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Abstract: FR-PO535

Neuropeptide FF Increases Na+/K+-ATPase Activity in Live Renal Proximal Tubule Cells

Session Information

Category: Fluid, Electrolytes, and Acid-Base Disorders

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

Authors

  • Lee, Hewang, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Polzin, Jacob Quentin Mullins, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Amatya, Bibhas, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Asico, Laureano D., The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Armando, Ines, The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
  • Felder, Robin Allen, UVA Health, Charlottesville, Virginia, United States
  • Jose, Pedro A., The George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, United States
Background

Neuropeptide FF (NPFF), an amidated peptide, acts as a pain-modulator. However, the effects of NPFF on renal Na+ transport and blood pressure are unknown.

Methods

Intracellular Na+ concentration in renal proximal tubule cells (RPTCs) was measured using the lifetime Na+-binding of NaTRIUM Green-2 (Green-2), monitored by fluorescence lifetime imaging (FLIM) and Na+ green tetraacetate, monitored by spectrometry. Blood pressure and sodium excretion were also studied in mice.

Results

In glass bottom-cultured RPTCs, Green-2 (5 µM/1 hr) had a biexponential decay in time-resolved fluorescence measurements in randomly selected regions of interest (ROI) in the cytoplasm of these RPTCs. In the basal state, the lifetime t2 (Na+-binding decay time component, nanoseconds) was 3.44±0.05 (n=8). NPFF (100 nM) sequentially decreased the lifetime t2: 3.44±0.04 (5 min), 3.35±0.07 (10 min), 3.31±0.07 (15 min), and 3.25±0.06 (20 min), analyzed from the ROIs (n=8-20). The decrease in t2, Na+-binding decay time component, due to NPFF was associated with a decrease in intracellular Na+ concentration (NPFF: 79.0±4.7%, n=4 vs vehicle: 100.0±9.4%, n=4). The NPFF-mediated decrease in intracellular Na+ was due to an increase in Na+/K+-ATPase activity, because ouabain (50 μM) which inhibits Na+/K+-ATPase activity increased intracellular Na+ (125.9±6.1%, n=4) and prevented the NPFF-mediated decrease in intracellular Na+ concentration (124.3±4.9%, n=4). The effect of NPFF on other sodium transporters and exchangers was not determined. The stimulatory effect of NPFF on Na+ transport has physiological significance because in anesthetized C57Bl/6 mice (n=4), the acute bilateral renal subcapsular injection of NPFF (10 µg/100 µL) increased systolic blood pressure (SBP, mm Hg), 15 min post injection (138.5±12.8 vs basal, 99.5±1.3, P < 0.05), peaked at 25 min (148.3±8.3), and gradually returned to baseline at 60 min. The chronic bilateral renal subcapsular infusion of NPFF (9.25 mM, 0.5 mL/hr, n=4) for 7 days also increased SBP and decreased urinary sodium excretion that were prevented by RF9, an NPFF antagonist.

Conclusion

NPFF decreased intracellular Na+ concentration in RPTCs by stimulating Na+/K+-ATPase activity, decreased Na+ excretion, and increased blood pressure in C57Bl/6 mice.

Funding

  • NIDDK Support