The levels of 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand, and other molecular species of monoacylglycerols in rat brain were examined. In this study, we sacrificed the animals in liquid nitrogen to minimize postmortem changes. We found that rat brain contains 0.23 nmol/g tissue of 2-arachidonoylglycerol, which accounts for 10.5% of the total monoacylglycerol present in this tissue. We next investigated the:level of 2-arachidonoylglycerol after in vivo stimulation with picrotoxinin. We found that the level of 2-arachidonoylglycerol was elevated markedly in picrotoxinin-administered rat brain (4- to 6-fold over the control level). Changes in the levels of other molecular species were relatively small or negligible. Several cannabimimetic molecules as well as Delta(9)-tetrahydrocannabinol are known to depress neurotransmission and to exert anticonvulsant activities; endogenous 2-arachidonoylglycerol produced during neural excitation may play a regulatory role in calming the enhanced synaptic transmission.