Abstract
The most important risk factor for the development of glaucoma is elevated intraocular pressure (IOP). Hypotensive drugs decrease IOP, preventing optic nerve damage and further vision loss. The balance between aqueous humor (AH) production and drainage determines IOP, and problems in AH outflow pathways are associated with open-angle glaucoma development. Previous studies have shown the presence of diadenosine tetraphosphate (Ap4A) and pentaphosphate (Ap5A) in the AH. Topic application of Ap4A to the cornea decreased IOP, whereas Ap5A increased it. Because dinucleoside polyphosphates stimulate P2Y purinergic receptors, we studied their presence in trabecular meshwork (TM) cells. Additionally, the effects of diadenosine polyphosphates (ApnAs; n = 3–5) and Up4U (P1,P4-(diuridine 5′)-tetraphosphate; INS365) in outflow facility were tested. P2Y1, P2Y2, and P2Y4 receptors were detected in TM cells by Western blot and immunocytochemistry. In TM cells, Ap3A, Ap4A, and Ap5A induced discrete intracellular calcium concentration ([Ca2+]i) mobilizations compared with higher and more sustained [Ca2+]i mobilizations after Up4U application. In bovine ocular anterior segments perfused at constant pressure, 1 μM Ap3A or Ap4A increased outflow facility, whereas Up4U or Ap5A did not modify it. 2-MeSADP, a selective P2Y1 agonist, induced outflow facility increases similar to those obtained after Ap3A and Ap4A, and these were prevented by addition of the selective P2Y1 receptor antagonist MRS-2179 (2′-deoxy-N6-methyladenosine-3′,5′-diphosphate). Our results demonstrate that the hypotensive effect of Ap4A and other dinucleotides is mediated, at least in part, by increasing trabecular outflow facility through activation of P2Y1 receptors. The latter would seem to be an interesting target in the development of antiglaucomatous drugs to selectively increase AH outflow.
Footnotes
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This work was supported by BFI2002-01202, BFI2003-01190, PI031495, SAF2003-00338, and FEDER UCOM03-33-024, Spain.
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doi:10.1124/jpet.105.085274.
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ABBREVIATIONS: IOP, intraocular pressure; AH, aqueous humor; TM, trabecular meshwork; Ap4A, P1,P4-diadenosine tetraphosphate; Ap5A, P1,P5-diadenosine pentaphosphate; PL, phospholipase; IP3, inositol 1,4,5-trisphosphate; PKC, protein kinase C; BSA, bovine serum albumin; DMEM, Dulbecco's modified Eagle's medium; PBS, phosphate-buffered saline; TRITC, tetramethylrhodamine B isothiocyanate; HPLC, high-performance liquid chromatography; Ap3A, P1,P3-diadenosine triphosphate; 2-MeSADP, 2-(methylthio) adenosine 5′-diphosphate; MRS-2179, 2′-deoxy-N6-methyladenosine-3′,5′-diphosphate; PPADS, pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid; Up4U, P1,P4-(diuridine 5′)-tetraphosphate; ANOVA, analysis of variance; [Ca2+]i, intracellular calcium concentration; INS37217, P1-(uridine 5′)-P4-(2′-deoxycytidine-5′)-tetraphosphate.
- Received February 22, 2005.
- Accepted June 7, 2005.
- The American Society for Pharmacology and Experimental Therapeutics
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