenetic lobules. The number of Purkinje cell in a given volume was larger in the archicerebellum, but synapse density was higher in the neocerebellum.These data suggest that cellular and synaptic organization of the Purkinje cell is different according to their phylogenetic background.

We investigated the effects of lead (Pb) and ascorbic acid co-administration on rat cerebellar development. Prior to mating, rats were randomly divided into control, Pb, and Pb plus ascorbic acid (PA) groups. Pregnant rats were administered Pb in drinking water (0.3% Pb acetate), and ascorbic acid (100 mg/kg) via oral intubation until the end of the experiment. Offspring were sacrificed at postnatal day 21, the age at which the morphology of the cerebellar cortex in developing pups is similar to that of the adult brain. In the cerebellum, Pb exposure significantly reduced Purkinje cells and ascorbic acid prevented their reduction. Along with the change of the Purkinje cells, long-term Pb exposure significantly reduced the expression of the synaptic marker (synaptophysin), γ-aminobutyric acid (GABA)-synthesizing enzyme (glutamic acid decarboxylase 67), and axonal myelin basic protein while ascorbic acid co-treatment attenuated Pb-mediated reduction of these proteins in the cerebellum of pups. However, glutamatergic N-methyl-D-aspartate receptor subtype 1 (NMDAR1), anchoring postsynaptic density protein 95 (PSD95), and antioxidant superoxide dismutases (SODs) were adversely changed; Pb exposure increased the expression of NMDAR1, PSD95, and SODs while ascorbic acid co-administration attenuated Pb-mediated induction. Although further studies are required about the neurotoxicity of the Pb exposure, the results presented here suggest that developmental Pb exposure disrupted normal development of Purkinje cells by increasing glutamatergic and oxidative stress in the cerebellum. Additionally, ascorbic acid co-treatment is beneficial in attenuating prenatal and postnatal Pb exposure-induced maldevelopment of Purkinje cells in the developing cerebellum.

The disruptor of telomeric silencing 1-like (DOT1L) mediates methylation of histone H3 at position lysine 79 (H3K79). Conditional knockout of Dot1l in mouse cerebellar granule cells (Dot1l-cKOAtoh1) led to a smaller external granular layer with fewer precursors of granule neurons. Dot1l-cKOAtoh1 mice had impaired proliferation and differentiation of granular progenitors, which resulted in a smaller cerebellum. Mutant mice showed mild ataxia in motor behavior tests. In contrast, Purkinje cell-specific conditional knockout mice showed no obvious phenotype. Genome-wide transcription analysis of Dot1l-cKOAtoh1 cerebella using microarrays revealed changes in genes that function in cell cycle, cell migration, axon guidance, and metabolism. To identify direct DOT1L target genes, we used genome-wide profiling of H3K79me2 and transcriptional analysis. Analysis of differentially methylated regions (DR) and differentially expressed genes (DE) revealed in total 12 putative DOT1L target genes in Dot1l-cKOAtoh1 affecting signaling (Tnfaip8l3, B3galt5), transcription (Otx1), cell migration and axon guidance (Sema4a, Sema5a, Robo1), cholesterol and lipid metabolism (Lss, Cyp51), cell cycle (Cdkn1a), calcium-dependent cell-adhesion or exocytosis (Pcdh17, Cadps2), and unknown function (Fam174b). Dysregulated expression of these target genes might be implicated in the ataxia phenotype observed in Dot1l-cKOAtoh1. Electronic supplementary material The online version of this article (10.1007/s12035-018-1377-1) contains supplementary material, which is available to authorized users.

Cerebellar Purkinje cells arborize unique dendrites that exhibit a planar, fan shape. The dendritic branches fill the space of their receptive field with little overlap. This dendritic arrangement is well-suited to form numerous synapses with the afferent parallel fibers of the cerebellar granule cells in a non-redundant manner. Purkinje cell dendritic arbor morphology is achieved by a combination of dynamic local branch growth behaviors, including elongation, branching, and retraction. Impacting these behaviors, the self-avoidance of each branch terminal is essential to form the non-overlapping dendritic configuration. This review outlines recent advances in our understanding of the cellular and molecular mechanisms of dendrite formation during cerebellar Purkinje cell development.

Climbing fiber-driven long-term depression (LTD) of parallel fiber synapses onto cerebellar Purkinje cells has long been investigated as a putative mechanism of motor learning. We recently discovered that the rules governing the induction of LTD at these synapses vary across different regions of the cerebellum. Here, we discuss the design of LTD induction protocols in light of this heterogeneity in plasticity rules. The analytical advantages of the cerebellum provide an opportunity to develop a deeper understanding of how the specific plasticity rules at synapses support the implementation of learning.

Cerebellar Purkinje cells (PCs) and cerebellar pathways are primarily affected in many autosomal dominant cerebellar ataxias. PCs generate complex spikes (CS) in vivo when activated by climbing fiber (CF) which rise from the inferior olive. In this study, we investigated the functional state of the CF-PC circuitry in the transgenic mouse model of spinocerebellar ataxia type 2 (SCA2), a polyglutamine neurodegenerative genetic disease. In our experiments, we used an extracellular single-unit recording method to compare the PC activity pattern and the CS shape in age-matched wild-type mice and SCA2-58Q transgenic mice. We discovered no alterations in the CS properties of PCs in aging SCA2 mice. To examine the integrity of the olivocerebellar pathway, we applied harmaline, an alkaloid that acts directly on the inferior olive neurons. The pharmacological stimulation of olivocerebellar circuit by harmaline uncovered disturbances in SCA2-58Q PC activity pattern and in the complex spike shape when compared with age-matched wild-type cells. The abnormalities in the CF-PC circuitry were aggravated with age. We propose that alterations in CF-PC circuitry represent one of potential causes of ataxic symptoms in SCA2 and in other SCAs.

Zebrin II/aldolase C expression in the normal cerebellum is restricted to a Purkinje cell subset and is the canonical marker for stripes and zones. This spatial restriction has been confirmed in over 30 species of mammals, birds, fish, etc. In a transgenic mouse model in which the Neurogenin 2 gene has been disrupted (Neurog2-/-), the cerebellum is smaller than normal and Purkinje cell dendrites are disordered, but the basic zone and stripe architecture is preserved. Here, we show that in the Neurog2-/- mouse, in addition to the normal Purkinje cell expression, zebrin II is also expressed in a population of cells with a morphology characteristic of microglia. This identity was confirmed by double immunohistochemistry for zebrin II and the microglial marker, Iba1. The expression of zebrin II in cerebellar microglia is not restricted by zone or stripe or lamina. A second zone and stripe marker, PLCβ4, does not show the same ectopic expression. When microglia are compared in control vs. Neurog2-/- mice, no difference is seen in apparent number or distribution, suggesting that the ectopic zebrin II immunoreactivity in Neurog2-/- cerebellum reflects an ectopic expression rather than the invasion of a new population of microglia from the periphery. This ectopic expression of zebrin II in microglia is unique as it is not seen in numerous other models of cerebellar disruption, such as in Acp2-/- mice and in human pontocerebellar hypoplasia. The upregulation of zebrin II in microglia is thus specific to the disruption of Neurog2 downstream pathways, rather than a generic response to a cerebellar disruption.

Previous genetic fate-mapping studies have indicated that embryonic glial fibrillary acidic protein-positive (GFAP(+)) cells are multifunctional progenitor/neural stem cells that can produce astrocytes as well as neurons and oligodendrocytes throughout the adult mouse central nervous system (CNS). However, emerging evidence from recent studies indicates that GFAP(+) cells adopt different cell fates and generate different cell types in different regions. Moreover, the fate of GFAP(+) cells in the young adult mouse CNS is not well understood. In the present study, hGFAP-Cre/R26R transgenic mice were used to investigate the lineage of embryonic GFAP(+) cells in the young adult mouse CNS. At postnatal day 21, we found that GFAP(+) cells mainly generated NeuN(+) neurons in the cerebral cortex (both ventral and dorsal), hippocampus, and cerebellum. Strangely, these cells were negative for the Purkinje cell marker calbindin in the cerebellum and the neuronal marker NeuN in the thalamus. Thus, contrary to previous studies, our genetic fate-mapping revealed that the cell fate of embryonic GFAP(+) cells at the young adult stage is significantly different from that at the adult stage.

The primary output cells of the cerebellar cortex, Purkinje cells, make kinematic predictions about ongoing movements via high-frequency simple spikes, but receive sensory error information about that movement via low-frequency complex spikes (CS). How is the vector space of sensory errors encoded by this low-frequency signal? Here we measured Purkinje cell activity in the oculomotor vermis of animals during saccades, then followed the chain of events from experience of visual error, generation of CS, modulation of simple spikes, and ultimately change in motor output. We found that while error direction affected the probability of CS, error magnitude altered its temporal distribution. Production of CS changed the simple spikes on the next trial, but regardless of the actual visual error, this change biased the movement only along a vector that was parallel to the Purkinje cell's preferred error. From these results, we inferred the anatomy of a sensory-to-motor adaptive controller that transformed visual error vectors into motor-corrections.

Bone marrow-derived cells are known to infiltrate the adult brain and fuse with cerebellar Purkinje cells. Histological observations that such heterotypic cell fusion events are substantially more frequent following cerebellar injury suggest they could have a role in the protection of mature brain neurons. To date, the possibility that cell fusion can preserve or restore the structure and function of adult brain neurons has not been directly addressed; indeed, though frequently suggested, the possibility of benefit has always been rather speculative. Here we report, for the first time, that fusion of a bone marrow-derived cell with a neuron in vivo, in the mature brain, results in the formation of a spontaneously firing neuron. Notably, we also provide evidence supporting the concept that heterotypic cell fusion acts as a biological mechanism to repair pathological changes in Purkinje cell structure and electrophysiology. We induced chronic central nervous system inflammation in chimeric mice expressing bone marrow cells tagged with enhanced green fluorescent protein. Subsequent in-depth histological analysis revealed significant Purkinje cell injury. In addition, there was an increased incidence of cell fusion between bone marrow-derived cells and Purkinje cells, revealed as enhanced green fluorescent protein-expressing binucleate heterokaryons. These fused cells resembled healthy Purkinje cells in their morphology, soma size, ability to synthesize the neurotransmitter gamma-aminobutyric acid, and synaptic innervation from neighbouring cells. Extracellular recording of spontaneous firing ex vivo revealed a shift in the predominant mode of firing of non-fused Purkinje cells in the context of cerebellar inflammation. By contrast, the firing patterns of fused Purkinje cells were the same as in healthy control cerebellum, indicating that fusion of bone marrow-derived cells with Purkinje cells mitigated the effects of cell injury on electrical activity. Together, our histological and electrophysiological results provide novel fundamental insights into physiological processes by which nerve cells are protected in adult life. Electronic supplementary material The online version of this article (10.1007/s00401-018-1833-z) contains supplementary material, which is available to authorized users.

The calcium-sensing protein synaptotagmin 7 mediates facilitation that is masked by depression, but supports frequency-invariant transmission in mouse cerebellar and vestibular synapses. In most parts of the brain, synaptic strength varies as a function of neuronal firing frequency, a phenomenon known as 'short-term synaptic plasticity'. This fundamental nonlinearity in synaptic transmission has to be compensated for to achieve frequency-independent transmission in some auditory, vestibular and cerebellar circuits. Now Wade Regehr and colleagues show that synaptotagmin 7, a calcium-sensing protein, supports a short-term facilitation that increases with firing frequency and thus compensates for use-dependent depression at synapses in the cerebellum and in the vestibular system. The resulting constant synaptic strength over a wide range of firing frequencies allows for efficient encoding of neuronal firing rates. At most synapses in the brain, short-term plasticity dynamically modulates synaptic strength. Rapid frequency-dependent changes in synaptic strength have key roles in sensory adaptation, gain control and many other neural computations1,2. However, some auditory, vestibular and cerebellar synapses maintain constant strength over a wide range of firing frequencies3,4,5, and as a result efficiently encode firing rates. Despite its apparent simplicity, frequency-invariant transmission is difficult to achieve because of inherent synaptic nonlinearities6. Here we study frequency-invariant transmission at synapses from Purkinje cells to deep cerebellar nuclei and at vestibular synapses in mice. Prolonged activation of these synapses leads to initial depression, which is followed by steady-state responses that are frequency invariant for their physiological activity range. We find that synaptotagmin 7 (Syt7), a calcium sensor for short-term facilitation7, is present at both synapses. It was unclear why a sensor for facilitation would be present at these and other depressing synapses. We find that at Purkinje cell and vestibular synapses, Syt7 supports facilitation that is normally masked by depression, which can be revealed in wild-type mice but is absent in Syt7 knockout mice. In wild-type mice, facilitation increases with firing frequency and counteracts depression to produce frequency-invariant transmission. In Syt7-knockout mice, Purkinje cell and vestibular synapses exhibit conventional use-dependent depression, weakening to a greater extent as the firing frequency is increased. Presynaptic rescue of Syt7 expression restores both facilitation and frequency-invariant transmission. Our results identify a function for Syt7 at synapses that exhibit overall depression, and demonstrate that facilitation has an unexpected and important function in producing frequency-invariant transmission.

Dystrophic or swollen axons occur in a variety of neurological disorders. Although the structures are often viewed as an incidental or age-related phenomenon, there are few reports that actually described axonal and dendritic swellings on cerebellar Purkinje cells in domestic animals with no apparent neurological clinical signs. A 12-year-old Holstein-Friesian dairy cow, which exhibited no apparent cerebellar or other significant clinical signs, was culled from old age. On histopathology of the cerebellum, there were Purkinje cell axonal torpedoes and axonal spheroids involving nerve fibers in the folial and medullary white matter, which were identified by Bodian’s silver impregnation technique and anti-neuron-specific enolase (NSE) immunohistochemical stain. Less frequently, dendritic swellings on Purkinje cells were found in the molecular layer. Cell bodies of Purkinje cells appeared otherwise normal and were not accompanied by Bergmann glia proliferation. Immunohistochemistry for glial fibrillary acid protein (GFAP) failed to demonstrate significant fibrillary gliosis. Deposition of lipofuscin pigments was seen in the cortical and medullary neurons and glial cells. Any sites of the cerebellum showed no evidence of neuronal cell degeneration, glial reaction, cellular infiltrates, or vascular lesions. Morphometrically, there was no significant decrease in number of Purkinje cells in comparison with younger control cows (n = 2). Purkinje cell axonal torpedoes or axonal spheroids were absent in the cerebellum of younger control cows (n = 10). The precise mechanism for the development of swollen Purkinje cell axons and dendrites remains to be determined, although an association with advanced age was considered one of the likeliest possibilities.

BACKGROUND: Essential tremor (ET) is among the most common neurological diseases. Postmortem studies have noted a series of pathological changes in the ET cerebellum. Heterotopic Purkinje cells (PCs) are those whose cell body is mis-localized in the molecular layer. In neurodegenerative settings, these are viewed as a marker of the progression of neuronal degeneration. OBJECTIVE: We (1) quantify heterotopias in ET cases vs. controls, (2) compare ET cases to other cerebellar degenerative conditions (Spinocerebellar ataxia [SCA] 1, 2, 3, and 6), (3) compare these SCAs to one another, and (4) assess heterotopia within the context of associated PC loss in each disease. METHODS: Heterotopic PCs were quantified using a standard LH&E stained section of neocerebellum. Counts were normalized to PC layer length (n-heterotopia count). It is also valuable to consider PC counts when assessing heterotopia, as loss of PCs extends both to normally located as well as heterotopic PCs. Therefore, we divided n-heterotopias by PC counts. RESULTS: There were 96 brains (43 ET, 31 SCA [12 SCA1, 7 SCA2, 7 SCA3, 5 SCA6] and 22 controls). The median number of n-heterotopias in ET cases was two times higher than that of controls (2.6 vs. 1.2, p

Cerebellar participation in timing and sensory-motor sequences has been supported by several experimental and clinical studies. A relevant role of the cerebellum in timing of conditioned responses in the range of milliseconds has been demonstrated, but less is known regarding the role of the cerebellum in supra-second timing of operant responses. A dissociated role of the cerebellum and striatum in timing in the millisecond and second range had been reported, respectively. The climbing fibre-Purkinje cell synapse is crucial in timing models; thus, the aberrant connection between these cellular elements is a suitable model for evaluating the contribution of the cerebellum in timing in the supra-second range. The aberrant connection between climbing fibres and Purkinje cells was induced by administration of the antagonist of NMDA receptors MK-801 to Sprague-Dawley rats at postnatal days 7-14. The timing of an operant response with two fixed intervals (5 and 8 s) and egocentric sequential learning was evaluated in 60-day-old adult rats. The aberrant connections caused a reduced accuracy in the timing of the instrumental response that was more evident in the 8-s interval and a reduced number of successive correct responses (responses emitted in the correct second without any other response between them) in the 8-s interval. In addition, an inability to incorporate new information in a sequence previously learned in egocentric-based sequence learning was apparent in rats with aberrant CF-PC synapses. These results support a relevant role for the cerebellum in the fine-tuning of the timing of operant responses in the supra-second range.

Neurons communicate through excitatory and inhibitory synapses. Both lines of communication are adjustable and allow the fine tuning of signal exchange required for learning processes in neural networks. Several distinct modes of plasticity modulate glutamatergic and GABAergic synapses in Purkinje cells of the cerebellar cortex to promote motor control and learning. In the present paper, we present evidence for a role of short-term ionic plasticity in the cerebellar circuit activity. This type of plasticity results from altered chloride driving forces at the synapses that molecular layer interneurons form on Purkinje cell dendrites. Previous studies have provided evidence for transiently diminished chloride gradients at these GABAergic synapses following climbing fiber activity. Electrical stimulation of climbing fibers in acute slices caused a decline of inhibitory postsynaptic currents recorded from Purkinje cells. Dendritic calcium-gated chloride channels of the type anoctamin 2 (ANO2) were proposed to mediate this short-term modulation of inhibition, but the significance of this process for motor control has not been established yet. Here, we report results of behavioral studies obtained from Ano2 −/− mice, a mouse line that was previously shown to lack this particular mode of ionic plasticity. The animals display motor coordination deficits that constitute a condition of mild ataxia. Moreover, motor learning is severely impaired in Ano2 −/− mice, suggesting cerebellar dysfunction. This reduced motor performance of Ano2 −/− mice highlights the significance of inhibitory control for cerebellar function and introduces calcium-dependent short-term ionic plasticity as an efficient control mechanism for neural inhibition.

We investigated the neuron-specific enolase (NSE) promoter in terms of its promoter strength and neuronal specificity in the cerebellum in vivo. The 1.8 kb rat NSE promoter was divided into three regions, A (0.8 kb), B (0.7 kb), and C (0.3 kb), starting from the 5' side. Then, we made various deletion constructs and assessed them by virally expressing GFP under the control of one of the deleted promoters. Removing region A reduced GFP expression to ~6% of that of the original 1.8 kb promoter. Further deletion of region B (presence of region C alone) did not influence the promoter strength, but removing region B from the original 1.8 kb promoter reduced the GFP expression to ~6% of the original level, similar to the level observed after deletion of region A. Immunohistochemistry showed robust GFP expression in Purkinje cells and modest expression in interneurons by the original promoter. Removing region A and/or region B abolished the GFP expression in Purkinje cells in most cerebellar lobules, with the expression in interneurons almost unchanged. These results suggest that region C, which is a proximal 0.3 kb sequence, contains cis-acting elements that drive transcription predominantly in interneurons. The addition of either region A or B onto region C does not alter the promoter properties; however, the addition of both regions A and B to region C drastically enhanced the promoter activity in Purkinje cells, suggesting the synergistic action of cis-acting regulatory elements in regions A and B for strong activation in Purkinje cells.

Comparative neuroanatomy provides insights into the evolutionary functional adaptation of specific mammalian cerebellar lobules, in which the lobulation pattern and functional localization are conserved. However, accurate identification of homologous lobules among mammalian species is challenging. In this review, we discuss the inter-species homology of crus I and II lobules which occupy a large volume in the posterior cerebellar hemisphere, particularly in humans. Both crus I/II in humans are homologous to crus I/II in non-human primates, according to Paxinos and colleagues; however, this area has been defined as crus I alone in non-human primates, according to Larsell and Brodal. Our neuroanatomical analyses in humans, macaques, marmosets, rats, and mice demonstrate that both crus I/II in humans are homologous to crus I/II or crus I alone in non-human primates, depending on previous definitions, and to crus I alone in rodents. Here, we refer to the region homologous to human crus I/II lobules as "ansiform area (AA)" across animals. Our results show that the AA's olivocerebellar climbing fiber and Purkinje cell projections as well as aldolase C gene expression patterns are both distinct and conserved in marmosets and rodents. The relative size of the AA, as represented by the AA volume fraction in the whole cerebellum was 0.34 in human, 0.19 in macaque, and approximately 0.1 in marmoset and rodents. These results indicate that the AA reflects an evolutionarily conserved structure in the mammalian cerebellum, which is characterized by distinct connectivity from neighboring lobules and a massive expansion in skillful primates.

Familial and sporadic essential tremor (ET) cases differ in several respects. Whether they differ with respect to cerebellar pathologic changes has yet to be studied. We quantified a broad range of postmortem features (Purkinje cell (PC) counts, PC axonal torpedoes, a host of associated axonal changes, heterotopic PCs, and hairy basket ratings) in 60 ET cases and 30 controls. Familial ET was defined using both liberal criteria (n = 27) and conservative criteria (n = 20). When compared with controls, ET cases had lower PC counts, more torpedoes, more heterotopic PCs, a higher hairy basket rating, an increase in PC axonal collaterals, an increase in PC thickened axonal profiles, and an increase in PC axonal branching. Familial and sporadic ET had similar postmortem changes, with few exceptions, regardless of the definition criteria. The PC counts were marginally lower in familial than sporadic ET (respective p values = 0.059 [using liberal criteria] and 0.047 [using conservative criteria]). The PC thickened axonal profile count was marginally lower in familial ET than sporadic ET (respective p values = 0.037 [using liberal criteria] and 0.17 [using conservative criteria]), and the PC axonal branching count was marginally lower in familial than sporadic ET (respective p values = 0.045 [using liberal criteria] and 0.079 [using conservative criteria]). After correction for multiple comparisons, however, there were no significant differences. Overall, familial and sporadic ET cases share very similar cerebellar postmortem features. These data indicate that pathological changes in the cerebellum are a part of the pathophysiological cascade of events in both forms of ET.

Cerebellar MR imaging has several challenging aspects, due to the fine, repetitive layered structure of cortical folia with underlying axonal pathways. In this MR study, we imaged with high-angular resolution diffusion imaging (HARDI) abnormal cerebellar cortical structure (gray matter) and myelinated axonal pathways (white matter) of a mouse spontaneous mutation, Purkinje cell degeneration (pcd), in which almost all Purkinje neurons degenerate, mainly between postnatal days 20 and 35. Mouse brains at postnatal day 20 (P20) and at 8 months were scanned, and known or expected abnormalities, such as reduction of the white matter volume, disorganized pathways likely linked to parallel fibers, mossy fibers, and other fibers running from/to the cerebellar cortex were observed in mutant mice. Such abnormalities were detected at both an early and a fully advanced degeneration stage. These results suggest that our diffusion MR tractography is useful for early detection and tracking of neuropathology in the cerebellum.

Results of recent genome-wide association studies (GWAS) and whole genome sequencing (WGS) highlighted type II cadherins as risk genes for autism spectrum disorders (ASD). To determine whether these cadherins may be linked to the morphogenesis of ASD-relevant brain regions, in situ hybridization (ISH) experiments were carried out to examine the mRNA expression profiles of two ASD-associated cadherins, Cdh9 and Cdh11, in the developing cerebellum. During the first postnatal week, both Cdh9 and Cdh11 were expressed at high levels in segregated sub-populations of Purkinje cells in the cerebellum, and the expression of both genes was declined as development proceeded. Developmental expression of Cdh11 was largely confined to dorsal lobules (lobules VI/VII) of the vermis as well as the lateral hemisphere area equivalent to the Crus I and Crus II areas in human brains, areas known to mediate high order cognitive functions in adults. Moreover, in lobules VI/VII of the vermis, Cdh9 and Cdh11 were expressed in a complementary pattern with the Cdh11-expressing areas flanked by Cdh9-expressing areas. Interestingly, the high level of Cdh11 expression in the central domain of lobules VI/VII was correlated with a low level of expression of the Purkinje cell marker calbindin, coinciding with a delayed maturation of Purkinje cells in the same area. These findings suggest that these two ASD-associated cadherins may exert distinct but coordinated functions to regulate the wiring of ASD-relevant circuits in the cerebellum. Electronic supplementary material The online version of this article (10.1186/s13041-019-0461-4) contains supplementary material, which is available to authorized users.

In the version of this article initially published, in the section “Wiring diagram” the sentence beginning “These newly discovered recurrent pathways…” was repeated; and in the x paragraph of the section “Acquisition versus consolidation” the sentence beginning “A genetic modification…” should have read, “A genetic modification that causes climbing fibers to be directed to the ipsilateral, rather than contralateral, cerebellar cortex, reverses the direction selectivity of the climbing fiber responses of their Purkinje cell targets and, concomitantly, reverses the direction selectivity of their simple spike responses.” These errors have been corrected in the PDF and HTML versions of this article.

Early onset and late onset essential tremor (ET) cases differ in several respects. Whether they differ with respect to cerebellar pathologic changes remains to be determined. We quantified a broad range of postmortem features (Purkinje cell (PC) counts, PC axonal torpedoes and associated axonal changes, heterotopic PCs, and hairy basket ratings) in 30 ET cases with age of tremor onset 0.05), heterotopic PC counts (9.90 ± 11.55 [median =6.00] vs. 5.40 ± 5.10 [median =3.50], p = 0.092), and hairy basket ratings (1.95 ± 0.62 [median =2.00] vs. 2.05 ± 0.92 [median =2.00], p = 0.314). When using the age of onset cut-points of 40 or 60 years, results were similar. Early onset and late onset ET cases share similar cerebellar postmortem features. These data do not support the notion that these age-of-onset related forms of ET represent distinct clinical-pathological entities.

Ultrastructural changes in synaptic boutons of parallel fibers (PF) and the spines of Purkinje cell dendrites (PCD) in the frog cerebellum were stdied on exposure to high concentrations (1 mM) of glutamate (Glu) and NO-generating compounds, creating a model of stroke. Exposure to Glu led to envelopment of terminal boutons by spines, while NO-generating compounds, conversely, led to envelopment of spines by boutons. Morphological studies showed that in Glu solution, there was a dominance of synapses in which glial cells were surrounded by spines, while boutons were dominant in the presence of NO. On electrical stimulation of PF, the relative content of synapses whose boutons were enveloped by glial cells was greater than the proportion of synapses in which glial cells were enveloped by spines, by a factor of 10. These morphological changes reflect the functional state of PF and PCD synapses in response to the harmful influences of excess Glu and NO, apparent as different forms of synaptic contacts and neuron-glial structures.

Autosomal-recessive cerebellar atrophy is usually associated with inactivating mutations and early-onset presentation. The underlying molecular diagnosis suggests the involvement of neuronal survival pathways, but many mechanisms are still lacking and most patients elude genetic diagnosis. Using whole exome sequencing, we identified homozygous p.Val55Ala in the THG1L (tRNA-histidine guanylyltransferase 1 like) gene in three siblings who presented with cerebellar signs, developmental delay, dysarthria, and pyramidal signs and had cerebellar atrophy on brain MRI. THG1L protein was previously reported to participate in mitochondrial fusion via its interaction with MFN2. Abnormal mitochondrial fragmentation, including mitochondria accumulation around the nuclei and confinement of the mitochondrial network to the nuclear vicinity, was observed when patient fibroblasts were cultured in galactose containing medium. Culturing cells in galactose containing media promotes cellular respiration by oxidative phosphorylation and the action of the electron transport chain thus stimulating mitochondrial activity. The growth defect of the yeast thg1Δ strain was rescued by the expression of either yeast Thg1 or human THG1L; however, clear growth defect was observed following the expression of the human p.Val55Ala THG1L or the corresponding yeast mutant. A defect in the protein tRNAHis guanylyltransferase activity was excluded by the normal in vitro G-1 addition to either yeast tRNAHis or human mitochondrial tRNAHis in the presence of the THG1L mutation. We propose that homozygosity for the p.Val55Ala mutation in THG1L is the cause of the abnormal mitochondrial network in the patient fibroblasts, likely by interfering with THG1L activity towards MFN2. This may result in lack of mitochondria in the cerebellar Purkinje dendrites, with degeneration of Purkinje cell bodies and apoptosis of granule cells, as reported for MFN2 deficient mice.

Iodine is a significant micronutrient. Iodine deficiency (ID)-induced hypothyroxinemia and hypothyroidism during developmental period can cause cerebellar dysfunction. However, mechanisms are still unclear. Therefore, the present research aims to study effects of developmental hypothyroxinemia caused by mild ID and hypothyroidism caused by severe ID or methimazole (MMZ) on parallel fiber-Purkinje cell (PF-PC) synapses in filial cerebellum. Maternal hypothyroxinemia and hypothyroidism models were established in Wistar rats using ID diet and deionized water supplemented with different concentrations of potassium iodide or MMZ water. Birth weight and cerebellum weight were measured. We also examined PF-PC synapses using immunofluorescence, and western blot analysis was conducted to investigate the activity of Neurexin1/cerebellin1 (Cbln1)/glutamate receptor d2 (GluD2) tripartite complex. Our results showed that hypothyroxinemia and hypothyroidism decreased birth weight and cerebellum weight and reduced the PF-PC synapses on postnatal day (PN) 14 and PN21. Accordingly, the mean intensity of vesicular glutamate transporter (VGluT1) and Calbindin immunofluorescence was reduced in mild ID, severe ID, and MMZ groups. Moreover, maternal hypothyroxinemia and hypothyroidism reduced expression of Neurexin1/Cbln1/GluD2 tripartite complex. Our study supports the hypothesis that developmental hypothyroxinemia and hypothyroidism reduce PF-PC synapses, which may be attributed to the downregulation of Neurexin1/Cbln1/GluD2 tripartite complex.

Autism spectrum disorders (ASDs) are a group of neurodevelopmental disorders characterized by impaired and disordered language, decreased social interactions, stereotyped and repetitive behaviors, and impaired fine and gross motor skills. It has been well established that cerebellar abnormalities are one of the most common structural changes seen in the brains of people diagnosed with autism. Common cerebellar pathology observed in autistic individuals includes variable loss of cerebellar Purkinje cells (PCs) and increased numbers of reactive neuroglia in the cerebellum and cortical brain regions. The Lc/+ mutant mouse loses 100 % of cerebellar PCs during the first few weeks of life and provided a valuable model to study the effects of developmental PC loss on underlying structural and functional changes in cerebellar neural circuits. Lurcher (Lc) chimeric mice were also generated to explore the link between variable cerebellar pathology and subsequent changes in the structure and function of cerebellar neurons and neuroglia. Chimeras with the most severe cerebellar pathology (as quantified by cerebellar PC counts) had the largest changes in cFos expression (an indirect reporter of neural activity) in cerebellar granule cells (GCs) and cerebellar nucleus (CN) neurons. In addition, Lc chimeras with the fewest PCs also had numerous reactive microglia and Bergmann glia located in the cerebellar cortex. Structural and functional abnormalities observed in the cerebella of Lc chimeras appeared to be along a continuum, with the degree of pathology related to the number of PCs in individual chimeras.

Background Inflammation plays a critical role in accelerating the progression of neurodegenerative diseases, such as Alzheimer’s disease (AD) and ataxia telangiectasia (A-T). In A-T mouse models, LPS-induced neuroinflammation advances the degenerative changes found in cerebellar Purkinje neurons both in vivo and in vitro. In the current study, we ask whether ibuprofen, a non-steroidal anti-inflammatory drug (NSAID), can have the opposite effect and delay the symptoms of the disease. Methods We tested the beneficial effects of ibuprofen in both in vitro and in vivo models. Conditioned medium from LPS stimulated primary microglia (LM) applied to cultures of dissociated cortical neurons leads to numerous degenerative changes. Pretreatment of the neurons with ibuprofen, however, blocked this damage. Systemic injection of LPS into either adult wild-type or adult Atm−/− mice produced an immune challenge that triggered profound behavioral, biochemical, and histological effects. We used a 2-week ibuprofen pretreatment regimen to investigate whether these LPS effects could be blocked. We also treated young presymptomatic Atm−/− mice to determine if ibuprofen could delay the appearance of symptoms. Results Adding ibuprofen directly to neuronal cultures significantly reduced LM-induced degeneration. Curiously, adding ibuprofen to the microglia cultures before the LPS challenge had little effect, thus implying a direct effect of the NSAID on the neuronal cultures. In vivo administration of ibuprofen to Atm−/− animals before a systemic LPS immune challenge suppressed cytological damage. The ibuprofen effects were widespread as microglial activation, p38 phosphorylation, DNA damage, and neuronal cell cycle reentry were all reduced. Unfortunately, ibuprofen only slightly improved the LPS-induced behavioral deficits. Yet, while the behavioral symptoms could not be reversed once they were established in adult Atm−/− animals, administration of ibuprofen to young mutant pups prevented their symptoms from appearing. Conclusion Inflammatory processes impact the normal progression of A-T implying that modulation of the immune system can have therapeutic benefit for both the behavioral and cellular symptoms of this neurodegenerative disease. Electronic supplementary material The online version of this article (10.1186/s12974-018-1338-7) contains supplementary material, which is available to authorized users.

Circuit refinement during postnatal development is finely regulated by neuron–neuron interactions. Recent studies suggest participation of microglia in this process but it is unclear how microglia cooperatively act with neuronal mechanisms. To examine roles of microglia, we ablate microglia by microglia-selective deletion of colony-stimulating factor 1 receptor (Csf1r) by crossing floxed-Csf1r and Iba1-iCre mice (Csf1r-cKO). In Csf1r-cKO mice, refinement of climbing fiber (CF) to Purkinje cell (PC) innervation after postnatal day 10 (P10)–P12 is severely impaired. However, there is no clear morphological evidence suggesting massive engulfment of CFs by microglia. In Csf1r-cKO mice, inhibitory synaptic transmission is impaired and CF elimination is restored by diazepam, which suggests that impairment of CF elimination is caused by a defect of GABAergic inhibition on PCs, a prerequisite for CF elimination. These results indicate that microglia primarily promote GABAergic inhibition and secondarily facilitate the mechanism for CF elimination inherent in PCs., In the mammalian cerebellum, surplus synapses between climbing fibers (CF) and Purkinje cells (PC) are developmentally pruned. Here, Nakayama and colleagues show that ablation of microglia impairs pruning of CF-PC synapses because of dysfunction of GABAergic inhibition prerequisite for pruning.

Climbing and mossy fibers comprise two distinct afferent paths to the cerebellum. Climbing fibers directly evoke a large multispiked action potential in Purkinje cells termed a "complex spike" (CS). By logical exclusion, the other class of Purkinje cell action potential, termed "simple spike" (SS), has often been attributed to activity conveyed by mossy fibers and relayed to Purkinje cells through granule cells. Here, we investigate the relative importance of climbing and mossy fiber pathways in modulating neuronal activity by recording extracellularly from Purkinje cells, as well as from mossy fiber terminals and interneurons in folia 8-10. Sinusoidal roll-tilt vestibular stimulation vigorously modulates the discharge of climbing and mossy fiber afferents, Purkinje cells, and interneurons in folia 9-10 in anesthetized mice. Roll-tilt onto the side ipsilateral to the recording site increases the discharge of both climbing fibers (CSs) and mossy fibers. However, the discharges of SSs decrease during ipsilateral roll-tilt. Unilateral microlesions of the beta nucleus (β-nucleus) of the inferior olive blocks vestibular modulation of both CSs and SSs in contralateral Purkinje cells. The blockage of SSs occurs even though primary and secondary vestibular mossy fibers remain intact. When mossy fiber afferents are damaged by a unilateral labyrinthectomy (UL), vestibular modulation of SSs in Purkinje cells ipsilateral to the UL remains intact. Two inhibitory interneurons, Golgi and stellate cells, could potentially contribute to climbing fiber-induced modulation of SSs. However, during sinusoidal roll-tilt, only stellate cells discharge appropriately out of phase with the discharge of SSs. Golgi cells discharge in phase with SSs. When the vestibularly modulated discharge is blocked by a microlesion of the inferior olive, the modulated discharge of CSs and SSs is also blocked. When the vestibular mossy fiber pathway is destroyed, vestibular modulation of ipsilateral CSs and SSs persists. We conclude that climbing fibers are primarily responsible for the vestibularly modulated discharge of both CSs and SSs. Modulation of the discharge of SSs is likely caused by climbing fiber-evoked stellate cell inhibition.

Several lines of evidence show that classical or Pavlovian conditioning of blink responses depends on the cerebellum. Recordings from cerebellar Purkinje cells that control the eyelid and the conditioned blink show that during training with a conditioning protocol, a Purkinje cell develops a pause response to the conditional stimulus. This conditioned cellular response has many of the properties that characterise the overt blink. The present paper argues that the learned Purkinje cell pause response is the memory trace and main driver of the overt conditioned blink and that it explains many well-known behavioural phenomena.

Retinoid-related orphan receptor α (RORα) is a critical master transcription factor that governs postnatal cerebellar development. An RORα-deficient cerebellum has a persistent external granular layer (EGL), rudimentary Purkinje cell (PC) dendrites, grossly reduced numbers of immature parallel fiber (PF)-PC synapses, and multiple climbing fibers (CF) innervating PCs in mice after 3 weeks of age when these features have disappeared in wild-type mice. Functionally, metabotropic glutamate receptor (mGluR)-mediated signaling in PCs is completely abrogated. Here we examined whether these defects could be corrected by lentivirally providing the RORα gene to 3-week-old PCs of RORα-deficient homozygous staggerer (sg/sg) mice. RORα expression in sg/sg PCs significantly increased the numbers of PF–PC synapses, spines on PC dendritic branchlets, and internal granule cells, concomitant with regression of the EGL, suggesting enhanced proliferation in the EGL and migration of post-mitotic progeny into the internal granular layer with augmented synaptogenesis between PFs and PC dendrites. However, the primary dendritic stems were only slightly extended, and mGluR signaling and the loss of redundant CF synapses in sg/sg PCs remained unrestored. These results suggest that the mitogenic and migratory potential of external granule cells in response to RORα was preserved in the >3-week-old sg/sg mouse cerebellum. Moreover, sg/sg PCs sprouted spines and formed synapses with PFs. However, lengthening of the primary dendritic stems, establishment of mGluR signaling, and removal of CF synapses in sg/sg PCs were regressed by 3 weeks of age.