Human Dorsal Root Ganglia Neuronal Cell Line to Study Nociceptive Signaling: A New Pipeline for Pain Therapy.

Nociceptive afferent neurons within the dorsal root ganglion (DRG) detect and relay painful peripheral stimuli, and the malfunctioning of this process leads to sustained pain states. Animal model studies have been invaluable for demonstrating the importance of the DRG nociceptor in pain sensation and the development of related analgesic targets. However, a human in vitro model of nociception is essential to confirming the relevance of preclinical findings for therapeutic drug development. We characterized the nociceptive properties of differentiated cells from the human DRG-derived immortalized cell line HD10.6 and developed their use into an in vitro model of human nociceptive signaling and therapy. Within differentiated HD10.6 cells, we confirmed the abundance and function of machinery linked with pain sensation, including key ion channels (TRPV1, NaV1.7) and afferent peptides (CGRP, Substance P), by immunofluorescence and calcium influx assays. Through whole-cell patch clamp, including current clamp and voltage clamp, we recorded the baseline electrophysiological parameters of differentiated HD10.6 cells. We further found that differentiated HD10.6 cells express the mu opioid receptor 1 protein, and that mu agonist DAMGO blocks depolarization-evoked calcium influx in a naloxone-reversible fashion. Importantly, excitation and peripheral sensitization were induced within HD10.6 cells in response to an inflammatory cocktail, mirroring nociceptors in a pain state during and after tissue damage or inflammation. HD10.6 cells were also cultured into dual-chambered microfluidic devices to mirror the physiological anatomy of the nociceptor. Within this system, genetic therapy adeno-associated-virus was successfully taken up by the peripheral terminals and transported to the soma.