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Abstract #18507 Published in IGR 3-3
Cannabinoid CB(1) receptor expression, activation and detection of endogenous ligand in trabecular meshwork and ciliary process tissues
Stamer WD; Golightly SF; Hosohata Y; Ryan EP; Porter AC; Varga E; Noecker RJ; Felder CC; Yamamura HIEuropean Journal of Pharmacology 2001; 23: 277-286
Elevated intraocular pressure (IOP) is the primary risk factor for glaucoma. Cannabinoids interact with molecular targets in the eye and lower IOP by an unknown mechanism. The purpose of the present study was to examine eye tissues for functional cannabinoid receptors of the neuronal, CB(1) class, and an endogenous ligand, anandamide. The trabecular meshwork and ciliary processes are the primary structures of the eye that contribute to IOP and thus were the focus. Total RNA, frozen sections, cellular membranes, and primary cultures of cells were prepared from both bovine and cadaveric human tissues. Using cannabinoid CB(1) receptor-specific oligodeoxynucleotide primers, cannabinoid CB(1) receptor antiserum, and cannabinoid-specific compounds (CP-55,940, WIN55,212-2 and SR-141716A), the presence of cannabinoid CB(1) receptors in the ciliary processes and trabecular meshwork was determined. Using reverse transcription-polymerase chain reaction, the authors identified mRNA encoding cannabinoid CB(1) receptor protein in ciliary process and trabecular meshwork cells. Specific binding of anti-CB(1) immunoglobulin-G in tissue sections localized cannabinoid CB(1) receptor protein to the non-pigmented epithelial cells of the ciliary process and cells of the trabecular meshwork. While CP-55,940 and WIN55,212-2 failed to stimulate [(35)S]GTP gamma S binding in membrane preparations from trabecular meshwork and ciliary process, CP-55,940 significantly stimulated whole cell [(35)S]GTP gamma S binding by 51% over basal in ciliary process epithelial cells and 69% over basal in trabecular meshwork cells permeabilized with 5 μm digitonin (p < 0.001). Specificity of agonist stimulation was verified by complete blockade with the specific cannabinoid CB(1) receptor antagonist, SR-141716A. Moreover, activation of cannabinoid CB(1) receptors by CP-55,940 resulted in a 2.3 ± 0.3 and 1.7 ± 0.3-fold stimulation of cAMP accumulation in trabecular meshwork and ciliary process cells, respectively (p < 0.01). Lastly, anandamide was detected in human trabecular meshwork (3.08 pmol/g), ciliary process (49.42 pmol/g), and neurosensory retinal (4.48 pmol/g) tissues. For the first time, these data demonstrate in a single study the presence of both CB(1) mRNA and protein in trabecular meshwork and ciliary processes from two different species. Activation of heterotrimeric G proteins and stimulation of cAMP accumulation by cannabinoids in vitro suggest that their intraocular pressure-lowering effects in vivo result from activation of cannabinoid CB(1) receptors in the trabecular meshwork and increase aqueous outflow.
Dr W.D. Stamer, Department of Ophthalmology, The University of Arizona Health Sciences Center, Tucson, AZ 85724, USA. dstamer@eyes.arizona.edu
Classification:
2.5 Meshwork (Part of: 2 Anatomical structures in glaucoma)