Heterozygous mutations in GBA, the gene encoding the lysosomal enzyme glucosylceramidase beta/β-glucocerebrosidase, comprise the most common genetic risk factor for Parkinson disease (PD), but the mechanisms underlying this association remain unclear. Here, we show that in GbaL444P/WT knockin mice, the L444P heterozygous Gba mutation triggers mitochondrial dysfunction by inhibiting autophagy and mitochondrial priming, two steps critical for the selective removal of dysfunctional mitochondria by autophagy, a process known as mitophagy. In SHSY-5Y neuroblastoma cells, the overexpression of L444P GBA impeded mitochondrial priming and autophagy induction when endogenous lysosomal GBA activity remained intact. By contrast, genetic depletion of GBA inhibited lysosomal clearance of autophagic cargo. The link between heterozygous GBA mutations and impaired mitophagy was corroborated in postmortem brain tissue from PD patients carrying heterozygous GBA mutations, where we found increased mitochondrial content, mitochondria oxidative stress and impaired autophagy. Our findings thus suggest a mechanistic basis for mitochondrial dysfunction associated with GBA heterozygous mutations. Abbreviations: AMBRA1: autophagy/beclin 1 regulator 1; BECN1: beclin 1, autophagy related; BNIP3L/Nix: BCL2/adenovirus E1B interacting protein 3-like; CCCP: carbonyl cyanide 3-chloroyphenylhydrazone; CYCS: cytochrome c, somatic; DNM1L/DRP1: dynamin 1-like; ER: endoplasmic reticulum; GBA: glucosylceramidase beta; GBA-PD: Parkinson disease with heterozygous GBA mutations; GD: Gaucher disease; GFP: green fluorescent protein; LC3B: microtubule-associated protein 1 light chain 3 beta; LC3B-II: lipidated form of microtubule-associated protein 1 light chain 3 beta; MitoGreen: MitoTracker Green; MitoRed: MitoTracker Red; MMP: mitochondrial membrane potential; MTOR: mechanistic target of rapamycin kinase; MYC: MYC proto-oncogene, bHLH transcription factor; NBR1: NBR1, autophagy cargo receptor; Non-GBA-PD: Parkinson disease without GBA mutations; PD: Parkinson disease; PINK1: PTEN induced putative kinase 1; PRKN/PARK2: parkin RBR E3 ubiquitin protein ligase; RFP: red fluorescent protein; ROS: reactive oxygen species; SNCA: synuclein alpha; SQSTM1/p62: sequestosome 1; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20; VDAC1/Porin: voltage dependent anion channel 1; WT: wild type.

Developmental alterations of excitatory synapses are implicated in autism spectrum disorders (ASDs). Here, we report increased dendritic spine density with reduced developmental spine pruning in layer V pyramidal neurons in postmortem ASD temporal lobe. These spine deficits correlate with hyperactivated mTOR and impaired autophagy. In Tsc2 ± ASD mice where mTOR is constitutively overactive, we observed postnatal spine pruning defects, blockade of autophagy, and ASD-like social behaviors. The mTOR inhibitor rapamycin corrected ASD-like behaviors and spine pruning defects in Tsc2 ± mice, but not in Atg7(CKO) neuronal autophagy-deficient mice or Tsc2 ± :Atg7(CKO) double mutants. Neuronal autophagy furthermore enabled spine elimination with no effects on spine formation. Our findings suggest that mTOR-regulated autophagy is required for developmental spine pruning, and activation of neuronal autophagy corrects synaptic pathology and social behavior deficits in ASD models with hyperactivated mTOR.

Autism spectrum disorder (ASD) consists of a group of complex developmental disabilities characterized by impaired social interactions, deficits in communication and repetitive behavior. Multiple lines of evidence implicate mitochondrial dysfunction in ASD. In postmortem BA21 temporal cortex, a region that exhibits synaptic pathology in ASD, we found that compared to controls, ASD patients exhibited altered protein levels of mitochondria respiratory chain protein complexes, decreased Complex I and IV activities, decreased mitochondrial antioxidant enzyme SOD2, and greater oxidative DNA damage. Mitochondrial membrane mass was higher in ASD brain, as indicated by higher protein levels of mitochondrial membrane proteins Tom20, Tim23 and porin. No differences were observed in either mitochondrial DNA or levels of the mitochondrial gene transcription factor TFAM or cofactor PGC1α, indicating that a mechanism other than alterations in mitochondrial genome or mitochondrial biogenesis underlies these mitochondrial abnormalities. We further identified higher levels of the mitochondrial fission proteins (Fis1 and Drp1) and decreased levels of the fusion proteins (Mfn1, Mfn2 and Opa1) in ASD patients, indicating altered mitochondrial dynamics in ASD brain. Many of these changes were evident in cortical pyramidal neurons, and were observed in ASD children but were less pronounced or absent in adult patients. Together, these findings provide evidence that mitochondrial function and intracellular redox status are compromised in pyramidal neurons in ASD brain and that mitochondrial dysfunction occurs during early childhood when ASD symptoms appear.

A promising new approach, transcranial direct current stimulation (tDCS) has recently been used as a therapeutic modality for cerebellar ataxia. However, the strength of the conclusions drawn from individual studies in the current literature may be constrained by the small sample size of each trial. Following a systematic literature retrieval of studies, meta-analyses were conducted by pooling the standardized mean differences (SMDs) using random-effects models to assess the efficacy of tDCS on cerebellar ataxia, measured by standard clinical rating scales. Domain-specific effects of tDCS on gait and hand function were further evaluated based on 8-m walk and 9-hole peg test performance times, respectively. To determine the safety of tDCS, the incidences of adverse effects were analyzed using risk differences. Out of 293 citations, 5 randomized controlled trials involving a total of 72 participants with cerebellar ataxia were included. Meta-analysis indicated a 26.1% (p = 0.003) improvement in ataxia immediately after tDCS with sustained efficacy over months (28.2% improvement after 3 months, p = 0.04) when compared with sham stimulation. tDCS seems to be domain-specific as the current analysis suggested a positive effect on gait (16.3% improvement, p = 0.04) and failed to reveal differences for hand function (p = 0.10) with respect to sham. The incidence of adverse events in tDCS and sham groups was similar. tDCS is an effective intervention for mitigating ataxia symptoms with lasting results that can be sustained for months. This treatment shows preferential effects on gait ataxia and is relatively safe.

The mTORC1 node plays a major role in autophagy modulation. We report a role of the ubiquitous Gαq subunit, a known transducer of plasma membrane G protein-coupled receptors signaling, as a core modulator of mTORC1 and autophagy. Cells lacking Gαq/11 display higher basal autophagy, enhanced autophagy induction upon different types of nutrient stress along with a decreased mTORC1 activation status. They are also unable to reactivate mTORC1 and thus inactivate ongoing autophagy upon nutrient recovery. Conversely, stimulation of Gαq/11 promotes sustained mTORC1 pathway activation and reversion of autophagy promoted by serum or amino acids removal. Gαq is present in autophagic compartments and lysosomes and is part of the mTORC1 multi-molecular complex, contributing to its assembly and activation via its nutrient status-sensitive interaction with p62, which displays features of a Gαq effector. Gαq emerges as a central regulator of the autophagy machinery required to maintain cellular homeostasis upon nutrient fluctuations.

The most common genetic risk factors for Parkinson's disease (PD) are a set of heterozygous mutant (MT) alleles of the GBA1 gene that encodes β-glucocerebrosidase (GCase), an enzyme normally trafficked through the ER/Golgi apparatus to the lysosomal lumen. We found that half of the GCase in lysosomes from postmortem human GBA-PD brains was present on the lysosomal surface and that this mislocalization depends on a pentapeptide motif in GCase used to target cytosolic protein for degradation by chaperone-mediated autophagy (CMA). MT GCase at the lysosomal surface inhibits CMA, causing accumulation of CMA substrates including α-synuclein. Single-cell transcriptional analysis and proteomics of brains from GBA-PD patients confirmed reduced CMA activity and proteome changes comparable to those in CMA-deficient mouse brain. Loss of the MT GCase CMA motif rescued primary substantia nigra dopaminergic neurons from MT GCase-induced neuronal death. We conclude that MT GBA1 alleles block CMA function and produce α-synuclein accumulation.

The activation of mostly quiescent haematopoietic stem cells (HSCs) is a prerequisite for life-long production of blood cells1. This process requires major molecular adaptations to allow HSCs to meet the regulatory and metabolic requirements for cell division2-4. The mechanisms that govern cellular reprograming upon stem-cell activation, and the subsequent return of stem cells to quiescence, have not been fully characterized. Here we show that chaperone-mediated autophagy (CMA)5, a selective form of lysosomal protein degradation, is involved in sustaining HSC function in adult mice. CMA is required for protein quality control in stem cells and for the upregulation of fatty acid metabolism upon HSC activation. We find that CMA activity in HSCs decreases with age and show that genetic or pharmacological activation of CMA can restore the functionality of old mouse and human HSCs. Together, our findings provide mechanistic insights into a role for CMA in sustaining quality control, appropriate energetics and overall long-term HSC function. Our work suggests that CMA may be a promising therapeutic target for enhancing HSC function in conditions such as ageing or stem-cell transplantation.

Accumulating evidence from genetic and biochemical studies implicates dysfunction of the autophagic-lysosomal pathway as a key feature in the pathogenesis of Parkinson's disease (PD). Most studies have focused on accumulation of neurotoxic α-synuclein secondary to defects in autophagy as the cause of neurodegeneration, but abnormalities of the autophagic-lysosomal system likely mediate toxicity through multiple mechanisms. To further explore how endolysosomal dysfunction causes PD-related neurodegeneration, we generated a murine model of Kufor-Rakeb syndrome (KRS), characterized by early-onset Parkinsonism with additional neurological features. KRS is caused by recessive loss-of-function mutations in the ATP13A2 gene encoding the endolysosomal ATPase ATP13A2. We show that loss of ATP13A2 causes a specific protein trafficking defect, and that Atp13a2 null mice develop age-related motor dysfunction that is preceded by neuropathological changes, including gliosis, accumulation of ubiquitinated protein aggregates, lipofuscinosis, and endolysosomal abnormalities. Contrary to predictions from in vitro data, in vivo mouse genetic studies demonstrate that these phenotypes are α-synuclein independent. Our findings indicate that endolysosomal dysfunction and abnormalities of α-synuclein homeostasis are not synonymous, even in the context of an endolysosomal genetic defect linked to Parkinsonism, and highlight the presence of α-synuclein-independent neurotoxicity consequent to endolysosomal dysfunction.

The P140 peptide, a 21-mer linear peptide (sequence 131-151) generated from the spliceosomal SNRNP70/U1-70K protein, contains a phosphoserine residue at position 140. It significantly ameliorates clinical manifestations in autoimmune patients with systemic lupus erythematosus and enhances survival in MRL/lpr lupus-prone mice. Previous studies showed that after P140 treatment, there is an accumulation of autophagy markers sequestosome 1/p62 and MAP1LC3-II in MRL/lpr B cells, consistent with a downregulation of autophagic flux. We now identify chaperone-mediated autophagy (CMA) as a target of P140 and demonstrate that its inhibitory effect on CMA is likely tied to its ability to alter the composition of HSPA8/HSC70 heterocomplexes. As in the case of HSPA8, expression of the limiting CMA component LAMP2A, which is increased in MRL/lpr B cells, is downregulated after P140 treatment. We also show that P140, but not the unphosphorylated peptide, uses the clathrin-dependent endo-lysosomal pathway to enter into MRL/lpr B lymphocytes and accumulates in the lysosomal lumen where it may directly hamper lysosomal HSPA8 chaperoning functions, and also destabilize LAMP2A in lysosomes as a result of its effect on HSP90AA1. This dual effect may interfere with the endogenous autoantigen processing and loading to major histocompatibility complex class II molecules and as a consequence, lead to lower activation of autoreactive T cells. These results shed light on mechanisms by which P140 can modulate lupus disease and exert its tolerogenic activity in patients. The unique selective inhibitory effect of the P140 peptide on CMA may be harnessed in other pathological conditions in which reduction of CMA activity would be desired.

Microglia-mediated neuroinflammation contributes to multiple neurodegenerative disorders, including PD. Therefore, the regulation of microglial activation probably has the therapeutic potential. This study is aimed to determine whether NBP could suppress microglial activation and protect dopaminergic neurons from excessive neuroinflammation. In the present study, MPTP-induced PD model was established to explore the neuroprotective and anti-inflammatory effect of NBP. We assessed motor deficits, dopaminergic neurodegeneration and microglial activation in PD mice. In vitro, the anti-inflammatory activity of NBP was confirmed by cell viability assay of SH-SY5Y cells after being treated with conditioned medium from LPS-stimulated BV-2 cells and from 1-Methyl-4-phenylpyridinium iodide (MPP+)-stimulated BV-2 cells. The expression of pro-inflammatory molecules was determined by RT-PCR, Western Blot and ELISA assay. The generation of NO and ROS were also assessed. The involvement of signaling pathways such as MAPK, NF-κB, and PI3k/Akt were further investigated by Western Blot and immunofluorescence assay. The neuroprotective effect of NBP was demonstrated in vivo as shown by the improvement of dopaminergic neurodegeneration, motor deficits and microglial activation in MPTP-induced mouse model of PD. The expression of pro-inflammatory mediators was also reduced by NBP administration. In vitro, NBP also protected dopaminergic neurons from neurotoxicity induced by activated microglia. NBP pretreatment not only reduced pro-inflammatory molecules, but also suppressed NO release and ROS generation in BV-2 cells. Further mechanism research suggested that the inactivation of MAPK, NF-κB and PI3K/Akt may involve in anti-neuroinflammation role of NBP. In conclusion, our results revealed that NBP exerted dopaminergic neuroprotection through inhibition of microglia-mediated neuroinflammation, suggesting the promising therapeutic effect of NBP for PD.

Chaperone-mediated autophagy (CMA) serves as quality control during stress conditions through selective degradation of cytosolic proteins in lysosomes. Humanin (HN) is a mitochondria-associated peptide that offers cytoprotective, cardioprotective, and neuroprotective effects in vivo and in vitro. In this study, we demonstrate that HN directly activates CMA by increasing substrate binding and translocation into lysosomes. The potent HN analogue HNG protects from stressor-induced cell death in fibroblasts, cardiomyoblasts, neuronal cells, and primary cardiomyocytes. The protective effects are lost in CMA-deficient cells, suggesting that they are mediated through the activation of CMA. We identified that a fraction of endogenous HN is present at the cytosolic side of the lysosomal membrane, where it interacts with heat shock protein 90 (HSP90) and stabilizes binding of this chaperone to CMA substrates as they bind to the membrane. Inhibition of HSP90 blocks the effect of HNG on substrate translocation and abolishes the cytoprotective effects. Our study provides a novel mechanism by which HN exerts its cardioprotective and neuroprotective effects.

Tremor is the most common movement disorder; however, the pathophysiology of tremor remains elusive. While several neuropathological alterations in tremor disorders have been observed in post-mortem studies of human brains, a full understanding of the relationship between brain circuitry alterations and tremor requires testing in animal models. Additionally, tremor animal models are critical for our understanding of tremor pathophysiology, and/or to serve as a platform for therapy development.

Both silent information regulator 1 (SIRT1) and hypoxia inducible factor 1 (HIF-1) have been found to play important roles in the pathophysiology of Parkinson's disease (PD). However, their mechanisms and their relationship still require further study. In the present study, we focused on the change and relationship of SIRT1 and HIF-1α in PD. PD cell models were established by using methyl-4-phenylpyridinium (MPP(+)), which induced inhibition of cell proliferation, cell cycle arrest and apoptosis. We found that the expression of HIF-1α and its target genes VEGFA and LDHA increased and that SIRT1 expression was inhibited in MPP(+) treated cells. With further analysis, we found that the acetylation of H3K14 combined with the HIF-1α promoter was dramatically increased in cells treated with MPP(+), which resulted in the transcriptional activation of HIF-1α. Moreover, the acetylation of H3K14 and the expression of HIF-1α increased when SIRT1 was knocked down, suggesting that SIRT1 was involved in the epigenetic regulation of HIF-1α. At last, phenformin, another mitochondrial complex1 inhibitor, was used to testify that the increased HIF-1a was not due to off target effects of MPP(+). Therefore, our results support a link between PD and SIRT1/HIF-1α signaling, which may serve as a clue for understanding PD.

Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of familial Parkinson's disease. We found LRRK2 to be degraded in lysosomes by chaperone-mediated autophagy (CMA), whereas the most common pathogenic mutant form of LRRK2, G2019S, was poorly degraded by this pathway. In contrast to the behavior of typical CMA substrates, lysosomal binding of both wild-type and several pathogenic mutant LRRK2 proteins was enhanced in the presence of other CMA substrates, which interfered with the organization of the CMA translocation complex, resulting in defective CMA. Cells responded to such LRRK2-mediated CMA compromise by increasing levels of the CMA lysosomal receptor, as seen in neuronal cultures and brains of LRRK2 transgenic mice, induced pluripotent stem cell-derived dopaminergic neurons and brains of Parkinson's disease patients with LRRK2 mutations. This newly described LRRK2 self-perpetuating inhibitory effect on CMA could underlie toxicity in Parkinson's disease by compromising the degradation of α-synuclein, another Parkinson's disease-related protein degraded by this pathway.

The principal goal of this study was to determine the effect of the photoperiod on oxidative damage biomarkers in rats submitted to different light/darkness patterns, in a hyperlipidemic nephropathy model (induced by adriamycin), as well as its possible relationship with melatonin and leptin secretion rhythms. To test this hypothesis, six different groups were used (N = 6 rats per group): control (12 h/12h light:dark); exposure to permanent illumination (24 h light); exposure to darkness (22 h dark); injected with adriamycin, 12h/12h light:dark; injected with adriamycin + exposure to permanent illumination and injected with adriamycin + exposure to darkness (22 h dark). The different photoperiods were begun two weeks prior to medication and were maintained up to the day of the animal's sacrifice, ten days after medication. The following parameters were analysed: i) weight evolution; ii) in plasma: urea, creatinine, uric acid, total proteins, albumen, lactate dehydrogenase, creatinine-quinase, aspartate aminotransferase, alanine aminotransferase and total cholesterol; iii) in urine: urea, creatinine, total proteins and microalbumen; iv) biomarkers of oxidative damage in kidneys, heart, liver and brain: lipoperoxides, total glutathione, reduced glutathione, catalase, glutathione peroxidase, glutathione reductase and glutathione transferase; v) melatonin (pineal gland tissue and plasma) and leptin (plasma). From the results obtained it was concluded that the administration of adriamycin generated oxidative stress in renal, cerebral, hepatic and cardiac tissue. Additionally, in the healthy animal, but of a lesser relevance in the adriamycin animal, permanent light worsened the oxidative stress, whereas darkness improved it. This could be related to the circadian rhythm of the inverse release shown by melatonin and leptin, accentuating the release of melatonin in the darkness phase and that of leptin in the light phase. The correlation between melatonin and leptin in the healthy animal seemed to confirm the relationship between both variables and their influence on oxidative damage biomarkers.

Rapid eye movement sleep behavior disorder (RBD) and Parkinson's disease (PD) are two distinct clinical diseases but they share some common pathological and anatomical characteristics. This study aims to confirm the clinical features of RBD in Chinese PD patients.

The cerebellum is best known for its role in controlling motor behaviors. However, recent work supports the view that it also influences non-motor behaviors. The contribution of the cerebellum towards different brain functions is underscored by its involvement in a diverse and increasing number of neurological and neuropsychiatric conditions including ataxia, dystonia, essential tremor, Parkinson's disease (PD), epilepsy, stroke, multiple sclerosis, autism spectrum disorders, dyslexia, attention deficit hyperactivity disorder (ADHD), and schizophrenia. Although there are no cures for these conditions, cerebellar stimulation is quickly gaining attention for symptomatic alleviation, as cerebellar circuitry has arisen as a promising target for invasive and non-invasive neuromodulation. This consensus paper brings together experts from the fields of neurophysiology, neurology, and neurosurgery to discuss recent efforts in using the cerebellum as a therapeutic intervention. We report on the most advanced techniques for manipulating cerebellar circuits in humans and animal models and define key hurdles and questions for moving forward.

Pyramidal signs are common but often under-recognized in multiple system atrophy (MSA). The clinicopathological correlates of pyramidal signs in MSA are not well characterized. The present study aims to understand the role of pyramidal signs in MSA.

Damage to the cerebellum can affect neural structures involved in locomotion, causing gait and balance disorders. However, the integrity of cerebellum does not seem to be critical in managing sudden and unexpected environmental changes such as disturbances during walking. The cerebellum also plays a functional role in motor learning. Only a few effective therapies exist for individuals with cerebellar ataxia. With these in mind, we aimed at investigating: (1) corrective response of participants with cerebellar ataxia (CA) to unexpected gait perturbations; and (2) the effectiveness of a perturbation-based training to improve their dynamic stability during balance recovery responses and steady walking. Specifically, we hypothesized that: (1) CA group can show a corrective behavior similar to that of a healthy control group; (2) the exposure to a perturbation-based treatment can exploit residual learning capability, thus improving their dynamic stability during balance recovery responses and steady locomotion.

Loss of neuronal proteostasis, a common feature of the aging brain, is accelerated in neurodegenerative disorders, including different types of tauopathies. Aberrant turnover of tau, a microtubule-stabilizing protein, contributes to its accumulation and subsequent toxicity in tauopathy patients' brains. A direct toxic effect of pathogenic forms of tau on the proteolytic systems that normally contribute to their turnover has been proposed. In this study, we analyzed the contribution of three different types of autophagy, macroautophagy, chaperone-mediated autophagy, and endosomal microautophagy to the degradation of tau protein variants and tau mutations associated with this age-related disease. We have found that the pathogenic P301L mutation inhibits degradation of tau by any of the three autophagic pathways, whereas the risk-associated tau mutation A152T reroutes tau for degradation through a different autophagy pathway. We also found defective autophagic degradation of tau when using mutations that mimic common posttranslational modifications in tau or known to promote its aggregation. Interestingly, although most mutations markedly reduced degradation of tau through autophagy, the step of this process preferentially affected varies depending on the type of tau mutation. Overall, our studies unveil a complex interplay between the multiple modifications of tau and selective forms of autophagy that may determine its physiological degradation and its faulty clearance in the disease context.