title: Pericellular Ca(2+) recycling potentiates thrombin-evoked Ca(2+) signals in human platelets.
abstract: We have previously demonstrated that Na(+)/Ca(2+) exchangers (NCXs) potentiate Ca(2+) signaling evoked by thapsigargin in human platelets, via their ability to modulate the secretion of autocoids from dense granules. This link was confirmed in platelets stimulated with the physiological agonist, thrombin, and experiments were performed to examine how Ca(2+) removal by the NCX modulates platelet dense granule secretion. In cells loaded with the near-membrane indicator FFP-18, thrombin stimulation was observed to elicit an NCX-dependent accumulation of Ca(2+) in a pericellular region around the platelets. To test whether this pericellular Ca(2+) accumulation might be responsible for the influence of NCXs over platelet function, platelets were exposed to fast Ca(2+) chelators or had their glycocalyx removed. Both manipulations of the pericellular Ca(2+) rise reduced thrombin-evoked Ca(2+) signals and dense granule secretion. Blocking Ca(2+)-permeable ion channels had a similar effect, suggesting that Ca(2+) exported into the pericellular region is able to recycle back into the platelet cytosol. Single cell imaging with extracellular Fluo-4 indicated that thrombin-evoked rises in extracellular [Ca(2+)] occurred within the boundary described by the cell surface, suggesting their presence within the open canalicular system (OCS). FFP-18 fluorescence was similarly distributed. These data suggest that upon thrombin stimulation, NCX activity creates a rise in [Ca(2+)] within the pericellular region of the platelet from where it recycles back into the platelet cytosol, acting to both accelerate dense granule secretion and maintain the initial rise in cytosolic [Ca(2+)].
authors.affiliation: Department of Physiology, Development and Neuroscience, University of Cambridge Cambridge, U.K.
authors.forename: Stewart O
authors.initials: SO
authors.lastname: Sage
authors.forename: Nicholas
authors.initials: N
authors.lastname: Pugh
authors.forename: Richard W
authors.initials: RW
authors.lastname: Farndale
authors.forename: Alan G S
authors.initials: AG
authors.lastname: Harper
keywords: Ca2+; FFP-18; nanojunction; open canalicular system; pericellular; platelets