VOLUME REGULATION IN CORNEAL ENDOTHELIUM

细胞体积的维持是在所有动物中表达的稳态过程。 细胞必要的水运动通过质膜耦合 溶质运输机制，也涉及其他细胞 活动，如葡萄糖的积累，pHi调节或矢量 液体运输角膜内皮被赋予活性离子 负责矢量流体输送的泵机构，因此 维持角膜水化，这是角膜透明度的决定因素。 由于这种液体运输和细胞体积调节必须交织在一起， 必须建立数量保持机制， 调节，以获得流体输送的完整理解。在 此外，了解液体如何转运具有临床意义， 在渗透应力期间例如在接触透镜佩戴期间受到影响 和糖尿病本研究的重点是确立 在急性和持续性干扰下的音量调节， 和长期交易量规定/短期交易量 将通过以下细胞体积的变化来研究调节， 细胞内离子活动，以确定机制和第二 信使参与。实验将采用各向异性扰动， 酸中毒、缺氧、离子置换和转运特异性抑制剂。 长期的量调节将在适应的背景下进行审查 通过调节有机渗透剂的水平来维持各向异性负荷 (such如氨基酸、肌醇和山梨糖醇）。离体兔 角膜和培养的牛内皮细胞融合， 将使用在盖玻片或可渗透支持物上极化的。 将使用敏感光测量细胞体积的动态变化 散射技术，并通过以下浓度的荧光 用作体积标记的染料。本研究的长期目标是 了解液体运输，pHi调节和 正常角膜内皮细胞的体积调节活性 条件和代谢应激期间，高血糖和各向异性 载荷

Maintenance of cell volume is a homeostatic process expressed in all animal cells. The necessary water movement across the plasma membrane is coupled to solute transport mechanisms that are also involved in other cellular activities such as the accumulation of glucose, pHi regulation or vectorial fluid transport. The corneal endothelium is endowed with an active ion pump mechanism responsible for vectorial fluid transport and hence the maintenance of corneal hydration, a determinant of corneal transparency. Since this fluid transport and cell volume regulation must be intertwined, it is imperative to establish the mechanisms of volume maintenance and regulation to obtain a complete understanding of the fluid transport. In addition, it is of clinical significance to understand how fluid transport is affected during times of osmotic stress such as during contact lens wear and diabetes. The focus of this study is to establish the principles of volume regulation upon acute and sustained disturbances leading to short and long term volume regulations, respectively/ Short term volume regulation will be studied by following changes in cell volume and intracellular ionic activities to identify the mechanisms and second messengers involved. Experiments will employ anisosmotic perturbations, acidosis, hypoxia, ion substitutions and transport-specific inhibitors. Long term volume regulation will be examined in the context of adaptation to sustained anisosmotic loads by adjusting the levels of organic osmolytes (such as amino acids, myo-inositol and sorbitol). Isolated in vitro rabbit corneas and cultured bovine endothelial cells that are confluent and polarized on either glass coverslips or permeable supports will be used. The dynamics of cell volume will be measured using athe sensitive light scattering technique and by following the concentration of a fluorescent dye used as a volume marker. The long term goal of this study is to understand the interaction of the fluid transport, pHi regulation and volume regulation activities of the corneal endothelium under normal conditions and during metabolic stress, hyperglycemia and anisosmotic loads.