Abstract Context: Skeletal muscle endocannabinoids and sphingolipids (particularly sphingomyelins) are inversely associated with sleeping energy expenditure (SLEEP) in humans. The endocannabinoid system may increase sphingolipid synthesis via cannabinoid receptor-1. Objective: To investigate in human skeletal muscle whether endocannabinoids are responsible for the effect of sphingomyelins on SLEEP. Design: Muscle endocannabinoid [anandamide (AEA), 2-arachidonoylglycerol (2-AG)], endocannabinoid congeners [oleoylethanolamide (OEA), palmitoylethanolamide (PEA)], and sphingomyelin content were measured with liquid chromatography/mass spectrometry. SLEEP was assessed in a whole-room indirect calorimeter. Mediation analyses tested whether the inverse associations between sphingomyelins and SLEEP depended on endocannabinoids and endocannabinoid-related OEA and PEA. Setting: Inpatient study. Participants: Fifty-three Native Americans who...