title: Characterization of Na+-K+-2Cl- Cotransporter Activity in Rabbit Lacrimal Gland Duct Cells.
abstract: OBJECTIVE
  We recently reported that isolated duct segments from rabbit lacrimal gland (LG) were able to secrete fluid in response to secretagogues, which were blocked completely by bumetanide. This suggests the functional involvement of Na+-K+-2Cl- cotransporter (NKCC1) in ductal fluid secretion. Therefore, the aim of this study was to investigate the activity profile of NKCC1 in isolated rabbit LG duct segments.
  
  
  METHODS
  Interlobular ducts were isolated from fresh rabbit LG tissue. Microfluorometry with the ammonium (NH4+)-pulse technique was used to elicit pH changes in duct cells, and the rate of bumetanide-sensitive cytosolic acidification after addition of NH4+ was used to quantify the activity of NKCC1.
  
  
  RESULTS
  While basal activity of NKCC1 was undetectable, low cytosolic chloride (Cl-) level and hyperosmotic challenge (390 mOsm) were able to increase the activity of NKCC1. Carbachol (100 μM) had no significant effect on NKCC1 activity. Elevation of cytosolic calcium (Ca2+) level with Ca2+-ionophore (A 23187, 1 μM) did not cause any alteration in the activity of the cotransporter while direct activation of protein kinase C (phorbol myristate acetate, 100 nM) increased its activity slightly but in a significant manner. Addition of either forskolin (10 μM), cell-permeable cAMP analogue (8-bromo cAMP, 100 μM) or vasoactive intestinal peptide (200 nM) resulted in a significant increase in the activity of NKCC1.
  
  
  CONCLUSIONS
  These results highlight the functional involvement of NKCC1 in LG duct secretion. These findings may facilitate our understanding of LG function and may contribute to the development of targeted pharmacologic interventions in case of dry eye disease.
authors.affiliation: Department of Ophthalmology, University of Szeged, Szeged, Hungary.
authors.forename: Eszter
authors.initials: E
authors.lastname: Vizvári
authors.affiliation: 1st Department of Internal Medicine, University of Szeged, Szeged, Hungary.
authors.forename: Máté
authors.initials: M
authors.lastname: Katona
authors.affiliation: Department of Pharmacology and Pharmacotherapy, University of Szeged, Szeged, Hungary.
authors.forename: Péter
authors.initials: P
authors.lastname: Orvos
authors.affiliation: Department of Ophthalmology, University of Szeged, Szeged, Hungary.
authors.forename: Orsolya
authors.initials: O
authors.lastname: Berczeli
authors.affiliation: Department of Ophthalmology, University of Szeged, Szeged, Hungary.
authors.forename: Andrea
authors.initials: A
authors.lastname: Facskó
authors.affiliation: Department of Medical Physics and Informatics, University of Szeged, Szeged, Hungary.
authors.forename: Ferenc
authors.initials: F
authors.lastname: Rárosi
authors.affiliation: Department of Pharmacology and Pharmacotherapy, University of Szeged, Szeged, Hungary.
authors.forename: Viktória
authors.initials: V
authors.lastname: Venglovecz
authors.affiliation: 1st Department of Internal Medicine, University of Szeged, Szeged, Hungary 5Department of Pathophysiology, University of Szeged, Szeged, Hungary.
authors.forename: Zoltán
authors.initials: Z
authors.lastname: Rakonczay
authors.affiliation: 1st Department of Internal Medicine, University of Szeged, Szeged, Hungary.
authors.forename: Péter
authors.initials: P
authors.lastname: Hegyi
authors.affiliation: Pharmacology & Pharmaceutical Sciences, Ophthalmology, University of Southern California, Los Angeles, California, United States.
authors.forename: Chuanqing
authors.initials: C
authors.lastname: Ding
authors.affiliation: Department of Ophthalmology, University of Szeged, Szeged, Hungary 3Department of Pharmacology and Pharmacotherapy, University of Szeged, Szeged, Hungary.
authors.forename: Edit
authors.initials: E
authors.lastname: Tóth-Molnár
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology; D015352:Dry Eye Syndromes / Q000209:etiology / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007765:Lacrimal Apparatus / Q000378:metabolism*; D008297:Male; D009883:Ophthalmic Solutions / Q000494:pharmacology; D009994:Osmolar Concentration; D011817:Rabbits; D064506:Solute Carrier Family 12, Member 2 / Q000378:metabolism / Q000502:physiology*; D013666:Tears / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology
chemical_list: D002273:Carcinogens; D009883:Ophthalmic Solutions; D064506:Solute Carrier Family 12, Member 2; D005576:Colforsin; C016276:8-bromocyclic GMP; D014660:Vasoactive Intestinal Peptide; D002217:Carbachol; D006152:Cyclic GMP; D013755:Tetradecanoylphorbol Acetate