Endocannabinoids (eCBs) function as retrograde signaling molecules at synapses throughout the brain, regulate axonal growth and guidance during development, and drive adult neurogenesis. There remains a lack of genetic evidence as to the identity of the enzyme(s) responsible for the synthesis of eCBs in the brain. Diacylglycerol lipase-␣ (DAGL␣) and -␤ (DAGL␤) synthesize 2-arachidonoyl-glycerol (2-AG), the most abundant eCB in the brain. However, their respective contribution to this and to eCB signaling has not been tested. In the present study, we show ϳ80% reductions in 2-AG levels in the brain and spinal cord in DAGL␣ Ϫ/Ϫ mice and a 50% reduction in the brain in DAGL␤ Ϫ/Ϫ mice. In contrast, DAGL␤ plays a more important role than DAGL␣ in regulating 2-AG levels in the liver, with a 90% reduction seen in DAGL␤ Ϫ/Ϫ mice. Levels of arachidonic acid decrease in parallel with 2-AG, suggesting that DAGL activity controls the steady-state levels of both lipids. In the hippocampus, the postsynaptic release of an eCB results in the transient suppression of GABAmediated transmission at inhibitory synapses; we now show that this form of synaptic plasticity is completely lost in DAGL␣ Ϫ/Ϫ animals and relatively unaffected in DAGL␤ Ϫ/Ϫ animals. Finally, we show that the control of adult neurogenesis in the hippocampus and subventricular zone is compromised in the DAGL␣ Ϫ/Ϫ and/or DAGL␤ Ϫ/Ϫ mice. These findings provide the first evidence that DAGL␣ is the major biosynthetic enzyme for 2-AG in the nervous system and reveal an essential role for this enzyme in regulating retrograde synaptic plasticity and adult neurogenesis.

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Endocannabinoids are lipid molecules that serve as natural ligands for the cannabinoid receptors CB1 and CB2. They modulate a diverse set of physiological processes such as pain, cognition, appetite, and emotional states, and their levels and functions are tightly regulated by enzymatic biosynthesis and degradation. 2-Arachidonoylglycerol (2-AG) is the most abundant endocannabinoid in the brain and is believed to be hydrolyzed primarily by the serine hydrolase monoacylglycerol lipase (MAGL). Although 2-AG binds and activates cannabinoid receptors in vitro, when administered in vivo, it induces only transient cannabimimetic effects as a result of its rapid catabolism. Here we show using a mouse model with a targeted disruption of the MAGL gene that MAGL is the major modulator of 2-AG hydrolysis in vivo. Mice lacking MAGL exhibit dramatically reduced 2-AG hydrolase activity and highly elevated 2-AG levels in the nervous system. A lack of MAGL activity and subsequent long-term elevation of 2-AG levels lead to desensitization of brain CB1 receptors with a significant reduction of cannabimimetic effects of CB1 agonists. Also consistent with CB1 desensitization, MAGL-deficient mice do not show alterations in neuropathic and inflammatory pain sensitivity. These findings provide the first genetic in vivo evidence that MAGL is the major regulator of 2-AG levels and signaling and reveal a pivotal role for 2-AG in modulating CB1 receptor sensitization and endocannabinoid tone.

The vanilloid receptor 1 (VR1) is a cation channel expressed predominantly by nociceptive sensory neurons and is activated by a wide array of pain-producing stimuli, including capsaicin, noxious heat, and low pH. Although the behavioral effects of injected capsaicin and the VR1 antagonist capsazepine have indicated a potential role for VR1 in the generation and maintenance of persistent pain states, species differences in the molecular pharmacology of VR1 and a limited number of selective ligands have made VR1 difficult to study in vivo. N-(4-Tertiarybutylphenyl)-4-(3-cholorphyridin-2-yl)tetrahydropryazine-1(2H)-carbox-amide (BCTC) is a recently described inhibitor of capsaicin-and acidmediated currents at rat VR1. Here, we report the effects of BCTC on acute, inflammatory, and neuropathic pain in rats. Administration of BCTC (30 mg/kg p.o.) significantly reduced both mechanical and thermal hyperalgesia induced by intraplantar injection of 30 g of capsaicin. In rats with Freund's complete adjuvantinduced inflammation, BCTC significantly reduced the accompanying thermal and mechanical hyperalgesia (3 mg/kg and 10 mg/kg p.o., respectively). BCTC also reduced mechanical hyperalgesia and tactile allodynia 2 weeks after partial sciatic nerve injury (10 and 30 mg/kg p.o.). BCTC did not affect motor performance on the rotarod after administration of doses up to 50 mg/kg p.o. These data suggest a role for VR1 in persistent and chronic pain arising from inflammation or nerve injury.The vanilloid receptor type 1 (VR1) is a pivotal molecular integrator of noxious stimuli that is expressed on somatic and autonomic primary afferent neurons. VR1 has been confirmed as a ligand-gated ion channel after its cloning from rat and human tissues, and has been shown to be highly expressed in small-diameter primary afferent neurons (Caterina et al., 1997;Hayes et al., 2000;McIntyre et al., 2001). In vitro studies have shown that, like the native vanilloid receptor, recombinant VR1 can be activated by a variety of chemical as well as physical stimuli. In vitro, VR1 responds to plant-derived compounds, including capsaicin, a pungent component of chili peppers, lipid mediators such as anandamide , the lipoxygenase product 12-(S)-hydroperoxyeicosatetraenoic acid (Hwang et al., 2000), as well as noxious heat (Caterina et al., 1997) and low pH (Tominaga et al., 1998).A potential role for VR1 in nociception has been evident for some time because injection of the VR1 agonist capsaicin induces nocifensive and hyperalgesic behaviors in rodents and pain in humans (Szolcsanyi, 1977;Carpenter and Lynn, 1981;Simone et al., 1987Simone et al., , 1989Gilchrist et al., 1996). Further support for VR1 as a therapeutic target arose from experiments involving capsazepine. Capsazepine is a VR1 antagonist that has been shown to competitively inhibit capsaicin-mediated responses in isolated dorsal root ganglion (DRG) neurons (Bevan et al., 1992a) and tissues from rat (Bevan et al., 1992b;Cholewinski et al., 1993;Maggi et al., 1993;Santicioli et al., 19...

Subchondral bone remodeling plays an important role in nociception and the pathobiology of the MIA model with osteoclasts being implicated in both bone and cartilage resorption. Inhibition of osteoclastic activity when initiated early leads to improved efficacy.