Epithelial Systems Biology Laboratory, NHLBI, NIH

Isolated Perfused Tubule Technique

The Epithelial Systems Biology Laboratory investigates mechanisms responsible for regulation of transport across the epithelia of nephrons and collecting ducts in the kidney. Although the laboratory employs a wide variety of modern, large-scale data acquisition techniques for these studies, functional data remain of prime importance. Invented by Maurice Burg in the 1960's (1), the isolated perfused tubule technique continues to be utilized in the laboratory to provide a functional readout. Tubule segments, typically consisting of 200-400 cells, can be microdissected from rat or mouse kidneys and mounted on miniature glass micropipettes to allow measurement of transepithelial transport rates for water and solutes (2). The isolated perfused tubule technique has been particularly useful in studies of various knockout mouse models (3-5). Microdissected tubules can also be used for a variety of other experimental measurements (6) including, most recently, identification of complete transcriptomes for all 14 renal tubule segments using RNA-seq (7). Click here for blueprints for isolated perfused tubule apparatus.

References:

1. Burg M, Grantham J, Abramow M, Orloff J. Preparation and study of fragments of single rabbit nephrons. Am J Physiol. 1966; 210: 1293-8. PMID: 5923067.

2. Burg MB, Knepper MA. Single tubule perfusion techniques. Kidney Int. 1986; 30: 166-70. PMID: 3761862.

3. Chou CL, Ma T, Yang B, Knepper MA, Verkman AS. Fourfold reduction of water permeability in inner medullary collecting duct of aquaporin-4 knockout mice. Am J Physiol. 1998; 274: C549-54. PMID: 9486146.

4. Fenton RA, Chou CL, Stewart GS, Smith CP, Knepper MA. Urinary concentrating defect in mice with selective deletion of phloretin-sensitive urea transporters in the renal collecting duct. Proc Natl Acad Sci U S A. 2004; 101: 7469-74. PMID: 15123796.

5. Li JH, Chou CL, Li B, Gavrilova O, Eisner C, Schnermann J, Anderson SA, Deng CX, Knepper MA, Wess J. A selective EP4 PGE2 receptor agonist alleviates disease in a new mouse model of X-linked nephrogenic diabetes insipidus. J Clin Invest. 2009; 119: 3115-26. PMID: 19729836.

6. Wright PA, Burg MB, Knepper MA. Microdissection of kidney tubule segments. Methods Enzymol. 1990; 191: 226-31. PMID: 2074757.

7. Lee JW, Chou CL, Knepper MA. Deep sequencing in microdissected renal tubules identifies nephron segment-specific transcriptomes. J Am Soc Nephrol. 2015 Mar 27. [Epub ahead of print] PMID: 25817355.