FLUOROPHOTOMETRY OF ION TRANSPORT IN CILIARY BODY

稳态眼内压是以下因素之间平衡的结果： 流体产生和流出。 眼内压（IOP）升高，a 可能对视力造成破坏性影响的疾病， 退化，当流出设施减少时发生。 然而，在这方面， 缓解IOP升高的临床干预主要依赖于 药物操纵的房水生产。 这种幽默是 由覆盖睫状体的双层上皮产生。 两者，细胞之间的细胞旁过滤（由血液驱动， 水流体静压差）和流体的主动运动， 细胞可以促进血流。 细胞主动运输， 这是一个主要的过程，但人们对它的了解很少。 在 在很大程度上，这种有限的知识是由于遇到的困难 在应用经典的电和生理方法， 研究离子和液体在具有复杂结构的组织中的传输 解剖学和复杂的细胞排列。 该提案旨在应用新颖的 用于细胞内离子追踪的技术（基于细胞可捕获， 显示离子敏感光谱的荧光染料）以产生彻底的 表征1）存在于两种细胞中的单个易位系统， 上皮细胞层; 2）之间的通信的性质， 层;和3）这些活动的药理学调节。 专业的设备和解剖技术， 单个（或小组）细胞中H+、Na+、K+、Cl-和Ca 2+的测量 将使用完整睫状体条中的每种类型的细胞。 条带 将安装在灌注室中，允许控制细胞外 组织两侧的离子环境以及 由细胞外变化或由 药理学效应物将用于识别和表征每种 个体运输机。 新兴的知识将被应用于产生 合理的模型为房水分泌过程。 这些模型和 收集的关于个体的药理学调节的信息 运输功能，将提高我们的能力， 在青光眼患者或个体中操纵房水分泌 表现出眼压升高

The steady state intraocular pressure is the result of a balance between fluid production and outflow. Elevated intraocular pressure (IOP), a condition which could have devastating effects for vision due to tissue degeneration, occurs when the outflow facility is reduced. However, clinical intervention to relieve elevated IOP relies mostly in pharmacological manipulation of aqueous humor production. This humor is generated by the dual layered epithelium that covers the ciliary body. Both, paracellular filtration between the cells (driven by the blood to aqueous hydrostatic pressure difference) and active movement of fluid by the cells could contribute to the flow. The cellular active transport, which has been shown to be the primary process, is poorly understood. In great part, this limited knowledge is due to the difficulties encountered in the application of classical electrical and physiological approaches to the study of ion and fluid transport in a tissue endowed with an intricate anatomy and a complex cell arrangement. This proposal aims to apply novel techniques for intracellular ion tracking (based on cell entrapable, fluorescent dyes exhibiting ion sensitive spectra) to generate a thorough characterization of 1) the individual translocation systems present in both epithelial cell layers; 2) the nature of the communication between the layers; and 3) the pharmacological regulation of these activities. Specialized equipment and dissection techniques allowing intracellular measurements of H+, Na+, K+, Cl- and Ca2+ in single (or small groups of) cells of each type in intact ciliary body strips will be used. The strips will be mounted in perfusion chambers allowing control of the extracellular ionic environment in each side of the tissue and the changes in intracellular concentrations induced by extracellular changes or by pharmacological effectors will be used to identify, and characterize each individual transporter. The emerging knowledge will be applied to generate plausible models for the aqueous humor secretory process. These models and the information gathered on the pharmacological modulation of individual transport functions by hypotensive agents, will improve our capacity to manipulate aqueous humor secretion in glaucoma patients or individuals exhibiting an elevated IOP.