Molecular and Cellular Biochemistry

, Volume 350, Issue 1–2, pp 101–111 | Cite as

Salt modulates vascular response through adenosine A2A receptor in eNOS-null mice: role of CYP450 epoxygenase and soluble epoxide hydrolase

  • Mohammed A. NayeemEmail author
  • Darryl C. Zeldin
  • Matthew A. Boegehold
  • John R. Falck


High salt (HS) intake can change the arterial tone in mice, and the nitric oxide (NO) acts as a mediator to some of the receptors mediated vascular response. The main aim of this study was to explore the mechanism behind adenosine-induced vascular response in HS-fed eNOS+/+ and eNOS−/− mice The modulation of vascular response by HS was examined using aortas from mice (eNOS+/+ and eNOS−/−) fed 4% (HS) or 0.45% (NS) NaCl-diet through acetylcholine (ACh), NECA (adenosine-analog), CGS 21680 (A2A AR-agonist), MS-PPOH (CYP epoxygenase-blocker; 10−5 M), AUDA (sEH-blocker; 10−5 M), and DDMS (CYP4A-blocker; 10−5 M). ACh-response was greater in HS-eNOS+/+ (+59.3 ± 6.3%) versus NS-eNOS+/+ (+33.3 ± 8.0%; P < 0.05). However, there was no response in both HS-eNOS−/− and NS-eNOS−/−. NECA-response was greater in HS-eNOS−/− (+37.4 ± 3.2%) versus NS-eNOS−/− (+7.4.0 ± 3.8%; P < 0.05). CGS 21680-response was also greater in HS-eNOS−/− (+45.4 ± 5.2%) versus NS-eNOS−/−(+5.1 ± 5.0%; P < 0.05). In HS-eNOS−/−, the CGS 21680-response was reduced by MS-PPOH (+7.3 ± 3.2%; P < 0.05). In NS-eNOS−/−, the CGS 21680-response was increased by AUDA (+38.2 ± 3.3%; P < 0.05) and DDMS (+30.1 ± 4.1%; P < 0.05). Compared to NS, HS increased CYP2J2 in eNOS+/+ (35%; P < 0.05) and eNOS−/− (61%; P < 0.05), but decreased sEH in eNOS+/+ (74%; P < 0.05) and eNOS−/− (40%; P < 0.05). Similarly, CYP4A decreased in HS-eNOS+/+ (35%; P < 0.05) and HS-eNOS−/− (34%; P < 0.05). These data suggest that NS causes reduced-vasodilation in both eNOS+/+ and eNOS−/− via sEH and CYP4A. However, HS triggers possible A2AAR-induced relaxation through CYP epoxygenase in both eNOS+/+ and eNOS−/−.


Salt eNOS CYP2J2 sEH Relaxation Adenosine 



Endothelium-derived relaxing factors


Endothelium-derived hyperpolarizing factor


NG-nitro-l-arginine methyl ester


Soluble epoxide hydrolase


Endothelial nitric oxide synthase


Eicosatrienoic acid






12-(3-adamantan-1-yl-ureido)dodecanoic acid





The authors would like to thank Dr. Mustafa for his support (HL 027339 and MAN is Co-Investigator in HL 094447), AHA 2250298 (MAB), GM 31278 (JRF), and the Intramural Research Program of the NIEHS/NIH-Z01 ES025034 (DCZ).


  1. 1.
    Wong WT, Wong SL, Tian XY, Huang Y (2010) Endothelial dysfunction: the common consequence in diabetes and hypertension. J Cardiovasc Pharmacol 55:300–307PubMedCrossRefGoogle Scholar
  2. 2.
    Brandes RP, Schmitz-Winnenthal FH, Feletou M, Godecke A, Huang PL, Vanhoutte PM, Fleming I, Busse R (2000) An endothelium-derived hyperpolarizing factor distinct from NO and prostacyclin is a major endothelium-dependent vasodilator in resistance vessels of wild-type and endothelial NO synthase knockout mice. Proc Natl Acad Sci USA 97:9747–9752PubMedCrossRefGoogle Scholar
  3. 3.
    Nayeem MA, Ponnoth DS, Boegehold MA, Zeldin DC, Falck JR, Mustafa SJ (2009) High-salt diet enhances mouse aortic relaxation through adenosine A2A receptor via CYP epoxygenases. Am J Physiol Regul Integr Comp Physiol 296:R567–R574PubMedCrossRefGoogle Scholar
  4. 4.
    Nayeem MA, Poloyac SM, Falck JR, Zeldin DC, Ledent C, Ponnoth DS, Ansari HR, Mustafa SJ (2008) Role of CYP epoxygenases in A2A AR-mediated relaxation using A2A AR-null and wild-type mice. Am J Physiol Heart Circ Physiol 295:H2068–H2078PubMedCrossRefGoogle Scholar
  5. 5.
    Nayeem MA, Zeldin DC, Boegehold MA, Morisseau C, Marowsky A, Ponnoth DS, Roush KP, Falck JR (2010) Modulation by salt intake of the vascular response mediated through adenosine A2A receptor: role of CYP epoxygenase and soluble epoxide hydrolase. Am J Physiol Regul Integr Comp Physiol 299(1):R325–R333PubMedCrossRefGoogle Scholar
  6. 6.
    Elhusseiny A, Hamel E (2000) Muscarinic—but not nicotinic—acetylcholine receptors mediate a nitric oxide-dependent dilation in brain cortical arterioles: a possible role for the M5 receptor subtype. J Cereb Blood Flow Metab 20:298–305PubMedCrossRefGoogle Scholar
  7. 7.
    Rosenblum WI, Nishimura H, Nelson GH (1990) Endothelium-dependent l-Arg- and L-NMMA-sensitive mechanisms regulate tone of brain microvessels. Am J Physiol 259:H1396–H1401PubMedGoogle Scholar
  8. 8.
    Sobey CG, Faraci FM (1997) Effects of a novel inhibitor of guanylyl cyclase on dilator responses of mouse cerebral arterioles. Stroke 28:837–842 discussion 842-833PubMedGoogle Scholar
  9. 9.
    Faraci FM, Heistad DD (1998) Regulation of the cerebral circulation: role of endothelium and potassium channels. Physiol Rev 78:53–97PubMedGoogle Scholar
  10. 10.
    Faraci FM, Sobey CG (1999) Role of soluble guanylate cyclase in dilator responses of the cerebral microcirculation. Brain Res 821:368–373PubMedCrossRefGoogle Scholar
  11. 11.
    Huang PL (2000) Mouse models of nitric oxide synthase deficiency. J Am Soc Nephrol 11(Suppl 16):S120–S123PubMedGoogle Scholar
  12. 12.
    Shesely EG, Maeda N, Kim HS, Desai KM, Krege JH, Laubach VE, Sherman PA, Sessa WC, Smithies O (1996) Elevated blood pressures in mice lacking endothelial nitric oxide synthase. Proc Natl Acad Sci USA 93:13176–13181PubMedCrossRefGoogle Scholar
  13. 13.
    Huang PL, Huang Z, Mashimo H, Bloch KD, Moskowitz MA, Bevan JA, Fishman MC (1995) Hypertension in mice lacking the gene for endothelial nitric oxide synthase. Nature 377:239–242PubMedCrossRefGoogle Scholar
  14. 14.
    Godecke A, Decking UK, Ding Z, Hirchenhain J, Bidmon HJ, Godecke S, Schrader J (1998) Coronary hemodynamics in endothelial NO synthase knockout mice. Circ Res 82:186–194PubMedGoogle Scholar
  15. 15.
    Fagan KA, McMurtry I, Rodman DM (2000) Nitric oxide synthase in pulmonary hypertension: lessons from knockout mice. Physiol Res 49:539–548PubMedGoogle Scholar
  16. 16.
    Fagan KA, Fouty BW, Tyler RC, Morris KG Jr, Hepler LK, Sato K, LeCras TD, Abman SH, Weinberger HD, Huang PL, McMurtry IF, Rodman DM (1999) The pulmonary circulation of homozygous or heterozygous eNOS-null mice is hyperresponsive to mild hypoxia. J Clin Invest 103:291–299PubMedCrossRefGoogle Scholar
  17. 17.
    Steudel W, Scherrer-Crosbie M, Bloch KD, Weimann J, Huang PL, Jones RC, Picard MH, Zapol WM (1998) Sustained pulmonary hypertension and right ventricular hypertrophy after chronic hypoxia in mice with congenital deficiency of nitric oxide synthase 3. J Clin Invest 101:2468–2477PubMedCrossRefGoogle Scholar
  18. 18.
    Chataigneau T, Feletou M, Huang PL, Fishman MC, Duhault J, Vanhoutte PM (1999) Acetylcholine-induced relaxation in blood vessels from endothelial nitric oxide synthase knockout mice. Br J Pharmacol 126:219–226PubMedCrossRefGoogle Scholar
  19. 19.
    Ding Z, Godecke A, Schrader J (2002) Contribution of cytochrome P450 metabolites to bradykinin-induced vasodilation in endothelial NO synthase deficient mouse hearts. Br J Pharmacol 135:631–638PubMedCrossRefGoogle Scholar
  20. 20.
    Capdevila JH, Wei S, Yan J, Karara A, Jacobson HR, Falck JR, Guengerich FP, DuBois RN (1992) Cytochrome P-450 arachidonic acid epoxygenase. Regulatory control of the renal epoxygenase by dietary salt loading. J Biol Chem 267:21720–21726PubMedGoogle Scholar
  21. 21.
    Makita K, Takahashi K, Karara A, Jacobson HR, Falck JR, Capdevila JH (1994) Experimental and/or genetically controlled alterations of the renal microsomal cytochrome P450 epoxygenase induce hypertension in rats fed a high salt diet. J Clin Invest 94:2414–2420PubMedCrossRefGoogle Scholar
  22. 22.
    Hao CM, Breyer MD (2007) Physiologic and pathophysiologic roles of lipid mediators in the kidney. Kidney Int 71:1105–1115PubMedCrossRefGoogle Scholar
  23. 23.
    Zhao X, Pollock DM, Inscho EW, Zeldin DC, Imig JD (2003) Decreased renal cytochrome P450 2C enzymes and impaired vasodilation are associated with angiotensin salt-sensitive hypertension. Hypertension 41:709–714PubMedCrossRefGoogle Scholar
  24. 24.
    Dyke CK, Proctor DN, Dietz NM, Joyner MJ (1995) Role of nitric oxide in exercise hyperaemia during prolonged rhythmic handgripping in humans. J Physiol 488(Pt 1):259–265PubMedGoogle Scholar
  25. 25.
    Duffy SJ, New G, Tran BT, Harper RW, Meredith IT (1999) Relative contribution of vasodilator prostanoids and NO to metabolic vasodilation in the human forearm. Am J Physiol 276:H663–H670PubMedGoogle Scholar
  26. 26.
    Hirai T, Visneski MD, Kearns KJ, Zelis R, Musch TI (1994) Effects of NO synthase inhibition on the muscular blood flow response to treadmill exercise in rats. J Appl Physiol 77:1288–1293PubMedGoogle Scholar
  27. 27.
    Shoemaker JK, Halliwill JR, Hughson RL, Joyner MJ (1997) Contributions of acetylcholine and nitric oxide to forearm blood flow at exercise onset and recovery. Am J Physiol 273:H2388–H2395PubMedGoogle Scholar
  28. 28.
    Radegran G, Saltin B (1999) Nitric oxide in the regulation of vasomotor tone in human skeletal muscle. Am J Physiol 276:H1951–H1960PubMedGoogle Scholar
  29. 29.
    Persson MG, Gustafsson LE, Wiklund NP, Hedqvist P, Moncada S (1990) Endogenous nitric oxide as a modulator of rabbit skeletal muscle microcirculation in vivo. Br J Pharmacol 100:463–466PubMedGoogle Scholar
  30. 30.
    Yamada M, Ishikawa T, Fujimori A, Goto K (1997) Local neurogenic regulation of rat hindlimb circulation: role of calcitonin gene-related peptide in vasodilatation after skeletal muscle contraction. Br J Pharmacol 122:703–709PubMedCrossRefGoogle Scholar
  31. 31.
    Ray CJ, Marshall JM (2009) Nitric oxide (NO) does not contribute to the generation or action of adenosine during exercise hyperaemia in rat hindlimb. J Physiol 587:1579–1591PubMedCrossRefGoogle Scholar
  32. 32.
    McGiff JC, Carroll MA (1991) Cytochrome P450-dependent arachidonate metabolites, renal function and blood pressure regulation. Adv Prostaglandin Thromboxane Leukot Res 21B:675–682PubMedGoogle Scholar
  33. 33.
    Bauersachs J, Hecker M, Busse R (1994) Display of the characteristics of endothelium-derived hyperpolarizing factor by a cytochrome P450-derived arachidonic acid metabolite in the coronary microcirculation. Br J Pharmacol 113:1548–1553PubMedGoogle Scholar
  34. 34.
    Campbell WB, Gebremedhin D, Pratt PF, Harder DR (1996) Identification of epoxyeicosatrienoic acids as endothelium-derived hyperpolarizing factors. Circ Res 78:415–423PubMedGoogle Scholar
  35. 35.
    Fisslthaler B, Popp R, Kiss L, Potente M, Harder DR, Fleming I, Busse R (1999) Cytochrome P450 2C is an EDHF synthase in coronary arteries. Nature 401:493–497PubMedCrossRefGoogle Scholar
  36. 36.
    Gebremedhin D, Harder DR, Pratt PF, Campbell WB (1998) Bioassay of an endothelium-derived hyperpolarizing factor from bovine coronary arteries: role of a cytochrome P450 metabolite. J Vasc Res 35:274–284PubMedCrossRefGoogle Scholar
  37. 37.
    Miura H, Gutterman DD (1998) Human coronary arteriolar dilation to arachidonic acid depends on cytochrome P-450 monooxygenase and Ca2+-activated K+ channels. Circ Res 83:501–507PubMedGoogle Scholar
  38. 38.
    Mombouli JV, Vanhoutte PM (1997) Endothelium-derived hyperpolarizing factor(s): updating the unknown. Trends Pharmacol Sci 18:252–256PubMedCrossRefGoogle Scholar
  39. 39.
    Oltman CL, Weintraub NL, VanRollins M, Dellsperger KC (1998) Epoxyeicosatrienoic acids and dihydroxyeicosatrienoic acids are potent vasodilators in the canine coronary microcirculation. Circ Res 83:932–939PubMedGoogle Scholar
  40. 40.
    Popp R, Bauersachs J, Hecker M, Fleming I, Busse R (1996) A transferable, beta-naphthoflavone-inducible, hyperpolarizing factor is synthesized by native and cultured porcine coronary endothelial cells. J Physiol 497(Pt 3):699–709PubMedGoogle Scholar
  41. 41.
    Quilley J, McGiff JC (2000) Is EDHF an epoxyeicosatrienoic acid? Trends Pharmacol Sci 21:121–124PubMedCrossRefGoogle Scholar
  42. 42.
    Fang X, Kaduce TL, Weintraub NL, Harmon S, Teesch LM, Morisseau C, Thompson DA, Hammock BD, Spector AA (2001) Pathways of epoxyeicosatrienoic acid metabolism in endothelial cells. Implications for the vascular effects of soluble epoxide hydrolase inhibition. J Biol Chem 276:14867–14874PubMedCrossRefGoogle Scholar
  43. 43.
    Bauersachs J, Popp R, Hecker M, Sauer E, Fleming I, Busse R (1996) Nitric oxide attenuates the release of endothelium-derived hyperpolarizing factor. Circulation 94:3341–3347PubMedGoogle Scholar
  44. 44.
    Nishikawa Y, Stepp DW, Chilian WM (2000) Nitric oxide exerts feedback inhibition on EDHF-induced coronary arteriolar dilation in vivo. Am J Physiol Heart Circ Physiol 279:H459–H465PubMedGoogle Scholar
  45. 45.
    Cheng MK, Doumad AB, Jiang H, Falck JR, McGiff JC, Carroll MA (2004) Epoxyeicosatrienoic acids mediate adenosine-induced vasodilation in rat preglomerular microvessels (PGMV) via A2A receptors. Br J Pharmacol 141:441–448PubMedCrossRefGoogle Scholar
  46. 46.
    Node K, Ruan XL, Dai J, Yang SX, Graham L, Zeldin DC, Liao JK (2001) Activation of Galpha s mediates induction of tissue-type plasminogen activator gene transcription by epoxyeicosatrienoic acids. J Biol Chem 276:15983–15989PubMedCrossRefGoogle Scholar
  47. 47.
    Popp R, Brandes RP, Ott G, Busse R, Fleming I (2002) Dynamic modulation of interendothelial gap junctional communication by 11,12-epoxyeicosatrienoic acid. Circ Res 90:800–806PubMedCrossRefGoogle Scholar
  48. 48.
    Li PL, Chen CL, Bortell R, Campbell WB (1999) 11,12-Epoxyeicosatrienoic acid stimulates endogenous mono-ADP-ribosylation in bovine coronary arterial smooth muscle. Circ Res 85:349–356PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC. 2010

Authors and Affiliations

  • Mohammed A. Nayeem
    • 1
    Email author
  • Darryl C. Zeldin
    • 2
  • Matthew A. Boegehold
    • 1
  • John R. Falck
    • 3
  1. 1.Department of Physiology and PharmacologyCenter for Cardiovascular and Respiratory Sciences, West Virginia UniversityMorgantownUSA
  2. 2.Division of Intramural ResearchNIEHS/NIHDurhamUSA
  3. 3.Department of BiochemistryUTSWMCDallasUSA

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