We recently reported that activation of endocannabinoid receptors attenuates cardiac myocyte hypertrophy. Mitochondrial dysfunction has emerged as a critical determinant of aberrant myocyte energy production in cardiac hypertrophy. Thus, we determined endocannabinoid influence on mitochondrial function in the hypertrophied cardiac myocyte.
DESIGN AND METHOD:
The experimental paradigm of hypertrophy in this study was neonatal rat cardiac myocytes treated with endothelin-1 (ET1; 0.1 μM). Ligand activation of cannabinoid receptors was achieved using CB13 (1 μM), a peripherally-restricted dual agonist of cannabinoid receptor subtypes CB1 and CB2. Changes in mitochondrial membrane potential (δψm) were assessed by fluorescence microscopy using the potential sensitive dye, JC-1. Biochemical modulators of mitochondrial function (i.e. peroxisome proliferator-activated receptor-γ coactivator 1α [PGC-1α – a driver of mitochondrial biogenesis], carnitine palmitoyl transferase 1β [CPT-1β – facilitator of fatty acid uptake], and AMP-activated protein kinase [AMPK – energy sensor]) were assessed by real-time PCR and western blotting. The Seahorse Bioscience XF24 Analyzer was used to measure fatty acid oxidation-related bioenergetics parameters.
ET1 caused mitochondrial aberrations which included membrane depolarization (δψm 80 ± 5% vs. control; p < 0.05), reduced PGC-1α (59 ± 7% vs. control; p < 0.01) and CPT-1β (81 ± 5% vs. control; p < 0.05) expression, as well as depressed palmitate-dependent respiration (basal/maximal/reserved respiration respectively: 81 ± 5%, 78 ± 4%, 74 ± 5% vs. control; p < 0.05), coupling efficiency (83 ± 6% vs. control; p < 0.05), and respiratory control ratio (79 ± 5% vs. control; p < 0.01). CB13 treatment restored all mitochondrial parameters to normal. Incidentally, CB13 activated AMPK via phosphorylation at Thr172 (354 ± 58% vs. control; p < 0.01), and the ability of CB13 to improve mitochondrial membrane potential and PGC-1α was abolished by compound C (a chemical inhibitor of AMPK) or shRNA knockdown of AMPK. These data suggest that AMPK contributes to the mitochondrial protective effects of CB13.
The cardioprotective actions of liganded cannabinoid receptors extend to the mitochondrial level. Therefore, a cannabinoid-based treatment for cardiac disease remains a potential therapeutic strategy that warrants further study.
- [PubMed – as supplied by publisher]