Whole-brain analyses identify anterior cingulate μ-opioid signaling as a critical mediator of placebo analgesia in neuropathic pain

Rehal, S. K.; Boorman, D. C.; Cho, C.; Karimi, S. A.; Fazili, M. I.; Augusto, C. A.; Burek, J.; Khaled, F.; Gami, S. R.; Frankland, P. W.; Martin, L. J.

doi:10.64898/2026.06.04.729974

bioRxiv · 10.64898/2026.06.04.729974

Whole-brain analyses identify anterior cingulate μ-opioid signaling as a critical mediator of placebo analgesia in neuropathic pain

Rehal, S. K.·Boorman, D. C.·Cho, C.·Karimi, S. A.·Fazili, M. I.·Augusto, C. A.·Burek, J.·Khaled, F.·Gami, S. R.·Frankland, P. W.·Martin, L. J.

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Abstract

Placebo analgesia reflects the capacity of learning and expectation to engage endogenous pain-control systems, yet the neural circuits that support this phenomenon remain poorly understood. Here, we establish a conditioning-based placebo analgesia paradigm in mice following peripheral nerve injury and combine behavioral assessment with whole-brain activity mapping and targeted circuit manipulations to define its neural substrates. Conditioning with morphine produced robust placebo analgesia, expressed as reduced sensory sensitivity in the absence of drug. Brain-wide mapping of c-Fos expression revealed distributed changes across cortical and subcortical regions accompanied by a reorganization of functional connectivity, consistent with coordinated network-level engagement. Network analyses identified shifts in hub structure and selective strengthening and weakening of inter-regional interactions during placebo analgesia. Causal manipulations demonstrated a critical role for the anterior cingulate cortex, with excitatory activation of this region blocking placebo analgesia, whereas inhibitory manipulations had no effect. In contrast, perturbation of other candidate regions, including the basomedial amygdala and paraventricular thalamus, did not alter placebo responses. Finally, selective targeting of -opioid receptor-expressing neurons in the anterior cingulate cortex revealed that this cell population is necessary for the expression of placebo analgesia. Together, these findings reveal a brain-wide reorganization of network interactions underlying placebo analgesia and identify a specific cortical opioid circuit that gates its expression. One-Sentence SummaryUsing a mouse model of nerve injury, this study shows that placebo analgesia arises from coordinated brain-wide network reorganization and is gated by -opioid receptor-expressing neurons in the anterior cingulate cortex.

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Rehal, S. K., Boorman, D. C., Cho, C., Karimi, S. A., Fazili, M. I., Augusto, C. A., Burek, J., Khaled, F., Gami, S. R., Frankland, P. W., Martin, L. J.. 2026-06-09. Whole-brain analyses identify anterior cingulate μ-opioid signaling as a critical mediator of placebo analgesia in neuropathic pain. https://doi.org/10.64898/2026.06.04.729974

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