9 Optimization of Brain Slice Health for Ultrasonic Stimulation

Faculty Mentor: Jan Kubanek (Biomedical Engineering, University of Utah)

There is a critical need to develop noninvasive treatments for drug-resistant neurological disorders. Focused ultrasound is a noninvasive modern technology that can provide mm precision and target deep brain structures. Yet, it is unknown the effects of ultrasound on different cell types. Our goal is to develop a method which we can determine the calcium response in neurons to ultrasonic stimulation. When optimizing a method to image calcium response, it is crucial to ensure brain slice health as cell death can occur during preparation of brain slices and in the imaging environment. We established using Thy1-GCaMP6s animals because of its increased fluorescence in CA1 hippocampal cells (Dana et al., 2014) and greater detection of active neurons (Chen et al., 2013). Using a Zeiss confocal microscope imaging at 12 Hz of coronal sections of Thy1-GCaMP6s (n=2), we determined that the NMDG protective recovery method for slice preparation and our solution process was sufficient. This success is marked by imaging predominantly healthy cells with full soma. During imaging coronal Thy1GCaMP6s (n=2) with an Olympus BX51 illumination turret, 10x water immersion .3NA objective, and GFP filter set, we determined the recording temperature of 30-32 ̊C and 7.35 pH was sufficient for healthy brain slices as observed by non-bloated cells in the imaging environment. Additionally, we recorded continual brain tissue viability for a minimum of six hours. Optimization of slice health and the imaging environment for the tissue will allow us to create a novel setup for ultrasonic stimulation of brain slices.