4-acetamido-4--isothiocyanatostilbene-2-2--disulfonic-acid and Pancreatic-Neoplasms

Voltage- and Ca(2+)-activated whole cell currents were studied in AR42J cells, a clonal cell line derived from rat pancreatic acinar cells, using a patch electrode voltage-clamp technique. Four kinds of ionic currents were identified by their ionic dependencies, pharmacological properties, and kinetic parameters: 1) an outward current flow due mainly to a voltage-dependent K(+)-conductance increase, 2) an initial transient inward current due to an Na(+)-conductance increase, 3) transient and long-duration inward current due to a Ca(2+)-conductance increase, and 4) a slowly activating inward current that persists over the duration of the depolarizing pulse and deactivates slowly upon repolarization, producing a slow inward tail current. The slow inward tail current was particularly robust and was interpreted as due to a Ca(2+)-activated Cl(-)-conductance increase, since 1) the generation of this current was blocked by removing the extracellular Ca2+, applying Ca(2+)-channel blockers (Cd2+, nifedipine), or by lowering the intracellular Ca2+ concentration [( Ca2+]i) with EGTA; and 2) the reversal potential (Erev) of the slow inward tail current was close to 0 mV in the control condition (152 mM [Cl-]o/154 mM [Cl-]i), and changes of the [Cl-]o/[Cl )i ratio shifted the Erev toward the predicted Cl- equilibrium potential.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; Animals; Calcium; Calcium Channels; Cell Line; Chlorides; Electric Conductivity; Electrophysiology; Kinetics; Membrane Potentials; Pancreas; Pancreatic Neoplasms; Potassium Channels; Rats; Sodium

The addition of glucose to suspensions of HIT-T15 insulinoma cells caused a small, transient acidification followed by a gradual, progressive alkalinisation, as assessed by the fluorescent pH-sensitive dye 2',7'-biscarboxyethyl-5'-(6')-carboxyfluorescein (BCECF). Treatment of cells with acetate or lactate produced an immediate, marked acidification followed by recovery and a subsequent alkalinisation. In contrast, addition of NH4Cl caused a rapid rise in intracellular pH (pHi) and recovery to resting values. In cells where Na+/H+ exchange was inhibited, either with amiloride or by omission of Na+ from the medium, glucose caused a progressive acidification, whilst recovery from acetate- or lactate-induced acidification was prevented. Under sodium-free conditions, recovery from acidification could be initiated by addition of Na+. Inhibition of HCO3-/Cl- exchange by pretreatment with 4,4'-diisothiocyanatostilbene 2,2'-disulphonic acid (DIDS), or by omission of HCO3- or Cl- from the medium did not affect any of the changes in pHi elicited by the above agents. It is concluded that the principal mechanism responsible for pHi regulation in HIT-T15 cells is the Na+/H+ antiporter and that the HCO3-/Cl- exchange systems make little, if any, contribution.

Topics: 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; Acetates; Adenoma, Islet Cell; Ammonium Chloride; Bicarbonates; Carrier Proteins; Chlorides; Fluoresceins; Fluorescent Dyes; Hydrogen-Ion Concentration; Insulin; Insulin Secretion; Insulinoma; Lactates; Lactic Acid; Pancreatic Neoplasms; Sodium-Hydrogen Exchangers; Tumor Cells, Cultured