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Vasopressin regulates the phosphorylation state of AMP-activated protein kinase (AMPK) in MDCK-C7 cells

Charity Nofziger1, Kameljit Kalsi, T Aaron West

  • 1Department of Biology, Indiana University, Indianapolis, USA.

Cellular Physiology and Biochemistry : International Journal of Experimental Cellular Physiology, Biochemistry, and Pharmacology
|December 18, 2008
PubMed

Insights

AMP-activated protein kinase (AMPK) regulates cellular metabolism and ion transport. This study shows vasopressin dephosphorylates AMPK independently of PKA, and AICAR inhibits ion transport in MDCK-C7 cells.

Area of Science:

  • Cellular Biology
  • Physiology
  • Molecular Biology

Background:

  • AMP-activated protein kinase (AMPK) links cellular metabolism to ion transport.
  • Madin-Darby Canine Kidney-Clone 7 (MDCK-C7) cells mimic renal principal cells, expressing key ion channels and regulators.
  • [Arg(8)]-vasopressin stimulates cAMP production, activating protein kinase A (PKA) and increasing Cl(-) and Na(+) transport.

Purpose of the Study:

  • To investigate the role of AMPK in vasopressin-stimulated ion transport in MDCK-C7 cells.
  • To determine the relationship between vasopressin, PKA, AMPK phosphorylation, and ion transport.
  • To elucidate the mechanism by which AICAR affects vasopressin-induced cellular responses.

Main Methods:

  • Ussing-style electrophysiology to measure ion transport.
  • Inhibition of PKA with H89.
  • Treatment with AICAR, an AMPK activator.
  • Immunoblotting to assess AMPK phosphorylation status (pAMPK(thr172)).

Main Results:

  • PKA inhibition blocked vasopressin-stimulated Cl(-) and Na(+) transport.
  • [Arg(8)]-vasopressin caused pAMPK(thr172) dephosphorylation, independent of PKA.
  • AICAR blocked vasopressin-stimulated currents after 24-hour incubation.
  • AICAR did not increase pAMPK(thr172) levels but inhibited vasopressin-stimulated cAMP production.
  • Vasopressin induced pAMPK(thr172) dephosphorylation even in the presence of AICAR.

Conclusions:

  • Vasopressin regulates AMPK phosphorylation in MDCK-C7 cells through a PKA-independent pathway.
  • AICAR inhibits ion transport independently of AMPK activation in these cells.
  • The findings suggest complex crosstalk between vasopressin signaling, cAMP, AMPK, and ion transport in renal cells.

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