Gillis, A. et al. Critical assessment of G protein-biased agonism at the μ-opioid receptor. Trends Pharmacol. Sci. 41, 947–959 (2020).
Varga, B. R., Streicher, J. M. & Majumdar, S. Strategies towards safer opioid analgesics—a review of old and upcoming targets. Br. J. Pharmacol. 180, 975–993 (2023).
Kelly, E., Conibear, A. & Henderson, G. Biased agonism: lessons from studies of opioid receptor agonists. Annu. Rev. Pharmacol. Toxicol. 63, 491–515 (2023).
Volkow, N. D., Michaelides, M. & Baler, R. The neuroscience of drug reward and addiction. Physiol. Rev. 99, 2115–2140 (2019).
Altier, N. & Stewart, J. Dopamine receptor antagonists in the nucleus accumbens attenuate analgesia induced by ventral tegmental area substance P or morphine and by nucleus accumbens amphetamine. J. Pharmacol. Exp. Ther. 285, 208–215 (1998).
Altier, N. & Stewart, J. The role of dopamine in the nucleus accumbens in analgesia. Life Sci. 65, 2269–2287 (1999).
Wood, P. B. Role of central dopamine in pain and analgesia. Expert Rev. Neurother. 8, 781–797 (2008).
Taylor, N. E. et al. The rostromedial tegmental nucleus: a key modulator of pain and opioid analgesia. Pain 160, 2524–2534 (2019).
Wang, X. Q., Mokhtari, T., Zeng, Y. X., Yue, L. P. & Hu, L. The distinct functions of dopaminergic receptors on pain modulation: a narrative review. Neural Plast. 2021, 6682275 (2021).
Kishikawa, Y. et al. Dysregulation of dopamine neurotransmission in the nucleus accumbens in immobilization-induced hypersensitivity. Front. Pharmacol. 13, 988178 (2022).
Noursadeghi, E., Rashvand, M. & Haghparast, A. Nucleus accumbens dopamine receptors mediate the stress-induced analgesia in an animal model of acute pain. Brain Res. 1784, 147887 (2022).
Noursadeghi, E. & Haghparast, A. Modulatory role of intra-accumbal dopamine receptors in the restraint stress-induced antinociceptive responses. Brain Res. Bull. 195, 172–179 (2023).
Shahani, P. et al. The interaction effects between opioidergic and D1-like dopamine receptors in the nucleus accumbens on pain-related behaviors in the animal model of acute pain. Pharmacol. Biochem. Behav. 246, 173911 (2025).
Shields, B. C. et al. Deconstructing behavioral neuropharmacology with cellular specificity. Science 356, eaaj2161 (2017).
Shields, B. C. et al. DART.2: bidirectional synaptic pharmacology with thousandfold cellular specificity. Nat. Methods 21, 1288–1297 (2024).
Reynolds, J. N. J. et al. Coincidence of cholinergic pauses, dopaminergic activation and depolarisation of spiny projection neurons drives synaptic plasticity in the striatum. Nat. Commun. 13, 1296 (2022).
Jang, H. J., McMahon Ward, R., Golden, C. E. M. & Constantinople, C. M. Acetylcholine demixes heterogeneous dopamine signals for learning and moving. Nat. Neurosci. 29, 840–850 (2026).
Carroll, K. M., DeVito, E. E., Yip, S. W., Nich, C. & Sofuoglu, M. Double-blind placebo-controlled trial of galantamine for methadone-maintained individuals with cocaine use disorder: secondary analysis of effects on illicit opioid use. Am. J. Addict. 28, 238–245 (2019).
Javed, T. et al. Association of status of acetylcholinesterase and ACHE gene 3’ UTR variants (rs17228602, rs17228616) with drug addiction vulnerability in pakistani population. Chem. Biol. Interact. 308, 130–136 (2019).
Gawel, K., Labuz, K., Jenda, M., Silberring, J. & Kotlinska, J. H. Influence of cholinesterase inhibitors, donepezil and rivastigmine on the acquisition, expression, and reinstatement of morphine-induced conditioned place preference in rats. Behav. Brain Res. 268, 169–176 (2014).
Mei, D. et al. Cognitive enhancer donepezil attenuates heroin-seeking behavior induced by cues in rats. J. Integr. Neurosci. 22, 76 (2023).
Buccafusco, J. J. & Bain, J. N. A 24-h access I.V. self-administration schedule of morphine reinforcement and the estimation of recidivism: pharmacological modification by arecoline. Neuroscience 149, 487–498 (2007).
Zhou, W. et al. Role of acetylcholine transmission in nucleus accumbens and ventral tegmental area in heroin-seeking induced by conditioned cues. Neuroscience 144, 1209–1218 (2007).
Slatkin, N. E., Rhiner, M. & Bolton, T. M. Donepezil in the treatment of opioid-induced sedation: report of six cases. J. Pain Symptom Manage. 21, 425–438 (2001).
Wehrfritz, A. P. et al. Interaction of physostigmine and alfentanil in a human pain model. Br. J. Anaesth. 104, 359–368 (2010).
Sun, E. C., Darnall, B. D., Baker, L. C. & Mackey, S. Incidence of and risk factors for chronic opioid use among opioid-naive patients in the postoperative period. JAMA Intern. Med. 176, 1286–1293 (2016).
Sutherland, T. N. et al. Preoperative vs postoperative opioid prescriptions and prolonged opioid refills among US youths. JAMA Netw. Open 7, e2420370 (2024).
Lankenau, S. E. et al. Initiation into prescription opioid misuse amongst young injection drug users. Int. J. Drug Policy 23, 37–44 (2012).
Martinez-Rivera, A. et al. Elevating levels of the endocannabinoid 2-arachidonoylglycerol blunts opioid reward but not analgesia. Sci. Adv. 10, eadq4779 (2024).
Kumar, V. et al. Highly selective dopamine D3 receptor (D3R) antagonists and partial agonists based on eticlopride and the D3R crystal structure: new leads for opioid dependence treatment. J. Med. Chem. 59, 7634–7650 (2016).
Zaveri, N. T., Marquez, P. V., Meyer, M. E., Hamid, A. & Lutfy, K. The nociceptin receptor (NOP) agonist AT-312 blocks acquisition of morphine- and cocaine-induced conditioned place preference in mice. Front. Psychiatry 9, 638 (2018).
Coppola, J. J. & Disney, A. A. Is there a canonical cortical circuit for the cholinergic system? Anatomical differences across common model systems. Front. Neural Circuits 12, 8 (2018).
Moeller, S. J. & Abi-Dargham, A. Letter to the editor: a novel therapeutic for opioid use disorder targeting the cholinergic system. Am. J. Addict. 28, 235–237 (2019).
Jensen, K. P., DeVito, E. E., Yip, S., Carroll, K. M. & Sofuoglu, M. The cholinergic system as a treatment target for opioid use disorder. CNS Drugs 32, 981–996 (2018).
Navratilova, E. & Porreca, F. Reward and motivation in pain and pain relief. Nat. Neurosci. 17, 1304–1312 (2014).
Harris, H. N. & Peng, Y. B. Evidence and explanation for the involvement of the nucleus accumbens in pain processing. Neural Regen. Res. 15, 597–605 (2020).
Beauchamp, A. et al. Whole-brain comparison of rodent and human brains using spatial transcriptomics. eLife 11, e79418 (2022).
Spanagel, R., Herz, A. & Shippenberg, T. S. Opposing tonically active endogenous opioid systems modulate the mesolimbic dopaminergic pathway. Proc. Natl Acad. Sci. USA 89, 2046–2050 (1992).
Gonzales, K. K. & Smith, Y. Cholinergic interneurons in the dorsal and ventral striatum: anatomical and functional considerations in normal and diseased conditions. Ann. NY Acad. Sci. 1349, 1–45 (2015).
Vaccarino, F. J., Bloom, F. E. & Koob, G. F. Blockade of nucleus accumbens opiate receptors attenuates intravenous heroin reward in the rat. Psychopharmacology 86, 37–42 (1985).
Corrigall, W. A. & Vaccarino, F. J. Antagonist treatment in nucleus accumbens or periaqueductal grey affects heroin self-administration. Pharmacol. Biochem. Behav. 30, 443–450 (1988).
Martin, T. J., Kim, S. A., Lyupina, Y. & Smith, J. E. Differential involvement of mu-opioid receptors in the rostral versus caudal nucleus accumbens in the reinforcing effects of heroin in rats: evidence from focal injections of β-funaltrexamine. Psychopharmacology 161, 152–159 (2002).
Terashvili, M. et al. (+)-Morphine attenuates the (−)-morphine-produced conditioned place preference and the µ-opioid receptor-mediated dopamine increase in the posterior nucleus accumbens of the rat. Eur. J. Pharmacol. 587, 147–154 (2008).
Kaneko, S. et al. Synaptic integration mediated by striatal cholinergic interneurons in basal ganglia function. Science 289, 633–637 (2000).
Hikida, T., Kitabatake, Y., Pastan, I. & Nakanishi, S. Acetylcholine enhancement in the nucleus accumbens prevents addictive behaviors of cocaine and morphine. Proc. Natl Acad. Sci. USA 100, 6169–6173 (2003).
Witten, I. B. et al. Cholinergic interneurons control local circuit activity and cocaine conditioning. Science 330, 1677–1681 (2010).
Severino, A. L. et al. µ-Opioid receptors on distinct neuronal populations mediate different aspects of opioid reward-related behaviors. eNeuro https://doi.org/10.1523/ENEURO.0146-20.2020 (2020).
Sanchez, J. et al. Targeted inhibition of mu-opioid receptors in neuronal subpopulations by membrane-tethered Naloxo-DART antagonists. Cell Chem. Biol. 32, 1473–1485 (2025).
Ponterio, G. et al. Powerful inhibitory action of mu opioid receptors (MOR) on cholinergic interneuron excitability in the dorsal striatum. Neuropharmacology 75, 78–85 (2013).
Stoeber, M. et al. A genetically encoded biosensor reveals location bias of opioid drug action. Neuron 98, 963–976 (2018).
Radoux-Mergault, A., Oberhauser, L., Aureli, S., Gervasio, F. L. & Stoeber, M. Subcellular location defines GPCR signal transduction. Sci. Adv. 9, eadf6059 (2023).
Gonzales, K. K., Pare, J. F., Wichmann, T. & Smith, Y. GABAergic inputs from direct and indirect striatal projection neurons onto cholinergic interneurons in the primate putamen. J. Comp. Neurol. 521, 2502–2522 (2013).
Ma, Y. Y. et al. Regional and cell-type-specific effects of DAMGO on striatal D1 and D2 dopamine receptor-expressing medium-sized spiny neurons. ASN Neuro https://doi.org/10.1042/AN20110063 (2012).
Banghart, M. R., Neufeld, S. Q., Wong, N. C. & Sabatini, B. L. Enkephalin disinhibits mu opioid receptor-rich striatal patches via delta opioid receptors. Neuron 88, 1227–1239 (2015).
Cantor, C. R. & Schimmel, P. R. Biophysical Chemistry: Part III: The Behavior of Biological Macromolecules (Macmillan, 1980).
Bedard, M. L. et al. All hands on deck: we need multiple approaches to uncover the neuroscience behind the opioid overdose crisis. ACS Chem. Neurosci. 14, 1921–1929 (2023).
Schildein, S., Huston, J. P. & Schwarting, R. K. Open field habituation learning is improved by nicotine and attenuated by mecamylamine administered posttrial into the nucleus accumbens. Neurobiol. Learn. Mem. 77, 277–290 (2002).
Schildein, S., Huston, J. P. & Schwarting, R. K. Injections of tacrine and scopolamine into the nucleus accumbens: opposing effects of immediate vs delayed posttrial treatment on memory of an open field. Neurobiol. Learn. Mem. 73, 21–30 (2000).
Robinson, T. E. & Berridge, K. C. The neural basis of drug craving: an incentive-sensitization theory of addiction. Brain Res. Rev. 18, 247–291 (1993).
Urs, N. M., Daigle, T. L. & Caron, M. G. A dopamine D1 receptor-dependent β-arrestin signaling complex potentially regulates morphine-induced psychomotor activation but not reward in mice. Neuropsychopharmacology 36, 551–558 (2011).
Serrano, A., Aguilar, M. A., Manzanedo, C., Rodriguez-Arias, M. & Minarro, J. Effects of DA D1 and D2 antagonists on the sensitisation to the motor effects of morphine in mice. Prog. Neuropsychopharmacol. Biol. Psychiatry. 26, 1263–1271 (2002).
Vezina, P., Kalivas, P. W. & Stewart, J. Sensitization occurs to the locomotor effects of morphine and the specific mu opioid receptor agonist, DAGO, administered repeatedly to the ventral tegmental area but not to the nucleus accumbens. Brain Res. 417, 51–58 (1987).
Stevens, K. E., Mickley, G. A. & McDermott, L. J. Brain areas involved in production of morphine-induced locomotor hyperactivity of the C57B1/6J mouse. Pharmacol. Biochem. Behav. 24, 1739–1747 (1986).
Rada, P. V., Mark, G. P., Taylor, K. M. & Hoebel, B. G. Morphine and naloxone, IP or locally, affect extracellular acetylcholine in the accumbens and prefrontal cortex. Pharmacol. Biochem. Behav. 53, 809–816 (1996).
Fiserova, M., Consolo, S. & Krsiak, M. Chronic morphine induces long-lasting changes in acetylcholine release in rat nucleus accumbens core and shell: an in vivo microdialysis study. Psychopharmacology 142, 85–94 (1999).
Mercer Lindsay, N., Chen, C., Gilam, G., Mackey, S. & Scherrer, G. Brain circuits for pain and its treatment. Sci. Transl. Med. 13, eabj7360 (2021).
Millan, M. J. Descending control of pain. Prog. Neurobiol. 66, 355–474 (2002).
Hnasko, T. S., Sotak, B. N. & Palmiter, R. D. Morphine reward in dopamine-deficient mice. Nature 438, 854–857 (2005).
Threlfell, S. et al. Striatal dopamine release is triggered by synchronized activity in cholinergic interneurons. Neuron 75, 58–64 (2012).
Cachope, R. et al. Selective activation of cholinergic interneurons enhances accumbal phasic dopamine release: setting the tone for reward processing. Cell Rep. 2, 33–41 (2012).
Nelson, A. B. et al. Striatal cholinergic interneurons Drive GABA release from dopamine terminals. Neuron 82, 63–70 (2014).
Krok, A. C. et al. Intrinsic dopamine and acetylcholine dynamics in the striatum of mice. Nature 621, 543–549 (2023).
Chantranupong, L. et al. Dopamine and glutamate regulate striatal acetylcholine in decision-making. Nature 621, 577–585 (2023).
Mohebi, A., Collins, V. L. & Berke, J. D. Accumbens cholinergic interneurons dynamically promote dopamine release and enable motivation. eLife 12, e85011 (2023).
Taniguchi, J. et al. Comment on ‘Accumbens cholinergic interneurons dynamically promote dopamine release and enable motivation’. eLife 13, e95694 (2024).
Touponse, G. C. et al. Cholinergic modulation of dopamine release drives effortful behaviour. Nature 651, 1020–1029 (2026).
Wilkinson, D. & Murray, J. Galantamine: a randomized, double-blind, dose comparison in patients with Alzheimer’s disease. Int. J. Geriatr. Psychiatry 16, 852–857 (2001).
Weaver, I. A., Yousefzadeh, S. A. & Tadross, M. R. An open-source head-fixation and implant-protection system for mice. HardwareX 13, e00391 (2023).
Dunn, T. W. et al. Geometric deep learning enables 3D kinematic profiling across species and environments. Nat. Methods 18, 564–573 (2021).
Nath, T. et al. Using DeepLabCut for 3D markerless pose estimation across species and behaviors. Nat. Protoc. 14, 2152–2176 (2019).
