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 Ca(2+) entry in non-permeabilized, intact cells, measuring the intracellular Ca(2+) levels using fluorescence microplate technique. Ca(2+) influx induced via Ca(2+) 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 Ba(2+) entry indicating that the non-capacitive Ca(2+) channels may also be affected. In addition, UFH caused a significant and dose-dependent delay in Ca(2+), 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 Ca(2+) entry possibly via store operated Ca(2+) channels (SOCCs). Inhibitory activities of UFH and 2-aminoethyl diphenyl borate on the capacitive Ca(2+) 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 Ca(2+) influx after store depletion by thapsigargin. We presume that the hyperpolarizing property of this agent might contribute to the suppression of Ca(2+) influx. We concluded that UFH can negatively modulate SOCCs and also other non-capacitive Ca(2+) channels and these activities might also account for its multiple biological effects.