Abstract: FR-OR130

RNA-Seq in Microdissected Rat Cortical Collecting Ducts during Development of Lithium-Induced Nephrogenic Diabetes Insipidus

Session Information

Category: Fluid, Electrolytes, and Acid-Base

  • 702 Water/Urea/Vasopressin, Organic Solutes

Authors

  • Sung, Chih-chien, NHLBI/NIH, Bethesda, United States
  • Chou, Chung-Lin, NHLBI/NIH, Bethesda, Maryland, United States
  • Chen, Lihe, NHLBI/NIH, Bethesda, Maryland, United States
  • Jung, Hyun Jun, NHLBI/NIH, Bethesda, Maryland, United States
  • Knepper, Mark A., NHLBI/NIH, Bethesda, Maryland, United States
Background

Lithium has been widely used to treat bipolar disorder, but many patients develop nephrogenic diabetes insipidus (NDI). Our previous studies in rats have demonstrated rapid activation of ERK1/ERK2 in collecting ducts after oral lithium administration (Trepiccione et al., KI, 2014). Associated gene expression changes early in the development of lithium-induced NDI have been incompletely explored.

Methods

Cortical collecting ducts (CCDs) were microdissected from rats 72 hrs after beginning treatment with lithium (lithium chloride, 40 mmol/kg of dry food) versus time controls. Single-tubule RNA-Seq was carried out independently in 3-5 CCD samples per rat (4 lithium-treated rats versus 4 controls) using the methods of Lee et al. (JASN, 2015). Cortical thick ascending limbs (cTALs) were also microdissected for RNA-Seq at 72 hrs.

Results

Lithium-treated rats showed increased water intake within 24 hrs, concomitant with a fall in AQP2 protein abundance (western blotting). RNA-Seq data at 72 hr revealed moderate decreases in AQP2, AQP3, and AQP4 mRNA. The three subunits of ENaC showed more profound decreases. A large number of transcripts coding for other transporters and receptors were decreased with lithium treatment including the vasopressin V2 receptor and the chloride channel ClC-Kb. Several known aldosterone-regulated genes showed decreases in mRNA including Sgk1, αENaC, and Gadd45g. Immediate early genes (typical of MAP kinase activation) and transcriptional targets of NF-κB were significantly more frequent among transcripts increased with lithium than among all expressed genes (Chi-square). Similar over-representation among lithium-affected transcripts were seen for two other pathways, e.g. cell cycle signaling, and Wnt signaling. No association with lithium treatment was found for several pathways including cyclic AMP signaling, estrogen receptor signaling and insulin signaling. In contrast to the CCD, no significant changes were found in mRNA levels in cTALs in response to lithium, indicating the effect of lithium was selective for CCD.

Conclusion

Cellular signaling during development of lithium-induced NDI is consistent with known responses to increased MAP kinase activation and includes major gene expression changes characteristic of NF-κB-dependent inflammatory signaling.

Funding

  • Other NIH Support