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CFTR functions as a bicarbonate channel in pancreatic duct cells.

Ishiguro H, Steward MC, Naruse S, Ko SB, Goto H, Case RM, Kondo T, Yamamoto A - J. Gen. Physiol. (2009)

Bottom Line: Apical HCO(3)(-) fluxes activated by cyclic AMP were independent of Cl(-) and luminal Na(+), and substantially inhibited by the CFTR blocker, CFTR(inh)-172.From the changes in pH(i), membrane potential, and buffering capacity, the flux and electrochemical gradient of HCO(3)(-) across the apical membrane were determined and used to calculate the HCO(3)(-) permeability.This suggests that CFTR functions as a HCO(3)(-) channel in pancreatic duct cells, and that it provides a significant pathway for HCO(3)(-) transport across the apical membrane.

View Article: PubMed Central - PubMed

Affiliation: Human Nutrition, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan. ishiguro@htc.nagoya-u.ac.jp

ABSTRACT
Pancreatic duct epithelium secretes a HCO(3)(-)-rich fluid by a mechanism dependent on cystic fibrosis transmembrane conductance regulator (CFTR) in the apical membrane. However, the exact role of CFTR remains unclear. One possibility is that the HCO(3)(-) permeability of CFTR provides a pathway for apical HCO(3)(-) efflux during maximal secretion. We have therefore attempted to measure electrodiffusive fluxes of HCO(3)(-) induced by changes in membrane potential across the apical membrane of interlobular ducts isolated from the guinea pig pancreas. This was done by recording the changes in intracellular pH (pH(i)) that occurred in luminally perfused ducts when membrane potential was altered by manipulation of bath K(+) concentration. Apical HCO(3)(-) fluxes activated by cyclic AMP were independent of Cl(-) and luminal Na(+), and substantially inhibited by the CFTR blocker, CFTR(inh)-172. Furthermore, comparable HCO(3)(-) fluxes observed in ducts isolated from wild-type mice were absent in ducts from cystic fibrosis (Delta F) mice. To estimate the HCO(3)(-) permeability of the apical membrane under physiological conditions, guinea pig ducts were luminally perfused with a solution containing 125 mM HCO(3)(-) and 24 mM Cl(-) in the presence of 5% CO(2). From the changes in pH(i), membrane potential, and buffering capacity, the flux and electrochemical gradient of HCO(3)(-) across the apical membrane were determined and used to calculate the HCO(3)(-) permeability. Our estimate of approximately 0.1 microm sec(-1) for the apical HCO(3)(-) permeability of guinea pig duct cells under these conditions is close to the value required to account for observed rates of HCO(3)(-) secretion. This suggests that CFTR functions as a HCO(3)(-) channel in pancreatic duct cells, and that it provides a significant pathway for HCO(3)(-) transport across the apical membrane.

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Estimation of apical HCO3− permeability. (A) Calculation of the rate of change of pHi by linear regression at 1-min intervals after step changes in [K+]B. The fitted lines are overlaid on data from the second half of the experiment shown in Fig. 5 D. Together with the midpoint value of pHi and the intracellular buffering capacity, these data were used to calculate the HCO3− flux across the apical membrane. (B) Distribution of calculated HCO3− permeabilities for HCO3− influx (n = 18 from 5 ducts) and HCO3− efflux (n = 18 from 5 ducts) pooled from experiments like the one shown in A. The horizontal dotted lines indicate the mean values. (C) Predicted changes in pHi resulting from changes in Va induced by switching [K+]B from 1 to 70 mM, and then back to 1 mM. Theoretical curves based on three alternative apical HCO3− permeability values (0.05, 0.1, and 0.2 µm sec−1) are superimposed on averaged data from four experiments of the type shown in Fig. 5 D. The dotted lines indicate the mean ± SEM. The top panel shows the predicted changes in Va, assuming a time constant of 45 s.
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fig6: Estimation of apical HCO3− permeability. (A) Calculation of the rate of change of pHi by linear regression at 1-min intervals after step changes in [K+]B. The fitted lines are overlaid on data from the second half of the experiment shown in Fig. 5 D. Together with the midpoint value of pHi and the intracellular buffering capacity, these data were used to calculate the HCO3− flux across the apical membrane. (B) Distribution of calculated HCO3− permeabilities for HCO3− influx (n = 18 from 5 ducts) and HCO3− efflux (n = 18 from 5 ducts) pooled from experiments like the one shown in A. The horizontal dotted lines indicate the mean values. (C) Predicted changes in pHi resulting from changes in Va induced by switching [K+]B from 1 to 70 mM, and then back to 1 mM. Theoretical curves based on three alternative apical HCO3− permeability values (0.05, 0.1, and 0.2 µm sec−1) are superimposed on averaged data from four experiments of the type shown in Fig. 5 D. The dotted lines indicate the mean ± SEM. The top panel shows the predicted changes in Va, assuming a time constant of 45 s.

Mentions: The rate of change of pHi (dpHi/dt) during these transitions was calculated by linear regression at 1-min intervals (Fig. 6 A). At each time point, the HCO3− flux across the apical membrane, JHCO3, was calculated from dpHi/dt using the following expression:(1)where βt is the total intracellular buffering capacity (values from Szalmay et al., 2001), and h is the volume of the epithelium per unit area. Assuming that the volume of the lateral intercellular spaces is negligible, the epithelial volume per unit area is equivalent to the cell height, which was estimated to be 10 µm (Argent et al., 1986; Arkle et al., 1986).


CFTR functions as a bicarbonate channel in pancreatic duct cells.

Ishiguro H, Steward MC, Naruse S, Ko SB, Goto H, Case RM, Kondo T, Yamamoto A - J. Gen. Physiol. (2009)

Estimation of apical HCO3− permeability. (A) Calculation of the rate of change of pHi by linear regression at 1-min intervals after step changes in [K+]B. The fitted lines are overlaid on data from the second half of the experiment shown in Fig. 5 D. Together with the midpoint value of pHi and the intracellular buffering capacity, these data were used to calculate the HCO3− flux across the apical membrane. (B) Distribution of calculated HCO3− permeabilities for HCO3− influx (n = 18 from 5 ducts) and HCO3− efflux (n = 18 from 5 ducts) pooled from experiments like the one shown in A. The horizontal dotted lines indicate the mean values. (C) Predicted changes in pHi resulting from changes in Va induced by switching [K+]B from 1 to 70 mM, and then back to 1 mM. Theoretical curves based on three alternative apical HCO3− permeability values (0.05, 0.1, and 0.2 µm sec−1) are superimposed on averaged data from four experiments of the type shown in Fig. 5 D. The dotted lines indicate the mean ± SEM. The top panel shows the predicted changes in Va, assuming a time constant of 45 s.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2654087&req=5

fig6: Estimation of apical HCO3− permeability. (A) Calculation of the rate of change of pHi by linear regression at 1-min intervals after step changes in [K+]B. The fitted lines are overlaid on data from the second half of the experiment shown in Fig. 5 D. Together with the midpoint value of pHi and the intracellular buffering capacity, these data were used to calculate the HCO3− flux across the apical membrane. (B) Distribution of calculated HCO3− permeabilities for HCO3− influx (n = 18 from 5 ducts) and HCO3− efflux (n = 18 from 5 ducts) pooled from experiments like the one shown in A. The horizontal dotted lines indicate the mean values. (C) Predicted changes in pHi resulting from changes in Va induced by switching [K+]B from 1 to 70 mM, and then back to 1 mM. Theoretical curves based on three alternative apical HCO3− permeability values (0.05, 0.1, and 0.2 µm sec−1) are superimposed on averaged data from four experiments of the type shown in Fig. 5 D. The dotted lines indicate the mean ± SEM. The top panel shows the predicted changes in Va, assuming a time constant of 45 s.
Mentions: The rate of change of pHi (dpHi/dt) during these transitions was calculated by linear regression at 1-min intervals (Fig. 6 A). At each time point, the HCO3− flux across the apical membrane, JHCO3, was calculated from dpHi/dt using the following expression:(1)where βt is the total intracellular buffering capacity (values from Szalmay et al., 2001), and h is the volume of the epithelium per unit area. Assuming that the volume of the lateral intercellular spaces is negligible, the epithelial volume per unit area is equivalent to the cell height, which was estimated to be 10 µm (Argent et al., 1986; Arkle et al., 1986).

Bottom Line: Apical HCO(3)(-) fluxes activated by cyclic AMP were independent of Cl(-) and luminal Na(+), and substantially inhibited by the CFTR blocker, CFTR(inh)-172.From the changes in pH(i), membrane potential, and buffering capacity, the flux and electrochemical gradient of HCO(3)(-) across the apical membrane were determined and used to calculate the HCO(3)(-) permeability.This suggests that CFTR functions as a HCO(3)(-) channel in pancreatic duct cells, and that it provides a significant pathway for HCO(3)(-) transport across the apical membrane.

View Article: PubMed Central - PubMed

Affiliation: Human Nutrition, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan. ishiguro@htc.nagoya-u.ac.jp

ABSTRACT
Pancreatic duct epithelium secretes a HCO(3)(-)-rich fluid by a mechanism dependent on cystic fibrosis transmembrane conductance regulator (CFTR) in the apical membrane. However, the exact role of CFTR remains unclear. One possibility is that the HCO(3)(-) permeability of CFTR provides a pathway for apical HCO(3)(-) efflux during maximal secretion. We have therefore attempted to measure electrodiffusive fluxes of HCO(3)(-) induced by changes in membrane potential across the apical membrane of interlobular ducts isolated from the guinea pig pancreas. This was done by recording the changes in intracellular pH (pH(i)) that occurred in luminally perfused ducts when membrane potential was altered by manipulation of bath K(+) concentration. Apical HCO(3)(-) fluxes activated by cyclic AMP were independent of Cl(-) and luminal Na(+), and substantially inhibited by the CFTR blocker, CFTR(inh)-172. Furthermore, comparable HCO(3)(-) fluxes observed in ducts isolated from wild-type mice were absent in ducts from cystic fibrosis (Delta F) mice. To estimate the HCO(3)(-) permeability of the apical membrane under physiological conditions, guinea pig ducts were luminally perfused with a solution containing 125 mM HCO(3)(-) and 24 mM Cl(-) in the presence of 5% CO(2). From the changes in pH(i), membrane potential, and buffering capacity, the flux and electrochemical gradient of HCO(3)(-) across the apical membrane were determined and used to calculate the HCO(3)(-) permeability. Our estimate of approximately 0.1 microm sec(-1) for the apical HCO(3)(-) permeability of guinea pig duct cells under these conditions is close to the value required to account for observed rates of HCO(3)(-) secretion. This suggests that CFTR functions as a HCO(3)(-) channel in pancreatic duct cells, and that it provides a significant pathway for HCO(3)(-) transport across the apical membrane.

Show MeSH
Related in: MedlinePlus