Deficiency of parkin causes neurodegeneration and accumulation of pathological α-synuclein in monkey models

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Abstract

Parkinson’s disease (PD) is characterized by age-dependent neurodegeneration and the accumulation of toxic phosphorylated α-synuclein (pS129-α-syn). The mechanisms underlying these crucial pathological changes remain unclear. Mutations in parkin RBR E3 ubiquitin protein ligase (PARK2), the gene encoding parkin that is phosphorylated by PTEN-induced putative kinase 1 (PINK1) to participate in mitophagy, cause early onset PD. However, current parkin-KO mouse and pig models do not exhibit neurodegeneration. In the current study, we utilized CRISPR/Cas9 technology to establish parkin-deficient monkey models at different ages. We found that parkin deficiency leads to substantia nigra neurodegeneration in adult monkey brains and that parkin phosphorylation decreases with aging, primarily due to increased insolubility of parkin. Phosphorylated parkin is important for neuroprotection and the reduction of pS129-α-syn. Consistently, overexpression of WT parkin, but not a mutant form that cannot be phosphorylated by PINK1, reduced the accumulation of pS129-α-syn. These findings identify parkin phosphorylation as a key factor in PD pathogenesis and suggest it as a promising target for therapeutic interventions.

Authors

Rui Han, Qi Wang, Xin Xiong, Xiusheng Chen, Zhuchi Tu, Bang Li, Fei Zhang, Chunyu Chen, Mingtian Pan, Ting Xu, Laiqiang Chen, Zhifu Wang, Yanting Liu, Dajian He, Xiangyu Guo, Feng He, Peng Wu, Peng Yin, Yunbo Liu, Xiaoxin Yan, Shihua Li, Xiao-Jiang Li, Weili Yang

Reactive microglia partially envelop viable neurons in prion diseases

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Abstract

Microglia are recognized as the main cells in the central nervous system responsible for phagocytosis. The current study demonstrated that in prion disease, microglia effectively phagocytose prions or PrPSc during early preclinical stages. However, a critical shift occured in microglial activity during the late preclinical stage, transitioning from PrPSc uptake to establishing extensive neuron-microglia body-to-body cell contacts. This change was followed by a rapid accumulation of PrPSc in the brain. Microglia that enveloped neurons exhibited hypertrophic, cathepsin D-positive lysosomal compartments. However, most neurons undergoing envelopment were only partially encircled by microglia. Despite up to 40% of cortical neurons being partially enveloped at clinical stages, only a small percentage of envelopment proceeded to full engulfment. Partially enveloped neurons lacked apoptotic markers but showed signs of functional decline. Neuronal envelopment was independent of the CD11b pathway, previously associated with phagocytosis of newborn neurons during neurodevelopment. This phenomenon of partial envelopment was consistently observed across multiple prion-affected brain regions, various mouse-adapted strains, and different subtypes of sporadic Creutzfeldt-Jakob disease (sCJD) in humans. The current work describes a new phenomenon of partial envelopment of neurons by reactive microglia in the context of an actual neurodegenerative disease, not a disease model.

Authors

Natallia Makarava, Tarek Safadi, Olga Bocharova, Olga Mychko, Narayan P. Pandit, Kara Molesworth, Simone Baiardi, Li Zhang, Piero Parchi, Ilia V. Baskakov

Modeling primary microcephaly with human brain organoids reveals fundamental roles of CIT kinase activity

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Abstract

Brain size and cellular heterogeneity are tightly regulated by species-specific proliferation and differentiation of multipotent neural progenitor cells (NPCs). Errors in this process are among the mechanisms of primary hereditary microcephaly (MCPH), a group of disorders characterized by reduced brain size and intellectual disability. Biallelic CIT missense variants that disrupt kinase function (CITKI/KI) and frameshift loss-of-function variants (CITFS/FS) are the genetic basis for MCPH17; however, the function of CIT catalytic activity in brain development and NPC cytokinesis is unknown. Therefore, we created the CitKI/KI mouse model and found that it does not phenocopy human microcephaly, unlike biallelic CitFS/FS animals. Nevertheless, both Cit models exhibited binucleation, DNA damage, and apoptosis. To investigate human-specific mechanisms of CIT microcephaly, we generated CITKI/KI and CITFS/FS human forebrain organoids. We found that CITKI/KI and CITFS/FS organoids lose cytoarchitectural complexity, transitioning from pseudostratified to simple neuroepithelium. This change was associated with defects that disrupt polarity of NPC cytokinesis, in addition to elevating apoptosis. Together, our results indicate that both CIT catalytic and scaffolding functions in NPC cytokinesis are critical for human corticogenesis. Species differences in corticogenesis and the dynamic 3D features of NPC mitosis underscore the utility of human forebrain organoid models for understanding human microcephaly.

Authors

Gianmarco Pallavicini, Amanda Moccia, Giorgia Iegiani, Roberta Parolisi, Emily R. Peirent, Gaia Elena Berto, Martina Lorenzati, Rami Y. Tshuva, Alessia Ferraro, Fiorella Balzac, Emilia Turco, Shachi U. Salvi, Hedvig F. Myklebust, Sophia Wang, Julia Eisenberg, Maushmi Chitale, Navjit S. Girgla, Enrica Boda, Orly Reiner, Annalisa Buffo, Ferdinando Di Cunto, Stephanie L. Bielas

Stimulation of an entorhinal-hippocampal extinction circuit facilitates fear extinction in a post-traumatic stress disorder model

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Abstract

Effective psychotherapy of post-traumatic stress disorder (PTSD) remains challenging due to the fragile nature of fear extinction, for which ventral hippocampal CA1 (vCA1) region is considered as a central hub. However, neither the core pathway nor the cellular mechanisms involved in implementing extinction are known. Here, we unveil a direct pathway, where layer 2a fan cells in the lateral entorhinal cortex (LEC) target parvalbumin-expressing interneurons (PV-INs) in the vCA1 region to propel low gamma-band synchronization of the LEC-vCA1 activity during extinction learning. Bidirectional manipulations of either hippocampal PV-INs or LEC fan cells sufficed fear extinction. Gamma entrainment of vCA1 by deep brain stimulation (DBS) or noninvasive transcranial alternating current stimulation (tACS) of LEC persistently enhanced the PV-IN activity in vCA1, thereby promoting fear extinction. These results demonstrate that the LEC-vCA1 pathway forms a top-down motif to empower low gamma-band oscillations that facilitate fear extinction. Finally, application of low gamma DBS and tACS to a mouse model with persistent PTSD showed potent efficacy, suggesting that the dedicated LEC-vCA1 pathway can be stimulated for therapy to remove traumatic memory trace.

Authors

Ze-Jie Lin, Xue Gu, Wan-Kun Gong, Mo Wang, Yan-Jiao Wu, Qi Wang, Xin-Rong Wu, Xin-Yu Zhao, Michael X. Zhu, Lu-Yang Wang, Quanying Liu, Ti-Fei Yuan, Wei-Guang Li, Tian-Le Xu

CRISPR–Cas13d targeting suppresses repeat-associated non-AUG translation of C9orf72 hexanucleotide repeat RNA

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Abstract

A hexanucleotide GGGGCC repeat expansion in the non-coding region of C9orf72 gene is the most common genetic mutation identified in patients with amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The resulting repeat RNA and dipeptide repeat proteins from non-conventional repeat translation have been recognized as important markers associated with the diseases. CRISPR-Cas13d, a powerful RNA targeting tool, has faced challenges in effectively targeting RNA with stable secondary structures. Here we report that CRISPR-Cas13d can be optimized to specifically target GGGGCC repeat RNA. Our results demonstrate that the CRISPR-Cas13d system can be harnessed to significantly diminish the translation of poly-dipeptides originating from the GGGGCC repeat RNA. This efficacy has been validated in various cell types, including induced pluripotent stem cells and differentiated motor neurons originating from C9orf72-ALS patients, as well as in C9orf72 repeat transgenic mice. These findings demonstrate the application of CRISPR-Cas13d in targeting RNA with intricate higher-order structures and suggest a potential therapeutic approach for ALS and FTD.

Authors

Honghe Liu, Xiao-Feng Zhao, Yu-Ning Lu, Lindsey R. Hayes, Jiou Wang

GLP-1R–positive neurons in the lateral septum mediate the anorectic and weight-lowering effects of liraglutide in mice

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Abstract

Liraglutide, a glucagon-like peptide-1 (GLP-1) analog, is approved for obesity treatment, but the specific neuronal sites that contribute to its therapeutic effects remain elusive. Here, we show that GLP-1 receptor–positive (GLP-1R–positive) neurons in the lateral septum (LSGLP-1R) play a critical role in mediating the anorectic and weight-loss effects of liraglutide. LSGLP-1R neurons were robustly activated by liraglutide, and chemogenetic activation of these neurons dramatically suppressed feeding. Targeted knockdown of GLP-1 receptors within the LS, but not in the hypothalamus, substantially attenuated liraglutide’s ability to inhibit feeding and lower body weight. The activity of LSGLP-1R neurons rapidly decreased during naturalistic feeding episodes, while synaptic inactivation of LSGLP-1R neurons diminished the anorexic effects triggered by liraglutide. Together, these findings offer critical insights into the functional role of LSGLP-1R neurons in the physiological regulation of energy homeostasis and delineate their instrumental role in mediating the pharmacological efficacy of liraglutide.

Authors

Zijun Chen, Xiaofei Deng, Cuijie Shi, Haiyang Jing, Yu Tian, Jiafeng Zhong, Gaowei Chen, Yunlong Xu, Yixiao Luo, Yingjie Zhu

The NR4A2/VGF pathway fuels inflammation-induced neurodegeneration via promoting neuronal glycolysis

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Abstract

A disturbed balance between excitation and inhibition (E/I balance) is increasingly recognized as a key driver of neurodegeneration in multiple sclerosis (MS), a chronic inflammatory disease of the central nervous system. To understand how chronic hyperexcitability contributes to neuronal loss in MS, we transcriptionally profiled neurons from mice lacking inhibitory metabotropic glutamate signaling with shifted E/I balance and increased vulnerability to inflammation-induced neurodegeneration. This revealed a prominent induction of the nuclear receptor NR4A2 in neurons. Mechanistically, NR4A2 increased susceptibility to excitotoxicity by stimulating continuous VGF secretion leading to glycolysis-dependent neuronal cell death. Extending these findings to people with MS (pwMS), we observed increased VGF levels in serum and brain biopsies. Notably, neuron-specific deletion of Vgf in a mouse model of MS ameliorated neurodegeneration. These findings underscore the detrimental effect of a persistent metabolic shift driven by excitatory activity as a fundamental mechanism in inflammation-induced neurodegeneration.

Authors

Marcel S. Woo, Lukas C. Bal, Ingo Winschel, Elias Manca, Mark Walkenhorst, Bachar Sevgili, Jana K. Sonner, Giovanni Di Liberto, Christina Mayer, Lars Binkle-Ladisch, Nicola Rothammer, Lisa Unger, Lukas Raich, Alexandros Hadjilaou, Barbara Noli, Antonio L. Manai, Vanessa Vieira, Nina Meurs, Ingrid Wagner, Ole Pless, Cristina Cocco, Samuel B. Stephens, Markus Glatzel, Doron Merkler, Manuel A. Friese

Convergent generation of atypical prions in knockin mouse models of genetic prion disease

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Abstract

Most cases of human prion disease arise due to spontaneous misfolding of WT or mutant prion protein, yet recapitulating this event in animal models has proven challenging. It remains unclear whether spontaneous prion generation can occur within the mouse lifespan in the absence of protein overexpression and how disease-causing mutations affect prion strain properties. To address these issues, we generated knockin mice that express the misfolding-prone bank vole prion protein (BVPrP). While mice expressing WT BVPrP (I109 variant) remained free from neurological disease, a subset of mice expressing BVPrP with mutations (D178N or E200K) causing genetic prion disease developed progressive neurological illness. Brains from spontaneously ill knockin mice contained prion disease–specific neuropathological changes as well as atypical protease-resistant BVPrP. Moreover, brain extracts from spontaneously ill D178N- or E200K-mutant BVPrP–knockin mice exhibited prion seeding activity and transmitted disease to mice expressing WT BVPrP. Surprisingly, the properties of the D178N- and E200K-mutant prions appeared identical before and after transmission, suggesting that both mutations guide the formation of a similar atypical prion strain. These findings imply that knockin mice expressing mutant BVPrP spontaneously develop a bona fide prion disease and that mutations causing prion diseases may share a uniform initial mechanism of action.

Authors

Surabhi Mehra, Matthew E.C. Bourkas, Lech Kaczmarczyk, Erica Stuart, Hamza Arshad, Jennifer K. Griffin, Kathy L. Frost, Daniel J. Walsh, Surachai Supattapone, Stephanie A. Booth, Walker S. Jackson, Joel C. Watts

Exosomal TNF-α mediates voltage-gated Na⁺ channels 1.6 overexpression and contributes to brain-tumor induced neuronal hyperexcitability

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Abstract

Patients affected by glioma frequently suffer of epileptic discharges, however the causes of brain tumor-related epilepsy (BTRE) are still not completely understood. We investigated the mechanisms underlying BTRE by analyzing the effects of exosomes released by U87 glioma cells and by patient-derived glioma cells. Rat hippocampal neurons incubated for 24 h with these exosomes exhibited increased spontaneous firing, while their resting membrane potential shifted positively by 10-15 mV. Voltage clamp recordings demonstrated that the activation of the Na+ current shifted towards more hyperpolarized voltages by 10-15 mV. To understand the factors inducing hyperexcitability we focused on exosomal cytokines. Western Blot and ELISA assays show that TNF-α is present inside glioma-derived exosomes. Remarkably, incubation with TNF-α fully mimicked the phenotype induced by exosomes, with neurons firing continuously, while their resting membrane potential shifted positively. RT-PCR revealed that both exosomes and TNF-α induced over-expression of the voltage-gated Na channel Nav1.6, a low-threshold Na+ channel responsible for hyperexcitability. When neurons were preincubated with Infliximab, a specific TNF-α inhibitor, the hyperexcitability induced by exosomes and TNF-α were drastically reduced. We propose that Infliximab, an FDA approved drug to treat rheumatoid arthritis, could ameliorate the conditions of glioma patients suffering of BTRE.

Authors

Cesar Adolfo Sanchez Trivino, Renza Spelat, Federica Spada, Camilla D'Angelo, Ivana Manini, Irene Giulia Rolle, Tamara Ius, Pietro Parisse, Anna Menini, Daniela Cesselli, Miran Skrap, Fabrizia Cesca, Vincent Torre

Autism-associated neuroligin-3 deficiency in medial septum causes social deficits and sleep loss in mice

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Abstract

Patients with autism spectrum disorder (ASD) frequently experience sleep disturbance. Genetic mutations in Neuroligin-3 (NLG3) genes are highly correlative with ASD and sleep disturbance. However, the cellular and neural circuit bases of this correlation remain elusive. Here, we find the conditional knockout of NLG3 (NLG3-CKO) in the medial septum (MS) impairs social memory and reduces sleep. NLG3 knockout in MS causes hyperactivity of MS-GABA neurons during social avoidance and wakefulness. Activation of MSGABA neurons induces social memory deficits and sleep loss in C57BL/6 mice. In contrast, inactivation of these neurons ameliorates social memory deficits and sleep loss in NLG3-CKO mice. Sleep deprivation leads to social memory deficits, while social isolation causes sleep loss, both resulting in a reduction of NLG3 expression and an increase in activity of GABAergic neurons in MS from C57BL/6 mice. Furthermore, MS-GABA-innervated CA2 neurons specifically regulate social memory without impacting sleep, whereas MSGABA-innervating neurons in the preoptic area selectively control sleep without affecting social behavior. Together, these findings demonstrate that the hyperactive MS-GABA neurons impair social memory and disrupt sleep resulting from NLG3 knockout in MS, and achieve the modality specificity through their divergent downstream targets.

Authors

Haiyan Sun, Yu Shen, Pengtao Ni, Xin Liu, Yan Li, Zhentong Qiu, Jiawen Su, Yihan Wang, Miao Wu, Xiangxi Kong, Jun-Li Cao, Wei Xie, Shuming An