Functional presence of Na+‐K+‐2Cl‐ cotransporter in rabbit lacrimal gland ductal epithelial cells

Abstract

AbstractPurpose Capability of fluid secretion of rabbit lacrimal gland duct epithelial cells was recently demonstrated by the authors using isolated duct segment model developed earlier. Inhibition of basolateral Na+‐K+‐2Cl‐ cotransporter (NKCC1) with bumetanide resulted in a complete stoppage of fluid secretion. Aim of the present work was to determine the activity and the regulation of NKCC1.Methods Experiments were performed on isolated rabbit lacrimal gland interlobular duct segments. Intracellular fluorophotometry with ammonium pulse technique was used to measure cotransporter activity. The rate of bumetanide sensitive cytosolic acidification after addition of NH4‐ can be used to quantify the activity of the cotransporter. Role of various potentially stimulatory effects were investigated.Results Unstimulated activity of NKCC1 was determined. Elevated intracellular cAMP level evoked by forskolin stimulation resulted in marked increase in NKCC1 activity. Beside increased intracellular cAMP levels, low cytosolic Cl‐ concentration and cell shrinkage evoked by hyperosmotic environment were significant determinants of increased cotransporter activity.Conclusion Our earlier results suggested the dominant role of Cl‐ dependent mechanisms in lacrimal gland ductal fluid secretion. The presented experimental work proves the substantial role of NKCC1 in basolateral Cl‐ uptake. Intracellular regulation of Cl‐ handling and luminal Cl‐ secretion needs further investigation.