Role of adipose mitochondrial CB1 signaling in the regulation of energy balance – MitoCB1-Fat

We first pursued our preliminary studies to characterize the effects of CB1 receptor activation on mitochondrial activity from adipose tissue.
We also conducted anatomical studies in adpisoe tissue from mice to demonstrate the presence of the CB1 receptor within mitochondrial membranes. In fact we have set up a collaboration with the laboratory of Professor Antonio Giordano, from the University of Ancona (Italy).

We first continued our preliminary studies to characterize the effects of CB1 receptor activation on mitochondrial activity from adipose tissue. We have shown that the activation of this receptor is able to inhibit the activity of complex I of the respiratory chain and the consumption of oxygen coupled to the synthesis of ATP. Moreover, we showed that these processes are dependent on the mitochondrial localization of the receptor, as it is absent in mice deficient for the receptor, or whose receptor has been mutated to remain exclusively at the plasma membrane. We have also characterized the signaling pathway involved in this inhibition: in fact we have demonstrated that the activation of the CB1 receptor in the mitochondrial membrane activates the inhibitory G proteins present in these organelles, triggering an inhibition of the cyclic AMP and protein kinase A pathway, resulting in a minor activation of the complex I through phosphorylation.
We also conducted anatomical studies to demonstrate the presence of the CB1 receptor within mitochondrial membranes. In fact, we have set up a collaboration with the laboratory of Professor Antonio Giordano, from the University of Ancona (Italy). Thanks to this collaboration, we have developed an electron microscopy technique coupled with immunological markings. Using this technique, a specific signal for the CB1 receptor on mitochondria was shown, which is absent in mice deficient for the receptor, or whose receptor has been mutated to remain exclusively at the plasma membrane.

Due to the SARS-CoV 2 pandemic, the generation of transgenic animals has been halted, which has had a strong impact on the progress of the in vivo experiments included in this project. These experiments have been restarted in the year 2021 and are currently being processed and analyzed.

The role of mtCB1 in the regulation of several cellular processes in adipocytes, mainly related to mitochondrial activity, has been demonstrated. These data are part of a publication in the process of submission to the prestigious journal Molecular Metabolism.
We also characterized a new line of mutant mice in which the expression of the CB1 receptor is exclusively restricted to the plasma membrane, a tool that will allow us to study in depth the impact of mtCB1 activation in vivo. The characterization of this new mouse line is part of a publication in the prestigious journal Neuron.
(https://pubmed.ncbi.nlm.nih.gov/33770505/)

The endocannabinoid system (ECS) has recently emerged as an important modulator of food intake and energy balance. Cannabinoid type-1 (CB1) receptor and its main endogenous lipid ligands, 2-arachidonoyl-glycerol (2-AG) and anandamide (AEA), are largely present in the brain and in peripheral organs involved in the regulation of energy metabolism, such as liver, adipose tissue, skeletal muscle, pancreas and GI tract. Furthermore, there is evidence of correlations between pathological ECS up-regulation and metabolic diseases in both animals models and humans studies. However, several pieces of evidence indicate that the relationship between the ECS and energy intake and metabolism is more complex than previously believed, likely due to the different sites where the ECS could act in the body. For instance, neuronal cannabinoid type-1 (CB1)R signalling has been shown as a key determinant of the ECS action on energy balance. However, due to important side effects, targeting this receptor is deleterious in the treatment of obesity. At peripheral level, CB1 receptor in adipose tissue seems to be the new target for ECS regulation of energy metabolism. Indeed, conditional deletion of the CB1 receptor gene specifically in adipocytes is sufficient to protect adult mice from diet induced obesity and associated metabolic alterations by increasing energy expenditure and mitochondrial functions in adipose tissue, and to reverse the phenotype in already obese mice. Furthermore, mitochondrial activity and its products, such as reactive oxygen species (ROS), play a crucial role in the homeostasis of adipose tissue by directly regulating adipo-lipogenesis, lipolysis fatty acids and ketone body’s metabolism. Moreover, as already shown for brain and skeletal muscle, functional CB1 receptor is present in mitochondria of adipocytes, where it negatively regulates mitochondrial activity. Thus, we hypothesize that mtCB1 activation in adipose tissue, by interfering with oxidative phosphorylation, is able to induce metabolic alteration in this organ which may affect the whole body metabolism. This project will focus on the role of mitochondrial activity and GPCRs signalling in adipose tissue, with particular emphasis on mtCB1 receptor, in the context of physiological and pathological regulation of energy balance. To this aim, we will characterize the impact of mtCB1 signalling on adipocytes-related processes in vitro, as well as its role on the regulation of energy balance in normal and diet-induced obesity conditions in vivo. Since peripheral CB1R pools are key therapeutical targets for obesity and metabolic diseases, studying adipose mtCB1 signalling may provide the base for developing efficient and safe strategies for the treatment of obesity.