Abstract
PurposeCannabinoids, such as Δ9-THC, act through an endogenous signaling system in the vertebrate eye that reduces IOP via CB1 receptors. Endogenous cannabinoid (eCB) ligand, 2-arachidonoyl glycerol (2-AG), likewise activates CB1 and is metabolized by monoacylglycerol lipase (MAGL). We investigated ocular 2-AG and its regulation by MAGL and the therapeutic potential of harnessing eCBs to lower IOP.MethodsWe tested the effect of topical application of 2-AG and MAGL blockers in normotensive mice and examined changes in eCB-related lipid species in the eyes and spinal cord of MAGL knockout (MAGL−/−) mice using high performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS). We also examined the protein distribution of MAGL in the mouse anterior chamber.Results2-Arachidonoyl glycerol reliably lowered IOP in a CB1- and concentration-dependent manner. Monoacylglycerol lipase is expressed prominently in nonpigmented ciliary epithelium. The MAGL blocker KML29, but not JZL184, lowered IOP. The ability of CB1 to lower IOP is not desensitized in MAGL−/− mice. Ocular monoacylglycerols, including 2-AG, are elevated in MAGL−/− mice but, in contrast to the spinal cord, arachidonic acid and prostaglandins are not changed.ConclusionsOur data confirm a central role for MAGL in metabolism of ocular 2-AG and related lipid species, and that endogenous 2-AG can be harnessed to reduce IOP. The MAGL blocker KML29 has promise as a therapeutic agent, while JZL184 may have difficulty crossing the cornea. These data, combined with the relative specificity of MAGL for ocular monoacylglycerols and the lack of desensitization in MAGL−/− mice, suggest that the development of an optimized MAGL blocker offers therapeutic potential for treatment of elevated IOP.
Highlights
We tested the effect of topical application of 2arachidonoyl glycerol (2-AG) and monoacylglycerol lipase (MAGL) blockers in normotensive mice and examined changes in Endogenous cannabinoid (eCB)-related lipid species in the eyes and spinal cord of MAGL knockout (MAGLÀ/À) mice using high performance liquid chromatography/ tandem mass spectrometry (HPLC/MS/MS)
Our data confirm a central role for MAGL in metabolism of ocular 2-AG and related lipid species, and that endogenous 2-AG can be harnessed to reduce IOP
These exogenous cannabinoids act on an endogenous signaling system that consists of receptors, ligands, and enzymes that produce and break down these ligands.[2]
Summary
Experiments were conducted at the Indiana University campus. All mice used for IOP experiments were handled according to the guidelines of the Indiana University animal care committee, and in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. Mice were allowed to acclimatize to the animal care facility for at least a week before their use in experiments. Intraocular pressure was measured in mice by rebound tonometry, using a Tonolab (Icare Finland Oy, Helsinki, Finland) This instrument uses a light plastic-tipped probe to briefly make contact with the cornea; after the probe encounters the eye, the instrument measures the speed at which the probe rebounds to calculate IOP. Intraocular pressure measurements following drug administration were analyzed by unpaired t-tests comparing drugtreated to vehicle-treated eyes. An XDB-C18 (Agilent Technologies) reversed phase HPLC analytical column was used, which was kept at 408C by a column oven (HP, Palo Alto, CA, USA). Mobile phase A consisted of 20%/80% (vol/vol) methanol/water and 1 mM ammonium acetate (Sigma-Aldrich Corp., St. Louis, MO, USA).
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