Moreover, the ability of CB-13 to attenuate cardiomyocyte hypertrophy was abolished by disruption of AMPK signaling using a chemical inhibitor (compound C/dorsomorphin; Ki = 109 nM; 1 μM) or by shRNA knockdown of AMPKα1/2 when normalized to their respective controls, (Fig. 7). The selective eNOS inhibitor N5-(1-iminoethyl)-L-ornithine (L-NIO; IC50 = 500 nM; 10× and 5× less potent at neuronal nitric oxide synthase and iNOS, respectively; 1 μM)77 also ablated the antihypertrophic effects of CB-13 (Fig. 7). It bears mentioning that shRNA knockdown of AMPKα1/2 and L-NIO increased baseline cardiomyocyte size to 161% ± 20% (n = 3, P < 0.05) and 159% ± 17% (n = 5, P < 0.01) versus control, respectively, suggesting basal antigrowth activities of AMPK and eNOS in cardiomyocytes. Collectively, these findings indicate that CB-13 attenuates cardiomyocyte hypertrophy through AMPK-eNOS signaling.
To the best of our knowledge, this study shows for the first time that ligand activation of cannabinoid receptors attenuates hypertrophy of isolated cardiomyocytes through AMPK-eNOS signaling. Anandamide, a naturally occurring endocannabinoid,78 and 3 synthetic CB receptor ligands (R-methanandamide, JWH-133, and CB-13) prevented ET1-induced cardiomyocyte enlargement. As cardiac hypertrophy is a major risk factor for heart failure, we further investigated here the signaling mechanisms that underlie the antihypertrophic effects of CB receptors.
We first investigated the role of CB1 and CB2 receptors and found that distinct CB receptor subtypes mediate the antihypertrophic actions of R-methanandamide. In fact, selective antagonism of CB receptor subtypes uncoupled the inhibitory effects on ET1-induced myocyte growth from effects on hypertrophic gene expression. In particular, R-methanandamide prevents myocyte enlargement through CB2 receptors and was sensitive to AM630. In contrast, inhibition of fetal gene activation trafficked through CB1 receptors and was sensitive to AM251. We also considered the possibility that the ability of AM251 to block R-methanandamide effects might be due to agonism of GPR55 rather than antagonism of CB1.79 However, it is likely that AM251 was functioning here solely as a CB1 receptor antagonist. First, the concentration of AM251 that blocked R-methanandamide effects on BNP gene activation was 0.1 μM in accordance with its Ki at CB1 receptors (7.5 nM)80. In contrast, Kapur et al79 demonstrated GPR55-mediated effects of AM251 at 30 μM, with an EC50 of ∼10 μM, whereas at 0.1 μM, and even a logarithmic increment higher at 1.0 μM, AM251 failed to invoke GPR55 signaling. This information strongly suggests that the inhibitory effects of AM251 are attributable to antagonism of CB1 receptors. In support of this notion, we verified that AM281, a CB1 receptor antagonist lacking effects on GPR55, also abolished the ability of R-methanandamide to suppress BNP gene activation.
There exist literature precedents in which morphological changes are uncoupled from the hypertrophic gene program. For example, Thorburn et al81 reported that in isolated myocytes, ERK signaling mediates phenylephrine-dependent ANP promoter activation but not organization of contractile proteins such as actin. Furthermore, AP-1, and in particular c-Fos, is a key mediator of hypertrophic gene expression but not myocyte growth82. In vivo, activated GSK-3β can dissociate the expression of hypertrophic genes from cardiac growth overexpression; transgenic mice that overexpress calcineurin exhibit cardiac growth and BNP expression, whereas coexpression of activated GSK-3β inhibits cardiac growth but not BNP expression.83 Interestingly, there is evidence that endocannabinoids might interfere with c-Fos signaling84 through CB1 receptors85. We therefore speculate that activated CB1 receptors may attenuate BNP expression by suppressing ERK-AP-1 signaling, though this remains to be determined.
Selective agonism of CB2 receptors with JWH-133 suppressed myocyte enlargement but failed to prevent fetal gene activation. In contrast, we found that the antihypertrophic actions of a dual CB1/CB2 agonist, CB-13, extended beyond those of JWH-133, such that both myocyte enlargement and BNP gene expression were inhibited. These findings are in agreement with other reports of cardioprotective attributes that pertain predominantly to activation of CB2 receptors such as protection from ischemic insult,27–29,86 antiarrhythmic effects,87 and prevention of endothelial dysfunction of coronary arteries.7,88 However, although CB2 signaling may be cardioprotective, there is also evidence that activation of CB1 receptors is associated with dysfunction of vascular endothelium,89 exerts proatherosclerotic actions,90–92 and promotes oxidative stress, inflammation and/or cell death in vitro93 and in pathological conditions such as diabetic cardiomyopathy94 and doxorubicin-induced cardiomyopathy.95,96 Our data agree with the notion that sole activation of CB1 receptors is cardio-deleterious; in the context of cardiomyocyte hypertrophy, myocyte enlargement was suppressed by CB2 receptors, whereas only fetal gene activation was prevented by CB1 receptors. Thus, in the presence of sole activation of CB1 receptors and the absence of CB2 signaling, myocyte hypertrophy would persist and potentially give rise to adverse endpoints such as ischemia (vis-à-vis reduced capillary density), myocyte misalignment, and cardiac stiffening. Activation of CB2 receptors is therefore a necessary component of cannabinoid-based antihypertrophic therapy, which we speculate may ameliorate adverse effects of unopposed activation of CB1 receptors alone.
CB-13 stimulated phosphorylation of AMPKα (Thr172) and eNOS (Ser1177) at known activation sites (Figs. 6A, B),56,57,75,76 and we determined that eNOS phosphorylation is downstream of AMPK since it was inhibited by AMPKα knockdown (Figs. 6C, D). Inhibition of AMPK or eNOS signaling abolished the antihypertrophic effects of CB-13 (Fig. 7). Taken together, our findings suggest that signaling through AMPK-eNOS crosstalk might play a role in the antihypertrophic effects of CB receptors. This is consistent with reports by others, in which AMPK-eNOS crosstalk underlies the antigrowth effects of non-cannabinoid interventions such as metformin,97 calorie restriction,37 and resveratrol.98 Interestingly, phosphorylative activation of AMPK and eNOS peaked by 4 hours, and returned to baseline by 24 hours. This suggests that AMPK-eNOS crosstalk might occur at a pivotal, early point with downstream effects that will collectively prevent the hypertrophic response. Attenuation of prohypertrophic RhoA/RhoA kinase (ROCK) signaling might be one such downstream event. These members of the Rho family GTPases are well-established mediators of cardiac hypertrophy,99–101 and Hunter et al102 reported that NO inhibits ET1-induced cardiac myocyte hypertrophy by blocking the RhoA/ROCK cascade. Moreover, CB receptor activation inhibits RhoA signaling.103–105 We therefore speculate that CB-13, by transiently activating AMPK/eNOS crosstalk, elicits NO-dependent blockade of RhoA/ROCK, although the effects of CB-13 on RhoA/ROCK signaling remain to be determined.
The authors thank Lam Dang and Ping Lu for technical assistance.
1. Matsuda LA, Lolait SJ, Brownstein MJ, et al.. Structure of a cannabinoid receptor and functional expression of the cloned cDNA. Nature. 1990;346:561–564.
2. Bonz A, Laser M, Kullmer S, et al.. Cannabinoids acting on CB1 receptors decrease contractile performance in human atrial muscle. J Cardiovasc Pharmacol. 2003;41:657–664.
3. Gebremedhin D, Lange AR, Campbell WB, et al.. Cannabinoid CB1 receptor of cat cerebral arterial muscle functions to inhibit L-type Ca2+ channel current. Am J Physiol. 1999;276(6 pt 2):H2085–H2093.
4. Liu J, Gao B, Mirshahi F, et al.. Functional CB1 cannabinoid receptors in human vascular endothelial cells. Biochem J. 2000;346(pt 3):835–840.
5. Munro S, Thomas KL, Abu-Shaar M. Molecular characterization of a peripheral receptor for cannabinoids. Nature. 1993;365:61–65.
6. Valk PJ, Delwel R. The peripheral cannabinoid receptor, Cb2, in retrovirally-induced leukemic transformation and normal hematopoiesis. Leuk Lymphoma. 1998;32:29–43.
7. Bouchard JF, Lepicier P, Lamontagne D. Contribution of endocannabinoids in the endothelial protection afforded by ischemic preconditioning in the isolated rat heart. Life Sci. 2003;72:1859–1870.
8. Hillard CJ, Jarrahian A. The movement of N-arachidonoylethanolamine (anandamide
) across cellular membranes. Chem Phys Lipids. 2000;108:123–134.
9. Di Marzo V, Bisogno T, De Petrocellis L, et al.. Cannabimimetic fatty acid derivatives: the anandamide
family and other endocannabinoids. Curr Med Chem. 1999;6:721–744.
10. Beltramo M, Piomelli D. Carrier-mediated transport and enzymatic hydrolysis of the endogenous cannabinoid 2-arachidonylglycerol. Neuroreport. 2000;11:1231–1235.
11. Piomelli D, Beltramo M, Glasnapp S, et al.. Structural determinants for recognition and translocation by the anandamide
transporter. Proc Natl Acad Sci U S A. 1999;96:5802–5807.
12. Deutsch DG, Chin SA. Enzymatic synthesis and degradation of anandamide
, a cannabinoid receptor agonist. Biochem Pharmacol. 1993;46:791–796.
13. Maccarrone M, van der Stelt M, Rossi A, et al.. Anandamide
hydrolysis by human cells in culture and brain. J Biol Chem. 1998;273:32332–32339.
14. Dinh TP, Freund TF, Piomelli D. A role for monoglyceride lipase in 2-arachidonoylglycerol inactivation. Chem Phys Lipids. 2002;121:149–158.
15. Dinh TP, Carpenter D, Leslie FM, et al.. Brain monoglyceride lipase participating in endocannabinoid
inactivation. Proc Natl Acad Sci U S A. 2002;99:10819–10824.
16. Saario SM, Savinainen JR, Laitinen JT, et al.. Monoglyceride lipase-like enzymatic activity is responsible for hydrolysis of 2-arachidonoylglycerol in rat cerebellar membranes. Biochem Pharmacol. 2004;67:1381–1387.
17. Battista N, Fezza F, Finazzi-Agro A, et al.. The endocannabinoid
system in neurodegeneration. Ital J Biochem. 2006;55:283–289.
18. Pacher P, Batkai S, Kunos G. The endocannabinoid
system as an emerging target of pharmacotherapy. Pharmacol Rev. 2006;58:389–462.
19. Parker P, Patterson J, Johnson J. Pharmacotherapy. A Pathophysiologic Approach: Heart Failure. 6th ed. The McGraw-Hill Companies, Inc; 2005.
20. Levy D, Garrison RJ, Savage DD, et al.. Prognostic implications of echocardiographically determined left ventricular mass in the Framingham Heart Study. N Engl J Med. 1990;322:1561–1566.
21. Ho KK, Pinsky JL, Kannel WB, et al.. The epidemiology of heart failure: the Framingham Study. J Am Coll Cardiol. 1993;22(4 suppl A):6A–13A.
22. Berenji K, Drazner MH, Rothermel BA, et al.. Does load-induced ventricular hypertrophy progress to systolic heart failure? Am J Physiol Heart Circ Physiol. 2005;289:H8–H16.
23. Frey N, Katus HA, Olson EN, et al.. Hypertrophy of the heart: a new therapeutic target? Circulation. 2004;109:1580–1589.
24. Felder CC, Nielsen A, Briley EM, et al.. Isolation and measurement of the endogenous cannabinoid receptor agonist, anandamide
, in brain and peripheral tissues of human and rat. FEBS Lett. 1996;393:231–235.
25. Galiegue S, Mary S, Marchand J, et al.. Expression of central and peripheral cannabinoid receptors in human immune tissues and leukocyte subpopulations. Eur J Biochem. 1995;232:54–61.
26. Bilfinger TV, Salzet M, Fimiani C, et al.. Pharmacological evidence for anandamide
amidase in human cardiac and vascular tissues. Int J Cardiol. 1998;64(suppl 1):S15–S22.
27. Lagneux C, Lamontagne D. Involvement of cannabinoids in the cardioprotection induced by lipopolysaccharide. Br J Pharmacol. 2001;132:793–796.
28. Underdown NJ, Hiley CR, Ford WR. Anandamide
reduces infarct size in rat isolated hearts subjected to ischaemia-reperfusion by a novel cannabinoid mechanism. Br J Pharmacol. 2005;146:809–816.
29. Lepicier P, Bouchard JF, Lagneux C, et al.. Endocannabinoids protect the rat isolated heart against ischaemia. Br J Pharmacol. 2003;139:805–815.
30. Dolinsky VW, Dyck JR. Role of AMP-activated protein kinase in healthy and diseased hearts. Am J Physiol. 2006;291:H2557–H2569.
31. Chan AY, Soltys CL, Young ME, et al.. Activation of AMP-activated protein kinase inhibits protein synthesis associated with hypertrophy in the cardiac myocyte
. J Biol Chem. 2004;279:32771–32779.
32. Chen BL, Ma YD, Meng RS, et al.. Activation of AMPK
inhibits cardiomyocyte hypertrophy by modulating of the FOXO1/MuRF1 signaling pathway in vitro. Acta Pharmacol Sin. 2010;31:798–804.
33. Stuck BJ, Lenski M, Bohm M, et al.. Metabolic switch and hypertrophy of cardiomyocytes following treatment with angiotensin II are prevented by AMP-activated protein kinase. J Biol Chem. 2008;283:32562–32569.
34. Li HL, Yin R, Chen D, et al.. Long-term activation of adenosine monophosphate-activated protein kinase attenuates pressure-overload-induced cardiac hypertrophy
. J Cell Biochem. 2007;100:1086–1099.
35. Zhang P, Hu X, Xu X, et al.. AMP activated protein kinase-alpha2 deficiency exacerbates pressure-overload-induced left ventricular hypertrophy and dysfunction in mice. Hypertension. 2008;52:918–924.
36. Chan AY, Dolinsky VW, Soltys CL, et al.. Resveratrol inhibits cardiac hypertrophy
via AMP-activated protein kinase and Akt. J Biol Chem. 2008;283:24194–24201.
37. Dolinsky VW, Morton JS, Oka T, et al.. Calorie restriction prevents hypertension and cardiac hypertrophy
in the spontaneously hypertensive rat. Hypertension. 2010;56:412–421.
38. Kola B, Hubina E, Tucci SA, et al.. Cannabinoids and ghrelin have both central and peripheral metabolic and cardiac effects via AMP-activated protein kinase. J Biol Chem. 2005;280:25196–25201.
39. Lim CT, Kola B, Feltrin D, et al.. Ghrelin and cannabinoids require the ghrelin receptor to affect cellular energy metabolism. Mol Cell Endocrinol. 2013;365:303–308.
40. Dagon Y, Avraham Y, Ilan Y, et al.. Cannabinoids ameliorate cerebral dysfunction following liver failure via AMP-activated protein kinase. FASEB J. 2007;21:2431–2441.
41. Ortega-Gutierrez S, Molina-Holgado E, Guaza C. Effect of anandamide
uptake inhibition in the production of nitric oxide and in the release of cytokines in astrocyte cultures. Glia. 2005;52:163–168.
42. Vannacci A, Giannini L, Passani MB, et al.. The endocannabinoid
2-arachidonylglycerol decreases the immunological activation of Guinea pig mast cells: involvement of nitric oxide and eicosanoids. J Pharmacol Exp Ther. 2004;311:256–264.
43. Gasperi V, Fezza F, Pasquariello N, et al.. Endocannabinoids in adipocytes during differentiation and their role in glucose uptake. Cell Mol Life Sci. 2007;64:219–229.
44. Harris D, McCulloch AI, Kendall DA, et al.. Characterization of vasorelaxant responses to anandamide
in the rat mesenteric arterial bed. J Physiol. 2002;539(pt 3):893–902.
45. Lepicier P, Bibeau-Poirier A, Lagneux C, et al.. Signaling pathways involved in the cardioprotective effects of cannabinoids. J Pharmacol Sci. 2006;102:155–166.
46. Balligand JL, Cannon PJ. Nitric oxide synthases and cardiac muscle. Autocrine and paracrine influences. Arterioscler Thromb Vasc Biol. 1997;17:1846–1858.
47. Wollert KC, Drexler H. Regulation of cardiac remodeling by nitric oxide: focus on cardiac myocyte
hypertrophy and apoptosis. Heart Fail Rev. 2002;7:317–325.
48. Ziolo MT, Bers DM. The real estate of NOS signaling: location, location, location. Circ Res. 2003;92:1279–1281.
49. Barouch LA, Harrison RW, Skaf MW, et al.. Nitric oxide regulates the heart by spatial confinement of nitric oxide synthase isoforms. Nature. 2002;416:337–339.
50. Barouch LA, Cappola TP, Harrison RW, et al.. Combined loss of neuronal and endothelial nitric oxide synthase causes premature mortality and age-related hypertrophic cardiac remodeling in mice. J Mol Cell Cardiol. 2003;35:637–644.
51. Khan SA, Skaf MW, Harrison RW, et al.. Nitric oxide regulation of myocardial contractility and calcium cycling: independent impact of neuronal and endothelial nitric oxide synthases. Circ Res. 2003;92:1322–1329.
52. Arstall MA, Sawyer DB, Fukazawa R, et al.. Cytokine-mediated apoptosis in cardiac myocytes: the role of inducible nitric oxide synthase induction and peroxynitrite generation. Circ Res. 1999;85:829–840.
53. Mungrue IN, Gros R, You X, et al.. Cardiomyocyte overexpression of iNOS in mice results in peroxynitrite generation, heart block, and sudden death. J Clin Invest. 2002;109:735–743.
54. Sam F, Sawyer DB, Xie Z, et al.. Mice lacking inducible nitric oxide synthase have improved left ventricular contractile function and reduced apoptotic cell death late after myocardial infarction. Circ Res. 2001;89:351–356.
55. Feng Q, Lu X, Jones DL, et al.. Increased inducible nitric oxide synthase expression contributes to myocardial dysfunction and higher mortality after myocardial infarction in mice. Circulation. 2001;104:700–704.
56. Chen ZP, Mitchelhill KI, Michell BJ, et al.. AMP-activated protein kinase phosphorylation of endothelial NO synthase. FEBS Lett. 1999;443:285–289.
57. Morrow VA, Foufelle F, Connell JM, et al.. Direct activation of AMP-activated protein kinase stimulates nitric-oxide synthesis in human aortic endothelial cells. J Biol Chem. 2003;278:31629–31639.
58. Wu J, LaPointe MC, West BL, et al.. Tissue-specific determinants of human atrial natriuretic factor gene expression in cardiac tissue. J Biol Chem. 1989;264:6472–6479.
59. Huang Y, Zhang H, Shao Z, et al.. Suppression of endothelin-1-induced cardiac myocyte
hypertrophy by PPAR agonists: role of diacylglycerol kinase zeta. Cardiovasc Res. 2011;90:267–275.
60. Romano MR, Lograno MD. Cannabinoid agonists induce relaxation in the bovine ophthalmic artery: evidences for CB1 receptors, nitric oxide and potassium channels. Br J Pharmacol. 2006;147:917–925.
61. Lam FF, Luk PW, Ng ES. Pharmacological characterization of receptor types mediating the dilator action of anandamide
on blood vessels of the rat knee joint. Life Sci. 2007;80:1495–1502.
62. Alibin CP, Kopilas MA, Anderson HD. Suppression of cardiac myocyte
hypertrophy by conjugated linoleic acid: role of peroxisome proliferator-activated receptors alpha and gamma. J Biol Chem. 2008;283:10707–10715.
63. Chien KR, Knowlton KU, Zhu H, et al.. Regulation of cardiac gene expression during myocardial growth and hypertrophy: molecular studies of an adaptive physiologic response. FASEB J. 1991;5:3037–3046.
64. LaPointe MC. Molecular regulation of the brain natriuretic peptide gene. Peptides. 2005;26:944–956.
65. Abadji V, Lin S, Taha G, et al.. (R)-methanandamide: a chiral novel anandamide
possessing higher potency and metabolic stability. J Med Chem. 1994;37:1889–1893.
66. Lan R, Gatley J, Lu Q, et al.. Design and synthesis of the CB1 selective cannabinoid antagonist AM281: a potential human SPECT ligand. AAPS PharmSci. 1999;1:E4.
67. Ross RA, Brockie HC, Stevenson LA, et al.. Agonist-inverse agonist characterization at CB1 and CB2 cannabinoid receptors of L759633, L759656, and AM630. Br J Pharmacol. 1999;126:665–672.
68. Hosking RD, Zajicek JP. Therapeutic potential of cannabis in pain medicine. Br J Anaesth. 2008;101:59–68.
69. Kunos G, Osei-Hyiaman D, Batkai S, et al.. Should peripheral CB(1) cannabinoid receptors be selectively targeted for therapeutic gain? Trends Pharmacol Sci. 2009;30:1–7.
70. Palazuelos J, Aguado T, Egia A, et al.. Non-psychoactive CB2 cannabinoid agonists stimulate neural progenitor proliferation. FASEB J. 2006;20:2405–2407.
71. Gertsch J, Leonti M, Raduner S, et al.. Beta-caryophyllene is a dietary cannabinoid. Proc Natl Acad Sci U S A. 2008;105:9099–9104.
72. Huffman JW, Liddle J, Yu S, et al.. 3-(1′,1′-Dimethylbutyl)-1-deoxy-delta8-THC and related compounds: synthesis of selective ligands for the CB2 receptor. Bioorg Med Chem. 1999;7:2905–2914.
73. Dziadulewicz EK, Bevan SJ, Brain CT, et al.. Naphthalen-1-yl-(4-pentyloxynaphthalen-1-yl)methanone: a potent, orally bioavailable human CB1/CB2 dual agonist with antihyperalgesic properties and restricted central nervous system penetration. J Med Chem. 2007;50:3851–3856.
74. Cluny NL, Keenan CM, Duncan M, et al.. Naphthalen-1-yl-(4-pentyloxynaphthalen-1-yl)methanone (SAB378), a peripherally restricted cannabinoid CB1/CB2 receptor agonist, inhibits gastrointestinal motility but has no effect on experimental colitis in mice. J Pharmacol Exp Ther. 2010;334:973–980.
75. Witters LA, Kemp BE, Means AR. Chutes and ladders: the search for protein kinases that act on AMPK
. Trends Biochem Sci. 2006;31:13–16.
76. Beauloye C, Bertrand L, Horman S, et al.. AMPK
activation, a preventive therapeutic target in the transition from cardiac injury to heart failure. Cardiovasc Res. 2011;90:224–233.
77. Moore WM, Webber RK, Jerome GM, et al.. L-N6-(1-iminoethyl)lysine: a selective inhibitor of inducible nitric oxide synthase. J Med Chem. 1994;37:3886–3888.
78. Devane WA, Hanus L, Breuer A, et al.. Isolation and structure of a brain constituent that binds to the cannabinoid receptor. Science. 1992;258:1946–1949.
79. Kapur A, Zhao P, Sharir H, et al.. Atypical responsiveness of the orphan receptor GPR55 to cannabinoid ligands. J Biol Chem. 2009;284:29817–29827.
80. Lan R, Liu Q, Fan P, et al.. Structure-activity relationships of pyrazole derivatives as cannabinoid receptor antagonists. J Med Chem. 1999;42:769–776.
81. Thorburn J, Frost JA, Thorburn A. Mitogen-activated protein kinases mediate changes in gene expression, but not cytoskeletal organization associated with cardiac muscle cell hypertrophy. J Cell Biol. 1994;126:1565–1572.
82. Jeong MY, Kinugawa K, Vinson C, et al.. AFos dissociates cardiac myocyte
hypertrophy and expression of the pathological gene program. Circulation. 2005;111:1645–1651.
83. Antos CL, McKinsey TA, Frey N, et al.. Activated glycogen synthase-3 beta suppresses cardiac hypertrophy
in vivo. Proc Natl Acad Sci U S A. 2002;99:907–912.
84. Greco R, Gasperi V, Sandrini G, et al.. Alterations of the endocannabinoid
system in an animal model of migraine: evaluation in cerebral areas of rat. Cephalalgia. 2010;30:296–302.
85. Soderstrom K, Tian Q. CB(1) cannabinoid receptor activation dose dependently modulates neuronal activity within caudal but not rostral song control regions of adult zebra finch telencephalon. Psychopharmacology (Berl). 2008;199:265–273.
86. Di Filippo C, Rossi F, Rossi S, et al.. Cannabinoid CB2 receptor activation reduces mouse myocardial ischemia-reperfusion injury: involvement of cytokine/chemokines and PMN. J Leukoc Biol. 2004;75:453–459.
87. Krylatov AV, Ugdyzhekova DS, Bernatskaya NA, et al.. Activation of type II cannabinoid receptors improves myocardial tolerance to arrhythmogenic effects of coronary occlusion and reperfusion. Bull Exp Biol Med. 2001;131:523–525.
88. Wagner JA, Hu K, Karcher J, et al.. CB(1) cannabinoid receptor antagonism promotes remodeling and cannabinoid treatment prevents endothelial dysfunction and hypotension in rats with myocardial infarction. Br J Pharmacol. 2003;138:1251–1258.
89. Tiyerili V, Zimmer S, Jung S, et al.. CB1 receptor inhibition leads to decreased vascular AT1 receptor expression, inhibition of oxidative stress and improved endothelial function. Basic Res Cardiol. 2010;105:465–477.
90. Molica F, Burger F, Thomas A, et al.. Endogenous cannabinoid receptor CB1 activation promotes vascular smooth-muscle cell proliferation and neointima formation. J Lipid Res. 2013;54:1360–1368.
91. Sugamura K, Sugiyama S, Nozaki T, et al.. Activated endocannabinoid
system in coronary artery disease and antiinflammatory effects of cannabinoid 1 receptor blockade on macrophages. Circulation. 2009;119:28–36.
92. Dol-Gleizes F, Paumelle R, Visentin V, et al.. Rimonabant, a selective cannabinoid CB1 receptor antagonist, inhibits atherosclerosis in LDL receptor-deficient mice. Arterioscler Thromb Vasc Biol. 2009;29:12–18.
93. Rajesh M, Mukhopadhyay P, Hasko G, et al.. Cannabinoid-1 receptor activation induces reactive oxygen species-dependent and -independent mitogen-activated protein kinase activation and cell death in human coronary artery endothelial cells. Br J Pharmacol. 2010;160:688–700.
94. Rajesh M, Batkai S, Kechrid M, et al.. Cannabinoid 1 receptor promotes cardiac dysfunction, oxidative stress, inflammation, and fibrosis in diabetic cardiomyopathy. Diabetes. 2012;61:716–727.
95. Mukhopadhyay P, Batkai S, Rajesh M, et al.. Pharmacological inhibition of CB1 cannabinoid receptor protects against doxorubicin-induced cardiotoxicity. J Am Coll Cardiol. 2007;50:528–536.
96. Mukhopadhyay P, Rajesh M, Batkai S, et al.. CB1 cannabinoid receptors promote oxidative stress and cell death in murine models of doxorubicin-induced cardiomyopathy and in human cardiomyocytes. Cardiovasc Res. 2010;85:773–784.
97. Zhang CX, Pan SN, Meng RS, et al.. Metformin attenuates ventricular hypertrophy by activating the AMP-activated protein kinase-endothelial nitric oxide synthase pathway in rats. Clin and Exp Pharmacol Physiol. 2011;38:55–62.
98. Thandapilly SJ, Louis XL, Yang T, et al.. Resveratrol prevents norepinephrine induced hypertrophy in adult rat cardiomyocytes, by activating NO-AMPK
pathway. Eur J Pharmacol. 2011;668:217–224.
99. Loirand G, Guerin P, Pacaud P. Rho kinases in cardiovascular physiology and pathophysiology. Circ Res. 2006;98:322–334.
100. Zeidan A, Gan XT, Thomas A, et al.. Prevention of RhoA activation and cofilin-mediated actin polymerization mediates the antihypertrophic effect of adenosine receptor agonists in angiotensin II- and endothelin-1-treated cardiomyocytes. Mol Cell Biochem. 2014;385:239–248.
101. Brown JH, Del Re DP, Sussman MA. The Rac and Rho hall of fame: a decade of hypertrophic signaling hits. Circ Res. 2006;98:730–742.
102. Hunter JC, Zeidan A, Javadov S, et al.. Nitric oxide inhibits endothelin-1-induced neonatal cardiomyocyte hypertrophy via a RhoA-ROCK-dependent pathway. J Mol Cell Cardiol. 2009;47:810–818.
103. Kurihara R, Tohyama Y, Matsusaka S, et al.. Effects of peripheral cannabinoid receptor ligands on motility and polarization in neutrophil-like HL60 cells and human neutrophils. J Biol Chem. 2006;281:12908–12918.
104. Rajesh M, Mukhopadhyay P, Batkai S, et al.. CB2-receptor stimulation attenuates TNF-alpha-induced human endothelial cell activation, transendothelial migration of monocytes, and monocyte-endothelial adhesion. Am J Physiol. 2007;293:H2210–H2218.
105. Nithipatikom K, Gomez-Granados AD, Tang AT, et al.. Cannabinoid receptor type 1 (CB1) activation inhibits small GTPase RhoA activity and regulates motility of prostate carcinoma cells. Endocrinology. 2012;153:29–41.
106. Reisner Y, Meiry G, Zeevi-Levin N, et al.. Impulse conduction and gap junctional remodelling by endothelin-1 in cultured neonatal rat ventricular myocytes. J Cell Mol Med. 2009;13:562–573.
107. Yu L, Li M, She T, et al.. Endothelin-1 stimulates the expression of L-type Ca2+ channels in neonatal rat cardiomyocytes via the extracellular signal-regulated kinase 1/2 pathway. J Membr Biol. 2013;246:343–353.
108. Piomelli D. The molecular logic of endocannabinoid
signalling. Nat Rev Neurosci. 2003;4:873–884.
109. Howlett AC, Barth F, Bonner TI, et al.. International Union of Pharmacology. XXVII. Classification of cannabinoid receptors. Pharmacol Rev. 2002;54:161–202.