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

Steffen Madsen

doi:10.3389/fphys.2011.00056

Abstract

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 three 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−-co-transport 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 co-transporter. 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 three aquaporins in both apical and lateral membranes of salmonid enterocytes facilitates a model for transcellular water transport in the intestine of SW-acclimated salmonids.

Highlights

Animals living in a dehydrating environment need compensatory mechanisms in order to maintain osmotic homeostasis
This study has shown that Aqp1aa, -1ab, and -8ab are localized in the apical brush borders of enterocytes lining the entire intestine from the pyloric caeca through the anus, allowing entry of water from the lumen into the enterocytes
The pyloric caeca may have a particular significance in quantitative uptake of water

Summary

Introduction

Animals living in a dehydrating environment need compensatory mechanisms in order to maintain osmotic homeostasis. Cartilaginous species actively build up the internal osmotic pressure by means of organic osmolytes to slightly exceed the external osmotic pressure. Thereby they may gain water passively across the gill epithelium. Whether marine or euryhaline, maintain hypo-osmotic body fluids by drinking seawater (SW) and actively excreting surplus ions. It has been known since the classical paper by Smith (1932) that they swallow an appreciable amount of water (drinking rates typically in the range of 1–5 mL−1 kg−1 h−1, Perrott et al, 1992), which is processed and partially absorbed in the gastrointestinal tract (GIT).

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