Although exposure to exocannabinoids (e.g. marijuana) is associated with adverse pregnancy outcome, little is known about the biochemistry, physiology, and consequences of endocannabinoids in human pregnancy. In these studies, we measured the levels of the endocannabinoid anandamide (N-arachidonoylethanolamine, AEA) by HPLC-mass spectrometry in 77 pregnant and 25 nonpregnant women. The mean +/- sem plasma AEA levels in the first, second, and third trimesters were 0.89 +/- 0.14, 0.44 +/- 0.12, and 0.42 +/- 0.11 nm, respectively. The levels in the first trimester were significantly higher than those in either the second or third trimester. During labor, AEA levels were 3.7 times nonlaboring term levels (2.5 +/- 0.22 vs. 0.68 +/- 0.09 nm, P < 0.0001). During the menstrual cycle, levels in the follicular phase were significantly higher than those in the luteal phase (1.68 +/- 0.16 vs. 0.87 +/- 0.09 nm, P < 0.005). Postmenopausal and luteal-phase levels were similar to those in the first trimester. These findings suggest that successful pregnancy implantation and progression requires low levels of AEA. At term, AEA levels dramatically increase during labor and are affected by the duration of labor, suggesting a role for AEA in normal labor.