Modulation of pain sensitivity by the locus coeruleus–paraventricular thalamic nucleus–anterior cingulate cortex pathway in mice

Background: Noradrenergic projections from the locus coeruleus (LC) to the thalamus and anterior cingulate cortex (ACC) contribute to pain‒like behaviors, yet their hierarchical organization remains unclear. Here, we examined how LC‒derived norepinephrine (NE) inputs to the paraventricular thalamic nucleus (PVA) and ACC differentially regulate nociceptive sensitization. Methods: In adult male and female mice, complete Freund’s adjuvant (CFA) was used to induce pain‒like behaviors. To examine functional connectivity among LC, PVA, and ACC, we combined targeted recombination in active populations (Fos‒TRAP), in vivo recordings, and viral tracing. We used optogenetic and chemogenetic tools to selectively manipulate LC projections and assess their impact on neural activity and pain behaviors. Results: CFA led to enhanced c‒Fos expression in LC, PVA, and ACC (Cells per microscopic field; LC: 13.60 ± 2.24 vs. 44.50 ± 7.72; PVA: 8.00 ± 1.58 vs. 66.40 ± 9.45; ACC: 12.80 ± 2.28 vs. 36.70 ± 2.59; p < 0.001), alongside increased gamma‒band activity and single‒unit firing rates. Monosynaptic LC–ACC and polysynaptic LC–PVA–ACC circuits were identified. Notably, nociception‒related LC neurons preferentially projected to PVA, which subsequently targeted hyperactive ACC neurons. Under inflammatory pain conditions, activation of the LC–PVA–ACC circuits evoked greater ACC firing (Hz; LC–PVA–ACC vs. LC–ACC: 15.75 ± 2.88 vs. 9.72 ± 2.06; P < 0.001) and tactile‒evoked responses (Hz; 22.98 ± 2.60 vs. 15.34 ± 1.86; P < 0.001) than direct LC–ACC activation. Consistently, optogenetic or chemogenetic manipulation of the LC–PVA–ACC circuit produced stronger modulation of mechanical and thermal pain sensitivity than direct LC–ACC stimulation. Conclusions: We identify the LC–PVA–ACC pathway as a hierarchical noradrenergic circuit that modulates nociceptive sensitization via a thalamocortical relay, thereby revealing a circuit‒specific mechanism by which the LC–NE system regulates pain processing.