Endocannabinoids: getting the message across.

T he endocannabinoids have emerged as important intercellular signals in the nervous system. The fatty acid amide arachidonylethanolamide (anandamide) is the prototypical endocannabinoid, i.e., an endogenous ligand of the G proteincoupled cannabinoid receptor in the brain, CB1, which binds the main psychoactive component of marijuana and other derivatives of Cannabis sativa (1). After a rise in intracellular calcium or activation of certain neurotransmitter receptors, endocannabinoids are synthesized by cleavage of phospholipid precursors that are present in cellular membranes (2). They often act as retrograde messengers, emanating from morphologically undifferentiated regions of postsynaptic principal cells and traveling backwards across synapses, where they transiently inhibit the release of either the inhibitory neurotransmitter -aminobutyric acid (GABA) or the excitatory transmitter glutamate (see Fig. 1 and ref. 3 for a review). This powerful influence on synaptic transmission enables the endocannabinoids to help regulate behaviors, such as feeding (4), fear (5), and anxiety (6). As is true of other intercellular messengers, the time course and magnitude of endocannabinoid actions are strongly influenced by their lifetime in the extracellular space. Anandamide is enzymatically degraded by fatty acid amide hydrolase (FAAH), but FAAH is an intracellular enzyme, so anandamide must be internalized to be hydrolyzed (7). Most evidence has pointed to a sodiumand energy-independent transporter operating by facilitated diffusion as the endocannabinoid uptake mechanism (8). Support for this model included saturability of the uptake process, its selectivity for certain fatty acids, and its inhibition by anandamide analogs. However, the topic has been controversial (9). It has been suggested that passive diffusion alone is sufficient for anandamide uptake, with intracellular hydrolysis by FAAH maintaining the concentration gradient (10, 11). An article by Fegley et al. (12) in this issue of PNAS neatly disposes of the major components of this model and, in addition, introduces a new transport inhibitor that could serve as a basis for therapeutic drug development, thereby adding strong support for the transporter hypothesis. The Endocannabinoid Systems Anandamide and 2-arachidonyl glycerol (2-AG) are the main endocannabinoids in the brain (2). Once released, they can activate CB1, which, as the most abundant G protein-coupled receptor in the brain, is widely, although heterogeneously, distributed on particular axon terminals (2). (CB2 receptors are found in the periphery, often in association with cells of the immune system.) Where CB1 receptors are found on inhibitory axons, endocannabinoids mediate depolarization-induced suppression of inhibition (13, 14), and, where they are found on excitatory axons, they mediate depolarization-induced suppression of excitation (15). Endocannabinoids typically do not travel far from their source, usually 1 m, although distances up to 10 m are possible under some conditions (e.g., refs. 13 and 16). They generally activate receptors on axon terminals that synapse on the releasing cell, thereby serving as spatially and temporally restricted messengers. Anandamide Uptake A key challenge to the anandamide transporter hypothesis arose from the observation that the initial rate of anandamide accumulation in certain cell lines is not saturable although accumulation at later times is (11). Moreover, inhibitors of FAAH slowed anandamide uptake (10), and, conversely, in some cells, inhibitors of anandamide transport can inhibit FAAH (11). This constellation of effects is consistent with a hypothesis that passive anandamide influx is driven by a transmembrane anandamide gradient that is maintained by the intracellular action of FAAH (which would keep intracellular concentrations low). In testing central tenets of this idea in primary neuronal cultures, Fegley et al. (12) find that anandamide uptake is the same in wild-type and FAAH / mutant mice, and that puta-