Abstract 081: Liganded Cannabinoid Receptors Suppress Cardiac Myocyte Hypertrophy by AMP-activated protein kinase (AMPK) and endothelial nitric oxide synthase (eNOS) signaling

Abstract

Endocannabinoids are bioactive lipids that signal through CB1 and CB2 cannabinoid receptors. Despite an effort to avoid the psychoactive side effects mediated by central CB1 receptors, we previously found that selective activation of CB2 receptors is not sufficient to prevent cardiac myocyte hypertrophy. Thus, the objective of this study is to determine the effects of CB13, a peripherally-restricted agonist of CB1/CB2 receptors, on hypertrophy. The effects of CB-13 on endothelin-1 (ET1)-induced hypertrophy were determined using isolated neonatal rat myocytes. Hypertrophic indicators included myocyte enlargement, protein synthesis by [3H]-leucine incorporation, and fetal gene expression as brain natriuretic peptide (BNP) mRNA levels. AMPK and eNOS signaling were assessed by immunoblotting. ET1 increased myocyte size (122±4% vs. control; p<0.01), BNP expression (426±88% vs. control; p<0.01) and [3H]-leucine incorporation (147±13% vs. control; p<0.05). CB13 attenuated all three hypertrophic indicators (respectively, 104±3%, 210±32%, 90±8% vs. control; not significant). We next queried whether the anti-hypertrophic actions of CB13 were mediated by AMPK. CB13 increased total expression (222±45% vs. control; p<0.05) and phosphorylation (354±58% vs. control; p<0.01) of AMPK. Also, pretreatment with a chemical inhibitor of AMPK, compound C, attenuated the anti-hypertrophic actions of CB-13. AMPK is a regulator of cellular energy, so we determined that ET-1-induced mitochondrial depolarization, as assessed using the potential-sensitive dye, JC-1, was prevented by CB-13. In addition, CB-13 increased eNOS phosphorylation (237±51%; p<0.01), which suggests that AMPK-eNOS crosstalk, through anti-growth nitric oxide signaling, plays a role. These findings support the notion that agonism of CB1 and CB2 receptors by CB13, a synthetic endocannabinoid with poor brain-blood barrier penetration, prevents hypertrophy and mitochondrial dysfunction. These CB13 actions rely on AMPK signaling, and are associated with eNOS activation. In conclusion, cannabinoid-based treatment of heart disease remains a viable goal with therapeutic potential and warrants further study.