Ultrasonocoverslip: A High Throughput In-Vitro Platform for Investigating Neuron-Glia Response to Direct Stimulations using Low Intensity (<1W/cm²) Ultrasound

Molecular mechanistic studies on low intensity ultrasound brain stimulations have been spotlighted to have insights on reliable acoustic neuronal stimulations. While recent studies report different ultrasound effects showing both astrocytic TRPA1 mediated neuronal excitation and direct neuronal excitations through calcium-permeable mechanosensitive channels, further reliable in-vitro platforms to acquire the efficient stimulation parameters are lacking. To address the needs, ultrasonocoverslip, an ultrasound transducer integrated glass coverslip, for culturing and stimulating brain cells directly on the transducer is developed. Ultrasonocoverslip can deliver uniform ultrasonic longitudinal pressure to the cultured brain cells, and simultaneously allows to observe cellular signaling from neurons and astrocytes. The 'U' shaped electrode of the ultrasonic transducer separates sonicated and non-sonicated regions, respectively, the artifactual stimulations other than the longitudinal wave can be identified as a signal in the non-sonicated region. Due to direct culture on the transducer, the acoustic absorber could be loaded in the back side of the transducer, and reverberations causing elongated stimulation could be substantially reduced. With the reliable in-vitro platform, the minimum ultrasound pressure level and stimulation length which can solely activate astrocytic TRPA1 channel mediated-neuronal excitation with low intensity ultrasound stimulation can be acquired through pharmacologic studies made in the novel in-vitro platform.