Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2002 Jan;109(1):89-99.
doi: 10.1172/JCI13292.

Acid incubation reverses the polarity of intercalated cell transporters, an effect mediated by hensin

George J Schwartz  1 , Shuichi Tsuruoka, Soundarapandian Vijayakumar, Snezana Petrovic, Ayesa Mian, Qais Al-Awqati
Affiliations

Acid incubation reverses the polarity of intercalated cell transporters, an effect mediated by hensin

George J Schwartz et al. J Clin Invest. 2002 Jan.

Abstract

Metabolic acidosis causes a reversal of polarity of HCO(3)(-) flux in the cortical collecting duct (CCD). In CCDs incubated in vitro in acid media, beta-intercalated (HCO(3)(-)-secreting) cells are remodeled to functionally resemble alpha-intercalated (H(+)-secreting) cells. A similar remodeling of beta-intercalated cells, in which the polarity of H(+) pumps and Cl(-)/HCO(3)(-) exchangers is reversed, occurs in cell culture and requires the deposition of polymerized hensin in the ECM. CCDs maintained 3 h at low pH ex vivo display a reversal of HCO(3)(-) flux that is quantitatively similar to an effect previously observed in acid-treated rabbits in vivo. We followed intracellular pH in the same beta-intercalated cells before and after acid incubation and found that apical Cl/HCO(3) exchange was abolished following acid incubation. Some cells also developed basolateral Cl(-)/HCO(3)(-) exchange, indicating a reversal of intercalated cell polarity. This adaptation required intact microtubules and microfilaments, as well as new protein synthesis, and was associated with decreased size of the apical surface of beta-intercalated cells. Addition of anti-hensin antibodies prevented the acid-induced changes in apical and basolateral Cl(-)/HCO(3)(-) exchange observed in the same cells and the corresponding suppression of HCO(3)(-) secretion. Acid loading also promoted hensin deposition in the ECM underneath adapting beta-intercalated cells. Hence, the adaptive conversion of beta-intercalated cells to alpha-intercalated cells during acid incubation depends upon ECM-associated hensin.

PubMed Disclaimer

Figures

Figure 1
Figure 1
Acid incubation converts HCO3– secretion to HCO3– absorption, a process that is hensin-dependent. Upper panel: five CCDs (control serum, basal, white bars) secrete HCO3–. When Cl– was removed (Cl-free lumen), HCO3– absorption resulted; the difference is the rate of Cl–-dependent HCO3– secretion (HCO3 flux). After 3-hour incubation at pH 6.8 (black bars), net HCO3– absorption was observed (basal). Removal of luminal Cl– caused a small increase in HCO3– absorption (Cl–-free lumen), and the Cl–dependent HCO3– secretion (HCO3 flux) was substantially less than before acid incubation. Middle panel: five CCDs (anti-hensin antiserum) were incubated similarly, except anti-hensin Ab was included in the bath. Anti-hensin antiserum (black bars) blocked half of the acid-induced change in HCO3– transport (basal) but did not change the flux after removal of luminal Cl– (Cl-free lumen), so that the Cl–-dependent rate of HCO3– secretion (HCO3 flux) was only slightly smaller than preincubation values. Lower panel: four CCDs (anti-hensin antiserum + fusion protein) were incubated similarly, except that hensin Ab had been preincubated with the fusion protein that had been used as the immunogen. After acid incubation (black bars), the pattern of net HCO3– transport (basal), HCO3– absorption (Cl-free lumen), and HCO3– secretory flux was similar to that observed in the upper panel for control serum. *Significantly different from preincubation (P < 0.05); **significantly different from incubation in control serum (P < 0.05).
Figure 2
Figure 2
Identification of individual intercalated cells and their response to 3-hour incubation. (a) Perfusion of CCDs with BCECF-AM results in selective concentration of fluorescence in intercalated cells. Left tracing shows a β-intercalated cell, which alkalinizes with removal of luminal Cl– and acidifies upon removal of bath Cl–. Right tracing shows an α-intercalated cell, which is unresponsive to the removal of luminal Cl–, but alkalinizes upon the removal of bath Cl–. (b) Representative tracings of pH changes in β-intercalated cells presented to illustrate the variety of responses to Cl– removal under different incubation conditions. Top panel: Incubation at pH 7.4 has no effect on the response of β-intercalated cells to removal of Cl–. Middle panel: Incubation at pH 6.8 causes a loss of apical Cl–/HCO3– exchange activity and prevents acidification with the removal of bath Cl–. Moreover, two of four β-intercalated cells (diamonds and x’s) now alkalinize after basolateral Cl– removal. The responses of all cells are depicted in Figure 4. Lower panel: Anti-hensin Ab added with the pH 6.8 incubation prevented the loss of apical Cl–/HCO3– exchange and permits acidification with bath Cl– removal.
Figure 3
Figure 3
Loss of apical Cl–/HCO3– exchange activity in response to acid incubation is an adaptation that is prevented not only by hensin Ab, but also by cytoskeletal and protein synthesis inhibitors; this adaptation does not depend on changes in cell calcium. Mean (± SE) reversible change in cell pH upon removal of luminal Cl– in β-intercalated cells before incubation (white bars) and after incubation (black bars). *Significantly different from preincubation value.
Figure 4
Figure 4
Acid incubation induces a reversal of polarity of Cl–/HCO3– exchangers, a process that is hensin-dependent and mediated by intact microfilaments and protein synthesis; this adaptation is independent of changes in cell calcium. Histogram of changes in cell pH (pHi) induced by removal of bath Cl– before 3-hour incubation (white bars) and after incubation (black bars). The shaded vertical bar in each graph is a marker for no change in pHi with removal of bath Cl–.
Figure 5
Figure 5
Apical surface (cap length) of β-intercalated cells is diminished by incubation at low pH, in a process that depends on microtubules and protein synthesis. Apical surface of β-intercalated cells was measured from photographs of perfused CCDs labeled with PNA (upper graph) and B63 (lower graph), both specific markers of such cells. *Significantly different from control incubation (pH 7.4). 6.8, incubation at pH 6.8 for 3 hours; colch, colchicine (10 μM); aniso, anisomycin (10 μM).
Figure 6
Figure 6
Confocal immunofluorescent imaging of isolated CCDs stained for extracellular hensin (green) and peanut agglutinin (PNA, red); real magnification was ×100. Upper two panels show a CCD from control and 3-day acid-treated rabbit kidneys; there are three identified intercalated cells from the latter CCD. Lower three panels show successive 1-μm optical cuts for each of these cells.
Figure 7
Figure 7
Confocal immunofluorescent imaging of isolated collecting ducts stained for extracellular hensin (green) and band 3 (AE1, red); real magnification was ×100. Top panel: control CCD, middle panel: 3-day acid-treated CCD; bottom panel: control OMCD.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Kaissling B, Kriz W. Structural analysis of the rabbit kidney. Adv Anat Embryol Cell Biol. 1979; 56:1–121. - PubMed
    1. Madsen KM, Tisher CC. Structural-functional relationships along the distal nephron. Am J Physiol. 1986; 250:F1–F15. - PubMed
    1. Tsuruoka S, Schwartz GJ. Adaptation of rabbit cortical collecting duct HCO3–transport to metabolic acidosis in vitro. J Clin Invest. 1996; 97:1076–1084. - PMC - PubMed
    1. Schwartz GJ, Barasch J, Al-Awqati Q. Plasticity of functional epithelial polarity. Nature. 1985; 318:368–371. - PubMed
    1. Satlin LM, Schwartz GJ. Cellular remodeling of HCO3–-secreting cells in rabbit renal collecting duct in response to an acidic environment. J Cell Biol. 1989; 109:1279–1288. - PMC - PubMed

Publication types

MeSH terms