To characterize the background current in fetal human nasopharyngeal epithelial cells and clarify its relationship with volume activated Cl(-) currents (I(Cl,vol)), whole-cell patch clamp and cell imaging techniques were employed. Under isotonic conditions, a background current [(5.9+/-2.1) pA/pF at +80 mV, n=21] was detected. The current presented a weak outward rectification and a negligible time-dependent inactivation. The current-voltage relationship showed that the reversal potential of the background current [(-0.73+/-1.7) mV, n=21] was close to the calculated equilibrium potential for Cl(-)(-0.9 mV). Application of extracellular hypertonic stimulation (440 mOsmol/L) suppressed the current by (59.6+/-7.1)% and the inhibition was reversible after returned to isotonic conditions. Bathing the cells in hypotonic solution (160 mOsmol/L) induced a volume-sensitive Cl(-) current. The Cl(-) channel blockers, tamoxifen (20 micromol/L) and 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) (100 micromol/L), inhibited the background current by (74.0+/-5.2)% (P<0.01, n=5) and (60.9+/-8.9)% (P<0.01, n=6) at +80 mV and increased basal cell volume by (107.7+/-2.9)% (P<0.01, n=25) and (104.4+/-2.4)% (P<0.01, n=19), respectively. The data indicate that Cl(-) current is an important component of the background current in fetal human nasopharyngeal epithelial cells. The background Cl(-) current is involved in volume activated Cl(-) current and basal cell volume regulation.
Cells, Cultured ; Chloride Channels ; antagonists & inhibitors ; physiology ; Electrophysiology ; Epithelial Cells ; cytology ; metabolism ; physiology ; Fetus ; Humans ; Nasopharynx ; cytology ; Nitrobenzoates ; pharmacology ; Patch-Clamp Techniques ; Tamoxifen ; pharmacology

Whole-cell patch clamp and cell volume measurement techniques were used to investigate the ATP-activated chloride current and the ATP effect on cell volume in nasopharyngeal carcinoma cells. Extracellular application of ATP in micromolar concentrations activated a current with the properties of modest outward rectification and negligible time-dependent inactivation in a dose-dependent manner. The current reversed at a potential [(-0.05+/-0.03) mV] close to the Cl- equilibrium potential (-0.9 mV). Substitution of Cl- with gluconate in the extracellular solution decreased the ATP-activated current and shifted the reversal potential positively. NPPB, one of the chloride channel blockers, inhibited the current by (81.03+/-9.36)%. The current was also depressed by the P2Y purinoceptor antagonist, reactive blue 2, by (67.39+/-5.06)%. ATP (50 micromol/L) decreased the cell volume under the isotonic condition. Depletion of extracellular and intracellular Cl- abolished the ATP effect on cell volume. The results suggest that extracellular ATP of micromolar scales can induce a chloride current associated with cell volume regulation by activation of chloride channel through binding to purinoceptor P2Y.
Adenosine Triphosphate ; physiology ; Cell Size ; drug effects ; Chloride Channels ; antagonists & inhibitors ; metabolism ; physiology ; Humans ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Nitrobenzoates ; pharmacology ; Patch-Clamp Techniques ; Tumor Cells, Cultured

The transwell chamber migration assay and the patch-clamp technique were used to investigate the volume-activated Cl(-) current (I(Cl.vol)) in migrated nasopharyngeal carcinoma cells (CNE-2Z). 47% hypotonic solution activated a ICl.vol in the migrated CNE-2Z cells. Compared with the control cells (non-migrated), the properties of this current and the sensitivity to Cl(-) channel blockers were changed. The current density in migrated CNE-2Z cells was higher than that in non-migrated cells. The current was almost completely inhibited by extracellular application of adenosine-5'-triphosphate (ATP, 10 mmol/L), 5-nitro-2-3-phenylpropylamino benzoic acid (NPPB, 100 mmol/L) and tamoxifen (30 mmol/L) in all voltage steps applied. The inhibition of NPPB and tamoxifen on the current was stronger in migrated cells than that in non-migrated cells. The permeability sequence of the four anions was Br(-)>Cl(-)> I (-)>Gluconate. The sequence was different from that of the non-migrated cells (I(-)> Br(-)> Cl(-)> Gluconate). The results suggest that volume-activated chloride channels may be involved in the CNE-2Z cell migration.
Carcinoma ; drug therapy ; metabolism ; pathology ; Cell Cycle ; drug effects ; physiology ; Cell Division ; drug effects ; Cell Movement ; drug effects ; Cell Size ; drug effects ; Chloride Channels ; antagonists & inhibitors ; metabolism ; physiology ; Chlorides ; metabolism ; Humans ; Nasopharyngeal Neoplasms ; drug therapy ; metabolism ; pathology ; Nitrobenzoates ; pharmacology ; Patch-Clamp Techniques ; Tamoxifen ; pharmacology ; Tumor Cells, Cultured