A 3-D In Vitro Mini-Brain Model to Study Parkinson's Disease

Abstract

Research in Parkinson’s disease is experiencing an upswing at the moment, on one hand due to a lack of curative drugs for the large number of patients. Drug testing is nearly exclusively performed in vivo in the so-called MPP, methamphetamine, 6-hydroxydopamine and rotenone models requiring tens of thousands of animals. Human neurons, which would be most relevant, are not usually available and existing cell lines e.g. neuroblastoma cells are only a very poor substitute. During our project funded by NCATS, NIH (1U18TR000547) we have developed a human-induced pluripotent stem cell (iPSC) 3-D model recapitulating many aspect of the human brain. The model is now for the first time applied for the study of Parkinson’s disease with respect to mechanisms of degeneration and mechanisms and efficacy of drugs in a project funded by Alternative Research & Development (ARDF). The 3-D model has shown to recapitulate early in vivo human neurodevelopment by showing the emergence of different kinds of neurons and glial cells, induction of genes that play important roles in neurodevelopment as well as presence of electrical activity. Human 3 Daggregates stain positively for different neuronal types, astrocytes and oligodendrocytes markers after 2, 4, and 8 weeks of differentiation showing a maturation in the cells during the differentiation process. In addition, real-time PCR analyses show higher expression of various neuronal differentiation markers in 3-D aggregates compared to 2-D cultures, indicating a faster maturation in the 3-D system. We have used Rotenone, a pesticide known to induce neurotoxicity by inhibition of mitochondrial complex I, and one of the compounds used to study Parkinson’s disease. The model shows increased ROS production and decreased mitochondrial function after exposure to Rotenone. In addition, the model shows a decrease in sensitivity to Rotenone exposures with increasing maturation. Notably, such human brain models will represent a versatile tool to study CNS physiology, pathology, and neurological disorders such a Parkinson’s disease.