Advancing the functional maturity of brain organoids by synthetic afferentation.

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PROJECT SUMMARY Human pluripotent stem cells (hiPSCs) are a powerful tool for understanding the principles that govern the development of the human brain and for modeling diseases of the nervous system. One important recent application of this technology is the generation of tri-dimensional (3D) brain cell cultures or "organoids." Brain organoids have been shown to develop structural, transcriptional, and functional similarities up to the mid-to-late gestation human brain as compared to preterm human EEG recordings and human transcriptome profiles. Currently they represent the closest cellular model to native human brain tissue available. While a powerful system for probing mechanisms of prenatal development attempts to advance the maturation state of brain organoids have proved to be incremental, inconclusive, or poorly reproduced. As a result, brain organoid systems remain largely inappropriate for modeling the postnatal brain. There is a significant need to develop a new generation of brain organoid models that are appropriate for interrogating later developmental stages. This proposal focuses on understanding the role of physiological inputs as a necessary driver of more robust, reproducible, and mature physiological states of activity in brain organoids. This project will develop approaches for introducing developmentally-relevant inputs and for reading out neuronal activity in human brain organoids. Extensive work from other systems has established that afferent network activity serves to establish, maintain, and refine active functional brain circuits. Here, we will test a series of innovative strategies to replace or mimic missing external inputs via prolonged patterned stimulation of human brain organoids. We will then observe the resulting effects on network activity and gene expression. We will benchmark these observations to existing data sets from human cortical organoids as well as the human and rodent brain. The tools and approaches established here are intended to be readily adapted to cell culture models of other organ systems for which neuronal inputs are important. If successful, this project will establish a novel platform for interrogating activity-dependent maturation and will enable access to more advanced stages of human brain development and human neurological and neuropsychiatric disease states.