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AE2 isoforms in rat kidney: immunohistochemical localization and regulation in response to chronic NH₄Cl loading

¹The Water and Salt Research Center, University of Aarhus, DK-8000 Aarhus C; ²Institute of Anatomy, University of Aarhus, DK-8000 Aarhus C, Denmark; ³Department of Medicine, Harvard Medical School, Molecular and Vascular Medicine Unit and Renal Unit, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02215; and ⁴Emory University School of Medicine, Renal Division, Atlanta, Georgia 30322

Submitted 21 November 2003 ; accepted in final form 26 January 2004

Three splice variants of anion exchanger (AE)2 (AE2a, b, and c) have been described in the rat, but their relative distribution in rat kidney is not known. The purpose of this study was to describe the segmental and cellular distribution of the AE2 isoforms in the rat kidney and to evaluate whether the expression levels of these AE2 isoforms are regulated independently in response to chronic NH₄Cl loading. Two polyclonal antibodies were generated, respectively, recognizing a NH₂-terminal peptide unique to AE2a and an amino acid sequence common to AE2a and AE2b. Antibody specificities were tested using cells transfected separately with the AE2a, AE2b, and AE2c isoforms. Immunohistochemistry on sections of paraffin-embedded rat kidneys showed a distribution of AE2a/AE2b labeling in the kidney similar to the distribution of AE2 in the rat kidney reported previously. AE2 is highly expressed in the medullary thick ascending limb, cortical thick ascending limb (cTAL), and macula densa. The pattern of AE2a-specific labeling differed from the pattern of AE2a/AE2b labeling in that relatively more of the total immunolabel was observed in the terminal inner medullary collecting duct. NH₄Cl loading (0.033 mmol NH₄Cl/g body wt for 7 days) did not change the labeling of AE2 isoforms in the medulla, whereas the labeling in the cortex was intensified and included more distal parts of the cTAL. Immunoblotting confirmed upregulation of AE2a/b expression in the cortex. These results indicate that AE2a and AE2b are differentially expressed and regulated in the rat kidney. The regulation following NH₄Cl loading of AE2b in the cTAL suggests a role for AE2 in transepithelial bicarbonate reabsorption in this segment.

immunohistochemistry; collecting duct; nephron segments

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