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A hexanucleotide repeat expansion (HRE) in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Human brain imaging and experimental studies indicate early changes in brain structure and connectivity in C9-ALS/FTD, even before symptom onset. Because these early disease phenotypes remain incompletely understood, we generated iPSC-derived cerebral organoid models from C9-ALS/FTD patients, presymptomatic C9ORF72-HRE (C9-HRE) carriers, and controls. Our work revealed the presence of all three C9-HRE-related molecular pathologies and developmental stage-dependent size phenotypes in cerebral organoids from C9-ALS/FTD patients. In addition, single-cell RNA sequencing identified changes in cell type abundance and distribution in C9-ALS/FTD organoids, including a reduction in the number of deep layer cortical neurons and the distribution of neural progenitors. Further, molecular and cellular analyses and patch-clamp electrophysiology detected various changes in synapse structure and function. Intriguingly, organoids from all presymptomatic C9-HRE carriers displayed C9-HRE molecular pathology, whereas the extent to which more downstream cellular defects, as found in C9-ALS/FTD models, were detected varied for the different presymptomatic C9-HRE cases. Together, these results unveil early changes in 3D human brain tissue organization and synaptic connectivity in C9-ALS/FTD that likely constitute initial pathologies crucial for understanding disease onset and the design of therapeutic strategies., (© 2024. The Author(s).)

Intermediate-length repeat expansions in ATAXIN-2 (ATXN2) are the strongest genetic risk factor for amyotrophic lateral sclerosis (ALS). At the molecular level, ATXN2 intermediate expansions enhance TDP-43 toxicity and pathology. However, whether this triggers ALS pathogenesis at the cellular and functional level remains unknown. Here, we combine patient-derived and mouse models to dissect the effects of ATXN2 intermediate expansions in an ALS background. iPSC-derived motor neurons from ATXN2-ALS patients show altered stress granules, neurite damage and abnormal electrophysiological properties compared to healthy control and other familial ALS mutations. In TDP-43 Tg -ALS mice, ATXN2-Q33 causes reduced motor function, NMJ alterations, neuron degeneration and altered in vitro stress granule dynamics. Furthermore, gene expression changes related to mitochondrial function and inflammatory response are detected and confirmed at the cellular level in mice and human neuron and organoid models. Together, these results define pathogenic defects underlying ATXN2-ALS and provide a framework for future research into ATXN2-dependent pathogenesis and therapy., (© 2024. The Author(s).)

Despite their large societal burden, the development of therapeutic treatments for neurodegenerative diseases (NDDs) has been relatively unsuccessful. This is, in part, due to a lack of representative experimental models that reveal fundamental aspects of human brain pathology. Recently, assays for in vitro modeling of the human central nervous system (CNS) have significantly improved with the development of brain and spinal cord organoids. Coupled with induced-pluripotent stem cell and genome editing technologies, CNS organoids are a promising tool for studying neurodegeneration in a patient-specific manner. An extensive array of protocols for the generation of organoids for different brain regions has been developed and used for studying neurodegenerative and other brain diseases. However, their application in the field of motor neuron disease (MND) has been limited due to a lack of adequate organoid models. The development of protocols to derive spinal cord and trunk organoids and progress in the field of assembloids are providing new opportunities for modeling MND. In this study here we review recent advances in the development of CNS organoid models, their application in NDDs, and technical limitations. Finally, we discuss future perspectives for the development of organoid-based systems for MND and provide a framework for their development. Impact statement Animal models and two-dimensional cultures are currently the main platforms for studying neurodegenerative diseases (NDDs). However, central nervous system (CNS) organoid technology offers novel possibilities for studying these diseases. Organoid modeling in combination with emerging organ-on-a-chip approaches, induced-pluripotent stem cell technology, and genome editing render in vitro modeling of NDDs more robust and physiologically relevant. In this study, we review the principles underlying CNS organoid generation, their use in NDD research, and future perspectives in organoid technology. Finally, we discuss how advances in different fields could be combined to generate a multisystem organoid-on-a-chip model to investigate a specific class of NDDs, motor neuron diseases.