Direct and Intervertebral Disc-Mediated Sensitization of Dorsal Root Ganglion Neurons by Hypoxia and Low pH

Objective: Ischemia-related risk factors are consistently correlated with discogenic pain, but it remains unclear how the ischemia-associated hypoxia and acidosis influence the peripheral sensory nervous system, namely the dorsal root ganglion (DRG), either directly or indirectly via intervertebral disc (IVD) mediation. Methods: Bovine tail IVD organ cultures were preconditioned in different hypoxic and/or acidic conditions for 3 days to collect the conditioned medium (CM). The DRG-derived ND7/23 cells were either treated by the IVD CM or directly stimulated by hypoxic and/or acidic conditions. Neuronal sensitization was evaluated using calcium imaging (Fluo-4) after 3 days. Results: We found that direct exposure of DRG cell line to hypoxia and acidosis increased both spontaneous and bradykinin-stimulated calcium response compared to normoxia-neutral pH cultures. Hypoxia and low pH in combination showed stronger effect than either parameter on its own. Indirect exposure of DRG to hypoxia-acidosis-stressed IVD CM also increased spontaneous and bradykinin-stimulated response, but to a lower extent than direct exposure. The impact of direct hypoxia and acidosis on DRG was validated in a primary sheep DRG cell culture, showing the same trend. Conclusion: Our data suggest that targeting hypoxia and acidosis stresses both in IVD and DRG could be a relevant objective in discogenic pain treatment.