Fluid Movement across Conjunctiva: Stim. and Regulation

描述（由申请人提供）：我们之前已经将分离的兔结膜定性为Na吸收和CI分泌的上皮模型。我们的假设是，这些运输机制，通过提供液体在组织中运动的驱动力，可以使结膜要么从泪膜的水层吸收液体，要么向水层提供液体。用兔上皮直接实验证实了这一点。我们目前的假设是，人类结膜也可能拥有转运蛋白，使其能够吸收Na并分泌CI，从而吸收或分泌液体。要发生这种液体流动，结膜必须有水渠（和大多数液体运输上皮一样），水渠通常在顶区和底外侧区不同。因此，我们建议通过生物化学和组织化学方法，确定人结膜中的电解质转运蛋白和水通道（水通道蛋白）与液体流过上皮的运动有关。由于受体介导的反应在兔结膜中是不典型的，因此将在小鼠和猪组织中进行类似于人类组织的试验，以确定人类结膜中液体运输的最佳模型。

DESCRIPTION (provided by applicant): We have previously characterized the isolated rabbit conjunctiva as an epithelial model of Na absorption and CI secretion. Our hypothesis was that these transport mechanisms, by providing the driving forces for fluid movement across the tissue, could enable the conjunctiva to either absorb fluid from, or contribute fluid to, the aqueous layer of the tear film. This was demonstrated with direct experiments using the rabbit epithelium. Our current hypothesis is that the human conjunctiva may also possess transporters that enable it to absorb Na and secrete CI with the consequent fluid absorption or secretion. For this fluid flow to occur, the conjunctiva must have (as most fluid transporting epithelia do) water channels, which are usually different in the apical and basolateral domains. We thus propose to identify with biochemical and histochemical protocols, the electrolyte transporters and water channels (aquaporins) in the human conjunctiva relevant towards the movement of fluid flow across the epithelium. Because receptor-mediated responses are atypical in rabbit conjunctiva, assays similar to those in human will be done on murine and porcine tissues to determine the best model for fluid transport in the human conjunctiva. Since aquaporins (AQPs) play a crucial role in fluid transport, and our experiments have shown that anisotonic conditions and cAMP-elevating agents can modify water permeability, protocols to understand the regulation of water permeability in murine and porcine tissues will be implemented. Biochemical, histochemical and functional experiments will be conducted on mice, rat and pig conjunctivae. From the determination of the transporters, the AQPs, and the functional physiological experiments that can be done on the conjunctiva of the selected animal models, a theoretical model for the fluid movement across the human conjunctiva will be proposed. From these studies we expect to expand the understanding of the physiology of the mammalian conjunctival epithelium, and provide evidence that direct or receptor-mediated stimulation of ionic transport mechanisms will result in fluid secretion towards the tear film. These studies may provide the groundwork for the design of future treatment modalities for dry-eye syndromes.