Central nervous system (CNS) toxicity results in a wide variety of functional and behavioral changes in animal models such as cognitive impairments, learning/memory loss, hearing/vision loss, alterations in sensory/motor reflex, and several other dysfunctions. While behavioral tests and animal models are sensitive enough to predict human neurotoxicity, the expense and labor-intensiveness of animal testing make it challenging to screen large numbers of chemical compounds. While running behavioral tests is not possible in in vitro setting, some parameters that can be evaluated include cytotoxicity of neuronal and glial cell bodies, demyelination, or altered neuronal firing patterns, which many toxins are known to impact and ultimately results in behavioral/cognitive deficits. Recent advances in induced pluripotent stem cell technology have resulted in the development of human cells-based microphysiological systems that have shown tremendous promise as advanced cellular models that can provide high-throughput and high-content data useful for toxin screening. While human organoids are expected to predict human neurotoxicity better than animal models, the data generated using human organoids may not be reliable enough unless studies show that in vitro neurotoxicity successfully predicts in vivo cortical dysfunctions shown in the historical animal studies. To overcome these challenges and strengthen findings obtained by human brain organotypic culture systems, we have developed an innovative functionally mature 3D rodent brain spheroid model of consistent size and unvarying composition by culturing embryonic dissociated cortical tissue. The objective of this phase I study is to demonstrate that certain chemical toxins known for causing neurotoxicity in rodents will induce toxicity in micro-engineered neural tissue that can be quantified using 3D image analysis, neuronal firing patterns and histomorphometry. This goal will be achieved by quantifying discrete biological and electrical quantifiable metrics by which untreated brain spheroids can be demonstrated to differ significantly compared to treated controls. This goal will be accomplished in two phases: The research plan will consist of first developing and characterizing 3D rodent brain spheroids using high throughput confocal screening and microelectrode arrays for baseline biological and functional evaluation.
(Aim 1). Then, we will compare biological and electrophysiological characteristics of cultured untreated spheroids with spheroids treated with environmental toxins to validate the assay in the presence of an insult (Aim 2). Successful completion of these aims will strongly position this technology for a Phase II award, in which we will seek to scale up fabrication and testing and fully validate the specificity and sensitivity of the clinically- analogous metrics with a larger library of compounds, as compared with animal data. We will further perform ?omics? studies and will eventually use this assay for determining a large spectrum of toxicological parameters resulting in understanding mechanisms of action as well as improved understanding of biological processes.
Central nervous system (CNS) toxicity results in a wide variety of functional and behavioral changes in animal models such as cognitive impairments, learning/memory loss, hearing/vision loss, alterations in sensory/motor reflex, and several other dysfunctions. Several industrial pollutants and other toxins that can make their way into our environment are well known to cause neurotoxicity in humans. A high-throughput 3D in vitro assay capable of detecting changes in the fraction of neuronal and glial cells in 3D brain simulating cultures along with alterations in neuronal electrical activity holds the promise of bridging the gap between ex vivo to in vivo animal toxicity screening. We hypothesize that the 3D organotypic system proposed can detect neural toxicity parameters in ways that mimic clinical neuropathology and will be able to help us in understanding the capabilities of the Microphysiological system as compared to the historical animal data. The development of 3D organotypic cellular models utilizing animal cells is important for the validation of human ?organs-on-chips? systems by direct comparison to animal data, which should be undertaken before assuming that human tissue chips predict clinical drug safety and efficacy.
