Localization of the Na+/H+ exchanger isoform NHE-3 in rabbit and canine kidney

The distribution and subcellular localization of Na+/H+ exchanger isoform NHE-3 was studied in rabbit and canine kidney using polyclonal antibodies to an NHE-3 fusion protein. Western blot analyses were performed against microsomal, brush-border, and basolateral membranes isolated from rabbit kidney cortex, outer medulla, and inner medulla. Immunoblots indicated that NHE-3 antibody recognized a strong band with 95-100 kDa molecular mass in cortical microsomes. Subcellular localization studies showed that NHE-3 was expressed in brush-border membranes of kidney cortex. Expression of NHE-3 in the medullary regions was studied by immunoblot analysis of NHE-3 antibody against the microsomal membranes from the outer and inner medulla. NHE-3 antibody specifically labelled a 95-100 kDa protein in outer but not inner medulla. Subcellular localization studies demonstrated that NHE-3 is localized to the brush-border membranes of the outer medulla. Immunoblot analysis against brush-border membranes from canine kidney cortex and outer medulla demonstrated the presence of an 83-90 kDa protein. The above experiments suggest that NHE-3 in rabbit kidney is a 95-100 kDa protein and is expressed in brush-border membranes of the cortex and outer medulla. The canine kidney NHE-3 is a 83-90 kDa protein and is expressed in brush-border membranes of the cortex and outer medulla. Based on its subcellular localization, we conclude that NHE-3 may be involved in vectorial Na+ and HCO3- transport and pHo regulation.