Glucagon induces the plasma membrane insertion of functional aquaporin-8 water channels in isolated rat hepatocytes

Sergio A. Gradilone, Fabiana García, Robert C Huebert, Pamela S. Tietz, M. Cecilia Larocca, Arlinet Kierbel, Flavia I. Carreras, Nicholas F La Russo, Raúl A. Marinelli

Research output: Contribution to journalArticle

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Abstract

Although glucagon is known to stimulate the cyclic adenosine monophosphate (cAMP)-mediated hepatocyte bile secretion, the precise mechanisms accounting for this choleretic effect are unknown. We recently reported that hepatocytes express the water channel aquaporin-8 (AQP8), which is located primarily in intracellular vesicles, and its relocalization to plasma membranes can be induced with dibutyryl cAMP. In this study, we tested the hypothesis that glucagon induces the trafficking of AQP8 to the hepatocyte plasma membrane and thus increases membrane water permeability. Immunoblotting analysis in subcellular fractions from isolated rat hepatocytes indicated that glucagon caused a significant, dose-dependent increase in the amount of AQP8 in plasma membranes (e.g., 102% with 1 μmol/L glucagon) and a simultaneous decrease in intracellular membranes (e.g., 38% with 1 μmol/L glucagon). Confocal immunofluorescence microscopy in cultured hepatocytes confirmed the glucagon-induced redistribution of AQP8 from intracellular vesicles to plasma membrane. Polarized hepatocyte couplets showed that this redistribution was specifically to the canalicular domain. Glucagon also significantly increased hepatocyte membrane water permeability by about 70%, which was inhibited by the water channel blocker dimethyl sulfoxide (DMSO). The inhibitors of protein kinase A, H-89, and PKI, as well as the microtubule blocker colchicine, prevented the glucagon effect on both AQP8 redistribution to hepatocyte surface and cell membrane water permeability. In conclusion, our data suggest that glucagon induces the protein kinase A and microtubule-dependent translocation of AQP8 water channels to the hepatocyte canalicular plasma membrane, which in turn leads to an increase in membrane water permeability. These findings provide evidence supporting the molecular mechanisms of glucagon-induced hepatocyte bile secretion.

Original languageEnglish (US)
Pages (from-to)1435-1441
Number of pages7
JournalHepatology
Volume37
Issue number6
DOIs
StatePublished - Jun 1 2003

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Aquaporins
Glucagon
Hepatocytes
Cell Membrane
Permeability
Water
Cyclic AMP-Dependent Protein Kinases
Bile
Microtubules
Cyclic AMP
Membranes
Cholagogues and Choleretics
aquaporin 8
Cell Membrane Permeability
Intracellular Membranes
Subcellular Fractions
Colchicine
Dimethyl Sulfoxide
Fluorescence Microscopy
Immunoblotting

ASJC Scopus subject areas

  • Hepatology

Cite this

Gradilone, S. A., García, F., Huebert, R. C., Tietz, P. S., Larocca, M. C., Kierbel, A., ... Marinelli, R. A. (2003). Glucagon induces the plasma membrane insertion of functional aquaporin-8 water channels in isolated rat hepatocytes. Hepatology, 37(6), 1435-1441. https://doi.org/10.1053/jhep.2003.50241

Glucagon induces the plasma membrane insertion of functional aquaporin-8 water channels in isolated rat hepatocytes. / Gradilone, Sergio A.; García, Fabiana; Huebert, Robert C; Tietz, Pamela S.; Larocca, M. Cecilia; Kierbel, Arlinet; Carreras, Flavia I.; La Russo, Nicholas F; Marinelli, Raúl A.

In: Hepatology, Vol. 37, No. 6, 01.06.2003, p. 1435-1441.

Research output: Contribution to journalArticle

Gradilone, SA, García, F, Huebert, RC, Tietz, PS, Larocca, MC, Kierbel, A, Carreras, FI, La Russo, NF & Marinelli, RA 2003, 'Glucagon induces the plasma membrane insertion of functional aquaporin-8 water channels in isolated rat hepatocytes', Hepatology, vol. 37, no. 6, pp. 1435-1441. https://doi.org/10.1053/jhep.2003.50241
Gradilone, Sergio A. ; García, Fabiana ; Huebert, Robert C ; Tietz, Pamela S. ; Larocca, M. Cecilia ; Kierbel, Arlinet ; Carreras, Flavia I. ; La Russo, Nicholas F ; Marinelli, Raúl A. / Glucagon induces the plasma membrane insertion of functional aquaporin-8 water channels in isolated rat hepatocytes. In: Hepatology. 2003 ; Vol. 37, No. 6. pp. 1435-1441.
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AU - García, Fabiana

AU - Huebert, Robert C

AU - Tietz, Pamela S.

AU - Larocca, M. Cecilia

AU - Kierbel, Arlinet

AU - Carreras, Flavia I.

AU - La Russo, Nicholas F

AU - Marinelli, Raúl A.

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AB - Although glucagon is known to stimulate the cyclic adenosine monophosphate (cAMP)-mediated hepatocyte bile secretion, the precise mechanisms accounting for this choleretic effect are unknown. We recently reported that hepatocytes express the water channel aquaporin-8 (AQP8), which is located primarily in intracellular vesicles, and its relocalization to plasma membranes can be induced with dibutyryl cAMP. In this study, we tested the hypothesis that glucagon induces the trafficking of AQP8 to the hepatocyte plasma membrane and thus increases membrane water permeability. Immunoblotting analysis in subcellular fractions from isolated rat hepatocytes indicated that glucagon caused a significant, dose-dependent increase in the amount of AQP8 in plasma membranes (e.g., 102% with 1 μmol/L glucagon) and a simultaneous decrease in intracellular membranes (e.g., 38% with 1 μmol/L glucagon). Confocal immunofluorescence microscopy in cultured hepatocytes confirmed the glucagon-induced redistribution of AQP8 from intracellular vesicles to plasma membrane. Polarized hepatocyte couplets showed that this redistribution was specifically to the canalicular domain. Glucagon also significantly increased hepatocyte membrane water permeability by about 70%, which was inhibited by the water channel blocker dimethyl sulfoxide (DMSO). The inhibitors of protein kinase A, H-89, and PKI, as well as the microtubule blocker colchicine, prevented the glucagon effect on both AQP8 redistribution to hepatocyte surface and cell membrane water permeability. In conclusion, our data suggest that glucagon induces the protein kinase A and microtubule-dependent translocation of AQP8 water channels to the hepatocyte canalicular plasma membrane, which in turn leads to an increase in membrane water permeability. These findings provide evidence supporting the molecular mechanisms of glucagon-induced hepatocyte bile secretion.

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