János Almássy1, Gyula Diszházi2, Marianna Skaliczki3, Ildikó Márton4, Zsuzsanna Édua Magyar2, Péter P Nánási2, David I Yule5. 1. Department of Physiology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary. Electronic address: almassy.janos@med.unideb.hu. 2. Department of Physiology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary. 3. Department of Physiology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary; Department of Dentistry, University of Debrecen, Debrecen, Hungary. 4. Department of Dentistry, University of Debrecen, Debrecen, Hungary. 5. Department of Pharmacology and Physiology and the Centre for Oral Biology, University of Rochester, Medical Center, Rochester, USA.
Abstract
PURPOSE: Primary fluid secretion in secretory epithelia relies on the unidirectional transport of ions and water across a single cell layer. This mechanism requires the asymmetric apico-basal distribution of ion transporters and intracellular Ca2+ signaling. The primary aim of the present study was to verify the localization and the identity of Ca2+-dependent ion channels in acinar cells of the mouse lacrimal gland. METHODS: Whole-cell patch-clamp-electrophysiology, spatially localized flash-photolysis of Ca2+ and temporally resolved digital Ca2+-imaging was combined. Immunostaining of enzymatically isolated mouse lacrimal acinar cells was performed. RESULTS: We show that the Ca2+-dependent K+-conductance is paxilline-sensitive, abundant in the luminal, but negligible in the basal membrane; and co-localizes with Cl--conductance. These data suggest that both Cl- and K+ are secreted into the lumen and thus they account for the high luminal [Cl-] (∼141 mM), but not for the relatively low [K+] (<17 mM) of the primary fluid. Accordingly, these results also imply that K+ must be reabsorbed from the primary tear fluid by the acinar cells. We hypothesized that apically-localized Na+-K+ pumps are responsible for K+-reabsorption. To test this possibility, immunostaining of lacrimal acinar cells was performed using anti-Na+-K+ ATP-ase antibody. We found positive fluorescence signal not only in the basal, but in the apical membrane of acinar cells too. CONCLUSIONS: Based on these results we propose a new primary fluid-secretion model in the lacrimal gland, in which the paracellular pathway of Na+ secretion is supplemented by a transcellular pathway driven by apical Na+-K+ pumps.
PURPOSE: Primary fluid secretion in secretory epithelia relies on the unidirectional transport of ions and water across a single cell layer. This mechanism requires the asymmetric apico-basal distribution of ion transporters and intracellular Ca2+ signaling. The primary aim of the present study was to verify the localization and the identity of Ca2+-dependent ion channels in acinar cells of the mouse lacrimal gland. METHODS: Whole-cell patch-clamp-electrophysiology, spatially localized flash-photolysis of Ca2+ and temporally resolved digital Ca2+-imaging was combined. Immunostaining of enzymatically isolated mouse lacrimal acinar cells was performed. RESULTS: We show that the Ca2+-dependent K+-conductance is paxilline-sensitive, abundant in the luminal, but negligible in the basal membrane; and co-localizes with Cl--conductance. These data suggest that both Cl- and K+ are secreted into the lumen and thus they account for the high luminal [Cl-] (∼141 mM), but not for the relatively low [K+] (<17 mM) of the primary fluid. Accordingly, these results also imply that K+ must be reabsorbed from the primary tear fluid by the acinar cells. We hypothesized that apically-localized Na+-K+ pumps are responsible for K+-reabsorption. To test this possibility, immunostaining of lacrimal acinar cells was performed using anti-Na+-K+ ATP-ase antibody. We found positive fluorescence signal not only in the basal, but in the apical membrane of acinar cells too. CONCLUSIONS: Based on these results we propose a new primary fluid-secretion model in the lacrimal gland, in which the paracellular pathway of Na+ secretion is supplemented by a transcellular pathway driven by apical Na+-K+ pumps.
Authors: Shivaram Selvam; Padmaja B Thomas; Hovhannes J Gukasyan; Alan S Yu; Douglas Stevenson; Melvin D Trousdale; Austin K Mircheff; Joel E Schechter; Ronald E Smith; Samuel C Yiu Journal: Am J Physiol Cell Physiol Date: 2007-08-15 Impact factor: 4.249