The Na+2Cl-K+ cotransporter in the rectal gland of Squalus acanthias is activated by cell shrinkage.
Effects of cAMP on Cl- secretion, intracellular Cl- activity and cell volume were studied in isolated perfused rectal gland tubules (RGT) of Squalus acanthias with electrophysiological and fluorescence methods. Recording of equivalent short-circuit current (Isc) showed that cAMP stimulates Na+Cl- secretion in a biphasic manner. The first and rapid phase corresponds to Cl- exit via the respective protein-kinase-A- (PKA-) phosphorylated Cl- conductance. The inhibitory effect of the loop diuretic furosemide (0.5 mmol/l, n=12) indicates that second phase reflects the delayed (1-2 min) activation of the Na+2Cl-K+ cotransporter. During the first phase cytosolic Cl- activity, as monitored by 6-methoxy-N-(3-sulfopropyl) quinolinium (SPQ) fluorescence, fell to 78% (n=23) of the control value. Concomitantly, a transient fall in cell volume was recorded by calcein fluorescence to 92% (n=5) of the control value. Preincubation of the RGT with phalloidin (0.1 mmol/l, n=6) or cytochalasin D (0.1 mmol/l, n=4) almost completely prevented the development of the second phase of Isc activation. When cytosolic Cl- activity was increased by exposing the RGT to a high K+ concentration (25 mmol/l), in the presence of mannitol to prevent volume increases, stimulation was unaffected and biphasic. In contrast, when cell volume was clamped to an increased value (115%, n=8) by removing extracellular NaCl, the second phase was abolished completely (n=11). These data suggest that the primary and key process for triggering the Na+2Cl-K+ cotransport is transient cell shrinkage.
Greger
R
R
Physiologisches Institut, Albert-Ludwigs-Universität, Hermann-Herder-Strasse 7, D-79104 Freiburg, Germany. Greger@ruf.uni-freiburg.de
Heitzmann
D
D
Hug
M J
MJ
Hoffmann
E K
EK
Bleich
M
M
D000818:Animals; D001464:Barium / Q000378:metabolism; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism*; D048429:Cell Size / Q000502:physiology*; D002712:Chlorides / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D015638:Cytochalasin D / Q000378:metabolism; D004284:Dogfish; D008297:Male; D009637:Masoprocol / Q000494:pharmacology; D010590:Phalloidine / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D012491:Salt Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D028021:Sodium-Potassium-Chloride Symporters
D002352:Carrier Proteins; D002712:Chlorides; D047428:Protein Kinase Inhibitors; D028021:Sodium-Potassium-Chloride Symporters; D010590:Phalloidine; D005576:Colforsin; D015638:Cytochalasin D; D001464:Barium; D003994:Bucladesine; D009637:Masoprocol; D012964:Sodium; D011188:Potassium; D002118:Calcium