Animal models have been widely used to study neurological diseases including neurodegenerative, developmental, and rare CNS disorders. However, commonly used rodent models give limited information on physiology and disease pathophysiology, as there are significant differences between rodent and human brains. The human brain has over a thousand times more neurons than the mouse brain, and single cell RNA sequencing has shown that there are gene expression differences1.While several cell types are common between human and rodents, certain cell types are missing in rodents such as the outer radial glial cells that are progenitor cells for neocortex development2. Non-human primates are an alternative to rodent models as they recapitulate the structure and function of the human CNS and can also be used for behavioral studies. However, due to the COVID-19 pandemic, there has been a significant shortage of these models which has impacted the pace of biomedical research3. Not surprisingly, there is an increasing interest in developing three-dimensional brain organoids to study disease pathophysiology and test novel therapies. Brain organoids are defined as self-assembled aggregates of cells that include more than one cell type and are developed from either embryonic stem (ES) cells or induced pluripotent stem (iPS) cells. Several studies have shown that CNS organoids are able to recapitulate structural and functional characteristics including neurogenesis, cell migration and neural circuitry4.

One of the major issues with using CNS organoids is apoptosis and necrosis in the core. Organoids rely on diffusion for gas exchange, nutrients and waste removal, which is too inefficient to maintain viable long-term culture. The study of neurodegenerative diseases typically requires long term cultures to study disease progression, so it is essential to improve the duration of viable cultures. One solution is to introduce vasculature in the grafting organoids in adult mouse brains via the recruitment of mouse endothelial cells. The development of vascular organoids (vOrganoids) is another method to introducing vasculature in organoid culture where organoids are co-cultured with human vascular endothelial cells (HUVECs). The co-culture approach was shown to induce the formation of a tubular network of blood vessels allowing the culture of organoids for up to 200 days5.

CNS organoids can be generated using two different methods: unguided and guided6. Unguided development of CNS organoids depends on the intrinsic self-organization capabilities of pluripotent stem cells to form defined cellular structures without the aid of any external factors. One of the drawbacks with unguided organoid generation is the variation in cell populations and size of the organoids. Guided methods introduce more control in organoid assembly via exposure to specific patterning factors at specific times. The presence of specific factors triggers specific signaling pathways to generate organoids of specific lineages. For example, the use of a Smad inhibitor shifts the organoids away from mesoderm and endoderm lineages and towards ectoderm lineages7.

The use of iPS cells to generate CNS organoids has broadened the applications of CNS organoids as patient derived cells can be used to generate disease specific organoids. Brain organoids are also used to study neuronal development and diseases associated with neurodevelopment such as microcephaly, macrocephaly, and autism spectrum disorders. Another big area of interest is the study of signaling pathways associated with neuropsychiatric diseases4, so that more targeted and efficacious therapies can be developed. One of the largest and well-funded neuroscience research areas is neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease and the development of disease specific organoids that can be maintained in long term cultures will be very useful to test therapies to slow or reverse neurodegeneration.









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