Search Results (93)

Radiation exposure causes neuroinflammation, oxidative stress, and neuronal loss, leading to cognitive and behavioral impairments. This study aims to evaluate the effect of niacin interventions on whole-brain irradiation (WBI)-induced cognitive and behavioral impairment. Female Wistar rats were randomly assigned to Control (Group 1), Radiation +Saline (Group 2), and Radiation +niacin (Group 3) groups. Rats in the irradiated groups (Groups 2 and 3) received a single dose of 20 Gy photon irradiation. Group 2 received water seven days after irradiation, while Group 3 received niacin (60 mg/kg, 2 mL) oral gavage for 15 days. On days 22, 23, and 24, behavioral assessments were performed, including the Open Field Test, the Sociability Test, and the Passive Avoidance Learning (PAL) task. Biochemical analyses included MDA, BDNF, TNF-α, CREB), SIRT1, and SIRT6 measured by ELISA. Histological assessments included neuronal density and GFAP immunostaining in CA1 and CA3 regions of the hippocampus and cerebellar Purkinje neurons. Radiation exposure importantly increased MDA and TNF-α levels, while SIRT1, SIRT6, BDNF, and CREB were notably reduced. This was accompanied by neuronal loss in the cerebellum and hippocampus, astrogliosis, and behavioral and cognitive deficits. Niacin treatment significantly decreased MDA and TNF-α levels while increasing BDNF, CREB, SIRT1, and SIRT6 expression, attenuating neuronal apoptosis. Immunohistochemical analysis demonstrated that niacin treatment enhanced neuronal density in the CA1 and CA3 regions of the hippocampus and cerebellar Purkinje neurons while reducing GFAP immunoreactivity in the CA1, CA3, and cerebellum following WBI. Behaviorally, niacin treatment improved social interaction, locomotor activity, and memory performance, underscoring its neuroprotective potential against WBI-induced damage. These findings suggest that niacin may ameliorate behavioral and cognitive impairments following whole brain irradiation by activating the SIRT1/CREB/BDNF or SIRT1/SIRT6/MDA/TNF-α signaling pathway. Full article

CASK-related disorders are a form of female-restricted intellectual disabilities associated with cerebellar and pontine hypoplasia. The CASK gene is regulated by X-chromosome inactivation, which results in a mosaic distribution of CASK-expressing and CASK-deficient neurons in the female brain. This mosaic distribution is believed to play a key role in the pathophysiology of X-linked neurological disorders; however, the detailed brain structure has not been extensively characterized. In this study, we used CASK heterozygous knockout (CASK-hKO) mice combined with X-linked GFP reporter mice to investigate motor abilities and the distribution of CASK-expressing cells in the brains of female CASK-hKO mice. The CASK-hKO mice exhibited motor deficits and cerebellar hypoplasia similar to those observed in patients with CASK-related disorders. Interestingly, although half of the cerebellar granule cells were CASK-negative during early postnatal development, almost all Purkinje cells and cerebellar granule cells were CASK-positive in adulthood, suggesting that CASK expression may determine the survival of cerebellar granule cells during postnatal development. We also analyzed CASK-hypomorphic mice, which express 50% less CASK than wild-type mice, and compared hemizygous males and heterozygous females. The CASK-hypomorphic heterozygous females displayed a thinner cerebellar cortex and a higher probability of CASK-positive granule cells in CASK-hKO females, suggesting that the survival of cerebellar granule cells is regulated by a combination of cell-autonomous and cell-competitive mechanisms between CASK-expressing and CASK-deficient cells, which are generated by X-chromosome inactivation. These findings provide new insights into the relationship between the mosaic distribution of cells established by X-chromosome inactivation and the pathophysiology of CASK-related disorders. Full article

Spinocerebellar ataxia (SCA), an autosomal dominant neurodegenerative condition, is marked by a gradual deterioration of cerebellar function. To date, more than 40 distinct SCA subtypes have been identified, with some attributed to CAG repeat expansions and others to point mutations or deletions. Among these, spinocerebellar ataxia type 14 (SCA14) stems from missense mutations or deletions within the PRKCG gene, encoding protein kinase C gamma (PKCγ), a pivotal signaling molecule abundant in Purkinje cells. Despite its significance, the precise mechanisms underlying how genetic alterations trigger Purkinje cell malfunction and degeneration remain elusive. Given the prominent role and high expression of PKCγ in Purkinje cells, SCA14 presents a unique opportunity to unravel the underlying pathogenesis. A straightforward hypothesis posits that alterations in the biological activity of PKCγ underlie the disease phenotype, and there are hints that mutated PKCγ proteins exhibit altered enzymatic function. Our prior research focused on the PKCγ-G118D mutation, commonly found in SCA14 patients, located in the regulatory domain of the protein. While cellular assays demonstrated enhanced enzymatic activity for PKCγ-G118D, transgenic mice carrying this mutation failed to exhibit suppressed dendritic development in cerebellar cultures, raising questions about its impact within living Purkinje cells. One hypothesis is that endogenous PKCγ might interfere with the expression or effect of PKCγ-G118D. To further investigate, we leveraged CRISPR-Cas9 technology to generate a PKCγ knockout mouse model and integrated it with an L7-based, Purkinje cell-specific transfection system to analyze the effects of G118D protein expression on the dendritic morphology of developing Purkinje cells. Our findings reveal that, utilizing this approach, PKCγ-G118D exerts a detrimental effect on Purkinje cell growth, confirming its negative influence, indicating that the potential of the G118D mutation to contribute to SCA14 pathogenesis. Full article

Hypothermia (HT) is used clinically for neonatal hypoxic–ischemic encephalopathy (HIE); however, the brain protection is incomplete and selective regional vulnerability and lifelong consequences remain. Refractory damage and impairment with HT cooling/rewarming could result from unchecked or altered persisting cell death and proteinopathy. We tested two hypotheses: (1) HT modifies neurodegeneration type, and (2) intrinsically disordered proteins (IDPs) and encephalopathy cause toxic conformer protein (TCP) proteinopathy neonatally. We studied postmortem human neonatal HIE cases with or without therapeutic HT, neonatal piglets subjected to global hypoxia-ischemia (HI) with and without HT or combinations of HI and quinolinic acid (QA) excitotoxicity surviving for 29–96 h to 14 days, and human oligodendrocytes and neurons exposed to QA for cell models. In human and piglet encephalopathies with normothermia, the neuropathology by hematoxylin and eosin staining was similar; necrotic cell degeneration predominated. With HT, neurodegeneration morphology shifted to apoptosis-necrosis hybrid and apoptotic forms in human HIE, while neurons in HI piglets were unshifting and protected robustly. Oligomers and putative TCPs of α-synuclein (αSyn), nitrated-Syn and aggregated αSyn, misfolded/oxidized superoxide dismutase-1 (SOD1), and prion protein (PrP) were detected with highly specific antibodies by immunohistochemistry, immunofluorescence, and immunoblotting. αSyn and SOD1 TCPs were seen in human HIE brains regardless of HT treatment. αSyn and SOD1 TCPs were detected as early as 29 h after injury in piglets and QA-injured human oligodendrocytes and neurons in culture. Cell immunophenotyping by immunofluorescence showed αSyn detected with antibodies to aggregated/oligomerized protein; nitrated-Syn accumulated in neurons, sometimes appearing as focal dendritic aggregations. Co-localization also showed aberrant αSyn accumulating in presynaptic terminals. Proteinase K-resistant PrP accumulated in ischemic Purkinje cells, and their target regions had PrP-positive neuritic plaque-like pathology. Immunofluorescence revealed misfolded/oxidized SOD1 in neurons, axons, astrocytes, and oligodendrocytes. HT attenuated TCP formation in piglets. We conclude that HT differentially affects brain damage in humans and piglets. HT shifts neuronal cell death to other forms in human while blocking ischemic necrosis in piglet for sustained protection. HI and excitotoxicity also acutely induce formation of TCPs and prion-like proteins from IDPs globally throughout the brain in gray matter and white matter. HT attenuates proteinopathy in piglets but seemingly not in humans. Shifting of cell death type and aberrant toxic protein formation could explain the selective system vulnerability, connectome spreading, and persistent damage seen in neonatal HIE leading to lifelong consequences even after HT treatment. Full article

This study investigates age-related neurodegeneration in the cerebellar cortex, emphasizing the role of IL-6 deficiency in preserving Purkinje cells. We found that apoptosis plays a minimal role in Purkinje cell loss by using 4-month- and 24-month-old wild-type (WT) and IL-6 knockout (IL-6KO) mice. At 24 months, WT mice exhibited severe Purkinje cell degeneration, including atrophic cell bodies, eosinophilic cytoplasm, pyknotic nuclei, mitochondrial disruption, and increased levels of lipofuscin-rich lysosomes. In contrast, IL-6KO mice showed fewer lysosomes, reduced mitochondrial damage, and less neuronal atrophy, indicating a neuroprotective effect. Lower p53 expression and decreased levels of its downstream effectors (p21, and Bax) in IL-6KO mice correlated with reduced cellular stress. Minimal changes in apoptotic markers (Bax and caspase-3) further reinforce the limited role of apoptosis. Neuroinflammation, marked by elevated GFAP, was prominent in aged WT mice but attenuated in IL-6KO mice. Reduced p53 accumulation, less severe neuroinflammation, and preserved metabolic homeostasis in IL-6KO mice correlated with improved Purkinje cell survival. These findings suggest that IL-6 accelerates neurodegeneration via p53-associated stress and inflammation, while IL-6 deficiency mitigates these effects. Targeting IL-6 signaling through anti-inflammatory strategies or IL-6 inhibition may offer a therapeutic approach for age-related neurodegenerative disorders. Full article

Niemann–Pick Disease Type C (NPC) is a hereditary neurodegenerative disease characterized by selective cell vulnerability, particularly affecting cerebellar anterior Purkinje neurons. These neurons exhibit a distinctive pattern of degeneration due to the loss of NPC1 and/or NPC2 protein function, progressively extending towards posterior cerebellar regions. Our study aimed to explore the early factors influencing this selective vulnerability of anterior Purkinje neurons in NPC. Oxytosis/ferroptosis, a novel form of regulated cell death, has been implicated in neurodegenerative diseases, with its inhibition showing promising therapeutic potential. Our laboratory has previously identified parallels between NPC cellular pathology and ferroptotic markers, including elevated levels of lipid peroxidation and iron, mitochondrial dysfunction, and Ca²⁺ dyshomeostasis. However, whether oxytosis/ferroptosis underlies NPC cellular pathology remains unexplored. We hypothesize that loss of NPC1 function increases vulnerability to ferroptosis and that anti-ferroptotic compounds will reverse NPC cellular pathology. Through bioinformatic analyses of pre-symptomatic Npc1^−/− Purkinje neurons and in vitro studies using primary dermal fibroblasts derived from NPC patients, we provide evidence suggesting that oxytosis/ferroptosis may play a pathogenic role in NPC. These findings highlight the potential of anti-ferroptotic compounds as a promising therapeutic strategy to mitigate neurodegeneration in NPC and potentially other related disorders. Full article

Precisely localizing the spatial distribution of proteins within various brain cell types and subcellular compartments, such as the synapses, is essential for generating and testing hypotheses to elucidate their roles in brain function. While the fms-like tyrosine kinase-3 (Flt3) has been extensively studied in the context of blood cell development and leukemia pathogenesis, its role in the brain remains poorly understood. Previous efforts to address this issue were hindered by the low expression levels of Flt3 and the limited sensitivity of the standard immunolabeling method, which were insufficient to reliably detect Flt3 protein in brain tissue. In this study, we systematically characterized Flt3 protein localization during brain development using a highly sensitive immunolabeling method based on alkaline phosphatase (AP) polymer biochemistry. This approach revealed a previously unrecognized neuron-selective Flt3 expression pattern in both mouse and human cerebella, with a developmental increase in total protein levels accompanied by a shift from a cytosolic to a dendritic subcellular distribution. Combining AP-polymer-based immunohistochemistry (AP-IHC) for Flt3 with conventional immunostaining of cell type marker proteins revealed parvalbumin- and calbindin-positive Purkinje cells to be the main cell type expressing Flt3 in the cerebellum. To validate the versatility of the AP-IHC method for detecting low-abundance neuronal proteins, we demonstrated robust labeling of Kir2.1, a potassium channel protein, in brain tissue sections from mouse, pig, and human samples. We further applied the AP-IHC method to human stem cell-derived neurons, effectively visualizing the postsynaptic density scaffold protein PSD95 within synapses. To our knowledge, this is the first study to employ an AP-IHC method combined with other standard immunofluorescent staining to co-detect weakly expressed neuronal proteins and other cellular markers in brain tissue and cultured neurons. Additionally, our findings uncover a previously unrecognized neuron-specific pattern of Flt3 expression in the cerebellum, laying the foundation for future mechanistic studies on its role in normal brain development and neurological disorders. Full article

Semaphorin 3A (Sema3A) is an axon guidance molecule, which is also abundant in the adult central nervous system (CNS), particularly in perineuronal nets (PNNs). PNNs are extracellular matrix structures that restrict plasticity. The cellular sources of Sema3A in PNNs are unknown. Most Sema3A-bearing neurons do not express Sema3A mRNA, suggesting that Sema3A may be released from other neurons. Another potential source of Sema3A is the choroid plexus. To identify sources of PNN-associated Sema3A, we focused on the cerebellar nuclei, which contain Sema3A⁺ PNNs. Cerebellar nuclei neurons receive prominent input from Purkinje cells (PCs), which express high levels of Sema3A mRNA. By using a non-invasive viral vector approach, we overexpressed Cre in PCs, the choroid plexus, or throughout the CNS of Sema3A^fl/fl mice. Knocking out Sema3A in PCs or the choroid plexus was not sufficient to decrease the amount of PNN-associated Sema3A. Alternatively, knocking out Sema3A throughout the CNS induced a decrease in PNN-associated Sema3A. However, motor deficits, microgliosis, and neurodegeneration were observed, which were due to Cre toxicity. Our study represents the first attempt to unravel cellular sources of PNN-associated Sema3A and shows that non-invasive viral-mediated Cre expression throughout the CNS could lead to toxicity, complicating the interpretation of Cre-mediated Sema3A knock-out. Full article

In recent decades, the scientific community has faced a major challenge in the search for new therapies that can slow down or alleviate the process of neuronal death that accompanies neurodegenerative diseases. This study aimed to identify an effective therapy using neurotrophic factors to delay the rapid and aggressive cerebellar degeneration experienced by the Purkinje Cell Degeneration (PCD) mouse, a model of childhood-onset neurodegeneration with cerebellar atrophy (CONDCA). Initially, we analyzed the changes in the expression of several neurotrophic factors related to the degenerative process itself, identifying changes in insulin-like growth factor 1 (IGF-1) and Vascular Endothelial Growth Factor B (VEGF-B) in the affected animals. Then, we administered pharmacological treatments using human recombinant IGF-1 (rhIGF-1) or VEGF-B (rhVEGF-B) proteins, considering their temporal variations during the degenerative process. The effects of these treatments on motor, cognitive, and social behavior, as well as on cerebellar destructuration were analyzed. Whereas treatment with rhIGF-1 did not demonstrate any neuroprotective effect, rhVEGF-B administration at moderate dosages stopped the process of neuronal death and restored motor, cognitive, and social functions altered in PCD mice (and CONDCA patients). However, increasing the frequency of rhVEGF-B administration had a detrimental effect on Purkinje cell survival, suggesting an inverted U-shaped dose–response curve of this substance. Additionally, we demonstrate that this neuroprotective effect was achieved through a partial inhibition or delay of apoptosis. These findings provide strong evidence supporting the use of rhVEGF-B as a pharmacological agent to limit severe cerebellar neurodegenerative processes. Full article

Paraneoplastic cerebellar degeneration (PCD) is a rapidly progressive, immune-mediated syndrome characterized by the degeneration of Purkinje cells, often associated with the presence of antibodies targeting intracellular antigens within these cells. These autoantibodies are implicated in the induction of cytotoxicity, leading to Purkinje cell death, as demonstrated in in vitro models. However, the precise roles of antibodies and T lymphocytes in mediating neuronal injury remain a subject of ongoing research, with T cells appearing to be the main effectors of cerebellar injury. Notably, at least 50% of PCD cases involve anti-Yo autoantibodies, also referred to as anti-PCA1 (Purkinje cell antigen 1) antibodies, which specifically target cerebellar degeneration-related protein 2 (CDR2) and its paralogue, CDR2-like (CDR2L). Another recognized antigen is CDR 34, a 34 kDa Purkinje cell antigen characterized by tandem repeats and a B-cell epitope; its detection in non-cerebellar tissues necessitates further in situ hybridization studies. Onconeural antigens are expressed in both Purkinje cells and tumour cells, where they localize in the cytoplasm and associate with membrane-bound and free ribosomes, playing critical roles in regulating transcription and calcium homeostasis. Recent studies suggest that the breakdown of immune tolerance is linked to genetic alterations in tumour cell antigens, leading to the formation of neoantigens that can elicit autoreactive T cells, which may underscore the function of Yo antibodies. In vitro studies indicate that anti-Yo antibodies can induce cell death independent of T lymphocytes. The disease progresses by initial lymphocytic infiltration, followed by a rapid loss of Purkinje cells without significant inflammation. However, in vivo models showcase that anti-Yo PCD is primarily T-cell mediated, with antibodies serving as biomarkers rather than direct effectors of neuronal death. This review examines the mechanisms underlying PCD, focusing on the roles of CDR2 and CDR2L in tumour development and their potential role in the degeneration of cerebellar Purkinje neurons. A comprehensive understanding of these processes is essential for advancing diagnostic, prognostic, and therapeutic strategies for PCD and associated malignancies. Full article

The purpose of this study was to examine the relationship between demyelination and cellular reactions in the cerebellum of Canine Distemper Virus (CDV)-infected dogs. We subdivided the disease staging by adding the degree of demyelination determined by Luxol Fast Blue staining to the previously reported disease staging from the acute stage to the chronic stage, and investigated the relationship between demyelination in the cerebellum and the number and histological changes in astroglia, microglia, and Purkinje cells in each stage. Reactions of astrocytes and microglia were observed at an early stage when demyelination was not evident. Changes progressed with demyelination. Demyelination initially began in the medulla adjoining the fourth ventricle and gradually spread to the entire cerebellum, including the lobes. CDV immune-positive granules were seen from the early stage, and inclusion bodies also appeared at the same time. CDV immune-positive reaction and inclusion bodies were observed in astrocytes, microglia, neurons, ependymal cells, and even leptomeningeal mononuclear cells. On the other hand, infiltration of CDV-immunoreactive particles from the pia mater to the gray matter and further into the white matter through the granular layer was observed from an early stage. Purkinje cells decreased from the intermediate stage, and a decrease in cells in the granular layer was also observed. There was no clear association between age and each stage, and the stages did not progress with age. Full article

Physical overexertion surpassing the functional capacity of the nervous system causes the hyperactivation of the neural structures of the cerebellum. In turn, it causes the depletion of intracellular resources and progressive structural changes in cerebellar cells and fibers. These degenerative changes may lead to cerebellar dysfunction, including the worsening of coordination, balance, and motor functions. In order to maintain the health and functioning of the cerebellum and the nervous system in general, one needs to avoid physical overexertion and have enough time to recover. Three major types of Purkinje cells were identified in control group animals. After the forced swimming test, animals had significant morphological changes in pyriform cells, granule cells, internuncial neurons, and neuroglial cells. In particular, the extreme degeneration of granule cells was manifested via their fusion into conglomerates. These changes demonstrate that neurodegeneration in the cerebellum takes place in response to physical overexertion. Full article

Agathisflavone is a flavonoid that exhibits anti-inflammatory and anti-oxidative properties. Here, we investigated the neuroprotective effects of agathisflavone on central nervous system (CNS) neurons and glia in the cerebellar slice ex vivo model of neonatal ischemia. Cerebellar slices from neonatal mice, in which glial fibrillary acidic protein (GFAP) and SOX10 drive expression of enhanced green fluorescent protein (EGFP), were used to identify astrocytes and oligodendrocytes, respectively. Agathisflavone (10 μM) was administered preventively for 60 min before inducing ischemia by oxygen and glucose deprivation (OGD) for 60 min and compared to controls maintained in normal oxygen and glucose (OGN). The density of SOX-10⁺ oligodendrocyte lineage cells and NG2 immunopositive oligodendrocyte progenitor cells (OPCs) were not altered in OGD, but it resulted in significant oligodendroglial cell atrophy marked by the retraction of their processes, and this was prevented by agathisflavone. OGD caused marked axonal demyelination, determined by myelin basic protein (MBP) and neurofilament (NF70) immunofluorescence, and this was blocked by agathisflavone preventative treatment. OGD also resulted in astrocyte reactivity, exhibited by increased GFAP-EGFP fluorescence and decreased expression of glutamate synthetase (GS), and this was prevented by agathisflavone pretreatment. In addition, agathisflavone protected Purkinje neurons from ischemic damage, assessed by calbindin (CB) immunofluorescence. The results demonstrate that agathisflavone protects neuronal and myelin integrity in ischemia, which is associated with the modulation of glial responses in the face of ischemic damage. Full article

Introduction: Reelin is a neuropeptide responsible for the migration and positioning of pyramidal neurons, interneurons, and Purkinje cells. In adulthood, it still supports neuroplasticity, especially dendritic spines formation and glutamatergic neurotransmission. Genetic studies have confirmed the involvement of reelin system failure in the etiopathogenesis of mental diseases, including schizophrenia. Given the role of reelin in brain cytoarchitectonics and the regularly observed reduction in its activity in prefrontal areas in cases of schizophrenia, dysfunction of the reelin pathway fits the neurodevelopmental hypothesis of schizophrenia, both as a biochemical predisposition and/or the ultimate trigger of psychosis and as a biosocial factor determining the clinical course, and finally, as a potential target for disease monitoring and treatment. Aim: The purpose of this study was to examine associations of the reelin blood level with clinical and neurocognitive parameters during an intensive, structured neurofeedback therapy of patients with schizophrenia. Methods: Thirty-seven male patients with paranoid schizophrenia were randomly divided into two groups: a group with 3-month neurofeedback as an add-on to ongoing antipsychotic treatment (NF, N18), and a control group with standard social support and antipsychotic treatment (CON, N19). The reelin serum concentration, clinical and neurocognitive tests were compared between the groups. Results: After 3-month trial (T2), the reelin serum level increased in the NF group vs. the CON group. The negative and general symptoms of PANSS (Positive and Negative Syndrome Scale) were reduced significantly more in the NF group at T2, and the d2 (d2 Sustained Attention Test) and BCIS (Beck Cognitive Insight Scale) scores improved only in the NF group. The AIS scores improved more dynamically in the NF group, but not enough to differentiate them from the CON group at T2. Conclusions: The clinical and neurocognitive improvement within the 3-month NF add-on therapy trial was associated with a significant increase of reelin serum level in schizophrenia patients. Full article

We aimed to produce a mouse model of spinocerebellar ataxia type 3 (SCA3) using the mouse blood–brain barrier (BBB)-penetrating adeno-associated virus (AAV)-PHP.B. Four-to-five-week-old C57BL/6 mice received injections of high-dose (2.0 × 10¹¹ vg/mouse) or low-dose (5.0 × 10¹⁰ vg/mouse) AAV-PHP.B encoding a SCA3 causative gene containing abnormally long 89 CAG repeats [ATXN3(Q89)] under the control of the ubiquitous chicken β-actin hybrid (CBh) promoter. Control mice received high doses of AAV-PHP.B encoding ATXN3 with non-pathogenic 15 CAG repeats [ATXN3(Q15)] or phosphate-buffered saline (PBS) alone. More than half of the mice injected with high doses of AAV-PHP.B encoding ATXN3(Q89) died within 4 weeks after the injection. No mice in other groups died during the 12-week observation period. Mice injected with low doses of AAV-PHP.B encoding ATXN3(Q89) exhibited progressive motor uncoordination starting 4 weeks and a shorter stride in footprint analysis performed at 12 weeks post-AAV injection. Immunohistochemistry showed thinning of the molecular layer and the formation of nuclear inclusions in Purkinje cells from mice injected with low doses of AAV-PHP.B encoding ATXN3(Q89). Moreover, ATXN3(Q89) expression significantly reduced the number of large projection neurons in the cerebellar nuclei to one third of that observed in mice expressing ATXN3(Q15). This AAV-based approach is superior to conventional methods in that the required number of model mice can be created simply by injecting AAV, and the expression levels of the responsible gene can be adjusted by changing the amount of AAV injected. Moreover, this method may be applied to produce SCA3 models in non-human primates. Full article