functional characterization of water transport and cellular localization of three aquaporin paralogs in the salmonid intestine.

Intestinal water absorption is greatly enhanced in salmonids upon acclimation from freshwater (FW) to seawater (SW); however, the molecular mechanism for water transport is unknown. We conducted a pharmacological characterization of water absorption in the rainbow trout intestine along with an investigation of the distribution and cellular localization of 3 aquaporins (Aqp1aa, -1ab and -8ab) in pyloric caeca, middle (M) and posterior (P) intestine of the Atlantic salmon. In vitro iso-osmotic water absorption (Jv) was higher in SW than FW-trout and was inhibited by (mmol L-1): 0.1 KCN (41%), 0.1 ouabain (72%) and 0.1 bumetanide (82%) suggesting that active transport, Na+,K+-ATPase and Na+,K+,2Cl--cotransport are involved in establishing the driving gradient for water transport. Jv was also inhibited by 1 mmol L-1 HgCl2, serosally (23% in M and 44% in P), mucosally (27% in M) or both (61% in M and 58% in P), suggesting involvement of both apical and basolateral aquaporins in water transport. The inhibition was antagonized by 5 mmol L-1 mercaptoethanol. By comparison, 10 mmol L-1 mucosal tetraethylammonium, an inhibitor of certain aquaporins, inhibited Jv by 20%. In the presence of glucose, mucosal addition of phloridzin inhibited water transport by 20%, suggesting that water transport is partially linked to the Na+-glucose cotransporter. Using polyclonal antibodies against salmon Aqp1aa, -1ab and -8ab, we detected Aqp1aa, and -1ab immunoreactivity in the brush border and sub-apical region of enterocytes in all intestinal segments. The Aqp8ab antibody showed a particularly strong immunoreaction in the brush border and sub-apical region of enterocytes throughout the intestine and also stained lateral membranes and peri-nuclear regions though at lower intensity. The present localization of 3 aquaporins in both apical and lateral membranes of salmonid enterocytes facilitates a model for transcellular water transport in the intestine of SW-acclimated salmonids.