Introduction: Mechanical overloading can trigger a degenerative-like cascade in an organ culture of intervertebral disc (IVD). Whether the overloaded IVD can influence the activation of nociceptors (i.e., the damage sensing neurons) remains unknown. The study aims to investigate the influence of overloaded IVD conditioned medium (CM) on the activation of nociceptors. Methods: In the static loading regime, force-controlled loading of 0.2 MPa for 20 h/day representing "long-term sitting and standing" was compared with a displacement-controlled loading maintaining original IVD height. In the dynamic loading regime, high-frequency-intensity loading representing degenerative "wear and tear" was compared with a lower-frequency-intensity loading. CM of differently loaded IVDs were collected to stimulate the primary bovine dorsal root ganglion (DRG) cultures. Calcium imaging (Fluo-4) and calcitonin gene-related peptide (CGRP) immunofluorescent labeling were jointly used to record the calcium flickering in CGRP(+) nociceptors. Results: Force-controlled loading led to a higher IVD cell death compared to displacement-controlled loading. Both static and dynamic overloading (force-controlled and high-frequency-intensity loadings) elevated the frequency of calcium flickering in the subsurface space of CGRP(+) nociceptors compared to their mild loading counterparts. Conclusion: In the organ culture system, IVD overloading mediated an altered IVD-nociceptor communication suggesting a biological mechanism associated with discogenic pain.