oxonol-v and 2--7--bis(carboxyethyl)-5(6)-carboxyfluorescein

Topics: Animals; Biological Transport; Cell Line; Cricetinae; Dithiothreitol; Fluoresceins; Fluorescent Dyes; Glyceraldehyde; Guanidines; Insulin; Insulin Secretion; Islets of Langerhans; Isoxazoles; Membrane Potentials; Mesocricetus

L-Lactate, D-lactate, propionate and acetate (all 20 mM) caused a marked intracellular acidification in HIT-T15 cells loaded with 2'7'-bis(carboxyethyl)-5'(6')-carboxyfluorescein (BCECF), followed by recovery to more alkaline values. The effects of L- and D-lactate, but not propionate or acetate, were inhibited by 5 mM alpha-fluorocinnamate. Both L- and D-lactate caused a marked depolarisation and rise in cytosolic [Ca2+] in HIT cells as assessed by oxonol-V and quin2 fluorescence, respectively. Propionate had similar, though less marked, effects, whereas acetate exerted only a modest influence on membrane potential and cytosolic [Ca2+]. The rate of oxidation of L-lactate by HIT cells greatly exceeded that of D-lactate. alpha-Fluorocinnamate delayed, but did not prevent, the effects of L-lactate on HIT cell membrane potential or cytosolic [Ca2+]. L-lactate diminished the rate of efflux of 86Rb+ from preloaded HIT cells. Inhibition of calcium- and nucleotide-sensitive K+ channels with tetraethylammonium and tolbutamide also reduced the 86Rb+ efflux rate, and prevented any further reduction in response to L-lactate. However, such inhibition of K+ channels did not prevent a further depolarisation and rise in cytosolic [Ca2+] upon the subsequent addition of lactate. It is suggested that the activation of HIT-T15 cells by lactate is not the result of intracellular acidification or increased metabolic flux, and does not require diminished K+ permeability. An alternative mechanism is based upon the possible electrogenic flux of lactate across the plasma membrane.

Topics: Acetates; Aminoquinolines; Animals; Calcium; Cytosol; Fluoresceins; Hydrogen-Ion Concentration; Insulinoma; Islets of Langerhans; Isoxazoles; Lactates; Lactic Acid; Membrane Potentials; Propionates; Tumor Cells, Cultured