Timed cues create mini-cerebellum in culture

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Researchers can coax human stem cells to grow into layered structures that mimic the brain’s center for motor control, the cerebellum, according to a study published 27 January in Cell Reports¹. The self-organizing cells could shed light on cerebellar development — a process that appears to go awry in autism.

Some people with autism have abnormal connections between the cerebellum and other brain regions. They also have fewer large, branching neurons called Purkinje cells in their cerebellums than controls do.

Researchers have used mice to investigate how these abnormalities contribute to behaviors seen in autism, such as motor impairments. But mice are an imperfect proxy for people with the disorder.

To create a cellular model of the developing cerebellum, the researchers cultured human embryonic stem cells in V-shaped wells that force the cells to cluster together. They then added chemical cues, such as growth factors known to be involved in brain development, to differentiate the cells into specific subtypes of cerebellar neurons.

After approximately three weeks in culture, the cells began to organize into hollow tubes. The researchers then transferred the structures into Petri dishes, giving them room to spread out.

After five weeks in culture, the tubes morphed into large, flat ovals. These structures show varying patterns of gene expression in different layers, suggesting that they are differentiating into specialized cells.

The researchers then added two more growth factors. This encouraged the ovals to merge into one multilayered structure reminiscent of the developing cerebellum during the first trimester of pregnancy. The whole process takes about seven weeks.

A close look at the cultured structures reveals precursors for four different subtypes of neurons seen in the cerebellum: Purkinje cells, Golgi cells, Dorsal cochlear nucleus projection neurons and non-Golgi interneurons.

When the researchers extracted the Purkinje-like precursors and cultured them on their own, the cells took on the appearance of mature Purkinje cells with elaborately branching projections, called dendrites. The cells also behave like mature Purkinje cells, exhibiting distinct patterns of electrical activity, including spontaneous repetitive firing.

Although the study is not the first to create brain-like structures in culture, previous efforts have not been able to model the cerebellum specifically. The researchers say it may be possible to replace embryonic stem cells with stem cells reprogrammed from adult skin cells. These would carry the unique genetic signature of an individual, perhaps an individual with autism.