Evidence for the intracellular accumulation of anandamide in adiposomes

Anandamide is a lipid messenger that carries out a wide variety of biological functions. It has been suggested that anandamide accumulation involves binding to a saturable cellular component. To identify the structure(s) involved in this process, we analyzed the intracellular distribution of both biotinylated and radiolabeled anandamide, providing direct evidence that lipid droplets, also known as adiposomes, constitute a dynamic reservoir for the sequestration of anandamide. In addition, confocal microscopy and biochemical studies revealed that the anandamide-hydrolase is also spatially associated with lipid droplets, and that cells with a larger adiposome compartment have an enhanced catabolism of anandamide. Overall, these findings suggest that adiposomes may have a critical role in accumulating anandamide, possibly by connecting plasma membrane to internal organelles along the metabolic route of this endocannabinoid. S. Oddi, F. Fezza: These authors contributed equally to the study.     

Activation of type-2 cannabinoid receptor inhibits neuroprotective and antiinflammatory actions of glucocorticoid receptor α: when one is better than two

Endocannabinoids (eCBs) and glucocorticoids (GCs) are two distinct classes of signaling lipids that exert both neuroprotective and immunosuppressive effects; however, the possibility of an actual interaction of their receptors [i.e., type-2 cannabinoid (CB₂) and glucocorticoid receptor α (GRα), respectively] remains unexplored. Here, we demonstrate that the concomitant activation of CB₂ and GRα abolishes the neuroprotective effects induced by each receptor on central neurons and on glial cells in animal models of remote cell death. We also show that the ability of eCBs and GCs, used individually, to inhibit tumour necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) production from activated human T lymphocytes is lost when CB₂ and GRα are activated simultaneously. In addition, signal transduction pathways triggered by concomitant activation of both receptors led to increased levels of GRβ, heat-shock proteins-70 and -90, and p-JNK, as well as to reduced levels of p-STAT6. These effects were reversed only by selectively antagonizing CB₂, but not GRα. Overall, our study demonstrates for the first time the existence of a CB₂-driven negative cross-talk between eCB and GC signaling in both rats and humans, thus paving the way to the possible therapeutic exploitation of CB₂ as a new target for chronic inflammatory and neurodegenerative diseases.     

Endocannabinoids in adipocytes during differentiation and their role in glucose uptake

The molecular basis for the control of energy balance by the endocannabinoid anandamide (AEA) is still unclear. Here, we show that murine 3T3-L1 fibroblasts have the machinery to bind, synthesize and degrade AEA, and that their differentiation into adipocytes increases by approximately twofold the binding efficiency of cannabinoid receptors (CBR), and by approximately twofold and approximately threefold, respectively, the catalytic efficiency of the AEA transporter and AEA hydrolase. In contrast, the activity of the AEA synthetase and the binding efficiency of vanilloid receptor were not affected by the differentiation process. In addition, we demonstrate that AEA increases by approximately twofold insulin-stimulated glucose uptake in differentiated adipocytes, according to a CB1R-dependent mechanism that involves nitric oxide synthase, but not lipoxygenase or cyclooxygenase. We also show that AEA binding to peroxisome proliferator-activated receptor-γ, known to induce differentiation of 3T3-L1 fibroblasts into adipocytes, is not involved in the stimulation of glucose uptake. Received 11 October 2006; received after revision 9 November 2006; accepted 28 November 2006 V.Gasperi and F. Fezza equally contributed to the study.     

Leptin-regulated endocannabinoids are involved in maintaining food intake

Leptin is the primary signal through which the hypothalamus senses nutritional state and modulates food intake and energy balance. Leptin reduces food intake by upregulating anorexigenic (appetite-reducing) neuropeptides, such as alpha-melanocyte-stimulating hormone, and downregulating orexigenic (appetite-stimulating) factors, primarily neuropeptide Y. Genetic defects in anorexigenic signalling, such as mutations in the melanocortin-4 (ref. 5) or leptin receptors, cause obesity. However, alternative orexigenic pathways maintain food intake in mice deficient in neuropeptide Y. CB1 cannabinoid receptors and the endocannabinoids anandamide and 2-arachidonoyl glycerol are present in the hypothalamus, and marijuana and anandamide stimulate food intake. Here we show that following temporary food restriction, CB1 receptor knockout mice eat less than their wild-type littermates, and the CB1 antagonist SR141716A reduces food intake in wild-type but not knockout mice. Furthermore, defective leptin signalling is associated with elevated hypothalamic, but not cerebellar, levels of endocannabinoids in obese db/db and ob/ob mice and Zucker rats. Acute leptin treatment of normal rats and ob/ob mice reduces anandamide and 2-arachidonoyl glycerol in the hypothalamus. These findings indicate that endocannabinoids in the hypothalamus may tonically activate CB1 receptors to maintain food intake and form part of the neural circuitry regulated by leptin.