Glucagon induces the gene expression of aquaporin-8 but not that of aquaporin-9 water channels in the rat hepatocyte

Leandro R. Soria, Sergio A. Gradilone, M. Cecilia Larocca, Raúl A. Marinelli

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Glucagon stimulates the vesicle trafficking of aquaporin-8 (AQP8) water channels to the rat hepatocyte canalicular membranes, a process thought to be relevant to glucagoninduced bile secretion. In this study, we investigated whether glucagon is able to modulate the gene expression of hepatocyte AQP8. Glucagon was administered to rats at 0.2 mg/100 g body wt ip in 2, 3, or 6 equally spaced doses for 8, 16, and 36 h, respectively. Immunoblotting analysis showed that hepatic 34-kDa AQP8 was significantly increased by 79 and 107% at 16 and 36 h, respectively. Hepatic AQP9 protein expression remained unaltered. AQP8 mRNA expression, assessed by real-time PCR, was not modified over time, suggesting a posttranscriptional mechanism of AQP8 protein increase. Glucagon effects on AQP8 were directly studied in primary cultured rat hepatocytes. Immunoblotting and confocal immunofluorescence microscopy confirmed the specific glucagon-induced AQP8 upregulation. The RNA polymerase II inhibitor actinomycin D was unable to prevent glucagon effect, providing additional support to the nontranscriptional upregulation of AQP8. Cycloheximide also showed no effect, suggesting that glucagon-induced AQP8 expression does not depend on protein synthesis but rather on protein degradation. Inhibitory experiments suggest that a reduced calpain-mediated AQP8 proteolysis could be involved. The action of glucagon on hepatocyte AQP8 was mimicked by dibutyryl cAMP and suppressed by PKA or phosphatidylinositol-3-kinase (PI3K) inhibitors. In conclusion, our data suggest that glucagon induces the gene expression of rat hepatocyte AQP8 by reducing its degradation, a process that involves cAMP-PKA and PI3K signal pathways.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Regulatory Integrative and Comparative Physiology
Volume296
Issue number4
DOIs
StatePublished - Apr 2009
Externally publishedYes

Fingerprint

Aquaporins
Glucagon
Hepatocytes
Gene Expression
Phosphatidylinositol 3-Kinase
Immunoblotting
Proteolysis
aquaporin 8
Up-Regulation
Proteins
Calpain
RNA Polymerase II
Liver
Dactinomycin
Cycloheximide
Fluorescence Microscopy
Confocal Microscopy
Bile
Real-Time Polymerase Chain Reaction
Signal Transduction

Keywords

  • Aquaporins
  • Calpains
  • Liver
  • Phosphatidylinositol-3-kinase
  • Protein kinase A

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

Cite this

Glucagon induces the gene expression of aquaporin-8 but not that of aquaporin-9 water channels in the rat hepatocyte. / Soria, Leandro R.; Gradilone, Sergio A.; Larocca, M. Cecilia; Marinelli, Raúl A.

In: American Journal of Physiology - Regulatory Integrative and Comparative Physiology, Vol. 296, No. 4, 04.2009.

Research output: Contribution to journalArticle

@article{27b4da3ca99845d7bdd4e4e5216d7276,
title = "Glucagon induces the gene expression of aquaporin-8 but not that of aquaporin-9 water channels in the rat hepatocyte",
abstract = "Glucagon stimulates the vesicle trafficking of aquaporin-8 (AQP8) water channels to the rat hepatocyte canalicular membranes, a process thought to be relevant to glucagoninduced bile secretion. In this study, we investigated whether glucagon is able to modulate the gene expression of hepatocyte AQP8. Glucagon was administered to rats at 0.2 mg/100 g body wt ip in 2, 3, or 6 equally spaced doses for 8, 16, and 36 h, respectively. Immunoblotting analysis showed that hepatic 34-kDa AQP8 was significantly increased by 79 and 107{\%} at 16 and 36 h, respectively. Hepatic AQP9 protein expression remained unaltered. AQP8 mRNA expression, assessed by real-time PCR, was not modified over time, suggesting a posttranscriptional mechanism of AQP8 protein increase. Glucagon effects on AQP8 were directly studied in primary cultured rat hepatocytes. Immunoblotting and confocal immunofluorescence microscopy confirmed the specific glucagon-induced AQP8 upregulation. The RNA polymerase II inhibitor actinomycin D was unable to prevent glucagon effect, providing additional support to the nontranscriptional upregulation of AQP8. Cycloheximide also showed no effect, suggesting that glucagon-induced AQP8 expression does not depend on protein synthesis but rather on protein degradation. Inhibitory experiments suggest that a reduced calpain-mediated AQP8 proteolysis could be involved. The action of glucagon on hepatocyte AQP8 was mimicked by dibutyryl cAMP and suppressed by PKA or phosphatidylinositol-3-kinase (PI3K) inhibitors. In conclusion, our data suggest that glucagon induces the gene expression of rat hepatocyte AQP8 by reducing its degradation, a process that involves cAMP-PKA and PI3K signal pathways.",
keywords = "Aquaporins, Calpains, Liver, Phosphatidylinositol-3-kinase, Protein kinase A",
author = "Soria, {Leandro R.} and Gradilone, {Sergio A.} and Larocca, {M. Cecilia} and Marinelli, {Ra{\'u}l A.}",
year = "2009",
month = "4",
doi = "10.1152/ajpregu.90783.2008",
language = "English (US)",
volume = "296",
journal = "American Journal of Physiology - Renal Fluid and Electrolyte Physiology",
issn = "1931-857X",
publisher = "American Physiological Society",
number = "4",

}

TY - JOUR

T1 - Glucagon induces the gene expression of aquaporin-8 but not that of aquaporin-9 water channels in the rat hepatocyte

AU - Soria, Leandro R.

AU - Gradilone, Sergio A.

AU - Larocca, M. Cecilia

AU - Marinelli, Raúl A.

PY - 2009/4

Y1 - 2009/4

N2 - Glucagon stimulates the vesicle trafficking of aquaporin-8 (AQP8) water channels to the rat hepatocyte canalicular membranes, a process thought to be relevant to glucagoninduced bile secretion. In this study, we investigated whether glucagon is able to modulate the gene expression of hepatocyte AQP8. Glucagon was administered to rats at 0.2 mg/100 g body wt ip in 2, 3, or 6 equally spaced doses for 8, 16, and 36 h, respectively. Immunoblotting analysis showed that hepatic 34-kDa AQP8 was significantly increased by 79 and 107% at 16 and 36 h, respectively. Hepatic AQP9 protein expression remained unaltered. AQP8 mRNA expression, assessed by real-time PCR, was not modified over time, suggesting a posttranscriptional mechanism of AQP8 protein increase. Glucagon effects on AQP8 were directly studied in primary cultured rat hepatocytes. Immunoblotting and confocal immunofluorescence microscopy confirmed the specific glucagon-induced AQP8 upregulation. The RNA polymerase II inhibitor actinomycin D was unable to prevent glucagon effect, providing additional support to the nontranscriptional upregulation of AQP8. Cycloheximide also showed no effect, suggesting that glucagon-induced AQP8 expression does not depend on protein synthesis but rather on protein degradation. Inhibitory experiments suggest that a reduced calpain-mediated AQP8 proteolysis could be involved. The action of glucagon on hepatocyte AQP8 was mimicked by dibutyryl cAMP and suppressed by PKA or phosphatidylinositol-3-kinase (PI3K) inhibitors. In conclusion, our data suggest that glucagon induces the gene expression of rat hepatocyte AQP8 by reducing its degradation, a process that involves cAMP-PKA and PI3K signal pathways.

AB - Glucagon stimulates the vesicle trafficking of aquaporin-8 (AQP8) water channels to the rat hepatocyte canalicular membranes, a process thought to be relevant to glucagoninduced bile secretion. In this study, we investigated whether glucagon is able to modulate the gene expression of hepatocyte AQP8. Glucagon was administered to rats at 0.2 mg/100 g body wt ip in 2, 3, or 6 equally spaced doses for 8, 16, and 36 h, respectively. Immunoblotting analysis showed that hepatic 34-kDa AQP8 was significantly increased by 79 and 107% at 16 and 36 h, respectively. Hepatic AQP9 protein expression remained unaltered. AQP8 mRNA expression, assessed by real-time PCR, was not modified over time, suggesting a posttranscriptional mechanism of AQP8 protein increase. Glucagon effects on AQP8 were directly studied in primary cultured rat hepatocytes. Immunoblotting and confocal immunofluorescence microscopy confirmed the specific glucagon-induced AQP8 upregulation. The RNA polymerase II inhibitor actinomycin D was unable to prevent glucagon effect, providing additional support to the nontranscriptional upregulation of AQP8. Cycloheximide also showed no effect, suggesting that glucagon-induced AQP8 expression does not depend on protein synthesis but rather on protein degradation. Inhibitory experiments suggest that a reduced calpain-mediated AQP8 proteolysis could be involved. The action of glucagon on hepatocyte AQP8 was mimicked by dibutyryl cAMP and suppressed by PKA or phosphatidylinositol-3-kinase (PI3K) inhibitors. In conclusion, our data suggest that glucagon induces the gene expression of rat hepatocyte AQP8 by reducing its degradation, a process that involves cAMP-PKA and PI3K signal pathways.

KW - Aquaporins

KW - Calpains

KW - Liver

KW - Phosphatidylinositol-3-kinase

KW - Protein kinase A

UR - http://www.scopus.com/inward/record.url?scp=65949103390&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=65949103390&partnerID=8YFLogxK

U2 - 10.1152/ajpregu.90783.2008

DO - 10.1152/ajpregu.90783.2008

M3 - Article

C2 - 19193945

AN - SCOPUS:65949103390

VL - 296

JO - American Journal of Physiology - Renal Fluid and Electrolyte Physiology

JF - American Journal of Physiology - Renal Fluid and Electrolyte Physiology

SN - 1931-857X

IS - 4

ER -