Molecular and biophysical characterization of cultured DRG neurons in response to focused ultrasound

Dorsal root ganglion (DRG) neurons have a wide range of functions, including touch, pain and itch. These neurons have recently emerged as promising targets for non-invasive focused ultrasound (FUS) neuromodulation. However, our understanding of the molecular and physical mechanisms underlying FUS-evoked responses in DRG neurons remains limited. Here, we explore the neuromodulatory effects of FUS on cultured DRG neurons using calcium imaging to track neural responses. We find that a 20-MHz FUS burst of 1-ms duration at an acoustic pressure of 5 MPa elicited calcium responses in 52% of DRG neurons. Single-cell RNA sequencing reveals that more than half of FUS-sensitive neurons belong to two subsets: the TH-expressing C low-threshold mechanoreceptors (C-LTMRs) and the MRGPRD-expressing C high-threshold mechanoreceptors (C-HTMRs), both of which express the Gαi-interacting protein (GINIP). This finding was further confirmed by using a ginip mouse model. We demonstrate that FUS excites both GINIP + and GINIP- neurons through membrane deformation, likely mediated by mechanosensitive ion channels. Our findings identify specific FUS parameters that activate distinct subsets of DRG neurons, opening new possibilities for using FUS to modulate DRG neuron activity.