IntroductionChemical signals, or neurotransmitters, represent the fundamental mode for intercellular communication in the nervous system. (1) The classical model for neurotransmitter action involves the uptake and storage of these small molecules into synaptic vesicles, release of vesicular contents into the synaptic cleft in response to depolarization of the presynaptic terminal by an action potential, binding of released neurotransmitters to cognate protein receptors on the postsynaptic (and presynaptic) terminal, and, finally, termination of signaling by protein-mediated uptake and degradation of neurotransmitters from the synaptic cleft. This model applies to a large number of well-studied neurotransmitters, including glutamate, γ-amino butyric acid (GABA), acetylcholine, and the monoamines, all of which represent aqueous solution-soluble molecules. More recently, lipids have emerged as an important class of chemical messengers in the nervous system that operate by a distinct mechanism.The hydrophobic nature of lipids precludes their stable uptake and storage into synaptic vesicles. Instead, lipid messengers appear to be biosynthesized and released by neurons at the moment of their intended action, which is often referred to as "on-demand" production. Similarly, the capacity of lipids to freely cross cell membranes places the burden of signal termination largely on the action of degradative enzymes. Lipid signaling systems are thus embedded within an elaborate collection of metabolic pathways, the composition and regulation of which ultimately establish the magnitude and duration of transmitter action. Here, we will review these general concepts as they relate to a specific class of lipid transmitters, the endogenous cannabinoids (endocannabinoids), and highlight how delineation of their cognate metabolic enzymes has been translated into the development of chemical and genetic tools to test the role that the endocannabinoid system plays in nervous system signaling and behavior.Endocannabinoids are defined as endogenous small molecules that activate the cannabinoid receptors CB1 and CB2, which are G-protein-coupled receptors that also recognize Δ 9tetrahydrocannabinol (THC), the psychoactive component of marijuana. (2, 3) The CB1 receptor is the major cannabinoid receptor in the nervous system and is responsible for mediating most of the neurobehavioral effects of THC. (4, 5) The CB2 receptor is predominantly expressed in immune cells, (6) where it appears to play a role in mediating the immunosuppressive effects of cannabinoids. Two principal endocannabinoids have been identified in mammals, N-arachidonoyl ethanolamine (anandamide) (7) and 2-Anandamide was the first identified endogenous ligand for the CB1 receptor. (7) As will be described in the following section, anandamide and other NAEs are produced upon demand through activity-dependent cleavage of membrane lipid precursors. The biological activity of anandamide in the central nervous system and in peripheral tissues is terminated by its removal from th...

Endocannabinoids are lipid signaling molecules that regulate a wide range of mammalian behaviors, including pain, inflammation, and cognitive/emotional state. The endocannabinoid anandamide is principally degraded by the integral membrane enzyme fatty acid amide hydrolase (FAAH), and there is currently much interest in developing FAAH inhibitors to augment endocannabinoid signaling in vivo. Here we report the discovery and detailed characterization of a highly efficacious and selective FAAH inhibitor PF-3845. Mechanistic and structural studies confirm that PF-3845 is a covalent inhibitor that carbamylates FAAH's serine nucleophile. PF-3845 selectively inhibits FAAH in vivo as determined by activity-based protein profiling and raises brain anandamide levels for up to 24 hrs, resulting in profound cannabinoid receptor-dependent reductions in inflammatory pain. These data thus designate PF-3845 as a valuable pharmacological tool for in vivo characterization of the endocannabinoid system.

Fatty acid amide hydrolase (FAAH) is a mammalian integral membrane enzyme that degrades the fatty acid amide family of endogenous signaling lipids, which includes the endogenous cannabinoid anandamide and the sleep-inducing substance oleamide. FAAH belongs to a large and diverse class of enzymes referred to as the amidase signature (AS) family. Investigations into the structure and function of FAAH, in combination with complementary studies of other AS enzymes, have engendered provocative molecular models to explain how this enzyme integrates into cell membranes and terminates fatty acid amide signaling in vivo. These studies, as well as their biological and therapeutic implications, are the subject of this review.

Fatty acid amides constitute a large and diverse class of lipid transmitters that includes the endogenous cannabinoid anandamide and the sleep-inducing substance oleamide. The magnitude and duration of fatty acid amide signaling are controlled by enzymatic hydrolysis in vivo. Fatty acid amide hydrolase (FAAH) activity in mammals has been primarily attributed to a single integral membrane enzyme of the amidase signature (AS) family. Here, we report the functional proteomic discovery of a second membrane-associated AS enzyme in humans that displays FAAH activity. The gene that encodes this second FAAH enzyme was found in multiple primate genomes, marsupials, and more distantly related vertebrates, but, remarkably, not in a number of lower placental mammals, including mouse and rat. The two human FAAH enzymes, which share 20% sequence identity and are referred to hereafter as FAAH-1 and FAAH-2, hydrolyzed primary fatty acid amide substrates (e.g. oleamide) at equivalent rates, whereas FAAH-1 exhibited much greater activity with N-acyl ethanolamines (e.g. anandamide) and N-acyl taurines. Both enzymes were sensitive to the principal classes of FAAH inhibitors synthesized to date, including O-aryl carbamates and ␣-keto heterocycles. These data coupled with the overlapping, but distinct tissue distributions of FAAH-1 and FAAH-2 suggest that these proteins may collaborate to control fatty acid amide catabolism in primates. The apparent loss of the FAAH-2 gene in some lower mammals should be taken into consideration when extrapolating genetic or pharmacological findings on the fatty acid amide signaling system across species.The fatty acid amide family of bioactive lipids can be divided into at least three chemical classes: N-acylethanolamines (1) and modulates several neurobehavioral processes, including pain (4), feeding (5), and memory (6). Oleamide is a sleep-inducing lipid that accumulates in the cerebrospinal fluid of sleep-deprived animals (2, 7). Oleamide has been shown to affect several protein receptors, including serotonin (8), GABA (9, 10) and cannabinoid (11) receptors, as well as gap junctions (12), and at least a subset of these proteins appears to be critical for mediating the hypnotic effects of oleamide (13,14). NATs are representative members of a large family of N-acyl amino acids that vary in both acyl chain and amino acid content (3, 15-17). These lipids have been shown to modulate pain sensation (17) and activate TRP channels (16).The signaling function of fatty acid amides is terminated by enzymatic hydrolysis in vivo. A principal enzyme involved in this process is fatty acid amide hydrolase (FAAH) (18 -20). Mice with a targeted disruption in the FAAH gene (FAAH(Ϫ/Ϫ) mice) (21) or those treated with FAAH inhibitors (22, 23) are severely impaired in their ability to degrade fatty acid amides and show hypersensitivity to the pharmacological effects of these lipids. Blockade of FAAH activity also leads to highly elevated endogenous levels of fatty acid amides in the nervous system (21-23) and periph...