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Abstract: FR-PO540

Estimation of Plasma Vasopressin Activity by AQP2 in Urinary Extracellular Vesicles

Session Information

Category: Fluid, Electrolytes, and Acid-Base Disorders

  • 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic

Authors

  • Kawaguchi, Tamami, Laboratory of Veterinary Pharmacology, Faculty of Agriculture, University of Miyazaki, Miyazaki, Japan
  • Sonoda, Hiroko, Laboratory of Veterinary Pharmacology, Faculty of Agriculture, University of Miyazaki, Miyazaki, Japan
  • Ikeda, Masahiro, Laboratory of Veterinary Pharmacology, Faculty of Agriculture, University of Miyazaki, Miyazaki, Japan
Background

The antidiuretic hormone, vasopressin (AVP), activates protein kinase A (PKA) in the principal cells of the collecting duct via the V2 receptor. Activated PKA is known to phosphorylate the amino acid residues (S256 and S269) on the water channel, aquaporin 2 (AQP2). These phosphorylations are considered to promote water reabsorption by increasing the trafficking of AQP2 into the apical membrane. The measurement of the blood AVP concentration is difficult because of its instability and binding to the platelets. It has been shown that urinary extracellular vesicles (uEVs) contain AQP2 protein. Therefore, in this study, we investigated whether AVP activity could be inferred by measuring the total or phosphorylated form of AQP2 in uEVs.

Methods

In experiment I, male SD rats were divided into three groups: a control group (free-drinking tap water), a dehydration group (water deprivation; DH group), and a hydration group (free-drinking 20% sucrose; HY group). In experiment II, male SD rats were divided into two groups: a vehicle group (50% of 5% glucose, 50% of saline, s.c.) and a dDAVP group (300 ng/kg desmopressin, V2 agonist, s.c.). Urine samples were collected for 12-24 hrs, and blood and kidney samples were collected at 24 hrs after the treatment. Total and phosphorylated (pS256, pS269) AQP2 protein levels in uEVs were evaluated by immunoblot analysis.

Results

In experiment I, urine volume was decreased in the DH group and was increased in the HY group. Moreover, urine osmolality was elevated in the DH group and reduced in the HY group. Total and phosphorylated AQP2 levels in uEVs were increased in the DH group, and only the pS269-AQP2 level was decreased in the HY group. In experiment II, urine osmolality in the dDAVP group significantly increased in comparison with the control group. The total AQP2 level showed an increasing tendency, and the pS269-AQP2 level was significantly increased in the dDAVP group. On the other hand, the pS256-AQP2 level was not altered.

Conclusion

These results indicate that the pS269-AQP2 level in uEVs responded well to the physiological and pharmacological changes in V2 receptor activation. Therefore, pS269-AQP2 in uEVs may be a potential non-invasive biomarker to estimate plasma AVP activity.