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Abstract: TH-PO505

Lithium and Gsk3α Inhibition Reduces Aquaporin-2 Expression in Primary Cultured Inner Medullary Collecting Duct Cells Due to Independent Mechanisms

Session Information

Category: Fluid and Electrolytes

  • 901 Fluid and Electrolytes: Basic

Authors

  • Edemir, Bayram, Martin Luther University Halle-Wittenberg, Halle (Saale), Germany
  • Kaiser, Marc, Praxis am Markt, Froendenberg, Werl, Germany
Background

Lithium is a widely used drug for the treatment of bipolar disorders. However, it has severe side effects and up to 40 % of the treated patients develop a diabetes insipidus. Lithium affects also the activity of the glycogen synthase kinase 3 (Gsk3) and mice deficient for Gsk3β isoform in the renal collecting duct showed severe urine concentration abilities. The respective cellular and molecular mechanisms are still not fully understood.

Methods

We used primary cultured inner medullary collecting duct cells to analyze the underlying mechanisms. The cells were treated for different time points and with different concentrations of lithium, the pharmacological inhibitor of the Gsk3α/β (SB216763) or the Gsk3β specific Inhibitor TWS119. The cells were additionally incubated with inhibitors of the lysosomal (using bafilomycin) or proteasomal pathways (using MG132) alone or together with the above mentioned drugs. The expression of Aqp2 was analyzed on the mRNA level by qPCR and on the protein level by Western blot and immunofluorescence analysis.

Results

The qPCR experiments showed that only lithium induced a down regulation of Aqp2 mRNA expression while Gsk3 inhibition by SB216763 had no effect and surprisingly TWS119 led to increased expression. While inhibition of the proteasome with MG132 did not prevent the lithium or SB216763 mediated down regulation of Aqp2, the inhibition of the lysosomal activity with bafilomycin or chloroquine prevented lithium and SB216763 mediated down regulation of Aqp2 on the protein level. On the mRNA level lithium still induced down regulation of Aqp2. The treatment with bafilomycin and chloroquine induced accumulation of Aqp2 in lysosomal structures, which was prevented when the cells were treated with dbcAMP which led to phosphorylation and membrane localization of Aqp2. While down regulation of Aqp2 was also evident when lithium was applied together with dbcAMP, dbcAMP prevented the SB216763 induced down regulation of Aqp2. Interestingly the use of TWS119 induced the expression of Aqp2 on protein and mRNA level.

Conclusion

We showed that lithium and the Gsk3α/β inhibition by SB216763 induces down regulation of Aqp2 on different levels. The exclusive Gsk3β inhibitor TWS119 had the opposite effect indicating that Gsk3α inhibition is involved in Aqp2 down regulation.

Funding

  • Government Support - Non-U.S.