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Abstract: TH-OR23

Optical Clearing and 3D Imaging Reveal a Sexual Dimorphism in the Structure and Remodeling Response of the Distal Convoluted Tubule

Session Information

Category: Fluid, Electrolyte, and Acid-Base Disorders

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

Authors

  • Tahaei, Seyedmohammadebrahim, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States
  • Coleman, Richard A., Johns Hopkins University School of Medicine, Baltimore, Maryland, United States
  • Saritas, Turgay, University Hospital RWTH, Aakhen, Germany
  • Ellison, David H., Oregon Health & Science University Oregon Clinical & Translational Research Institute, Portland, Oregon, United States
  • Welling, Paul A., Johns Hopkins University School of Medicine, Baltimore, Maryland, United States
Background

The abundance of the distal convoluted tubule (DCT) thiazide-sensitive sodium chloride cotransporter (NCC) is greater in females than males. Because structural remodeling of the DCT is dependent on NCC activity, it has been generally assumed that there is a corresponding sexual dimorphism in DCT morphology. Until now, this has never been directly examined. Here, combining new optical clearing techniques with staining for markers of DCT segments, volumetric imaging we quantitatively assess DCT morphology in male and female mice and study DCT remodeling response to furosemide.

Methods

Male and Female (3-month-old) mice were treated with vehicle or Furosemide administered in the food (100 mg/kg per day) for seven days. Total NCC and phospho-active NCC abundance (pNCC) was evaluated by Western Blot in one kidney, the other kidney was perfused and fixed for imaging. Kidneys were cleared using the Clear, Unobstructed Brain/Body Imaging Cocktails and Computational Analysis (CUBIC) pipeline, co-stained with antibodies that label the early DCT (DCT1, parvalbumin) and the entire DCT (DCT1 & 2; NCC), imaged using a high-speed spinning disc confocal microscope, and processed with 3D rendering and analysis software (IMARIS).

Results

We confirmed previous studies that females have greater NCC abundance in the basal state. Surprisingly, the length of the DCT was longer in males (~620 µm) than female mice (~560 µm). Furosemide treatment significantly increased the abundance of NCC and pNCC in both sexes, and this was paralleled by an expansion of the DCT length and volume. The remodeling response to furosemide was more profound in females (~20% increase in DCT length and 50% in DCT1 volume) than males (~8% and 30%, respectively). The DCT elongation effect of furosemide treatment in females stemmed largely from an increase in DCT1 length. Furosemide expanded the DCT2 significantly in males but not females.

Conclusion

Our study reveals a surprising sexual dimorphism of the DCT. The greater NCC density in a shorter structure may provide a means for females to protect sodium balance in face of greater basal distal sodium delivery, yet have larger reserve and remodeling capacity to adapt to unique physiological stresses.

Funding

  • NIDDK Support