Original Report| Volume 12, ISSUE 2, P280-287, February 2011

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Intra-Periaqueductal Gray Matter Microinjection of Orexin-A Decreases Formalin-Induced Nociceptive Behaviors in Adult Male Rats

Published:December 13, 2010DOI:


      Intracerebroventricular injection of orexin-A (hypocretin-1) has been shown to elicit the analgesic responses. However, the locations of central sites that may mediate these effects have not been clearly elucidated. This study was performed using male Sprague Dawley rats to investigate the antinociceptive effects of intra-periaqueductal gray matter (PAG) administration of orexin-A, 5 minutes prior to formalin (50 μL of 2%) injection. Orexin-A had no effect on tail-flick test as thermal and acute model. In the formalin test, intra-PAG injection of orexin-A (10 nM) decreased the formalin-induced nociceptive behaviors in the interphase and phase 2, but not in phase 1, indicating an antinociceptive role of exogenous orexin-A in the PAG. While Orexin-A failed to produce a dose-dependent decrease in formalin-evoked behaviors in phase 1, it may have induced a dose-dependent decrease in formalin-evoked behaviors in early phase 2. Control injections of orexin-A into the sites near the PAG resulted in less or no reduction in pain, indicating that the analgesia observed is probably due to a site of action within the PAG rather than at surrounding neural structures. The antinociceptive effect of orexin-A was compared with intra-PAG administration of morphine (.5 μL of 20 mM, 5 minutes before the formalin injection). Morphine decreased the formalin-induced nociceptive behaviors in all phases. To investigate whether the orexin has a special action on the early part of the second phase, or its delayed effects are related to its pharmacokinetics, the orexin-A was injected into the PAG, 10 minutes before the formalin injection. No difference was observed between 5 and 10 minutes injection of orexin-A prior to formalin injection. The antinociceptive effect of orexin was blocked by intra-PAG injection of SB-334867, a putative type 1 orexin receptor antagonist, suggesting the involvement of orexin receptor type 1 in antinociception produced with intra-PAG injection of orexin-A. The results showed that the orexin-A plays an antinociceptive role in PAG in the interphase and the late phase of formalin test through type 1 orexin receptor dependent mechanism.


      Orexin is produced exclusively in the lateral hypothalamus, where it is known to modulate the pain processing through PAG. The antinociceptive effect of orexin in PAG may provide a role for this neurotransmitter in the up-down modulating pain system and further support the development of orexin-1 agonists for pain treatment.

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        • Azhdari-Zarmehri H.
        • Semnanian S.
        • Fathollahi Y.
        • Haghdoost-Yazdi H.
        • Khakpay R.
        • Azizi H.
        • Rohampour K.
        Periaqueductal gray matter is involved in orexin-A induced analgesia in formalin test.
        FENS Abstr. 2010; 5: 140.1
        • Abbott F.V.
        • Franklin K.B.
        • Westbrook R.F.
        The formalin test: Scoring properties of the first and second phases of the pain response in rats.
        Pain. 1995; 60: 91-102
        • Behbehani M.M.
        • Park M.R.
        • Clement M.E.
        Interactions between the lateral hypothalamus and the periaqueductal gray.
        J Neurosci. 1988; 8: 2780-2787
        • Bingham S.
        • Davey P.T.
        • Babbs A.J.
        • Irving E.A.
        • Sammons M.J.
        • Wyles M.
        • Jeffrey P.
        • Cutler L.
        • Riba I.
        • Johns A.
        • Porter R.A.
        • Upton N.
        • Hunter A.J.
        • Parsons A.A.
        Orexin-A, an hypothalamic peptide with analgesic properties.
        Pain. 2001; 92: 81-90
        • Cheng J.K.
        • Chou R.C.
        • Hwang L.L.
        • Chiou L.C.
        Antiallodynic effects of intrathecal orexins in a rat model of postoperative pain.
        J Pharmacol Exp Ther. 2003; 307: 1065-1071
        • de Lecea. L.
        • Kilduff T.S.
        • Peyron C.
        • Gao X.
        • Foye P.E.
        • Danielson P.E.
        • Fukuhara C.
        • Battenberg E.L.
        • Gautvik V.T.
        • Bartlett F.S.
        • Frankel W.N.
        • van den Pol A.N.
        • Bloom F.E.
        • Gautvik K.M.
        • Sutcliffe J.G.
        The hypocretins: hypothalamus-specific peptides with neuroexcitatory activity.
        Proc Natl Acad Sci U S A. 1998; 95: 322-327
        • Dubuisson D.
        • Dennis S.G.
        The formalin test: A quantitative study of the analgesic effects of morphine, meperidine, and brain stem stimulation in rats and cats.
        Pain. 1977; 4: 161-174
        • Ferguson A.V.
        • Samson W.K.
        The orexin/hypocretin system: A critical regulator of neuroendocrine and autonomic function.
        Front Neuroendocrinol. 2003; 24: 141-150
        • Franklin K.B.
        • Abbott F.V.
        Pentobarbital, diazepam, and ethanol abolish the interphase diminution of pain in the formalin test: Evidence for pain modulation by GABAA receptors.
        Pharmacol Biochem Behav. 1993; 46: 661-666
        • Paxinos G.
        • Watson C.
        The Rat Brain in Stereotaxic Coordinates.
        4th and 6th eds. Academic Press, New York, NY2005
        • Gaumond I.
        • Arsenault P.
        • Marchand S.
        The role of sex hormones on formalin-induced nociceptive responses.
        Brain Res. 2002; 958: 139-145
        • Henry J.L.
        • Yashpal K.
        • Pitcher G.M.
        • Coderre T.J.
        Physiological evidence that the ‘interphase’ in the formalin test is due to active inhibition.
        Pain. 1999; 82: 57-63
        • Horvath T.L.
        • Peyron C.
        • Diano S.
        • Ivanov A.
        • Aston-Jones G.
        • Kilduff T.S.
        • van den Pol A.N.
        Hypocretin (orexin) activation and synaptic innervation of the locus coeruleus noradrenergic system.
        J Comp Neurol. 1999; 415: 145-159
        • Hunskaar S.
        • Berge O.G.
        • Hole K.
        Dissociation between antinociceptive and anti-inflammatory effects of acetylsalicylic acid and indomethacin in the formalin test.
        Pain. 1986; 25: 125-132
        • Hunskaar S.
        • Hole K.
        The formalin test in mice: Dissociation between inflammatory and non-inflammatory pain.
        Pain. 1987; 30: 103-114
        • Lewis V.A.
        • Gebhart G.F.
        Evaluation of the periaqueductal central gray (PAG) as a morphine-specific locus of action and examination of morphine-induced and stimulation-produced analgesia at coincident PAG loci.
        Brain Res. 1977; 124: 283-303
        • Mobarakeh J.I.
        • Takahashi K.
        • Sakurada S.
        • Nishino S.
        • Watanabe H.
        • Kato M.
        • Yanai K.
        Enhanced antinociception by intracerebroventricularly and intrathecally-administered orexin A and B (hypocretin-1 and -2) in mice.
        Peptides. 2005; 26: 767-777
        • Mobarakeh J.I.
        • Takahashi K.
        • Sakurada S.
        • Nishino S.
        • Watanabe H.
        • Kato M.
        • Naghdi N.
        • Yanai K.
        Enhanced antinociception by intracerebroventricularly administered orexin A in histamine H1 or H2 receptor gene knockout mice.
        Pain. 2005; 188: 254-262
        • Peyron C.
        • Tighe D.K.
        • van den Pol A.N.
        • de L.L.
        • Heller H.C.
        • Sutcliffe J.G.
        • Kilduff T.S.
        Neurons containing hypocretin (orexin) project to multiple neuronal systems.
        J Neurosci. 1998; 18: 9996-10015
        • Sakurai T.
        Roles of orexin/hypocretin in regulation of sleep/wakefulness and energy homeostasis.
        Sleep Med Rev. 2005; 9: 231-241
        • Sakurai T.
        • Amemiya A.
        • Ishii M.
        • Matsuzaki I.
        • Chemelli R.M.
        • Tanaka H.
        • Williams S.C.
        • Richardson J.A.
        • Kozlowski G.P.
        • Wilson S.
        • Arch J.R.
        • Buckingham R.E.
        • Haynes A.C.
        • Carr S.A.
        • Annan R.S.
        • McNulty D.E.
        • Liu W.S.
        • Terrett J.A.
        • Elshourbagy N.A.
        • Bergsma D.J.
        • Yanagisawa M.
        Orexins and orexin receptors: A family of hypothalamic neuropeptides and G protein-coupled receptors that regulate feeding behavior.
        Cell. 1998; 92: 573-585
        • Samson W.K.
        • Taylor M.M.
        • Ferguson A.V.
        Non-sleep effects of hypocretin/orexin.
        Sleep Med Rev. 2005; 9: 243-252
        • Siegel J.M.
        The narcoleptic borderland.
        Sleep Med. 2003; 4: 3-4
        • Suyama H.
        • Kawamoto M.
        • Shiraishi S.
        • Gaus S.
        • Kajiyama S.
        • Yuge O.
        Analgesic effect of intrathecal administration of orexin on neuropathic pain in rats.
        In Vivo. 2004; 18: 119-123
        • Tao R.
        • Ma Z.
        • McKenna J.T.
        • Thakkar M.M.
        • Winston S.
        • Strecker R.E.
        • McCarley R.W.
        Differential effect of orexins (hypocretins) on serotonin release in the dorsal and median raphe nuclei of freely behaving rats.
        Neuroscience. 2006; 141: 1101-1105
        • Yamamoto T.
        • Nozaki-Taguchi N.
        • Chiba T.
        Analgesic effect of intrathecally administered orexin-A in the rat formalin test and in the rat hot plate test.
        Br J Pharmacol. 2002; 137: 170-176
        • Yamamoto T.
        • Saito O.
        • Shono K.
        • Hirasawa S.
        Activation of spinal orexin-1 receptor produces anti-allodynic effect in the rat carrageenan test.
        Eur J Pharmacol. 2003; 481: 175-180
        • Yeung J.C.
        • Yaksh T.L.
        • Rudy T.A.
        Concurrent mapping of brain sites for sensitivity to the direct application of morphine and focal electrical stimulation in the production of antinociception in the rat.
        Pain. 1977; 4: 23-40