The objective of this proposal is to learn the cellular and molecular characteristics of the transporting systems for aqueous secretion by studying transepithelial transport across the ciliary epithelium of the eye. A completely new sort of membrane preparation of the isolated bilayered ciliary epithelium has been developed to facilitate these studies. Studies of this bilayer would be compared to single layer preparations of the pigmented and non-pigmented epithelium that have also been obtained. The bioelectric characteristics of this preparation, the transepithelial potential difference, resistance and short circuit current would be determined. Electrical and tracer methods would be used to examine the mechanisms for ion permeation. For example, the separate contributions of sodium, chloride and bicarbonate to the net current across these cellular preparations and their interactions would be studied. What fractions of the short circuit current are accounted for by the separate ions? The likelihood of electrically silent pathways would then be explored. Is the response polarized? The nature of these different transport processes would be examined and quantified. What is the involvement of channel mechanisms and/or carrier-mediated processes? The characteristics of potentially important outward anion transport (kidney and liver-like) systems can and would be determined in this preparation. The preparation would be used to explore how these transporting systems are regulated. Extracellular regulatory pathways would be examined by activating appropriate receptors with neurohormones, mediators and local hormones or neuropeptides to learn the potential role of these substances in influencing secretion. In particular the cyclic nucleotide messenger system would be explored to learn its functional relationship to the vectorial transport pathways for ions. It is also planned to study the transfer of ascorbic acid and glucose. Data obtained from certain kinetic studies of transport would be modeled to several kinetic analyses and compared with past uptake studies when possible. The rate of aqueous humor formation and its composition determine the nutrition of the avascular parts of the eye; the interior cornea, the trabecular meshwork, crystalline lens and probably the anterior vitreous. Therefore this research would bear on certain corneal dystrophies, glaucoma, possibly cataract, and changes in the vitreous with age. Studies of drug mechanisms, e.g., beta adrenergic blockers, would improve the applicant's understanding of the intrinsic mechanisms of aqueous humor formation and help determine how these agents reduce aqueous flow in glaucoma.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37EY008879-07
Application #
2019761
Study Section
Special Emphasis Panel (NSS)
Project Start
1996-01-19
Project End
1998-12-31
Budget Start
1997-01-01
Budget End
1998-12-31
Support Year
7
Fiscal Year
1997
Total Cost
Indirect Cost
Name
Yale University
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
082359691
City
New Haven
State
CT
Country
United States
Zip Code
06520
Crook, R B; Takahashi, K; Mead, A et al. (2000) The role of NaKCl cotransport in blood-to-aqueous chloride fluxes across rabbit ciliary epithelium. Invest Ophthalmol Vis Sci 41:2574-83
Hirata, K; Nathanson, M H; Burgstahler, A D et al. (1999) Relationship between inositol 1,4,5-trisphosphate receptor isoforms and subcellular Ca2+ signaling patterns in nonpigmented ciliary epithelia. Invest Ophthalmol Vis Sci 40:2046-53
Sears, J; Nakano, T; Sears, M (1998) Adrenergic-mediated connexin43 phosphorylation in the ocular ciliary epithelium. Curr Eye Res 17:104-7
Hirata, K; Nathanson, M H; Sears, M L (1998) Novel paracrine signaling mechanism in the ocular ciliary epithelium. Proc Natl Acad Sci U S A 95:8381-6
Chen, S; Wan, X L; Sears, M (1998) pICln can regulate swelling-induced Cl- currents in either layer of rabbit ciliary epithelium. Biochem Biophys Res Commun 246:59-63
Chen, S; Sears, M (1997) A low conductance chloride channel in the basolateral membranes of the non-pigmented ciliary epithelium of the rabbit eye. Curr Eye Res 16:710-8
Wan, X L; Sears, J; Chen, S et al. (1997) Circadian aqueous flow mediated by beta-arrestin induced homologous desensitization. Exp Eye Res 64:1005-11
Suzuki, Y; Nakano, T; Sears, M (1997) Calcium signals from intact rabbit ciliary epithelium observed with confocal microscopy. Curr Eye Res 16:166-75
Wan, X L; Chen, S; Sears, M (1997) Cloning and functional expression of a swelling-induced chloride conductance regulatory protein, plCln, from rabbit ocular ciliary epithelium. Biochem Biophys Res Commun 239:692-6
Horio, B; Sears, M; Mead, A et al. (1996) Regulation and bioelectrical effects of cyclic adenosine monophosphate production in the ciliary epithelial bilayer. Invest Ophthalmol Vis Sci 37:607-12

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