The interlobular ducts of guinea-pig pancreas secrete HCO₃^– into a HCO₃^–-rich (125 mM) luminal fluid following secretin stimulation (Ishiguro et al. 1998). Under these conditions intracellular HCO₃^– ([HCO₃^–]_i) is ~20 mM and intracellular potential is ~-60 mV (Ishiguro et al. 2000, 2002) indicating that there is a luminally directed electrochemical gradient for HCO₃^–. To examine whether luminal HCO₃^– transport is mediated by an anion conductance, we measured changes in intracellular pH (pH_i) when the cells were de- or hyperpolarized by manipulation of extracellular K⁺.

Female Hartley guinea-pigs (350-450 g) were humanely killed by cervical dislocation and the interlobular ducts isolated from the pancreas by collagenase digestion and microdissection (Ishiguro et al. 1998). pH_i was measured with 2â,7â-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). The ducts were superfused with HCO₃^–/CO₂-free Hepes-buffered solution (140 mM Cl^–) and luminally perfused with 125 mM HCO₃^–, 24 mM Cl^–, and 5 % CO₂. Dihydro-4,4â-diisothiocyanatostilbene-2,2â-disulphonic acid (H₂DIDS, 500 µM) was used to inhibit HCO₃^– efflux across the basolateral membrane. Changes in the bath K⁺ concentration ([K⁺]_B) were achieved by replacement with N-methyl-D-glucamine and [Na⁺]_B was fixed at 60 mM. Tests for statistical significant differences were made with Student’s t test for paired data.

When [K⁺]_B was raised from 5 to 70 mM to depolarize the cells, pH_i in the unstimulated ducts changed only slightly. In the presence of dibutyryl cAMP, which stimulates HCO₃^– secretion, depolarization caused a large increase in pH_i from 6.83 ± 0.11 to 7.32 ± 0.09 (means ± S.E.M., n = 4, P < 0.01). When [K⁺]_B was reduced from 5 to 1 mM to hyperpolarize the cells, pH_i decreased by 0.11 ± 0.01 (P < 0.05). To deplete intracellular Cl^–, the ducts were perfused with Cl^–-free solutions (containing glucuronate) and stimulated with dibutyryl cAMP for 30 min. Under Cl^– free conditions, when [K⁺]_B was reduced from 5 to 1 mM and then raised to 70 mM, pH_i decreased from 7.15 ± 0.06 (n = 4) to 7.06 ± 0.07 (P < 0.05) and then increased to 7.54 ± 0.16 (P < 0.01).

In summary, de- and hyperpolarization caused changes in pH_i that most probably reflected the influx and efflux of HCO₃^– across the luminal membrane. These HCO₃^– movements were not dependent on the presence of Cl^– and may be attributed to the presence of a significant HCO₃^– conductance at the luminal membrane as we have previously suggested (Ishiguro et al. 2002).

This work was supported by the Japanese Ministry of Education, Science, Technology, Sports and Culture.