Abstract
Opioids and nitric oxide (NO) interact functionally in different systems. NO-generating agents decrease the activity of opioid agonists, prevent opioid tolerance, and are used in opioid withdrawal syndromes. There exist, however, few reports indicating a direct interaction of the two systems. T47D human breast cancer cells in culture express opioid receptors, and opioid agonists inhibit their growth, while they release high amounts of the NO-related molecules NO2−/NO3− to the culture medium. We have used this system to assay a possible direct interaction of opiergic and nitric oxide systems. Our results show that δ- or μ-acting opioid agonists do not modify the release of NO2−/NO3−. In contrast, κ-acting opioid agonists (ethylketocyclazocine, and αS1-casomorphine) decrease the release of NO2−/NO3−, in a time- and dose-dependent manner. The general opioid antagonist diprenorphine (10−6 M) produce a similar NO2−/NO3− release inhibition, indicating a possible non-opioid-receptor mediated phenomenon. In addition, ethylketocyclazocine, αS1-casomorphin and diprenorphine directly inhibit NOS activity: agonists, interact with both calcium-dependent and independent NOS-isoforms, while the antagonist diprenorphine modifies only the activity of the calcium-dependent fraction of the enzyme. Analysis of this interaction revealed that opioids modify the dimeric active form of NOS, through binding to the reductase part of the molecule, acting as non-competitive inhibitors of the enzyme. This interaction opens interesting new possibilities for tumor biology and breast cancer therapy. Cell Death and Differentiation (2001) 8, 943–952
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
Abbreviations
- EKC:
-
ethylketocyclazocine
- DSLET:
-
[D-Ser2, Leu5]-enkephalin-Thr6; DAGO, [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin; NO, nitric oxide
- NOS:
-
nitric oxide synthase
- PMA:
-
phorbol 12-myristate, 13-acetate
References
LaForge KS, Yuferov V, Kreek MJ . 2000 Opioid receptor and peptide polymorphisms: Potential implications for addictions Eur. J. Pharmacol. 410: 249–268
Kampa M, Loukas S, Hatzoglou A, Martin P, Martin P-M . 1996 Identification of a novel opioid peptide derived from alpha-S1 human casein (alpha-S1 casomorphin, and alpha-S1 casomorphin-amide) Biochem. J. 319: 903–908
Kampa M, Bakogeorgou E, Hatzoglou A, Damianaki A, Martin PM, Castanas E . 1997 Opioid alkaloids and casomorphin peptides decrease the proliferation of prostatic cancer cell lines (LNCaP, PC3 and DU145) through a partial interaction with opioid receptors Eur. J. Pharmacol. 335: 255–265
Panagiotou S, Bakogeorgou E, Papakonstanti E, Hatzoglou A, Wallet F, Dussert C, Stournaras C, Martin PM, Castanas E . 1999 Opioid agonists modify breast cancer cell proliferation by blocking cells to the G2/M phase of the cycle: involvement of cytoskeletal elements J. Cell. Biochem. 73: 204–211
Papakonstanti EA, Bakogeorgou E, Castanas E, Emmanouel DS, Hartig R, C S . 1998 Early alterations of actin cytoskeleton in OK cells by opioids J. Cell. Biochem. 70: 60–69
Zajac JM, Roques BP . 1983 Differential properties of mu and delta opiate binding sites studied with highly selective ligands Life Sci. 33 Suppl. 1: 155–158
Marletta MA, Hursman AR, Rusche KM . 1998 Catalysis by nitric oxide synthase Curr. Opinion Chem. Biol. 2: 656–663
Stuehr DJ . 1999 Mammalian nitric oxide synthases Biohim. Biophys. Acta. 1411: 217–230
Denninger JW, Marletta MA . 1999 Guanylate cyclase and the NO/cGMP signaling pathway Biochim. Biophys. Acta. 1411: 334–350
Colasanti M, Suzuki H . 2000 The dual personality of NO Trends Pharmacol. Sci. 21: 249–252
Wink DA, Vodovotz Y, Laval J, Laval F, Dewhirst MW, Mitchell JB . 1998 The multifaceted roles of nitric oxide in cancer Carcinogenesis. 19: 711–721
Adams ML, Meyer ER, Cicero TJ . 1996 Effects of nitric oxide-related agents on opioid regulation of rat testicular steroidogenesis Biol. Reprod. 54: 1128–1134
Budziszewska B, Leskiewicz M, Jaworska-Feil L, Lason W . 1999 The effect of N-nitro-L-arginine methyl ester on morphine-induced changes in the plasma corticosterone and testosterone levels in mice Exp. Clin. Endocrinol. Diabetes. 107: 75–79
Bhargava HN, Bian JT, Kumar S . 1997 Mechanism of attenuation of morphine antinociception by chronic treatment with L-arginine J. Pharmacol. Exp. Ther. 281: 707–712
Bhargava HN, Bian JT . 1998 Effects of acute administration of L-arginine on morphine antinociception and morphine distribution in central and peripheral tissues of mice Pharmacol. Biochem. Behav. 61: 29–33
Babey AM, Kolesnikov Y, Cheng J, Inturrisi CE, Trifilletti RR, Pasternak GW . 1994 Nitric oxide and opioid tolerance Neuropharmacology 33: 1463–1470
Bhargava HN, Cao YJ . 1997 Effect of chronic administration of morphine, U-50, 488H and [D-Pen2, D- Pen5]enkephalin on the concentration of cGMP in brain regions and spinal cord of the mouse Peptides 18: 1629–1634
Pu S, Horvath TL, Diano S, Naftolin F, Kalra PS, Kalra SP . 1997 Evidence showing that beta-endorphin regulates cyclic guanosine 3′,5′- monophosphate (cGMP) efflux: anatomical and functional support for an interaction between opiates and nitric oxide [published erratum appears in Endocrinology 1997 Jun; 138(6): 2609] Endocrinology 138: 1537–1543
Lomniczi A, Mastronardi CA, Faletti AG, Seilicovich A, De Laurentiis A, McCann SM, Rettori V . 2000 Inhibitory pathways and the inhibition of luteinizing hormone-releasing hormone release by alcohol Proc. Natl. Acad. Sci. USA 97: 2337–2342
Faletti AG, Mastronardi CA, Lomniczi A, Seilicovich A, Gimeno M, McCann SM, Rettori V . 1999 beta-Endorphin blocks luteinizing hormone-releasing hormone release by inhibiting the nitricoxidergic pathway controlling its release Proc. Natl. Acad. Sci. USA 96: 1722–1726
Bhat GK, Mahesh VB, Ping L, Chorich L, Wiedmeier VT, Brann DW . 1998 Opioid-glutamate-nitric oxide connection in the regulation of luteinizing hormone secretion in the rat Endocrinology 139: 955–960
Prevot V, Rialas CM, Croix D, Salzet M, Dupouy JP, Poulain P, Beauvillain JC, Stefano GB . 1998 Morphine and anandamide coupling to nitric oxide stimulates GnRH and CRF release from rat median eminence: neurovascular regulation Brain Res. 790: 236–244
Bilfinger TV, Salzet M, Fimiani C, Deutsch DG, Tramu G, Stefano GB . 1998 Pharmacological evidence for anandamide amidase in human cardiac and vascular tissues Int. J. Cardiol. 64 Suppl 1: S15–S22
Stefano GB, Goumon Y, Bilfinger TV, Welters ID, Cadet P . 2000 Basal nitric oxide limits immune, nervous and cardiovascular excitation: human endothelia express a mu opiate receptor Prog. Neurobiol. 60: 513–530
Stefano GB, Salzet M, Magazine HI, Bilfinger TV . 1998 Antagonism of LPS and IFN-gamma induction of iNOS in human saphenous vein endothelium by morphine and anandamide by nitric oxide inhibition of adenylate cyclase J. Cardiovaso. Pharmacol. 31: 813–820
Stefano GB, Hartman A, Bilfinger TV, Magazine HI, Liu Y, Casares F, Goligorsky MS . 1995 Presence of the mu3 opiate receptor in endothelial cells. Coupling to nitric oxide production and vasodilation J. Biol. Chem. 270: 30290–30293
Wilderman MJ, Armstead WM . 1998 Role of endothelial nitric oxide synthase in hypoxia-induced pial artery dilation J. Cereb. Blood Flow Metab. 18: 531–538
Wilderman MJ, Armstead WM . 1997 Role of neuronal NO synthase in relationship between NO and opioids in hypoxia-induced pial artery dilation Am. J. Physiol. 273: H1807–H1815
Lue WM, Su MT, Lin WB, Tao PL . 1999 The role of nitric oxide in the development of morphine tolerance in rat hippocampal slices Eur. J. Pharmacol. 383: 129–135
Powell KJ, Hosokawa A, Bell A, Sutak M, Milne B, Quirion R, Jhamandas K . 1999 Comparative effects of cyclo-oxygenase and nitric oxide synthase inhibition on the development and reversal of spinal opioid tolerance Br. J. Pharmacol. 127: 631–644
Aley KO, Levine JD . 1997 Different mechanisms mediate development and expression of tolerance and dependence for peripheral mu-opioid antinociception in rat J. Neurosci. 17: 8018–8023
Bhargava HN . 1995 Attenuation of tolerance to, and physical dependence on, morphine in the rat by inhibition of nitric oxide synthase [published erratum appears in Gen Pharmacol 1996 Apr;27(3):557] Gen. Pharmacol. 26: 1049–1053
Majeed NH, Przewlocka B, Machelska H, Przewlocki R . 1994 Inhibition of nitric oxide synthase attenuates the development of morphine tolerance and dependence in mice Neuropharmacology 33: 189–192
Pineda J, Torrecilla M, Martin-Ruiz R, Ugedo L . 1998 Attenuation of withdrawal-induced hyperactivity of locus coeruleus neurones by inhibitors of nitric oxide synthase in morphine-dependent rats Neuropharmacology 37: 759–767
Highfield DA, Grant S . 1998 Ng-nitro-L-arginine, an NOS inhibitor, reduces tolerance to morphine in the rat locus coeruleus Synapse. 29: 233–239
Hall S, Milne B, Jhamandas K . 1996 Nitric oxide synthase inhibitors attenuate acute and chronic morphine withdrawal response in the rat locus coeruleus: an in vivo voltammetric study Brain Res. 739: 182–191
Vaupel DB, Kimes AS, London ED . 1995 Nitric oxide synthase inhibitors. Preclinical studies of potential use for treatment of opioid withdrawal Neuropsychopharmacology 13: 315–322
Herman BH, Vocci F, Bridge P . 1995 The effects of NMDA receptor antagonists and nitric oxide synthase inhibitors on opioid tolerance and withdrawal. Medication development issues for opiate addiction Neuropsychopharmacology 13: 269–293
Vaupel DB, Kimes AS, London ED . 1995 Comparison of 7-nitroindazole with other nitric oxide synthase inhibitors as attenuators of opioid withdrawal Psychopharmacology (Berl). 118: 361–368
London ED, Kimes AS, Vaupel DB . 1995 Inhibitors of nitric oxide synthase and the opioid withdrawal syndrome NIDA Res. Monogr. 147: 170–181
Kolesnikov YA, Pick CG, Ciszewska G, Pasternak GW . 1993 Blockade of tolerance to morphine but not to kappa opioids by a nitric oxide synthase inhibitor Proc. Natl. Acad. Sci. USA 90: 5162–5166
Adams ML, Kalicki JM, Meyer ER, Cicero TJ . 1993 Inhibition of the morphine withdrawal syndrome by a nitric oxide synthase inhibitor, NG-nitro-L-arginine methyl ester Life Sci. 52: L245–249
Machelska H, Labuz D, Przewlocki R, Przewlocka B . 1997 Inhibition of nitric oxide synthase enhances antinociception mediated by mu, delta and kappa opioid receptors in acute and prolonged pain in the rat spinal cord J. Pharmacol. Exp. Ther. 282: 977–984
Przewlocki R, Machelska H, Przewlocka B . 1993 Inhibition of nitric oxide synthase enhances morphine antinociception in the rat spinal cord Life Sci. 53: L1–L5
Cuellar B, Fernandez AP, Lizasoain I, Moro MA, Lorenzo P, Bentura ML, Rodrigo J, Leza JC . 2000 Up-regulation of neuronal NO synthase immunoreactivity in opiate dependence and withdrawal Psychopharmacology (Berl). 148: 66–73
Deliconstantinos G, Villiotou V, Stavrides JC . 1995 Met-enkephalin receptor-mediated increase of membrane fluidity modulates nitric oxide (NO) and cGMP production in rat brain synaptosomes Neurochem. Res. 20: 217–224
Pataki I, Telegdy G . 1998 Further evidence that nitric oxide modifies acute and chronic morphine actions in mice Eur. J. Pharmacol. 357: 157–162
Kavaliers M, Choleris E, Prato FS, Ossenkopp K . 1998 Evidence for the involvement of nitric oxide and nitric oxide synthase in the modulation of opioid-induced antinociception and the inhibitory effects of exposure to 60-Hz magnetic fields in the land snail Brain Res. 809: 50–57
Granados-Soto V, Rufino MO, Gomes Lopes LD, Ferreira SH . 1997 Evidence for the involvement of the nitric oxide-cGMP pathway in the antinociception of morphine in the formalin test Eur. J. Pharmacol. 340: 177–180
Kolesnikov YA, Pan YX, Babey AM, Jain S, Wilson R, Pasternak GW . 1997 Functionally differentiating two neuronal nitric oxide synthase isoforms through antisense mapping: evidence for opposing NO actions on morphine analgesia and tolerance Proc. Natl. Acad. Sci. USA 94: 8220–8225
Ding YQ, Li JL, Lu BZ, Wang D, Zhang ML, Li JS . 1998 Co-localization of mu-opioid receptor-like immunoreactivity with substance P-LI, calcitonin gene-related peptide-LI and nitric oxide synthase-LI in vagal and glossopharyngeal afferent neurons of the rat Brain Res. 792: 149–153
Fox-Threlkeld JE, Daniel EE, Christinck F, Hruby VJ, Cipris S, Woskowska Z . 1994 Identification of mechanisms and sites of actions of mu and delta opioid receptor activation in the canine intestine J. Pharmacol. Exp. Ther. 268: 689–700
Menzies JR, Corbett AD . 2000 Nociceptin inhibits tonic nitric oxide release in the mouse isolated proximal colon Eur. J. Pharmacol. 388: 183–186
Hatzoglou A, Ouafik L, Bakogeorgou E, Thermos K, Castanas E . 1995 Morphine cross-reacts with somatostatin receptor SSTR2 in the T47D human breast cancer cell line and decreases cell growth Cancer Res. 55: 5632–5636
Hatzoglou A, Bakogeorgou E, Hatzoglou C, Martin PM, Castanas E . 1996 Antiproliferative and receptor binding properties of alpha- and beta- casomorphins in the T47D human breast cancer cell line Eur. J. Pharmacol. 310: 217–223
Hatzoglou A, Bakogeorgou E, Castanas E . 1996 The antiproliferative effect of opioid receptor agonists on the T47D human breast cancer cell line, is partially mediated through opioid receptors Eur. J. Pharmacol. 296: 199–207
Hatzoglou A, Kampa M, Castanas E . Wine antioxidants as antiproliferative agents in human hormone-dependent tumors. In: 6th International Symposium on Grapevine physiology and biotechnology Heraklion, Greece, 11–15 June 2000 pp. 7
Munro AW, Taylor P, Walkinshaw MD . 2000 Structure of redox enzymes Curr. Opinion Biotechnol. 11: 369–376
Barjavel MJ, Bhargava HN . 1994 Effect of opioid receptor, agonists on nitric oxide synthase activity in rat cerebral cortex homogenate Neurosci. Lett. 181: 27–30
Benyo Z, Szabo C, Velkei MH, Bohus B, Wahl M, Sandor P . 1996 Intravenous beta-endorphin administration fails to alter hypothalamic blood flow in rats expressing normal or reduced nitric oxide synthase activity Peptides 17: 733–736
Iuvone T, Capasso A, D'Acquisto F, Carnuccio R . 1995 Opioids inhibit the induction of nitric oxide synthase in J774 macrophages Biochem. Biophys. Res. Commun. 212: 975–980
Kowalski J . 1998 Augmenting effect of opioids on nitrite production by stimulated murine macrophages Neuropeptides 32: 287–291
Li X, David Clark J . 2000 Chronic morphine exposure and the expression of heme oxygenase type 2 Brain Res. Mol. Brain Res. 75: 179–184
Lysle DT, How T . 2000 Heroin modulates the expression of inducible nitric oxide synthase Immunopharmacology 46: 181–192
Bhargava HN . 1994 Nitric oxide synthase inhibition blocks tolerance to the analgesic action of kappa-opiate receptor agonist in the rat Pharmacology 48: 234–241
Thorat SN, Reddy PL, Bhargava HN . 1993 Evidence for the role of nitric oxide in kappa-opiate tolerance in mice Brain Res. 621: 171–174
Bhargava HN, Kumar S, Barjavel MJ . 1998 Kinetic properties of nitric oxide synthase in cerebral cortex and cerebellum of morphine tolerant mice Pharmacology 56: 252–256
Engel PC . 1981 Enzyme kinetics: The steady-state approach., 2nd edition p. 96 London: Chapman and Hall
Ko JL, Arvidsson U, Williams FG, Law PY, Elde R, Loh HH . 1999 Visualization of time-dependent redistribution of ä-opioid receptors in neuronal cells during prolonged agonist exposure Mol. Brain Res. 69: 171–185
Gaudriault G, Nouel D, Dal Farra C, Beaudet A, Vincent J-P . 1997 Receptor-induced internalization of selective peptidic ì and ä opioid ligands J. Biol. Chem. 272: 2880–2888
Schweigerer L, Schmidt W, Teschenmacher H, Gramsch C . 1985 â-endorphin: Surface binding and internalization in thymoma cells Proc. Natl. Acad. Sci. USA. 82: 5751–5755
Law PY, Hom DS, Loh HH . 1984 Down regulation of opiate receptor in neuroblastoma x glioma NG108-15 hybrid cells J. Biol. Chem. 259: 4096–4104
Li J-G, Luo L-Y, Krupnick JG, Benovic JL, Liu-Chen L-Y . 1999 U50, 488H-induced internalization of the human ê opioid receptor involves a â-arestin- and dynamin-dependent mechanism: ê receptor internalization is not required for mitogen-activated protein kinase activation J. Biol. Chem. 274: 12087–12094
Jordan BA, Cvejic S, Devi LA . 2000 Kappa opioid receptor endocytosis by dynorphin peptides DNA Cell. Biol. 19: 19–27
Li J-G, Benovic JL, Liu-Chen L-Y . 2000 Mechanisms of agonist-induced down-regulation of the human ê-opioid receptor: Internalization is required for down regulation Mol. Pharmacol. 58: 795–801
Maneckjee R, Biswas R, Vonderhaar BK . 1990 Binding of opioids to human MCF-7 breast cancer cells and their effects on growth Cancer Res. 50: 2234–2238
Grisham MB, Johnson GG, Lancaster JRJ . 1996 Quantitation of nitrate and nitrite in extracellular fluids Meth. Enzymol. 268: 237–246
Granger DL, Taintor RR, Boockvar KS, Hibbs JBJ . 1996 Measurement of nitrate and nitrite in biological samples using nitrate reductase and Griess reaction Meth. Enzymol. 268: 142–151
Kopec KK, Carroll RT . 2000 Phagocytosis is regulated by Nitric Oxide in Murine Microglia Nitric Oxide Biol. Med. 4: 103–111
Kojima H, Nakatsubo N, Kikuchi K, Kawahara S, Kirino Y, Nagoshi H, Hirata Y, Nagano T . 1998 Detection and imaging of nitric oxide with novel fluorescent indicators: Diaminofluorosceins. Anal. Chem. 70, 2446–2453 of the constitutive endothelial nitric oxide synthase gene in Alzheimer disease Mol. Chem. Neuropathol. 30: 139–159
Marletta MA . 1993 Nitric oxide synthase structure and mechanism J. Biol. Chem. 268: 12231–12234
Moncada S, Higgs A . 1993 The L-arginine-nitric oxide pathway N. Engl. J. Med. 329: 2002–2012
Dawes EA . 1972 Quantitative problems in biochemistry. 1st edition Edinburgh: Churchill Livingstone
Acknowledgements
Work partially supported by grants from the General Secretariat of Research and Technology (GGET), the Ministry of Health (KESY), and Varelas S.A.
Author information
Authors and Affiliations
Corresponding author
Additional information
Edited by G Melino
Rights and permissions
About this article
Cite this article
Kampa, M., Hatzoglou, A., Notas, G. et al. Opioids are non-competitive inhibitors of nitric oxide synthase in T47D human breast cancer cells. Cell Death Differ 8, 943–952 (2001). https://doi.org/10.1038/sj.cdd.4400893
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/sj.cdd.4400893
Keywords
This article is cited by
-
Local anaesthetics upregulate nitric oxide generation in cord blood and adult human neutrophils
Scientific Reports (2019)
-
The influence of opioids on matrix metalloproteinase-2 and -9 secretion and mRNA levels in MCF-7 breast cancer cell line
Molecular Biology Reports (2011)
