Noladin ether acts on trabecular meshwork cannabinoid (CB1) receptors to enhance aqueous humor outflow facility.
OBJECTIVE
To study the effects of 2-arachidonyl glyceryl ether (noladin ether), an endocannabinoid ligand selective for cannabinoid (CB)1 receptor, on aqueous humor outflow facility, to investigate the involvement of trabecular meshwork CB1 receptors and the p42/44 MAP kinase signaling pathway and to explore the cellular mechanisms of noladin ether-induced changes of outflow facility.


METHODS
The effects of noladin ether on aqueous humor outflow facility were measured in a porcine anterior-segment-perfused organ culture model. The expression of CB1 receptors on cultured porcine trabecular meshwork cells and the coupling of these receptors to p42/44 MAP kinase was determined by immunofluorescence microscopy and Western blot analysis. Both Western blot and zymography were used to monitor the effects of noladin ether on matrix metalloproteinase (MMP)-2. In morphologic studies, AlexaFluor 488-labeled phalloidin staining was used to examine actin filament, and immunohistochemistry with anti-paxillin antibodies was used to detect focal adhesions.


RESULTS
Within 1 hour after adding 3, 30, or 300 nM of noladin ether, the aqueous humor outflow facility increased concentration dependently. The effect of 30 nM of noladin ether was completely blocked by SR141716A, a selective CB1 antagonist. Positive signals were detected on cultured porcine trabecular meshwork cells with an anti-CB1 antibody in immunofluorescence microscopy and Western blot studies. Treatment of trabecular meshwork cells with 30 nM of noladin ether activated p42/44 MAP kinase, whereas pretreatment with SR141716A blocked the p42/44 MAP kinase-activating effects of noladin ether. In addition, the enhancement of outflow facility induced by noladin ether was blocked by pretreatment of porcine anterior segments with PD98059, an inhibitor of p42/44 MAP kinase pathway. Furthermore, noladin ether treatment caused rounding of trabecular meshwork cells, and there was a decrease of actin stress fibers, as well as a decrease in focal adhesions. These noladin ether-induced morphologic changes were also blocked by SR141716A and PD98059.


CONCLUSIONS
The results demonstrate for the first time that administration of noladin ether, an endocannabinoid agonist selective for the CB1 receptor, increases aqueous humor outflow facility. The data also show that noladin ether-induced enhancement of outflow facility is mediated through the trabecular meshwork CB1 receptor, with an involvement of p42/44 MAP kinase signaling pathway and changes in actin cytoskeletons.
Njie
Ya Fatou
YF
Department of Pharmacology and Toxicology, School of Medicine, University of Louisville, Louisville, Kentucky 40292, USA.
Kumar
Akhilesh
A
Qiao
Zhuanhong
Z
Zhong
Lichun
L
Song
Zhao-Hui
ZH
D000199:Actins / Q000378:metabolism; D000818:Animals; D001082:Aqueous Humor / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D005989:Glycerides / Q000494:pharmacology*; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D008856:Microscopy, Fluorescence; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism; D009924:Organ Culture Techniques; D010477:Perfusion; D010766:Phosphorylation; D010880:Piperidines / Q000494:pharmacology; D011720:Pyrazoles / Q000494:pharmacology; D043884:Receptor, Cannabinoid, CB1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000077285:Rimonabant; D015398:Signal Transduction; D013552:Swine; D014129:Trabecular Meshwork / Q000187:drug effects* / Q000378:metabolism
D000199:Actins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D005989:Glycerides; D010880:Piperidines; D011720:Pyrazoles; D043884:Receptor, Cannabinoid, CB1; C424442:noladin ether; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020778:Matrix Metalloproteinase 2; D000077285:Rimonabant; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one