Cerebellar Purkinje Cell Loss Research Articles

Iron accumulation and axonal damage of cerebellum in idiopathic cervical dystonia.

Postmortem brain study indicated that cerebellar Purkinje cell (PC) loss might be a pathological finding in patients with inherited and idiopathic cervical dystonia (ICD). The analysis of conventional magnetic resonance imaging brain scans failed to yield support for this finding. Previous studies have identified that iron overload can be the consequence of neuron death. The objectives of this study were to investigate iron distribution and demonstrate changes in axons in the cerebellum, providing evidence for PC loss in patients with ICD. Twenty-eight patients with ICD (20 females) and 28 age- and sex-matched healthy controls were recruited. A spatially unbiased infratentorial template was applied to perform cerebellum optimized quantitative susceptibility mapping and diffusion tensor analysis based on magnetic resonance imaging. Voxel-wise analysis was performed to assess cerebellar tissue magnetic susceptibility and fractional anisotropy (FA) alterations, and the clinical relevance of these findings was investigated in the patients with ICD. Increased susceptibility values revealed by quantitative susceptibility mapping in the right lobule CrusI, CrusII, VIIb, VIIIa, VIIIb and IX were found in the patients with ICD. A reduced FA value was found across almost all the cerebellum; an FA value of the significant clusters within the right lobule VIIIa significantly correlated with the motor severity of patients with ICD (r=-0.575, p=0.002). Our study provided evidence for cerebellar iron overload and axonal damage in patients with ICD, which may indicate PC loss and related axonal changes. These results provide evidence for the neuropathological findings in patients with ICD and further highlight the cerebellar involvement in the pathophysiology of dystonia.

The extra-cerebellar effects of spinocerebellar ataxia type 1 (SCA1): looking beyond the cerebellum.

Spinocerebellar ataxia type 1 (SCA1) is one of nine polyglutamine (polyQ) diseases and is characterized as an adult late-onset, progressive, dominantly inherited genetic disease. SCA1 is caused by an increase in the number of CAG repeats in the ATXN1 gene leading to an expanded polyQ tract in the ATAXIN-1 protein. ATAXIN-1 is broadly expressed throughout the brain. However, until recently, SCA1 research has primarily centered on the cerebellum, given the characteristic cerebellar Purkinje cell loss observed in patients, as well as the progressive motor deficits, including gait and limb incoordination, that SCA1 patients present with. There are, however, also other symptoms such as respiratory problems, cognitive defects and memory impairment, anxiety, and depression observed in SCA1 patients and mouse models, which indicate that there are extra-cerebellar effects of SCA1 that cannot be explained solely through changes in the cerebellar region of the brain alone. The existing gap between human and mouse model studies of extra-cerebellar regions in SCA1 makes it difficult to answer many important questions in the field. This review will cover both the cerebellar and extra-cerebellar effects of SCA1 and highlight the need for further investigations into the impact of mutant ATXN1 expression in these regions. This review will also discuss implications of extra-cerebellar effects not only for SCA1 but other neurodegenerative diseases showing diverse pathology as well.

The effects of gestational choline supplementation on cerebellar Purkinje cell number in the sheep model of binge alcohol exposure during the first trimester-equivalent.

Individuals with fetal alcohol spectrum disorders (FASD) incur enduring brain damage and neurodevelopmental impairments from prenatal alcohol exposure (PAE). Preclinical rodent models have demonstrated that choline supplementation during development can reduce the severity of adverse neurodevelopmental consequences of PAE. This study used the sheep model to evaluate dietary choline supplementation during pregnancy as a therapeutic intervention, testing the hypothesis that choline can ameliorate alcohol-induced cerebellar Purkinje cell loss. Pregnant ewes were randomly assigned either to a normal control [NC] group (n=8), or to groups given intravenous infusions of alcohol (or saline) from gestational days 4-41 (the first trimester-equivalent). A weekly binge-drinking pattern was modeled, with three consecutive days of infusions of saline [SAL], 1.75g/kg/day alcohol [1.75ALC], or 2.5g/kg/day alcohol [2.5ALC] followed by four days off. Infused ewes were randomly assigned to receive dietary supplements throughout pregnancy of choline (10mg/kg/day) or placebo (n=8 per group). Mean blood alcohol concentrations (BAC) were significantly higher in the 2.5ALC groups (287mg/dL) than the 1.75ALC groups (197mg/dL). Lamb cerebella were harvested on postnatal day 180 and processed for stereological counts of Purkinje cells. Both alcohol doses caused significant reductions in Purkinje number relative to NC and SAL-Placebo groups, confirming previous findings. Effects of choline supplementation depended on infusion group: it significantly protected against Purkinje cell loss in the 2.5ALC group, had no effect in the 1.75ALC group, and significantly reduced numbers in the SAL-Choline group (though neither the SAL-Choline nor the SAL-Placebo group differed from the NC group). The protection by choline evident only in the 2.5ALC group suggests that multiple, BAC-dependent mechanisms of cerebellar damage may be activated with alcohol exposure in the first trimester, and that choline may protect against pathogenic mechanisms that emerge at higher BACs. These outcomes extend the evidence that early choline supplementation can mitigate some neurodevelopmental defects resulting from binge-like PAE.

Social Deficits and Cerebellar Degeneration in Purkinje Cell Scn8a Knockout Mice.

Mutations in the SCN8A gene encoding the voltage-gated sodium channel α-subunit Nav1. 6 have been reported in individuals with epilepsy, intellectual disability and features of autism spectrum disorder. SCN8A is widely expressed in the central nervous system, including the cerebellum. Cerebellar dysfunction has been implicated in autism spectrum disorder. We investigated conditional Scn8a knockout mice under C57BL/6J strain background that specifically lack Scn8a expression in cerebellar Purkinje cells (Scn8aflox/flox, L7Cre+ mice). Cerebellar morphology was analyzed by immunohistochemistry and MR imaging. Mice were subjected to a battery of behavioral tests including the accelerating rotarod, open field, elevated plus maze, light-dark transition box, three chambers, male-female interaction, social olfaction, and water T-maze tests. Patch clamp recordings were used to evaluate evoked action potentials in Purkinje cells. Behavioral phenotyping demonstrated that Scn8aflox/flox, L7Cre+ mice have impaired social interaction, motor learning and reversal learning as well as increased repetitive behavior and anxiety-like behaviors. By 5 months of age, Scn8aflox/flox, L7Cre+ mice began to exhibit cerebellar Purkinje cell loss and reduced molecular thickness. At 9 months of age, Scn8aflox/flox, L7Cre+ mice exhibited decreased cerebellar size and a reduced number of cerebellar Purkinje cells more profoundly, with evidence of additional neurodegeneration in the molecular layer and deep cerebellar nuclei. Purkinje cells in Scn8aflox/flox, L7Cre+ mice exhibited reduced repetitive firing. Taken together, our experiments indicated that loss of Scn8a expression in cerebellar Purkinje cells leads to cerebellar degeneration and several ASD-related behaviors. Our study demonstrated the specific contribution of loss of Scn8a in cerebellar Purkinje cells to behavioral deficits characteristic of ASD. However, it should be noted that our observed effects reported here are specific to the C57BL/6 genome type.

An Autism-Associated Neuroligin-3 Mutation Affects Developmental Synapse Elimination in the Cerebellum.

Neuroligin is a postsynaptic cell-adhesion molecule that is involved in synapse formation and maturation by interacting with presynaptic neurexin. Mutations in neuroligin genes, including the arginine to cystein substitution at the 451st amino acid residue (R451C) of neuroligin-3 (NLGN3), have been identified in patients with autism spectrum disorder (ASD). Functional magnetic resonance imaging and examination of post-mortem brain in ASD patients implicate alteration of cerebellar morphology and Purkinje cell (PC) loss. In the present study, we examined possible association between the R451C mutation in NLGN3 and synaptic development and function in the mouse cerebellum. In NLGN3-R451C mutant mice, the expression of NLGN3 protein in the cerebellum was reduced to about 10% of the level of wild-type mice. Elimination of redundant climbing fiber (CF) to PC synapses was impaired from postnatal day 10–15 (P10–15) in NLGN3-R451C mutant mice, but majority of PCs became mono-innervated as in wild-type mice after P16. In NLGN3-R451C mutant mice, selective strengthening of a single CF relative to the other CFs in each PC was impaired from P16, which persisted into juvenile stage. Furthermore, the inhibition to excitation (I/E) balance of synaptic inputs to PCs was elevated, and calcium transients in the soma induced by strong and weak CF inputs were reduced in NLGN3-R451C mutant mice. These results suggest that a single point mutation in NLGN3 significantly influences the synapse development and refinement in cerebellar circuitry, which might be related to the pathogenesis of ASD.

Brain pathology and cerebellar purkinje cell loss in a mouse model of chronic neuronopathic Gaucher disease.

Gaucher disease (GD) is currently the focus of considerable attention due primarily to the association between the gene that causes GD (GBA) and Parkinson's disease. Mouse models exist for the systemic (type 1) and for the acute neuronopathic forms (type 2) of GD. Here we report the generation of a mouse that phenotypically models chronic neuronopathic type 3 GD. Gba-/-;Gbatg mice, which contain a Gba transgene regulated by doxycycline, accumulate moderate levels of the offending substrate in GD, glucosylceramide, and live for up to 10 months, i.e. significantly longer than mice which model type 2 GD. Gba-/-;Gbatg mice display behavioral abnormalities at ∼4 months, which deteriorate with age, along with significant neuropathology including loss of Purkinje neurons. Gene expression is altered in the brain and in isolated microglia, although the changes in gene expression are less extensive than in mice modeling type 2 disease. Finally, bone deformities are consistent with the Gba-/-;Gbatg mice being a genuine type 3 GD model. Together, the Gba-/-;Gbatg mice share pathological pathways with acute neuronopathic GD mice but also display differences that might help understand the distinct disease course and progression of type 2 and 3 patients.

Dual-Targeted Autoimmune Sword in Fatal Epilepsy: Patient's glutamate receptor AMPA GluR3B peptide autoimmune antibodies bind, induce Reactive Oxygen Species (ROS) in, and kill both human neural cells and T cells

Nodding Syndrome (NS) is a fatal pediatric epilepsy of unknown etiology, accompanied by multiple neurological impairments, and associated with Onchocerca volvulus (Ov), malnutrition, war-induced trauma, and other insults. NS patients have neuroinflammation, and ~50% have cross-reactive Ov/Leiomodin-1 neurotoxic autoimmune antibodies. ResultsStudying 30 South Sudanese NS patients and a similar number of healthy subjects from the same geographical region, revealed autoimmune antibodies to 3 extracellular peptides of ionotropic glutamate receptors in NS patients: AMPA-GluR3B peptide antibodies (86%), NMDA-NR1 peptide antibodies (77%) and NMDA-NR2 peptide antibodies (87%) (in either 1:10, 1:100 or 1:1000 serum dilution). In contrast, NS patients did not have 26 other well-known autoantibodies that target the nervous system in several autoimmune-mediated neurological diseases. We demonstrated high expression of both AMPA-GluR3 and NMDA-NR1 in human neural cells, and also in normal human CD3+ T cells of both helper CD4+ and cytotoxic CD8+ types. Patient's GluR3B peptide antibodies were affinity-purified, and by themselves precipitated short 70 kDa neuronal GluR3. NS patient's affinity-purified GluR3B peptide antibodies also bound to, induced Reactive Oxygen Species (ROS) in, and killed both human neural cells and T cells within 1-2 hours only. NS patient's purified IgGs, or serum (1:10 or 1:30), induced similar effects. In vivo video EEG experiments in normal mice, revealed that when NS patient's purified IgGs were released continuously (24/7 for 1 week) in normal mouse brain, they induced all the following: 1.Seizures, 2. Cerebellar Purkinje cell loss, 3. Degeneration in the hippocampus and cerebral cortex, and 4. Elevation of CD3+ T cells, and of activated Mac-2+microglia and GFAP+astrocytes in both the gray and white matter of the cerebral cortex, hippocampus, corpus calossum and cerebellum of mice. NS patient's serum cytokines: IL-1β, IL-2, IL-6, IL-8, TNFα, IFNγ, are reduced by 85–99% compared to healthy subjects, suggesting severe immunodeficiency in NS patients. This suspected immunodeficiency could be caused by combined effects of the: 1. Chronic Ov infection, 2. Malnutrition, 3. Killing of NS patient's T cells by patient's own GluR3B peptide autoimmune antibodies (alike the killing of normal human T cells by the NS patient's GluR3B peptide antibodies found herein in vitro). ConclusionsRegardless of NS etiology, NS patients suffer from ‘Dual-targeted Autoimmune Sword’: autoimmune AMPA GluR3B peptide antibodies that bind, induce ROS in, and kill both neural cells and T cells. These neurotoxic and immunotoxic GluR3B peptide autoimmune antibodies, and also NS patient's NMDA-NR1/NR2A and Ov/Leiomodin-1 autoimmune antibodies, must be silenced or removed. Moreover, the findings of this study are relevant not only to NS, but also to many more patients with other types of epilepsy, which have GluR3B peptide antibodies in serum and/or CSF. This claim is based on the following facts: 1. The GluR3 subunit is expressed in neural cells in crucial brains regions, in motor neurons in the spinal cord, and also in other cells in the body, among them T cells of the immune system, 2. The GluR3 subunit has diverse neurophysiological role, and its deletion or abnormal function can: disrupt oscillatory networks of both sleep and breathing, impair motor coordination and exploratory activity, and increase the susceptibility to generate seizures, 3. GluR3B peptide antibodies were found so far in ~27% of >300 epilepsy patients worldwide, which suffer from various other types of severe, intractable and enigmatic epilepsy, and which turned out to be 'Autoimmune Epilepsy'. Furthermore, the findings of this study could be relevant to different neurological diseases besides epilepsy, since other neurotransmitter-receptors autoantibodies are present in other neurological and psychiatric diseases, e.g. autoimmune antibodies against other GluRs, Dopamine receptors, GABA receptors, Acetylcholine receptors and others. These neurotransmitter-receptors autoimmune autoantibodies might also act as ‘Dual-targeted Autoimmune Sword’ and damage both neural cells and T cells (as the AMPA-GluR3B peptide antibodies induced in the present study), since T cells, alike neural cells, express most if not all these neurotransmitter receptors, and respond functionally to the respective neurotransmitters - a scientific and clinical topic we coined 'Nerve-Driven Immunity'.

Purkinje cell-specific Grip1/2 knockout mice show increased repetitive self-grooming and enhanced mGluR5 signaling in cerebellum

Cerebellar Purkinje cell (PC) loss is a consistent pathological finding in autism. However, neural mechanisms of PC-dysfunction in autism remain poorly characterized. Glutamate receptor interacting proteins 1/2 (Grip1/2) regulate AMPA receptor (AMPAR) trafficking and synaptic strength. To evaluate role of PC-AMPAR signaling in autism, we produced PC-specific Grip1/2 knockout mice by crossing Grip2 conventional and Grip1 conditional KO with L7-Cre driver mice. PCs in the mutant mice showed normal morphology and number, and a lack of Grip1/2 expression. Rodent behavioral testing identified normal ambulation, anxiety, social interaction, and an increase in repetitive self-grooming. Electrophysiology studies revealed normal mEPSCs but an impaired mGluR-LTD at the Parallel Fiber-PC synapses. Immunoblots showed increased expression of mGluR5 and Arc, and enhanced phosphorylation of P38 and AKT in cerebellum of PC-specific Grip1/2 knockout mice. Results indicate that loss of Grip1/2 in PCs contributes to increased repetitive self-grooming, a core autism behavior in mice. Results support a role of AMPAR trafficking defects in PCs and disturbances of mGluR5 signaling in cerebellum in the pathogenesis of repetitive behaviors.

Intermittent convection-enhanced delivery of GDNF into rhesus monkey putamen: absence of local or cerebellar toxicity

Glial cell line-derived neurotrophic factor (GDNF) has demonstrated neurorestorative and neuroprotective effects in rodent and nonhuman primate models of Parkinson’s disease. However, continuous intraputamenal infusion of GDNF (100 µg/day) resulted in multifocal cerebellar Purkinje cell loss in a 6-month toxicity study in rhesus monkeys. It was hypothesized that continuous leakage of GDNF into the cerebrospinal fluid compartment during the infusions led to down-regulation of GDNF receptors on Purkinje cells, and that subsequent acute withdrawal of GDNF then mediated the observed cerebellar lesions. Here we present the results of a 9-month toxicity study in which rhesus monkeys received intermittent intraputamenal infusions via convection-enhanced delivery. Animals were treated with GDNF (87.1 µg; N = 14) or vehicle (N = 6) once every 4 weeks for a total of 40 weeks (11 treatments). Four of the GDNF-treated animals were utilized in a satellite study assessing the impact of concomitant catheter repositioning prior to treatment. In the main study, eight animals (5 GDNF, 3 control) were euthanized at the end of the treatment period, along with the four satellite study animals, while the remaining eight animals (5 GDNF, 3 control) were euthanized at the end of a 12-week recovery period. There were no GDNF-related adverse effects and in particular, no GDNF-related microscopic findings in the brain, spinal cord, dorsal root ganglia, or trigeminal ganglia. Therefore, 87.1 µg/4 weeks is considered the no observed adverse effect level for GDNF in rhesus monkeys receiving intermittent, convection-enhanced delivery of GDNF for 9 months.

Effect of treadmill exercise on spatial navigation impairment associated with cerebellar Purkinje cell loss following chronic cerebral hypoperfusion.

In addition to roles in motor coordination, the cerebellum is also associated with cognitive function. The aim of the present study was to investigate the effect of treadmill exercise on spatial navigation deficit induced by chronic cerebral hypoperfusion (CCH). Furthermore, whether decreased loss of Purkinje cells, which contain the calcium-binding protein in the posterior lobe of the cerebellum, attenuates the spatial navigation deficit induced by CCH was also investigated. Wistar rats were randomly divided into three groups: Sham group, bilateral common carotid arteries occlusion (BCCAO) group and a BCCAO + exercise (Ex) group. The rats in the BCCAO + Ex group ran on a treadmill for 30 min once a day for 8 weeks, starting at 4 weeks post-birth. CCH was induced by performing BCCAO at 12 weeks post-birth. The Morris water maze test was performed to determine the spatial navigation function of the rats. To investigate the histological features of the cerebellum in all of the experimental groups post-treatment, terminal deoxynucleotidyl transferase dUTP nick end labeling staining, as well as immunohistochemical analysis revealing the expression of calbindin, parvalbumin, glial fibrillary acidic protein, ionized calcium-binding adaptor molecule 1 and caspase-3, was performed. The results of the present study revealed that treadmill exercise improved spatial navigation, decreased the expression of reactive astrocytes and microglial cells, and decreased apoptotic rates in the cerebellar vermis post-CCH. Treadmill exercise also attenuated the loss of Purkinje cells following CCH. The number of Purkinje cells was revealed to be negatively correlated with spatial navigation performance. These results indicate that treadmill exercise may attenuate spatial navigation impairment via inhibition of Purkinje cell loss in the posterior lobe of the cerebellum following CCH. Therefore, treadmill exercise may represent a therapeutic strategy for the treatment of patients with spatial navigation impairment following CCH.

Intrathecal 2-hydroxypropyl-β-cyclodextrin decreases neurological disease progression in Niemann-Pick disease, type C1: a non-randomised, open-label, phase 1–2 trial

Niemann-Pick disease, type C1 (NPC1) is a lysosomal storage disorder characterised by progressive neurodegeneration. In preclinical testing, 2-hydroxypropyl-β-cyclodextrins (HPβCD) significantly delayed cerebellar Purkinje cell loss, slowed progression of neurological manifestations, and increased lifespan in mouse and cat models of NPC1. The aim of this study was to assess the safety and efficacy of lumbar intrathecal HPβCD. In this open-label, dose-escalation phase 1-2a study, we gave monthly intrathecal HPβCD to participants with NPC1 with neurological manifestation at the National Institutes of Health (NIH), Bethesda, MD, USA. To explore the potential effect of 2-week dosing, three additional participants were enrolled in a parallel study at Rush University Medical Center (RUMC), Chicago, IL, USA. Participants from the NIH were non-randomly, sequentially assigned in cohorts of three to receive monthly initial intrathecal HPβCD at doses of 50, 200, 300, or 400 mg per month. A fifth cohort of two participants received initial doses of 900 mg. Participants from RUMC initially received 200 or 400 mg every 2 weeks. The dose was escalated based on tolerance or safety data from higher dose cohorts. Serum and CSF 24(S)-hydroxycholesterol (24[S]-HC), which serves as a biomarker of target engagement, and CSF protein biomarkers were evaluated. NPC Neurological Severity Scores (NNSS) were used to compare disease progression in HPβCD-treated participants relative to a historical comparison cohort of 21 NPC1 participants of similar age range. Between Sept 21, 2013, and Jan 19, 2015, 32 participants with NPC1 were assessed for eligibility at the National Institutes of Health. 18 patients were excluded due to inclusion criteria not met (six patients), declined to participate (three patients), pursued independent expanded access and obtained the drug outside of the study (three patients), enrolled in the RUMC cohort (one patient), or too late for the trial enrolment (five patients). 14 patients were enrolled and sequentially assigned to receive intrathecal HPβCD at a starting dose of 50 mg per month (three patients), 200 mg per month (three patients), 300 mg per month (three patients), 400 mg per month (three patients), or 900 mg per month (two patients). During the first year, two patients had treatment interrupted for one dose, based on grade 1 ototoxicity. All 14 patients were assessed at 12 months. Between 12 and 18 months, one participant had treatment interrupted at 17 months due to hepatocellular carcinoma, one patient had dose interruption for 2 doses based on caregiver hardship and one patient had treatment interrupted for 1 dose for mastoiditis. 11 patients were assessed at 18 months. Between Dec 11, 2013, and June 25, 2014, three participants were assessed for eligibility and enrolled at RUMC, and were assigned to receive intrathecal HPβCD at a starting dose of 200 mg every 2 weeks (two patients), or 400 mg every two weeks (one patient). There were no dropouts in this group and all 3 patients were assessed at 18 months. Biomarker studies were consistent with improved neuronal cholesterol homoeostasis and decreased neuronal pathology. Post-drug plasma 24(S)-HC area under the curve (AUC8-72) values, an indicator of neuronal cholesterol homoeostasis, were significantly higher than post-saline plasma 24(S)-HC AUC8-72 after doses of 900 mg (p=0·0063) and 1200 mg (p=0·0037). CSF 24(S)-HC concentrations in three participants given either 600 or 900 mg of HPβCD were increased about two fold (p=0·0032) after drug administration. No drug-related serious adverse events were observed. Mid-frequency to high-frequency hearing loss, an expected adverse event, was documented in all participants. When managed with hearing aids, this did not have an appreciable effect on daily communication. The NNSS for the 14 participants treated monthly increased at a rate of 1·22, SEM 0·34 points per year compared with 2·92, SEM 0·27 points per year (p=0·0002) for the 21 patient comparison group. Decreased progression was observed for NNSS domains of ambulation (p=0·0622), cognition (p=0·0040) and speech (p=0·0423). Patients with NPC1 treated with intrathecal HPβCD had slowed disease progression with an acceptable safety profile. These data support the initiation of a multinational, randomised, controlled trial of intrathecal HPβCD. National Institutes of Health, Dana's Angels Research Trust, Ara Parseghian Medical Research Foundation, Hope for Haley, Samantha's Search for the Cure Foundation, National Niemann-Pick Disease Foundation, Support of Accelerated Research for NPC Disease, Vtesse, Janssen Research and Development, a Johnson & Johnson company, and Johnson & Johnson.

N-butyldeoxynojirimycin delays motor deficits, cerebellar microgliosis, and Purkinje cell loss in a mouse model of mucolipidosis type IV

Mucolipidosis type IV (MLIV) is a lysosomal storage disease exhibiting progressive intellectual disability, motor impairment, and premature death. There is currently no cure or corrective treatment. The disease results from mutations in the gene encoding mucolipin-1, a transient receptor potential channel believed to play a key role in lysosomal calcium egress. Loss of mucolipin-1 and subsequent defects lead to a host of cellular aberrations, including accumulation of glycosphingolipids (GSLs) in neurons and other cell types, microgliosis and, as reported here, cerebellar Purkinje cell loss. Several studies have demonstrated that N-butyldeoxynojirimycin (NB-DNJ, also known as miglustat), an inhibitor of the enzyme glucosylceramide synthase (GCS), successfully delays the onset of motor deficits, improves longevity, and rescues some of the cerebellar abnormalities (e.g., Purkinje cell death) seen in another lysosomal disease known as Niemann-Pick type C (NPC). Given the similarities in pathology between MLIV and NPC, we examined whether miglustat would be efficacious in ameliorating disease progression in MLIV. Using a full mucolipin-1 knockout mouse (Mcoln1−/−), we found that early miglustat treatment delays the onset and progression of motor deficits, delays cerebellar Purkinje cell loss, and reduces cerebellar microgliosis characteristic of MLIV disease. Quantitative mass spectrometry analyses provided new data on the GSL profiles of murine MLIV brain tissue and showed that miglustat partially restored the wild type profile of white matter enriched lipids. Collectively, our findings indicate that early miglustat treatment delays the progression of clinically relevant pathology in an MLIV mouse model, and therefore supports consideration of miglustat as a therapeutic agent for MLIV disease in humans.

Treadmill exercise ameliorates motor dysfunction through inhibition of Purkinje cell loss in cerebellum of valproic acid-induced autistic rats.

Autism is a complex developmental disorder with impairments in social interaction, communication, repetitive behavior and motor skills. Exercise enhances cognitive function, ameliorates motor dysfunction, and provides protective profits against neurodegeneration. In the present study, we evaluated the effect of treadmill exercise on the motor coordination and Purkinje cell loss in relation with reactive astrocytes and microglial activation in the cerebellum using valproic acid (VPA)-induced autism rat model. On the 12th day of pregnancy, the pregnant rats in the VPA-exposed group received intraperitoneal injections of 600-mg/kg VPA. After birth, the rat pups were divided into four groups: the control group, the exercise group, the VPA-treated group, the VPA-treated and exercise group. The rat pups in the exercise groups were forced to run on a treadmill for 30 min once a day, 5 times a week for 4 weeks. In the present results, motor balance and coordination was disturbed by induction of autism, in contrast, treadmill exercise alleviated motor dysfunction in the autistic rats. Purkinje cell loss, reactive astrocytes, and microglial activation were occurred by induction of autism, in contrast, treadmill exercise enhanced survival rate of Purkinje neurons through inhibition of reactive astrocytes and microglia in the autistic rats. The present study showed that exercise may provide a potential therapeutic strategy for the alleviation of motor dysfunction in autistic patients.

The Cerebellum and SIDS: Disordered Breathing in a Mouse Model of Developmental Cerebellar Purkinje Cell Loss during Recovery from Hypercarbia.

The cerebellum assists coordination of somatomotor, respiratory, and autonomic actions. Purkinje cell alterations or loss appear in sudden infant death and sudden death in epilepsy victims, possibly contributing to the fatal event. We evaluated breathing patterns in 12 wild-type (WT) and Lurcher mutant mice with 100% developmental cerebellar Purkinje cell loss under baseline (room air), and recovery from hypercapnia, a concern in sudden death events. Six mutant and six WT mice were exposed to 4-min blocks of increasing CO2 (2, 4, 6, and 8%), separated by 4-min recovery intervals in room air. Breath-by-breath patterns, including depth of breathing and end-expiratory pause (EEP) durations during recovery, were recorded. No baseline genotypic differences emerged. However, during recovery, EEP durations significantly lengthened in mutants, compared to WT mice, following the relatively low levels of CO2 exposure. Additionally, mutant mice exhibited signs of post-sigh disordered breathing during recovery following each exposure. Developmental cerebellar Purkinje cell loss significantly affects compensatory breathing patterns following mild CO2 exposure, possibly by inhibiting recovery from elevated CO2. These data implicate cerebellar Purkinje cells in the ability to recover from hypercarbia, suggesting that neuropathologic changes or loss of these cells contribute to inadequate ventilatory recovery to increased environmental CO2. Multiple disorders, including sudden infant death syndrome (SIDS) and sudden unexpected death in epilepsy (SUDEP), appear to involve both cardiorespiratory failure and loss or injury to cerebellar Purkinje cells; the findings support the concept that such neuropathology may precede and exert a prominent role in these fatal events.

Heat Shock Protein Beta-1 Modifies Anterior to Posterior Purkinje Cell Vulnerability in a Mouse Model of Niemann-Pick Type C Disease.

Selective neuronal vulnerability is characteristic of most degenerative disorders of the CNS, yet mechanisms underlying this phenomenon remain poorly characterized. Many forms of cerebellar degeneration exhibit an anterior-to-posterior gradient of Purkinje cell loss including Niemann-Pick type C1 (NPC) disease, a lysosomal storage disorder characterized by progressive neurological deficits that often begin in childhood. Here, we sought to identify candidate genes underlying vulnerability of Purkinje cells in anterior cerebellar lobules using data freely available in the Allen Brain Atlas. This approach led to the identification of 16 candidate neuroprotective or susceptibility genes. We demonstrate that one candidate gene, heat shock protein beta-1 (HSPB1), promoted neuronal survival in cellular models of NPC disease through a mechanism that involved inhibition of apoptosis. Additionally, we show that over-expression of wild type HSPB1 or a phosphomimetic mutant in NPC mice slowed the progression of motor impairment and diminished cerebellar Purkinje cell loss. We confirmed the modulatory effect of Hspb1 on Purkinje cell degeneration in vivo, as knockdown by Hspb1 shRNA significantly enhanced neuron loss. These results suggest that strategies to promote HSPB1 activity may slow the rate of cerebellar degeneration in NPC disease and highlight the use of bioinformatics tools to uncover pathways leading to neuronal protection in neurodegenerative disorders.

Necroptosis in Niemann-Pick disease, type C1: a potential therapeutic target.

Niemann–Pick disease, type C1 (NPC1) is a neurodegenerative, lysosomal storage disorder due to mutation of the NPC1 gene. The NPC1 phenotype is characterized by progressive neuronal dysfunction, including cerebellar ataxia and dementia. There is histological evidence of neuroinflammation and progressive neuronal loss, with cerebellar Purkinje cells particularly vulnerable to loss of NPC1 function. Necroptosis was evaluated as a mechanism of neuronal loss. Receptor-interacting protein kinase 1 (RIP1) and RIP3 are key components of the necrosomal complex that regulates necroptotic cell death. We report increased expression of RIP1 and RIP3 in NPC1 fibroblasts, NPC1 iPS cell-derived neuronal precursors, and in cerebellar tissue from both NPC1 mice and patients. Our data suggest a positive correlation between NPC1 neurological disease severity and assembly of the necrosome complex. Furthermore, we demonstrate that pharmacological inhibition of RIP1 decreases cell death both in vitro and in vivo. Treatment of Npc1-mutant mice with necrostatin-1, an allosteric inhibitor of RIP1, significantly delayed cerebellar Purkinje cell loss, progression of neurological symptoms, and death. Collectively, our data identified necroptosis as a key component of the molecular network that contributes to neuronal loss in NPC1 and establish that inhibition of necroptosis is a potential therapeutic intervention.

Impaired hypercarbic and hypoxic responses from developmental loss of cerebellar Purkinje neurons: implications for sudden infant death syndrome.

Impaired responsivity to hypercapnia or hypoxia is commonly considered a mechanism of failure in sudden infant death syndrome (SIDS). The search for deficient brain structures mediating flawed chemosensitivity typically focuses on medullary regions; however, a network that includes Purkinje cells of the cerebellar cortex and its associated cerebellar nuclei also helps mediate responses to carbon dioxide (CO2) and oxygen (O2) challenges and assists integration of cardiovascular and respiratory interactions. Although cerebellar nuclei contributions to chemoreceptor challenges in adult models are well described, Purkinje cell roles in developing models are unclear. We used a model of developmental cerebellar Purkinje cell loss to determine if such loss influenced compensatory ventilatory responses to hypercapnic and hypoxic challenges. Twenty-four Lurcher mutant mice and wild-type controls were sequentially exposed to 2% increases in CO2 (0-8%) or 2% reductions in O2 (21-13%) over 4 min, with return to room air (21% O2/79% N2/0% CO2) between each exposure. Whole body plethysmography was used to continuously monitor tidal volume (TV) and breath frequency (f). Increased f to hypercapnia was significantly lower in mutants, slower to initiate, and markedly lower in compensatory periods, except for very high (8%) CO2 levels. The magnitude of TV changes to increasing CO2 appeared smaller in mutants but only approached significance. Smaller but significant differences emerged in response to hypoxia, with mutants showing smaller TV when initially exposed to reduced O2 and lower f following exposure to 17% O2. Since cerebellar neuropathology appears in SIDS victims, developmental cerebellar neuropathology may contribute to SIDS vulnerability.

Cerebellar Purkinje cell p75 neurotrophin receptor and autistic behavior.

The p75 neurotrophin receptor (p75NTR) is normally expressed in cerebellar Purkinje cells throughout the lifespan. Children with autism spectrum behavior exhibit apparent cerebellar Purkinje cell loss. Cerebellar transcriptome changes seen in the murine prenatal valproate exposure model of autism include all of the proteins known to constitute the p75NTR interactome. p75NTR is a modulator of cytoplasmic and mitochondrial redox potential, and others have suggested that aberrant response to oxidant stress has a major role in the pathogenesis of autism. We have created Purkinje cell-selective p75NTR knockout mice that are the progeny of hemizygous Cre-Purkinje cell protein 2 C57Bl mice and p75NTR floxed C57Bl mice. These Cre-loxP mice exhibit complete knockout of p75NTR in ~50% of the cerebellar Purkinje cells. Relative to Cre-only mice and wild-type C57Bl mice, this results in a behavioral phenotype characterized by less allogrooming of (P<0.05; one-way analysis of variance) and socialization or fighting with (each P<0.05) other mice; less (1.2-fold) non-ambulatory exploration of their environment than wild-type (P<0.01) or Cre only (P<0.01) mice; and almost twofold more stereotyped jumping behavior than wild-type (P<0.05) or Cre (P<0.02) mice of the same strain. Wild-type mice have more complex dendritic arborization than Cre-loxP mice, with more neurites per unit area (P<0.025, Student's t-test), more perpendicular branches per unit area (P<0.025) and more short branches/long neurite (P<0.0005). Aberrant developmental regulation of expression of p75NTR in cerebellar Purkinje cells may contribute to the pathogenesis of autism.

Essential tremor is not associated with cerebellar Purkinje cell loss

There has been controversy as to whether there is an underlying neurodegenerative process of the cerebellum in essential tremor (ET). The aim of this study was to examine whether ET is associated with Purkinje cell (PC) loss. Prospectively categorized ET and control subjects who were longitudinally examined in the Arizona Study for Aging and Neurodegenerative Disorders and came to autopsy between 1998 and 2013 underwent standardized neuropathological assessment of the brain. PC linear density of the cerebellar hemisphere was calculated in a blinded manner. There were 56 ET cases and 62 age-matched controls free of dementia and other neurodegenerative disorders included in the study. Mean PC linear density was 3.80 ± 0.81 cells per mm for tremor cases and 3.82 ± 0.91 cells per mm for controls (Δ 0.02; 95% confidence interval [CI]: -0.30-0.34). PC counts were not associated with tremor duration (r = 0.06; 95% CI: -0.21-0.32). These data demonstrate that ET is not associated with cerebellar PC loss.

Glutamate Dysfunction Associated with Developmental Cerebellar Damage: Relevance to Autism Spectrum Disorders

Neural abnormalities commonly associated with autism spectrum disorders include prefrontal cortex (PFC) dysfunction and cerebellar pathology in the form of Purkinje cell loss and cerebellar hypoplasia. It has been reported that loss of cerebellar Purkinje cells results in aberrant dopamine neurotransmission in the PFC which occurs via dysregulation of multisynaptic efferents from the cerebellum to the PFC. Using a mouse model, we investigated the possibility that developmental cerebellar Purkinje cell loss could disrupt glutamatergic cerebellar projections to the PFC that ultimately modulate DA release. We measured glutamate release evoked by local electrical stimulation using fixed-potential amperometry in combination with glutamate selective enzyme-based recording probes in urethane-anesthetized Lurcher mutant and wildtype mice. Target sites included the mediodorsal and ventrolateral thalamic nuclei, reticulotegmental nuclei, pedunculopontine nuclei, and ventral tegmental area. With the exception of the ventral tegmental area, the results indicated that in comparison to wildtype mice, evoked glutamate release was reduced in Lurcher mutants by between 9 and 72% at all stimulated sites. These results are consistent with the notion that developmental loss of cerebellar Purkinje cells drives reductions in evoked glutamate release in cerebellar efferent pathways that ultimately influence PFC dopamine release. Possible mechanisms whereby reductions in glutamate release could occur are discussed.