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 stimuli from the periphery to the brain, 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, there are functional biological differences between human and animal model nociceptors. Therefore, a complementary in vitro model of human nociception is critical 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. Within differentiated HD10.6 cells, we documented the abundance and localization of nociceptive machinery central to regulating excitability and linked with pain sensation including ion channels TRPV1 and NaV1.7 and afferent peptides CGRP and Substance P. Using calcium influx imaging assays, we confirmed the electrical functionality of TRPV1 and NaV1.7 in HD10.6 cells, and through whole-cell patch clamp, we found similar baseline electrophysiological parameters of HD10.6 cells to those previously observed in human patient DRGs. Further, we found that differentiated HD10.6 cells express the mu opioid receptor 1 protein, and DAMGO, a mu agonist, blocks depolarization-evoked calcium influx in a naloxone-reversible fashion. Importantly, using an inflammatory cocktail, excitation and peripheral sensitization are induced within HD10.6 cells, mirroring nociceptors in a pain state during or after tissue damage or inflammation. Finally, HD10.6 cells were also cultured into dual-chambered microfluidic devices to mirror the biological anatomy of the nociceptor. Within this system, we demonstrated the uptake of adeno-associated-virus (AAV) by the peripheral terminals and AAV transport to the soma. Altogether, we have developed the use of HD10.6 cells to create a system of human nociceptive signaling on a chip to study human nociceptor physiology and intervention. PerspectiveThere are essential differences between human and animal model nociceptors. Here, we develop a physiological model of "nociceptive signaling on a chip" using human-derived nociceptors to ultimately enhance the translatability of preclinical afferent signaling research to the human patient.