Experimental Brain Research

, Volume 236, Issue 12, pp 3203–3213 | Cite as

A3 adenosine receptor agonist attenuates neuropathic pain by suppressing activation of microglia and convergence of nociceptive inputs in the spinal dorsal horn

  • Ryuji TerayamaEmail author
  • Mitsuyasu Tabata
  • Kotaro Maruhama
  • Seiji Iida
Research Article


Peripheral nerve injuries cause glial activation and neuronal hyperactivity in the spinal dorsal horn. These changes have been considered to be involved in the underlying mechanisms for the development and maintenance of neuropathic pain. Using double immunofluorescence labeling, we previously demonstrated that spinal microglial activation induced by nerve injury enhanced convergence of nociceptive inputs in the spinal dorsal horn from uninjured afferents. The adenosine A3 receptor (A3AR) agonists have been shown to have antinociceptive activities in several experimental neuropathic pain models. However, the mechanisms underlying these antinociceptive actions of the A3AR agonist are still not fully explored. In this study, the effects of the A3AR agonist (i.e., IB-MECA) on microglial activation, enhancement of convergent nociceptive inputs, and nocifensive behaviors were examined after tibial nerve injury. Injury to the tibial nerve initially caused hyposensitivity to touch stimulus at 3 days, and then resulted in tactile allodynia at 14-day post-injury. The daily systemic administration of IB-MECA (0.1 mg/kg/day) for 8 days in a row starting on the day of nerve injury or 7 days after nerve injury prevented the development of behaviorally assessed hypersensitivities, and spinal microglial activation induced by nerve injury. These treatments also suppressed anomalous convergence of nociceptive primary inputs in the spinal dorsal horn. The present findings indicate that the A3AR agonist attenuates neuropathic pain states by suppressing enhanced microglial activation, and anomalous convergence of nociceptive inputs in the spinal dorsal horn from uninjured afferents after injury to the peripheral nerve.


Adenosine A3AR Microglia Nerve injury Spinal dorsal horn 



Adenosine A3 receptor


Analysis of variance


Brain-derived neurotrophic factor


Charge-coupled device


c-Fos protein-like immunoreactive


Central nervous system




Dimethyl sulfoxide


Electrical stimulation




Inducible nitric oxide synthase




Mitogen-activated protein kinase


Phosphorylated extracellular signal-regulated kinase


p-ERK immunoreactive


Phosphate buffer


Phosphate-buffered saline


Paw withdrawal threshold


Tissue necrosis factor-α



This study was supported by a Grant-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (16K11440).


  1. Abbracchio MP, Rainaldi G, Giammarioli AM et al (1997) The A3 adenosine receptor mediates cell spreading, reorganization of actin cytoskeleton, and distribution of Bcl-XL: studies in human astroglioma cells. Biochem Biophys Res Commun 241:297–304. CrossRefPubMedPubMedCentralGoogle Scholar
  2. Biggs JE, Lu VB, Stebbing MJ, Balasubramanyan S, Smith PA (2010) Is BDNF sufficient for information transfer between microglia and dorsal horn neurons during the onset of central sensitization? Mol Pain 6:44. CrossRefPubMedPubMedCentralGoogle Scholar
  3. Boison D (2013) Adenosine kinase: exploitation for therapeutic gain. Pharmacol Rev 65:906–943. CrossRefPubMedPubMedCentralGoogle Scholar
  4. Chaplan SR, Bach FW, Pogrel JW, Chung JM, Yaksh TL (1994) Quantitative assessment of tactile allodynia in the rat paw. J Neurosci Methods 53:55–63CrossRefGoogle Scholar
  5. Chen L, Jiang M, Pei L (2012) Comparison of three methods of drug delivery in the rat lumbar spinal subarachnoid space. Anat Rec (Hoboken) 295:1212–1220. CrossRefGoogle Scholar
  6. Choi IY, Lee JC, Ju C et al (2011) A3 adenosine receptor agonist reduces brain ischemic injury and inhibits inflammatory cell migration in rats. Am J Pathol 179:2042–2052. CrossRefPubMedPubMedCentralGoogle Scholar
  7. Coull JA, Boudreau D, Bachand K et al (2003) Trans-synaptic shift in anion gradient in spinal lamina I neurons as a mechanism of neuropathic pain. Nature 424:938–942. CrossRefPubMedGoogle Scholar
  8. Coull JA, Beggs S, Boudreau D et al (2005) BDNF from microglia causes the shift in neuronal anion gradient underlying neuropathic pain. Nature 438:1017–1021. CrossRefPubMedGoogle Scholar
  9. DeLeo JA, Yezierski RP (2001) The role of neuroinflammation and neuroimmune activation in persistent pain. Pain 90:1–6CrossRefGoogle Scholar
  10. Devor M, Govrin-Lippmann R (1983) Axoplasmic transport block reduces ectopic impulse generation in injured peripheral nerves. Pain 16:73–85CrossRefGoogle Scholar
  11. Dixon WJ (1980) Efficient analysis of experimental observations. Annu Rev Pharmacol Toxicol 20:441–462CrossRefGoogle Scholar
  12. Dworkin RH, O’Connor AB, Backonja M et al (2007) Pharmacologic management of neuropathic pain: evidence-based recommendations. Pain 132:237–251. CrossRefPubMedGoogle Scholar
  13. Fishman P, Bar-Yehuda S, Liang BT, Jacobson KA (2012) Pharmacological and therapeutic effects of A3 adenosine receptor agonists. Drug Discov Today 17:359–366. CrossRefPubMedGoogle Scholar
  14. Ford A, Castonguay A, Cottet M et al (2015) Engagement of the GABA to KCC2 signaling pathway contributes to the analgesic effects of A3AR agonists in neuropathic pain. J Neurosci 35:6057–6067. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Fujisawa N, Terayama R, Yamaguchi D, Omura S, Yamashiro T, Sugimoto T (2012) Fos protein-like immunoreactive neurons induced by electrical stimulation in the trigeminal sensory nuclear complex of rats with chronically injured peripheral nerve. Exp Brain Res 219:191–201. CrossRefPubMedGoogle Scholar
  16. Garrison CJ, Dougherty PM, Kajander KC, Carlton SM (1991) Staining of glial fibrillary acidic protein (GFAP) in lumbar spinal cord increases following a sciatic nerve constriction injury. Brain Res 565:1–7CrossRefGoogle Scholar
  17. Gyllenhammar E, Nordfors LO (2001) Systemic adenosine infusions alleviated neuropathic pain. Pain 94:121–122CrossRefGoogle Scholar
  18. Hains BC, Waxman SG (2006) Activated microglia contribute to the maintenance of chronic pain after spinal cord injury. J Neurosci 26:4308–4317CrossRefGoogle Scholar
  19. Hanisch UK (2002) Microglia as a source and target of cytokines. Glia 40:140–155. CrossRefPubMedGoogle Scholar
  20. Hughes AS, Averill S, King VR, Molander C, Shortland PJ (2008) Neurochemical characterization of neuronal populations expressing protein kinase C gamma isoform in the spinal cord and gracile nucleus of the rat. Neuroscience 153:507–517. CrossRefPubMedGoogle Scholar
  21. Jacobson KA (1998) Adenosine A3 receptors: novel ligands and paradoxical effects. Trends Pharmacol Sci 19:184–191CrossRefGoogle Scholar
  22. Janes K, Esposito E, Doyle T, Cuzzocrea S, Tosh DK, Jacobson KA, Salvemini D (2014) A3 adenosine receptor agonist prevents the development of paclitaxel-induced neuropathic pain by modulating spinal glial-restricted redox-dependent signaling pathways. Pain 155:2560–2567. CrossRefPubMedPubMedCentralGoogle Scholar
  23. Janes K, Wahlman C, Little JW, Doyle T, Tosh DK, Jacobson KA, Salvemini D (2015) Spinal neuroimmune activation is independent of T-cell infiltration and attenuated by A3 adenosine receptor agonists in a model of oxaliplatin-induced peripheral neuropathy. Brain Behav Immun 44:91–99. CrossRefPubMedGoogle Scholar
  24. Janes K, Symons-Liguori AM, Jacobson KA, Salvemini D (2016) Identification of A3 adenosine receptor agonists as novel non-narcotic analgesics. Br J Pharmacol 173:1253–1267. CrossRefPubMedPubMedCentralGoogle Scholar
  25. Jin SX, Zhuang ZY, Woolf CJ, Ji RR (2003) p38 mitogen-activated protein kinase is activated after a spinal nerve ligation in spinal cord microglia and dorsal root ganglion neurons and contributes to the generation of neuropathic pain. J Neurosci 23:4017–4022CrossRefGoogle Scholar
  26. Kalla R, Liu Z, Xu S et al (2001) Microglia and the early phase of immune surveillance in the axotomized facial motor nucleus: impaired microglial activation and lymphocyte recruitment but no effect on neuronal survival or axonal regeneration in macrophage-colony stimulating factor-deficient mice. J Comp Neurol 436:182–201CrossRefGoogle Scholar
  27. Kim SY, Bae JC, Kim JY, Lee HL, Lee KM, Kim DS, Cho HJ (2002) Activation of p38 MAP kinase in the rat dorsal root ganglia and spinal cord following peripheral inflammation and nerve injury. Neuroreport 13:2483–2486. CrossRefPubMedGoogle Scholar
  28. Kim Y, Kwon SY, Jung HS et al (2018) Amitriptyline inhibits MAPK/ERK, CREB pathway and proinflammatory cytokines through A3AR activation in rat neuropathic pain models. Korean J Anesthesiol. CrossRefPubMedPubMedCentralGoogle Scholar
  29. Lavand’homme PM, Eisenach JC (1999) Exogenous and endogenous adenosine enhance the spinal antiallodynic effects of morphine in a rat model of neuropathic pain. Pain 80:31–36CrossRefGoogle Scholar
  30. Ledeboer A, Sloane EM, Milligan ED, Frank MG, Mahony JH, Maier SF, Watkins LR (2005) Minocycline attenuates mechanical allodynia and proinflammatory cytokine expression in rat models of pain facilitation. Pain 115:71–83. CrossRefPubMedGoogle Scholar
  31. Little JW, Ford A, Symons-Liguori AM et al (2015) Endogenous adenosine A3 receptor activation selectively alleviates persistent pain states. Brain 138:28–35. CrossRefPubMedGoogle Scholar
  32. Lopes LV, Rebola N, Pinheiro PC, Richardson PJ, Oliveira CR, Cunha RA (2003) Adenosine A3 receptors are located in neurons of the rat hippocampus. Neuroreport 14:1645–1648. CrossRefPubMedGoogle Scholar
  33. McNicol E, Horowicz-Mehler N, Fisk RA et al (2003) Management of opioid side effects in cancer-related and chronic noncancer pain: a systematic review. J Pain 4:231–256CrossRefGoogle Scholar
  34. Molander C, Hongpaisan J, Grant G (1992) Changing pattern of c-FOS expression in spinal cord neurons after electrical stimulation of the chronically injured sciatic nerve in the rat. Neuroscience 50:223–236CrossRefGoogle Scholar
  35. Murashov AK, Haq IU, Hill C et al (2001) Crosstalk between p38, Hsp25 and Akt in spinal motor neurons after sciatic nerve injury. Brain Res Mol Brain Res 93:199–208CrossRefGoogle Scholar
  36. Piao ZG, Cho IH, Park CK et al (2006) Activation of glia and microglial p38 MAPK in medullary dorsal horn contributes to tactile hypersensitivity following trigeminal sensory nerve injury. Pain 121:219–231. CrossRefPubMedGoogle Scholar
  37. Post C (1984) Antinociceptive effects in mice after intrathecal injection of 5′-N-ethylcarboxamide adenosine. Neurosci Lett 51:325–330CrossRefGoogle Scholar
  38. Raghavendra V, Tanga F, DeLeo JA (2003) Inhibition of microglial activation attenuates the development but not existing hypersensitivity in a rat model of neuropathy. J Pharmacol Exp Ther 306:624–630. CrossRefPubMedGoogle Scholar
  39. Sawynok J (1998) Adenosine receptor activation and nociception. Eur J Pharmacol 347:1–11CrossRefGoogle Scholar
  40. Segerdahl M, Irestedt L, Sollevi A (1997) Antinociceptive effect of perioperative adenosine infusion in abdominal hysterectomy. Acta Anaesthesiol Scand 41:473–479CrossRefGoogle Scholar
  41. Shortland P, Molander C (1998) The time-course of abeta-evoked c-fos expression in neurons of the dorsal horn and gracile nucleus after peripheral nerve injury. Brain Res 810:288–293CrossRefGoogle Scholar
  42. Sugimoto T, Ichikawa H, Hijiya H, Mitani S, Nakago T (1993) c-Fos expression by dorsal horn neurons chronically deafferented by peripheral nerve section in response to spared, somatotopically inappropriate nociceptive primary input. Brain Res 621:161–166CrossRefGoogle Scholar
  43. Swett JE, Woolf CJ (1985) The somatotopic organization of primary afferent terminals in the superficial laminae of the dorsal horn of the rat spinal cord. J Comp Neurol 231:66–77. CrossRefPubMedGoogle Scholar
  44. Terayama R, Omura S, Fujisawa N, Yamaai T, Ichikawa H, Sugimoto T (2008) Activation of microglia and p38 mitogen-activated protein kinase in the dorsal column nucleus contributes to tactile allodynia following peripheral nerve injury. Neuroscience 153:1245–1255. CrossRefPubMedGoogle Scholar
  45. Terayama R, Fujisawa N, Yamaguchi D, Omura S, Ichikawa H, Sugimoto T (2011) Differential activation of mitogen-activated protein kinases and glial cells in the trigeminal sensory nuclear complex following lingual nerve injury. Neurosci Res 69:100–110. CrossRefPubMedGoogle Scholar
  46. Terayama R, Kishimoto N, Yamamoto Y et al (2015a) Convergent nociceptive input to spinal dorsal horn neurons after peripheral nerve injury. Neurochem Res 40:438–445. CrossRefPubMedGoogle Scholar
  47. Terayama R, Yamamoto Y, Kishimoto N, Maruhama K, Mizutani M, Iida S, Sugimoto T (2015b) Peripheral nerve injury activates convergent nociceptive input to dorsal horn neurons from neighboring intact nerve. Exp Brain Res 233:1201–1212. CrossRefPubMedGoogle Scholar
  48. Tokunaga A, Kondo E, Fukuoka T, Miki K, Dai Y, Tsujino H, Noguchi K (1999) Excitability of spinal cord and gracile nucleus neurons in rats with chronically injured sciatic nerve examined by c-fos expression. Brain Res 847:321–331CrossRefGoogle Scholar
  49. Tsuda M, Mizokoshi A, Shigemoto-Mogami Y, Koizumi S, Inoue K (2004) Activation of p38 mitogen-activated protein kinase in spinal hyperactive microglia contributes to pain hypersensitivity following peripheral nerve injury. Glia 45:89–95. CrossRefPubMedGoogle Scholar
  50. Tsuda M, Inoue K, Salter MW (2005) Neuropathic pain and spinal microglia: a big problem from molecules in “small” glia. Trends Neurosci 28:101–107. CrossRefPubMedGoogle Scholar
  51. Wahlman C, Doyle TM, Little JW et al (2018) Chemotherapy-induced pain is promoted by enhanced spinal adenosine kinase levels through astrocyte-dependent mechanisms. Pain 159:1025–1034. CrossRefPubMedGoogle Scholar
  52. Watkins LR, Milligan ED, Maier SF (2001a) Glial activation: a driving force for pathological pain. Trends Neurosci 24:450–455CrossRefGoogle Scholar
  53. Watkins LR, Milligan ED, Maier SF (2001b) Spinal cord glia: new players in pain. Pain 93:201–205CrossRefGoogle Scholar
  54. Wen YR, Suter MR, Kawasaki Y et al (2007) Nerve conduction blockade in the sciatic nerve prevents but does not reverse the activation of p38 mitogen-activated protein kinase in spinal microglia in the rat spared nerve injury model. Anesthesiology 107:312–321. CrossRefPubMedGoogle Scholar
  55. Yamaguchi D, Terayama R, Omura S, Tsuchiya H, Sato T, Ichikawa H, Sugimoto T (2014) Effect of adenosine A1 receptor agonist on the enhanced excitability of spinal dorsal horn neurons after peripheral nerve injury. Int J Neurosci 124:213–222. CrossRefPubMedGoogle Scholar
  56. Yamamoto Y, Terayama R, Kishimoto N, Maruhama K, Mizutani M, Iida S, Sugimoto T (2015) Activated microglia contribute to convergent nociceptive inputs to spinal dorsal horn neurons and the development of neuropathic pain. Neurochem Res 40:1000–1012. CrossRefPubMedGoogle Scholar
  57. Yan H, Zhang E, Feng C, Zhao X (2016) Role of A3 adenosine receptor in diabetic neuropathy. J Neurosci Res 94:936–946. CrossRefPubMedGoogle Scholar
  58. Zhou LJ, Yang T, Wei X et al (2011) Brain-derived neurotrophic factor contributes to spinal long-term potentiation and mechanical hypersensitivity by activation of spinal microglia in rat. Brain Behav Immun 25:322–334. CrossRefPubMedGoogle Scholar
  59. Zylka MJ (2011) Pain-relieving prospects for adenosine receptors and ectonucleotidases. Trends Mol Med 17:188–196. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Oral Function and AnatomyOkayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesOkayamaJapan
  2. 2.Department of Oral and Maxillofacial Reconstructive SurgeryOkayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesOkayamaJapan
  3. 3.Department of Maxillofacial Anatomy and NeuroscienceHiroshima University Graduate School of Biomedical and Health SciencesHiroshimaJapan

Personalised recommendations