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Na/K-ATPase in intercalated cells along the rat nephron revealed by antigen retrieval

The Na/K-ATPase plays a fundamental role in the physiology of various mammalian cells. In the kidney, previous immunocytochemical studies have localized this protein to the basolateral membrane in different tubule segments. However, intercalated cells (IC) of the collecting duct (CD) in rat and mouse were unlabeled with anti-Na/K-ATPase antibodies. We have recently described an antigen retrieval technique in which tissue sections are pre-treated with sodium dodecyl sulfate (SDS) prior to immunostaining. This procedure was used to re-examine the presence of Na/K-ATPase in IC along the rat nephron using a monoclonal antibody against Na/K-ATPase holoprotein. Subtypes of IC along the nephron were identified by their distinctive staining with polyclonal and monoclonal antibodies to the 31 kDa vacuolar H+-ATPase subunit, whereas principal cells (PC) were labeled with a polyclonal antibody to the water channel aquaporin-4 (AQP-4). In PC, the Na/K-ATPase and AQP-4 staining co-localized basolaterally. In contrast to previous reports, we found that IC of all types showed basolateral labeling with the anti-Na/K-ATPase antibody. The staining was quantified by fluorescence image analysis: it was weak to moderate in IC of cortical and outer-medullary collecting ducts and most intense in IC of inner-medullary collecting ducts. IC in the inner medulla showed a staining intensity that was equivalent or stronger to that in adjacent principal cells. Models of ion transport at the cellular and epithelial level in rat kidney must, therefore, take into account the potential role of a basolateral Na/K-ATPase in intercalated cell function.

sodium/potassium-ATPase; collecting duct; basolateral membrane; nephron; rat

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