Abstract
Cannabinoid CB1 receptors peripherally modulate energy metabolism. Here, we investigated the role of CB1 receptors in the expression of glucose/pyruvate/tricarboxylic acid (TCA) metabolism in rat abdominal muscle. Dihydrolipoamide dehydrogenase (DLD), a flavoprotein component (E3) of α-ketoacid dehydrogenase complexes with diaphorase activity in mitochondria, was specifically analyzed. After assessing the effectiveness of the CB1 receptor antagonist AM251 (3 mg kg-1, 14 days) on food intake and body weight, we could identified seven key enzymes from either glycolytic pathway or TCA cycle—regulated by both diet and CB1 receptor activity—through comprehensive proteomic approaches involving two-dimensional electrophoresis and MALDI-TOF/LC-ESI trap mass spectrometry. These enzymes were glucose 6-phosphate isomerase (GPI), triosephosphate isomerase (TPI), enolase (Eno3), lactate dehydrogenase (LDHa), glyoxalase-1 (Glo1) and the mitochondrial DLD, whose expressions were modified by AM251 in hypercaloric diet-induced obesity. Specifically, AM251 blocked high-carbohydrate diet (HCD)-induced expression of GPI, TPI, Eno3 and LDHa, suggesting a down-regulation of glucose/pyruvate/lactate pathways under glucose availability. AM251 reversed the HCD-inhibited expression of Glo1 and DLD in the muscle, and the DLD and CB1 receptor expression in the mitochondrial fraction. Interestingly, we identified the presence of CB1 receptors at the membrane of striate muscle mitochondria. DLD over-expression was confirmed in muscle of CB 1 -/- mice. AM251 increased the pyruvate dehydrogenase and glutathione reductase activity in C2C12 myotubes, and the diaphorase/oxidative activity in the mitochondria fraction. These results indicated an up-regulation of methylglyoxal and TCA cycle activity. Findings suggest that CB1 receptors in muscle modulate glucose/pyruvate/lactate pathways and mitochondrial oxidative activity by targeting DLD.
Highlights
The endocannabinoid signaling system (ECS) plays a fundamental role in the onset of obesity and metabolic disorders, implicating central and peripheral mechanisms predominantly via the activation of the cannabinoid cannabinoid receptor type 1 (CB1) receptors [1,2,3,4]
AM251 administered at 3 and 10 mg kg-1 were the most effective doses on reducing cumulative food intake over 240 minutes of standard diet (STD) feeding in rats that had been previously food-deprived for 24 hours feeding (Fig 1A)
We demonstrated that the expression of relevant metabolic enzymes involved in the regulation of glucose (GPI, triosephosphate isomerase (TPI)), pyruvate (Eno3, pyruvate kinase isozyme M1 (PKM1), lactate dehydrogenase (LDHa)), glyoxalase-1 (Glo1) and tricarboxylic acid (TCA) and amino acid catabolism (DLD) pathways in the abdominal rat muscle was altered in a hypercaloric diet-dependent manner
Summary
The endocannabinoid signaling system (ECS) plays a fundamental role in the onset of obesity and metabolic disorders, implicating central and peripheral mechanisms predominantly via the activation of the cannabinoid CB1 receptors [1,2,3,4]. SR141716A, which revealed metabolic benefits and body weight reduction in overweight and obese human subjects [19,20], induced psychiatric side effects, such as depression and anxiety, likely derived from a central CB1 receptor inverse agonist activity and leading to the withdrawal of the drug from the market [21,22] These adverse effects highlighted the importance of limiting CB1 receptor antagonism to peripheral organs in order to reduce potential central risks and enhance peripheral energy balance [23]. Evaluated in conjunction with changes in the diaphorase/oxidative, pyruvate dehydrogenase and glutathione reductase activity in an in vitro model, using mytube-differentiated C2C12 cells and its mitochondria These results were interpreted regarding the subcellular localization of the CB1 receptors in the striate muscle and its potential role in the mitochondrial oxidative metabolism
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