Oxygenation of the Endocannabinoid, 2-Arachidonylglycerol, to Glyceryl Prostaglandins by Cyclooxygenase-2

Abstract

Cyclooxygenases (COX) play an important role in lipid signaling by oxygenating arachidonic acid to endoperoxide precursors of prostaglandins and thromboxane. Two cyclooxygenases exist which differ in tissue distribution and regulation but otherwise carry out identical chemical functions. The neutral arachidonate derivative, 2-arachidonylglycerol (2-AG), is one of two described endocannabinoids and appears to be a ligand for both the central (CB1) and peripheral (CB2) cannabinoid receptors. Here we report that 2-AG is a substrate for COX-2 and that it is metabolized as effectively as arachidonic acid. COX-2-mediated 2-AG oxygenation provides the novel lipid, prostaglandin H 2 glycerol ester (PGH 2 -G), in vitro and in cultured macrophages. PGH 2 -G produced by macrophages is a substrate for cellular PGD synthase, affording PGD 2 -G. Pharmacological studies reveal that macrophage production of PGD 2 -G from endogenous sources of 2-AG is calcium-dependent and mediated by diacylglycerol lipase and COX-2. These results identify a distinct function for COX-2 in endocannabinoid metabolism and in the generation of a new family of prostaglandins derived from diacylglycerol and 2-AG. Cyclooxygenase (COX 1; prostaglandin endoperoxide synthase, EC 1.14.99.1) catalyzes the bis-dioxygenation of arachidonic acid, generating prostaglandin (PG) H 2 , the precursor to a diverse family of lipid mediators including PGs, thromboxane, and prostacyclin (1). The discovery of a second COX isoform, COX-2, has provided important insights into the molecular basis of inflammation, hyperalgesia, and cancer and has established a novel pharmacological target for their treatment (2-4). The major functional differences between COX-1 and COX-2 are believed to be related to their differential regulation and tissue distribution (5). COX-1 is a constitutive enzyme, whereas COX-2 is inducible and highly regulated by a range of agonists (6, 7). COX-1 activity accounts for PG and thromboxane production in gastric mucosa, kidney, and platelets (3). COX-2 activity is primarily responsible for PG biosynthesis in the central nervous system and inflammatory cells (8 -10). These observations suggest that COX-1 and COX-2 serve different physiological and pathophysiological functions.The possibility that COX-2 has distinct biochemical functions has not been explored extensively. There are conserved structural differences between the active sites of COX-1 and COX-2 which have been exploited in the development of selective COX-2 inhibitors. It is possible that these or other structural differences may have evolved to support separate biochemical functions for the two COX isoforms. The first indication of a separate biochemical function for COX-2 was provided by the observation that it selectively oxygenates the neutral ethanolamide derivative of arachidonic acid, anandamide (11). Anandamide and 2-arachidonylglycerol (2-AG) are endogenous ligands for the cannabinoid receptors that bind ⌬ 9 -tetrahydrocannabinol and mediate its pharmacological effects (12). The ca...

Competition and allostery govern substrate selectivity of cyclooxygenase-2

“…Specifically, the active site of COX-2 is larger than that of COX-1, allowing COX-2 to oxygenate bulkier amide and ester analogs of AA that are poor COX-1 substrates. Among these COX-2-selective substrates are the endocannabinoids 2-arachidonoylglycerol (2-AG) and arachidonoylethanolamide (3)(4)(5). COX-2-dependent oxygenation of these substrates leads to the biosynthesis of prostaglandin glyceryl esters (PG-Gs) and prostaglandin ethanolamides (prostamides), respectively (3,4).…”

“…Among these COX-2-selective substrates are the endocannabinoids 2-arachidonoylglycerol (2-AG) and arachidonoylethanolamide (3)(4)(5). COX-2-dependent oxygenation of these substrates leads to the biosynthesis of prostaglandin glyceryl esters (PG-Gs) and prostaglandin ethanolamides (prostamides), respectively (3,4). In vitro, COX-2 uses 2-AG and AA with similar kinetic efficiencies (3); however, PG-G production in intact cells is much lower than would be expected based on the relative amounts of cellular AA and 2-AG (6).…”

Altered GABAergic neurotransmission is associated with increased kainate-induced seizure in prostaglandin-endoperoxide synthase-2 deficient mice

“…Previous studies have suggested that activation of the CB1 receptors on GABA interneurons by retrograde endocannabinoid signaling induced by depolarization results in an increase in synaptic excitation due to a depolarization-induced disinhibition [3,28]. Since it is known that PTGS-2, but not PTGS-1, can catabolize endocannabinoids [30][31][32], we hypothesized that PTGS-2 -/-mice experience an excess of endocannabinoid production which would be expected to increase synaptic excitation [3,28] and that inhibition of the CB1 endocannabinoid receptor by AM-251 pretreatment before KA-injection would abrogate the enhanced sensitivity of PTGS-2 -/-mice to KA by blocking the CB1-mediated disinhibition. Contrary to our hypothesis, antagonism of the CB1 receptor prior to KA administration augmented, rather than diminished, the KA-induced seizure activity and neuronal damage, suggesting that the fraction of endocannabinoid that undergoes PTGS-2 metabolism does not significantly contribute to this process of seizure susceptibility.…”

“…Specifically, the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was decreased in the CA1 pyramidal neurons of PTGS-2 -/-mice, suggesting an alteration of GABAergic function. Augmented susceptibility to excitotoxicity was not mediated by endocannabinoid CB1 receptor signaling due to AA metabolites catabolized by PTGS-2, but not PTGS-1 [30][31][32]70], that are known to alter neuronal excitability [3,28].…”

Role of the Cannabinoid System in Pain Control and Therapeutic Implications for the Management of Acute and Chronic Pain Episodes

“…COX-2 exerts a negative influence on endocannabinoids because it catabolises them (as anandamide and 2-AG, that have shown neuroprotective properties in the injured brain) [83]. In a traumatic brain injury model, COX-2 inhibitor treatment protected 2-AG levels, enhanced functional recovery, and reduced cell death and inflammation [45], confirming an interaction between the endocannabinoid 2-AG and COX-2 enzyme.…”