Suppression of Ca2+ influx by unfractionated heparin in non-excitable intact cells via multiple mechanisms

Effect of unfractionated heparin (UFH), described as a cell-impermeant IP3 receptor antagonist, was studied on the capacitive Ca2+ entry in non-permeabilized, intact cells, measuring the intracellular Ca2+ levels using fluorescence microplate technique. Ca2+ influx induced via Ca2+ mobilization by histamine in Hela cells or evoked by store depletion with thapsigargin in RBL-2H3 cells was dose-dependently suppressed by UFH added either before or after the stimuli. UFH also prevented the spontaneous Ba2+ entry indicating that the non-capacitive Ca2+ channels may also be affected. In addition, UFH caused a significant and dose-dependent delay in Ca2+, and other bivalent cation inflow after treatment of the cells with Triton X-100, but it did not diminish the amount of these cations indicating that UFH did not act simply as a cation chelator, but modulated the capacitive Ca2+ entry possibly via store operated Ca2+ channels (SOCCs). Inhibitory activities of UFH and 2-aminoethyl diphenyl borate on the capacitive Ca2+ influx was found reversible, but the time courses of their actions were dissimilar suggesting distinct modes of action. It was also demonstrated using a fluorescence potentiometric dye that UFH had a considerable hyperpolarizing effect and could alter the changes of membrane potential during Ca2+ influx after store depletion by thapsigargin. We presume that the hyperpolarizing property of this agent might contribute to the suppression of Ca2+ influx. We concluded that UFH can negatively modulate SOCCs and also other non-capacitive Ca2+ channels and these activities might also account for its multiple biological effects.