Ligustilide inhibits Purkinje cell ferritinophagy via the ULK1/NCOA4 pathway to attenuate valproic acid-induced autistic features.

BACKGROUND: Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder in which social impairment is the core symptom. Presently, there are no definitive medications to cure core symptoms of ASD, and most therapeutic strategies ameliorate ASD symptoms. Treatments with proven efficacy in autism are imminent. Ligustilide (LIG), an herbal monomer extracted from Angelica Sinensis and Chuanxiong, is mainly distributed in the cerebellum and widely used in treating neurological disorders. However, there are no studies on its effect on autistic-like phenotypes and its mechanism of action. PURPOSE: Investigate the efficacy and mechanism of LIG in treating ASD using two Valproic acid(VPA)-exposed and BTBR T + Itpr3tf/J (BTBR) mouse models of autism. METHODS: VPA-exposed mice and BTBR mice were given LIG for treatment, and its effect on autistic-like phenotype was detected by behavioral experiments, which included a three-chamber social test. Subsequently, RNA-Sequence(RNA-Seq) of the cerebellum was performed to observe the biological changes to search target pathways. The autophagy and ferroptosis pathways screened were verified by WB(Western Blot) assay, and the cerebellum was stained by immunofluorescence and examined by electron microscopy. To further explore the therapeutic mechanism, ULK1 agonist BL-918 was used to block the therapeutic effect of LIG to verify its target effect. RESULTS: Our work demonstrates that LIG administration from P12-P14 improved autism-related behaviors and motor dysfunction in VPA-exposed mice. Similarly, BTBR mice showed the same improvement. RNA-Seq data identified ULK1 as the target of LIG in regulating ferritinophagy in the cerebellum of VPA-exposed mice, as evidenced by activated autophagy, increased ferritin degradation, iron overload, and lipid peroxidation. We found that VPA exposure-induced ferritinophagy occurred in the Purkinje cells, with enhanced NCOA4 and Lc3B expressions. Notably, the therapeutic effect of LIG disappeared when ULK1 was activated. CONCLUSION: LIG treatment inhibits ferritinophagy in Purkinje cells via the ULK1/NCOA4-dependent pathway. Our study reveals for the first time that LIG treatment ameliorates autism symptoms in VPA-exposed mice by reducing aberrant Purkinje ferritinophagy. At the same time, our study complements the pathogenic mechanisms of autism and introduces new possibilities for its therapeutic options.

Cerebellar Purkinje cell firing promotes conscious recovery from anesthesia state through coordinating neuronal communications with motor cortex.

Background: The neurobiological basis of gaining consciousness from unconscious state induced by anesthetics remains unknown. This study was designed to investigate the involvement of the cerebello-thalamus-motor cortical loop mediating consciousness transitions from the loss of consciousness (LOC) induced by an inhalational anesthetic sevoflurane in mice. Methods: The neural tracing and fMRI together with opto-chemogenetic manipulation were used to investigate the potential link among cerebello-thalamus-motor cortical brain regions. The fiber photometry of calcium and neurotransmitters, including glutamate (Glu), γ-aminobutyric acid (GABA) and norepinephrine (NE), were monitored from the motor cortex (M1) and the 5th lobule of the cerebellar vermis (5Cb) during unconsciousness induced by sevoflurane and gaining consciousness after sevoflurane exposure. Cerebellar Purkinje cells were optogenetically manipulated to investigate their influence on consciousness transitions during and after sevoflurane exposure. Results: Activation of 5Cb Purkinje cells increased the Ca2+ flux in the M1 CaMKIIα+ neurons, but this increment was significantly reduced by inactivation of posterior and parafascicular thalamic nucleus. The 5Cb and M1 exhibited concerted calcium flux, and glutamate and GABA release during transitions from wakefulness, loss of consciousness, burst suppression to conscious recovery. Ca2+ flux and Glu release in the M1, but not in the 5Cb, showed a strong synchronization with the EEG burst suppression, particularly, in the gamma-band range. In contrast, the Glu, GABA and NE release and Ca2+ oscillations were coherent with the EEG gamma band activity only in the 5Cb during the pre-recovery of consciousness period. The optogenetic activation of Purkinje cells during burst suppression significantly facilitated emergence from anesthesia while the optogenetic inhibition prolonged the time to gaining consciousness. Conclusions: Our data indicate that cerebellar neuronal communication integrated with motor cortex through thalamus promotes consciousness recovery from anesthesia which may likely serve as arousal regulation.

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], 1.75 g/kg/day alcohol [1.75ALC], or 2.5 g/kg/day alcohol [2.5ALC] followed by four days off. Infused ewes were randomly assigned to receive dietary supplements throughout pregnancy of choline (10 mg/kg/day) or placebo (n = 8 per group). Mean blood alcohol concentrations (BAC) were significantly higher in the 2.5ALC groups (287 mg/dL) than the 1.75ALC groups (197 mg/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.

Sphingolipid metabolism governs Purkinje cell patterned degeneration in Atxn1[82Q]/+ mice.

Patterned degeneration of Purkinje cells (PCs) can be observed in a wide range of neuropathologies, but mechanisms behind nonrandom cerebellar neurodegeneration remain unclear. Sphingolipid metabolism dyshomeostasis typically leads to PC neurodegeneration; hence, we questioned whether local sphingolipid balance underlies regional sensitivity to pathological insults. Here, we investigated the regional compartmentalization of sphingolipids and their related enzymes in the cerebellar cortex in healthy and pathological conditions. Analysis in wild-type animals revealed higher sphingosine kinase 1 (Sphk1) levels in the flocculonodular cerebellum, while sphingosine-1-phosphate (S1P) levels were higher in the anterior cerebellum. Next, we investigated a model for spinocerebellar ataxia type 1 (SCA1) driven by the transgenic expression of the expanded Ataxin 1 protein with 82 glutamine (82Q), exhibiting severe PC degeneration in the anterior cerebellum while the flocculonodular region is preserved. In Atxn1[82Q]/+ mice, we found that levels of Sphk1 and Sphk2 were region-specific decreased and S1P levels increased, particularly in the anterior cerebellum. To determine if there is a causal link between sphingolipid levels and neurodegeneration, we deleted the Sphk1 gene in Atxn1[82Q]/+ mice. Analysis of Atxn1[82Q]/+; Sphk1-/- mice confirmed a neuroprotective effect, rescuing a subset of PCs in the anterior cerebellum, in domains reminiscent of the modules defined by AldolaseC expression. Finally, we showed that Sphk1 deletion acts on the ATXN1[82Q] protein expression and prevents PC degeneration. Taken together, our results demonstrate that there are regional differences in sphingolipid metabolism and that this metabolism is directly involved in PC degeneration in Atxn1[82Q]/+ mice.

The protective effect of nnano-selenium against cadmium-induced cerebellar injury via the heat shock protein pathway in chicken.

Cadmium (Cd) is one of the toxic environmental heavy metals that poses health hazard to animals due to its toxicity. Nano-Selenium (Nano-Se) is a Nano-composite form of Se, which has emerged as a promising therapeutic agent for its protective roles against heavy metals-induced toxicity. Heat shock proteins (HSPs) play a critical role in cellular homeostasis. However, the potential protective effects of Nano-Se against Cd-induced cerebellar toxicity remain to be illustrated. To investigate the toxic effects of Cd on chicken's cerebellum, and the protective effects of Nano-Se against Cd-induced cerebellar toxicity, a total of 80 male chicks were divided into four groups and treated as follows: (A) 0 mg/kg Cd, (B) 1 mg/kg Nano-Se (C) 140 mg/kg Cd + 1 mg/kg Nano-Se (D) 140 mg/kg Cd for 90 days. We tested heat shock protein pathway-related factors including heat shock factors (HSFs) HSF1, HSF2, HSF3 and heat shock proteins (HSPs) HSP10, HSP25, HSP27, HSP40, HSP60, HSP70 and HSP90 expressions. Histopathological results showed that Cd treatment caused degradation of Purkinje cells. In addition, HSFs and HSPs expression decreased significantly in the Cd group. Nano-Se co-treatment with Cd enhanced the expression of HSFs and HSPs. In summary, our findings explicated a potential protective effect of Nano-Se against Cd-induced cerebellar injury in chicken, suggesting that Nano-Se is a promising therapeutic agent for the treatment of Cd toxicity.

The Effect of Induced Diabetes Mellitus on the Cerebellar Cortex of Adult Male Rat and the Possible Protective Role of Oxymatrine: A Histological, Immunohistochemical and Biochemical Study.

Diabetes mellitus (DM) represents a widespread metabolic disease with a well-known neurotoxicity in both central and peripheral nervous systems. Oxymatrine is a traditional Chinese herbal medicine that has various pharmacological activities including: anti-oxidant, anti-apoptotic and anti-inflammatory potentials. The present work aimed to study the impact of diabetes mellitus on the cerebellar cortex of adult male albino rat and to evaluate the potential protective role of oxymatrine. Fifty-five adult male rats were randomly divided into three groups: group I served as control, group II was given oxymatrine (80 mg/kg/day) orally for 8 weeks and group III was given a single dose of streptozotocin (50 mg/kg) intaperitoneally to induce diabetes. Then diabetic rats were subdivided into two subgroups: subgroup IIIa that received no additional treatment and subgroup IIIb that received oxymatrine similar to group II. The diabetic group revealed numerous changes in the Purkinje cell layer in the form of multilayer arrangement of Purkinje cells, shrunken cells with deeply stained nuclei as well as focal loss of the Purkinje cells. A significant increment in glial fibrillary acidic protein (GFAP) and synaptophysin expression were reported in immunohistochemistry compared with the control group. Transmission electron microscopy showed irregularity and splitting of myelin sheaths in the molecular layer, dark shrunken Purkinje cells with ill-defined nuclei, dilated Golgi saccules and dense granule cells with irregular nuclear outlines in the granular layer. In contrast, these changes were less evident in diabetic rats that received oxymatrine. In conclusion, Oxymatrine could protect the cerebellar cortex against changes induced by DM.

Withania coagulans extract attenuates oxidative stress-mediated apoptosis of cerebellar purkinje neurons after ischemia/reperfusion injury.

Cerebral ischemia/reperfusion (I/R) is known to increase reactive oxygen species (ROS) generation, consequences of oxidative stress (OS), and neuronal death in the susceptible brain areas including the cerebellum. Newly, remarkable attention has been paid to a natural diet with the capability to scavenge ROS. Withania coagulans root extract (WCE) is rich in components with antioxidants properties. Therefore, this study aimed to evaluate the effect of WCE on cerebellar Purkinje cells (PCs) against OS-mediated apoptosis after I/R injury. In this experimental study 64 male adult Wistar rats were randomly divided into 4 groups (n = 16) as follows: control, sham, I/R, and WCE 1000 + I/R. I/R animals were pretreated with daily administration of hydro-alcoholic WCE (1000 mg/kg) or distilled water as a vehicle for 30 days before I/R injury. After 72 h, the animals were sacrificed, the cerebellum tissue was removed and used for biochemical (CAT, SOD, GPx, and MDA levels) and histopathological (Nissl and TUNEL staining) assays. Findings showed that the MDA level and the number of apoptotic neurons significantly increased and viable Purkinje neurons decreased in I/R injury (p < 0.05). Administration of 1000 mg/kg WCE reduced MDA level and enhanced antioxidants activity including CAT, SOD, and GPx significantly. In addition, intact surviving PCs increased. At the same time, TUNEL-positive neurons decreased significantly in the WCE pre-treated group (p < 0.05). These findings suggest that WCE can counteract cerebral I/R-induced OS and associated neuronal death by enhancement of ROS scavenging and antioxidant capacity. It appears that pre-treatment with 1000 mg/kg WCE for thirty days can protect PCs against OS-mediated apoptosis after I/R injury.

Kola nut from Cola nitida vent. Schott administered to pregnant rats induces histological alterations in pups' cerebellum.

Kola nut (from Cola nitida) is popular in Nigeria and West Africa and is commonly consumed by pregnant women during the first trimester to alleviate morning sickness and dizziness. There is, however, a dearth of information on its effects on the developing brain. This study, therefore, investigated the potential effects of kola nut on the structure of the developing neonatal and juvenile cerebellum in the rat. Pregnant Wistar rats were administered water (as control) or crude (aqueous) kola nut extract at 400, 600, and 800 mg/kg body weight orally, from pregnancy to day 21 after birth. On postnatal days 1, 7, 14, 21 and 28, the pups were weighed, anaesthetised, sacrificed and perfused with neutral buffered formalin. Their brains were dissected out, weighed and the cerebellum preserved in 10% buffered formalin. Paraffin sections of the cerebellum were stained with haematoxylin and eosin for cerebellar cytoarchitecture, cresyl violet stain for Purkinje cell count, Glial Fibrillary Acidic Protein (GFAP) immunohistochemistry (IHC) for estimation of gliosis, and B-cell lymphoma 2 (Bcl-2) IHC for apoptosis induction. The kola nut-treated rats exhibited initial reduction in body and brain weights, persistent external granular layer, increased molecular layer thickness, and loss of Bergmann glia. Their Purkinje cells showed reduction in density, loss of dendrites and multiple layering, and their white matter showed neurodegeneration (spongiosis) and GFAP and Bcl-2 over-expression, with evidence of reactive astrogliosis. This study, therefore, demonstrates that kola nut, administered repeatedly at certain doses to pregnant dams, could disrupt normal postnatal cerebellar development in their pups. The findings suggest potential deleterious effects of excessive kola nut consumption on human brain and thus warrant further studies to understand the wider implications for human brain development.

From Molecules to Behavior in Long-Term Inorganic Mercury Intoxication: Unraveling Proteomic Features in Cerebellar Neurodegeneration of Rats.

Mercury is a severe environmental pollutant with neurotoxic effects, especially when exposed for long periods. Although there are several evidences regarding mercury toxicity, little is known about inorganic mercury (IHg) species and cerebellum, one of the main targets of mercury associated with the neurological symptomatology of mercurial poisoning. Besides that, the global proteomic profile assessment is a valuable tool to screen possible biomarkers and elucidate molecular targets of mercury neurotoxicity; however, the literature is still scarce. Thus, this study aimed to investigate the effects of long-term exposure to IHg in adult rats' cerebellum and explore the modulation of the cerebellar proteome associated with biochemical and functional outcomes, providing evidence, in a translational perspective, of new mercury toxicity targets and possible biomarkers. Fifty-four adult rats were exposed to 0.375 mg/kg of HgCl2 or distilled water for 45 days using intragastric gavage. Then, the motor functions were evaluated by rotarod and inclined plane. The cerebellum was collected to quantify mercury levels, to assess the antioxidant activity against peroxyl radicals (ACAPs), the lipid peroxidation (LPO), the proteomic profile, the cell death nature by cytotoxicity and apoptosis, and the Purkinje cells density. The IHg exposure increased mercury levels in the cerebellum, reducing ACAP and increasing LPO. The proteomic approach revealed a total 419 proteins with different statuses of regulation, associated with different biological processes, such as synaptic signaling, energy metabolism and nervous system development, e.g., all these molecular changes are associated with increased cytotoxicity and apoptosis, with a neurodegenerative pattern on Purkinje cells layer and poor motor coordination and balance. In conclusion, all these findings feature a neurodegenerative process triggered by IHg in the cerebellum that culminated into motor functions deficits, which are associated with several molecular features and may be related to the clinical outcomes of people exposed to the toxicant.

Role of aqueous extract of saffron in ameliorating effect of sofosbuvir on the cerebellar cortex in rat.

Sofosbuvir is a promising antiviral drug against chronic hepatitis C virus. Although it is characterized by its high efficacy, its adverse effects on nervous tissue are still unclear. Saffron is known for its neuroprotective property. This is a biochemical, histological and immunohistochemical study of the effect of sofosbuvir on the cerebellar cortex of rat and the possible ameliorating role of saffron's aqueous extract. Twenty-four adult male Wistar albino rats were equally divided into four groups; control, saffron extract-treated, sofosbuvir-treated (41.1 mg/kg/day for 6 weeks) and group concomitantly treated with saffron extract and sofosbuvir. Sofosbuvir-treated group recorded a significant increase in cerebellar malondialdehyde level coupling with a significant decrease in tissue glutathione and superoxide dismutase. Light microscopy revealed reduced number of Purkinje cells. The granular layer depicted many granular cells and Bergmann astrocytes with nuclear and cytoplasmic alterations. Electron microscopy revealed disorganized molecular layer with disarranged myelinated axons and disrupted mitochondria. Few shrunken Purkinje cells showed electron-dense cytoplasm and rarefied nuclei, indistinct nuclear envelope and dilated perinuclear space, areas of vacuolated cytoplasm, fragmented rough endoplasmic reticulum and few dark mitochondria. Some axons with tiny mitochondria were detected. A significant upregulation in immunohistochemical expression of GFAP-positive astrocytes was recorded. Concomitant administration of saffron extract significantly improved all studied parameters. Saffron extract is beneficial in ameliorating sofosbuvir-induced cerebellar morphological changes mainly through its antioxidant and neuroprotective properties.

F-phenibut (ß-(4-Fluorophenyl)-GABA), a potent GABAB receptor agonist, activates an outward-rectifying K+ current and suppresses the generation of action potentials in mouse cerebellar Purkinje cells.

The GABA analog phenibut (ß-Phenyl-GABA) is a GABAB receptor agonist that has been licensed for various uses in Russia. Phenibut is also available as a dietary supplement from online vendors worldwide, and previous studies have indicated that phenibut overdose results in intoxication, withdrawal symptoms, and addiction. F-phenibut (ß-(4-Fluorophenyl)-GABA), a derivative of phenibut, has not been approved for clinical use. However, it is also available as a nootropic supplement from online suppliers. F-phenibut binds to GABAB with a higher affinity than phenibut; therefore, F-phenibut may lead to more serious intoxication than phenibut. However, the mechanisms by which F-phenibut acts on GABAB receptors and influences neuronal function remain unknown. In the present study, we compared the potency of F-phenibut, phenibut, and the GABAB agonist (±)-baclofen (baclofen) using in vitro patch-clamp recordings obtained from mouse cerebellar Purkinje cells slice preparations Our findings indicate that F-phenibut acted as a potent GABAB agonist. EC50 of outward current density evoked by the three GABAB agonists decreased in the following order: phenibut (1362 µM) > F-phenibut (23.3 µM) > baclofen (6.0 µM). The outward current induced by GABAB agonists was an outward-rectifying K+ current, in contrast to the previous finding that GABAB agonists activates an inward-rectifying K+ current. The K+ current recorded in the present study was insensitive to extracellular Ba2+, intra- or extracellular Cs+, and intra- or extracellular tetraethylammonium-Cl. Moreover, F-phenibut suppressed action potential generation in Purkinje cells. Thus, abuse of F-phenibut may lead to severe damage by inhibiting the excitability of GABAB-expressing neurons.

Ginseng Gintonin Attenuates Lead-Induced Rat Cerebellar Impairments during Gestation and Lactation.

Gintonin, a novel ginseng-derived lysophosphatidic acid receptor ligand, improves brain functions and protects neurons from oxidative stress. However, little is known about the effects of gintonin against Pb-induced brain maldevelopment. We investigated the protective effects of gintonin on the developing cerebellum after prenatal and postnatal Pb exposure. Pregnant female rats were randomly divided into three groups: control, Pb (0.3% Pb acetate in drinking water), and Pb plus gintonin (100 mg/kg, p.o.). Blood Pb was increased in dams and pups; gintonin treatment significantly decreased blood Pb. On postnatal day 21, the number of degenerating Purkinje cells was remarkably increased while the number of calbindin-, GAD67-, NMDAR1-, LPAR1-immunoreactive intact Purkinje cells, and GABA transporter 1-immunoreactive pinceau structures were significantly reduced in Pb-exposed offspring. Following Pb exposure, gintonin ameliorated cerebellar degenerative effects, restored increased pro-apoptotic Bax, and decreased anti-apoptotic Bcl2. Gintonin treatment attenuated Pb-induced accumulation of oxidative stress (Nrf2 and Mn-SOD) and inflammation (IL-1ß and TNFα,), restoring the decreased cerebellar BDNF and Sirt1. Gintonin ameliorated Pb-induced impairment of myelin basic protein-immunoreactive myelinated fibers of Purkinje cells. Gintonin attenuated Pb-induced locomotor dysfunctions. The present study revealed the ameliorating effects of gintonin against Pb, suggesting the potential use of gintonin as a preventive agent in Pb poisoning during pregnancy and lactation.

Sunao Tawara : further musings on his tribulations in providing the basis for the modern-day understanding of cardiac electrophysiology.

Sunao Tawara, who was born in 1873 and died in 1952, is considered the father of modern cardiac electrophysiology. He published his monumental monograph describing the atrioventricular conduction axis in 1906. He achieved this task in the face of multiple tribulations as a doctoral student working in a cultural environment that was not his own. Although his letters underscoring the publication of the monograph have been published, little emphasis has been placed on the potential problems he encountered in bringing his task to fruition. For example, it was not until the final 6 months of his studies that he resolved the issue of the connection between the atrioventricular bundle and the so called "Purkinje cardiomyocytes". His exchanges with his mentor, Ludwig Aschoff, emphasized that the difficulties he encountered in making the connection caused him quite some turmoil. We believe that this issue, and others that he identified in his correspondence, are worthy of further attention.

Intrinsic Excitability Increase in Cerebellar Purkinje Cells after Delay Eye-Blink Conditioning in Mice.

Cerebellar-based learning is thought to rely on synaptic plasticity, particularly at synaptic inputs to Purkinje cells. Recently, however, other complementary mechanisms have been identified. Intrinsic plasticity is one such mechanism, and depends in part on the downregulation of calcium-dependent SK-type K+ channels, which contribute to a medium-slow afterhyperpolarization (AHP) after spike bursts, regulating membrane excitability. In the hippocampus, intrinsic plasticity plays a role in trace eye-blink conditioning; however, corresponding excitability changes in the cerebellum in associative learning, such as in trace or delay eye-blink conditioning, are less well studied. Whole-cell patch-clamp recordings were obtained from Purkinje cells in cerebellar slices prepared from male mice ∼48 h after they learned a delay eye-blink conditioning task. Over a period of repeated training sessions, mice received either paired trials of a tone coterminating with a periorbital shock (conditioning) or trials in which these stimuli were randomly presented in an unpaired manner (pseudoconditioning). Purkinje cells from conditioned mice show a significantly reduced AHP after trains of parallel fiber stimuli and after climbing fiber evoked complex spikes. The number of spikelets in the complex spike waveform is increased after conditioning. Moreover, we find that SK-dependent intrinsic plasticity is occluded in conditioned, but not pseudoconditioned mice. These findings show that excitability is enhanced in Purkinje cells after delay eye-blink conditioning, and point toward a downregulation of SK channels as a potential underlying mechanism. The observation that this learning effect lasts at least up to 2 d after training shows that intrinsic plasticity regulates excitability in the long term.SIGNIFICANCE STATEMENT Plasticity of membrane excitability ("intrinsic plasticity") has been observed in invertebrate and vertebrate neurons, coinduced with synaptic plasticity or in isolation. Although the cellular phenomenon per se is well established, it remains unclear what role intrinsic plasticity plays in learning and if it even persists long enough to serve functions in engram physiology beyond aiding synaptic plasticity. Here, we demonstrate that cerebellar Purkinje cells upregulate excitability in delay eye-blink conditioning, a form of motor learning. This plasticity is observed 48 h after training and alters synaptically evoked spike firing and integrative properties of these neurons. These findings show that intrinsic plasticity enhances the spike firing output of Purkinje cells and persists over the course of days.

Antisense Oligonucleotide Reverses Leukodystrophy in Canavan Disease Mice.

Marked elevation in the brain concentration of N-acetyl-L-aspartate (NAA) is a characteristic feature of Canavan disease, a vacuolar leukodystrophy resulting from deficiency of the oligodendroglial NAA-cleaving enzyme aspartoacylase. We now demonstrate that inhibiting NAA synthesis by intracisternal administration of a locked nucleic acid antisense oligonucleotide to young-adult aspartoacylase-deficient mice reverses their pre-existing ataxia and diminishes cerebellar and thalamic vacuolation and Purkinje cell dendritic atrophy. Ann Neurol 2020;87:480-485.

Curcumin Prevents Cerebellar Hypoplasia and Restores the Behavior in Hyperbilirubinemic Gunn Rat by a Pleiotropic Effect on the Molecular Effectors of Brain Damage.

Bilirubin toxicity to the central nervous system (CNS) is responsible for severe and permanent neurologic damage, resulting in hearing loss, cognitive, and movement impairment. Timely and effective management of severe neonatal hyperbilirubinemia by phototherapy or exchange transfusion is crucial for avoiding permanent neurological consequences, but these therapies are not always possible, particularly in low-income countries. To explore alternative options, we investigated a pharmaceutical approach focused on protecting the CNS from pigment toxicity, independently from serum bilirubin level. To this goal, we tested the ability of curcumin, a nutraceutical already used with relevant results in animal models as well as in clinics in other diseases, in the Gunn rat, the spontaneous model of neonatal hyperbilirubinemia. Curcumin treatment fully abolished the landmark cerebellar hypoplasia of Gunn rat, restoring the histological features, and reverting the behavioral abnormalities present in the hyperbilirubinemic rat. The protection was mediated by a multi-target action on the main bilirubin-induced pathological mechanism ongoing CNS damage (inflammation, redox imbalance, and glutamate neurotoxicity). If confirmed by independent studies, the result suggests the potential of curcumin as an alternative/complementary approach to bilirubin-induced brain damage in the clinical scenario.

Curcumin protects purkinje neurons, ameliorates motor function and reduces cerebellar atrophy in rat model of cerebellar ataxia induced by 3-AP.

BACKGROUND: Cerebellar ataxias comprise a group of terminal illnesses with ataxia as the main symptom. Curcumin as a yellow polyphenol was extracted from the rhizome ofCurcuma longa. Owing to its antioxidant, anti-inflammatory, anti-fibrotic and anti-tumor features, curcumin is considered as a potential therapeutic agent. AIM: In this study, we aim to investigate the neuroprotective effects of oral administration of curcumin on a rat model of cerebellar ataxia induced by neurotoxin 3-acetylpyridine. METHODS: The animals were randomly separated into three groups (control, 3-acetylpyridine, and curcumin + 3-acetylpyridine). Next, motor performance and muscle electromyography activity were assessed. Then, in the molecular part of the study, the anti-apoptotic role of curcumin in cerebellar ataxia and its relationship to protection of Purkinje cells were investigated. RESULTS: Curcumin treatment improved motor coordination and muscular activity, reduced cleaved caspase-3, and increased glutathione level in 3-AP-lesioned rats as well as total volumes of cerebellar granular and molecular layers. CONCLUSION: the present study implies that curcumin might have neuroprotective effects to counteract neurotoxicity of 3-AP-induced ataxia.

Developmental synapse remodeling in the cerebellum and visual thalamus.

Functional neural circuits of mature animals are shaped during postnatal development by eliminating early-formed redundant synapses and strengthening of necessary connections. In the nervous system of newborn animals, redundant synapses are only transient features of the circuit. During subsequent postnatal development, some synapses are strengthened whereas other redundant connections are weakened and eventually eliminated. In this review, we introduce recent studies on the mechanisms of developmental remodeling of climbing fiber-to-Purkinje cell synapses in the cerebellum and synapses from the retina to neurons in the dorsal lateral geniculate nucleus of the visual thalamus (retinogeniculate synapses). These are the two representative models of developmental synapse remodeling in the brain and they share basic principles, including dependency on neural activity. However, recent studies have disclosed that, in several respects, the two models use different molecules and strategies to establish mature synaptic connectivity. We describe similarities and differences between the two models and discuss remaining issues to be tackled in the future in order to understand the general schemes of developmental synapse remodeling.

Cerebellar Contribution to Preparatory Activity in Motor Neocortex.

In motor neocortex, preparatory activity predictive of specific movements is maintained by a positive feedback loop with the thalamus. Motor thalamus receives excitatory input from the cerebellum, which learns to generate predictive signals for motor control. The contribution of this pathway to neocortical preparatory signals remains poorly understood. Here, we show that, in a virtual reality conditioning task, cerebellar output neurons in the dentate nucleus exhibit preparatory activity similar to that in anterolateral motor cortex prior to reward acquisition. Silencing activity in dentate nucleus by photoactivating inhibitory Purkinje cells in the cerebellar cortex caused robust, short-latency suppression of preparatory activity in anterolateral motor cortex. Our results suggest that preparatory activity is controlled by a learned decrease of Purkinje cell firing in advance of reward under supervision of climbing fiber inputs signaling reward delivery. Thus, cerebellar computations exert a powerful influence on preparatory activity in motor neocortex.

Conversion of Graded Presynaptic Climbing Fiber Activity into Graded Postsynaptic Ca2+ Signals by Purkinje Cell Dendrites.

The brain must make sense of external stimuli to generate relevant behavior. We used a combination of in vivo approaches to investigate how the cerebellum processes sensory-related information. We found that the inferior olive encodes contexts of sensory-associated external cues in a graded manner, apparent in the presynaptic activity of their axonal projections (climbing fibers) in the cerebellar cortex. Individual climbing fibers were broadly responsive to different sensory modalities but relayed sensory-related information to the cortex in a lobule-dependent manner. Purkinje cell dendrites faithfully transformed this climbing fiber activity into dendrite-wide Ca2+ signals without a direct contribution from the mossy fiber pathway. These results demonstrate that the size of climbing-fiber-evoked Ca2+ signals in Purkinje cell dendrites is largely determined by the firing level of climbing fibers. This coding scheme emphasizes the overwhelming role of the inferior olive in generating salient signals useful for instructing plasticity and learning.