Title : Transmembrane transport of water and urea in rat corneal endothelial cells

Objectives: In mammals corneal transparency is achieved through mechanisms that maintain osmotic balance in the stroma. Corneal endothelium can maintain stromal thickness and hydration levels by transporting water, ions and urea. Cells need to regulate their plasma membrane permeability to water and solutes, processes that in cornea endothelial cells are largely unknown. The study investigated permeability of the apical and basolateral membranes of rat corneal endothelial cells to water and urea.

Methods: To obtain preparation of cells, a fragment of a Descemet’s membrane with cells was placed on a coverslip and placed in the flow chamber of the microscope, where the fragment was mechanically held using a nylon mesh. Cell volume kinetics was assessed by means of biophysical measurements applying the Calcein Fluorescence Quenching method. Calculations of the P_f and P_u were done using a biophysical mathematical model of cell volume regulation.

Results: We demonstrated that the apparent water permeability of the basolateral membrane of endothelial cells (4.43E−02 ± 7.57E−04 cm/s) is more than three times higher than that of the apical membrane (1.21E−02 ± 1.03E−04 cm/s). Permeability of the basolateral membrane to urea (1.23E−04 ± 1.56E−06 cm/s) was statistically significantly higher than that of the apical membrane (9.52E−05 ± 1.02E−06 cm/s) by approximately 30%. Phloretin at concentration of 0.1 mM significantly reduced urea permeability by more than 20% through both the apical and basolateral membranes.

Conclusions: The results suggest that the compositions of transporters involved in water transport in the apical and basolateral membranes differ significantly. Urea is the major product of protein metabolism in mammals. It is not completely compatible osmotic agent for cell because it can influence conformation of macromolecules. One can suggest that cellular transport of urea is important for removing urea from the cells. Presence of the phloretin-sensitive urea transporters in the plasma membrane of endothelial cells is demonstrated. The results indicate potential significance of urea transport for corneal function.

Dr. Solenov Evgeniy major staff scientist of Institute of Cytology and Genetics of the Academy of Sciences of Russia, 630090 Novosibirsk. Dr. Solenov works in the field of  the cell volume regulation, development  of vasopressin antidiuretic action in mammalian kidney.