The underlying mechanisms of brain stimulation in humans are poorly understood, especially at the level of gene expression. To address this gap in knowledge, we propose a series of three experiments that take advantage of the opportunity to obtain high-quality human neural tissue from neurosurgical patients in order to measure the impact of brain stimulation on gene expression. Our experiments will generate data to explicate changes at the level of gene expression that underlie brain circuit changes elicited by stimulation. Our study team has seven years of experience analyzing gene expression using an established pipeline for studying human cortical tissue from neurosurgical patients, including application of cutting-edge methods for measuring gene expression. These methods include single nuclei RNA-sequencing (snRNA-seq) and the exciting addition of single nuclei ATAC-sequencing (snATAC-seq) to understand stimulation-related changes in transcription factors and chromatin remodeling. Our hypotheses regarding specific gene classes were developed from our published data correlating gene expression changes with neurophysiological signatures (brain oscillations) linked with successful memory formation. In this proposal, our experiments address the complex problem of how stimulation alters neural circuits using three complementary approaches. First, we will use direct cortical stimulation in vivo immediately prior to resection of brain tissue in temporal lobectomy patients, followed by gene expression analysis. Our plans are supported by preliminary data showing differences in expression of immediate early genes (IEGs) following cortical stimulation, in line with predictions drawn from animal models. Second, we will build on techniques we have implemented for culture of human neural tissue (from neurosurgical patients) to measure gene expression changes elicited by chronic ex vivo stimulation. This experiment will elucidate the temporal dynamics of gene expression in the setting of stimulation, including transcription factor changes, using our experience with time series modeling of gene information. Finally, we will use multi-electrode arrays (MEAs) to measure the impact of ex vivo stimulation on networks of co-firing neurons, directly testing models of stimulation-induced changes in local circuits. We will connect these electrophysiological measures with gene expression changes elicited by stimulation. This experiment builds on our published work studying network activity in single unit recordings in humans, as well as our preliminary data demonstrating the ability to record electrophysiological signals from human neural tissue in culture. The stimulation parameters were developed to be aligned across in vivo and in vitro experiments to facilitate comparison. Taken together, our experiments will provide ground-breaking data elucidating the genetic underpinnings of how brain stimulation elicits neuromodulation. The experience of research team and proven record in publishing data using neurosurgical tissue specimens supports our expectations of success.

We propose to study gene expression changes elicited by brain stimulation using tissue obtained from neurosurgical patients. First, we will compare gene expression profiles between stimulation and non-stimulated tissue from the lateral temporal cortex in vivo. Second, using neural tissue cultures of human cortex, we will examine gene expression changes elicited by ex vivo stimulation over six days, and connect stimulation-related changes in neural connectivity with gene expression data.