Journal article
Harvest and primary culture of the murine aldosterone-sensitive distal nephron
American journal of physiology. Renal physiology, Vol.308(11), pp.F1306-F1315
06/01/2015
DOI: 10.1152/ajprenal.00668.2014
PMCID: PMC4451330
PMID: 25810438
Abstract
The aldosterone-sensitive distal nephron (ASDN) exhibits axial heterogeneity in structure and function from the distal convoluted tubule to the medullary collecting duct. Ion and water transport is primarily divided between the cortex and medulla of the ASDN, respectively. Transcellular transport in this segment is highly regulated in health and disease and is integrated across different cell types. We currently lack an inexpensive, high-yield, and tractable technique to harvest and culture cells for the study of gene expression and physiological properties of mouse cortical ASDN. To address this need, we harvested tubules bound to Dolichos biflorus agglutinin lectin-coated magnetic beads from the kidney cortex and characterized these cell preparations. We determined that these cells are enriched for markers of distal convoluted tubule, connecting tubule, and cortical collecting duct, including principal and intercalated cells. In primary culture, these cells develop polarized monolayers with high resistance (1,000-1,500 Omega* cm(2)) and maintain expression and activity of key channels. These cells demonstrate an amiloride-sensitive short-circuit current that can be enhanced with aldosterone and maintain measurable potassium and anion secretion. Our method can be easily adopted to study the biology of the ASDN and to investigate phenotypic differences between wild-type and transgenic mouse models.
Details
- Title: Subtitle
- Harvest and primary culture of the murine aldosterone-sensitive distal nephron
- Creators
- Mariana Labarca - Stanford UniversityJonathan M. Nizar - Stanford UniversityElisabeth M. Walczak - Stanford UniversityWuxing Dong - Stanford UniversityAlan C. Pao - Stanford UniversityVivek Bhalla - Stanford University
- Resource Type
- Journal article
- Publication Details
- American journal of physiology. Renal physiology, Vol.308(11), pp.F1306-F1315
- Publisher
- Amer Physiological Soc
- DOI
- 10.1152/ajprenal.00668.2014
- PMID
- 25810438
- PMCID
- PMC4451330
- ISSN
- 1931-857X
- eISSN
- 1522-1466
- Number of pages
- 10
- Grant note
- T32DK007357; R03DK083613; R01DK091565 / NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Institute of Diabetes & Digestive & Kidney Diseases (NIDDK) T32 DK007357; 5R03 DK083613; 1R01 DK091565 / NIH; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA Child Health Research Institute at Stanford University Becas Chile scholarship American Society of Nephrology Tashia and John Morgridge Endowed Postdoctoral Fellowship
- Language
- English
- Date published
- 06/01/2015
- Academic Unit
- Nephrology; Internal Medicine
- Record Identifier
- 9984359804002771
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