title: Thrombin stimulation of Cl-/HCO3- exchange in human platelets.
abstract: The presence of one acidifying Cl-/HCO3- exchange mechanism in human platelets has not been previously reported. This paper demonstrates that this mechanism does function and that it increases its activity after stimulation with thrombin. On resuspension of BCECF-loaded platelets in a chloride-free medium (gluconate replaced) that contains bicarbonate, cytosolic pH (pHi) increased and stabilized after 10 min at an alkaline value. After addition of 50 mM NaCl, pHi fell rapidly reaching steady state in the succeeding 5 min. The stilbene derivative 4-acetamido-4'-isothiocyanato stilbene-2,2' disulfonic acid (SITS) inhibited both, the alkalization in chloride-poor solution and the recovery from the alkaline load after chloride enrichment. The decline in pHi was observed whether chloride was delivered to the solution in the form of LiCl or NaCl, or when the later was applied after blockage of the Na+/H+ exchanger. The recovery in chloride-containing solution was in contrast to the effect of a similar change in osmolarity by addition of 50 mM sodium gluconate that did not produced a significant variation of pHi. Posterior addition of NaCl after 5 min in high gluconate reproduced the pHi fall of the control experiment. Alkali loads produced by 25 mM trimethylamine hydrochloride (TMA) were also counteracted by HCO(3-)-equivalent efflux via Cl-/HCO3- exchange. One of the major observations of the present study is that HCO3- equivalent efflux was twice as high when the platelets were previously stimulated with 0.1 IU of thrombin, but thrombin did not produce significant changes of the pHi recovery rate in a bicarbonate-free solution. The increase of the decline in pHi elicited by preexposure to thrombin was still observed in the presence of an inhibitor of the Na+/H+ exchange or in sodium-free solutions. It is concluded that a Na-independent Cl-/HCO3- exchange mechanism mediates the recovery of pHi from alkalosis in platelets and that thrombin activates this exchanger by a direct regulatory pathway.
authors.affiliation: Centro de Investigaciones Cardiovasculares, Facultad de Ciencias Médicas, Universidad Nacional de La Plata 60 y 120, 1900 La Plata, Argentina. ogende@atlas.med.unlp.edu.ar
authors.forename: Oscar A
authors.initials: OA
authors.lastname: Gende
mesh_terms: D000471:Alkalosis; D001639:Bicarbonates / Q000378:metabolism; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D027963:Chloride-Bicarbonate Antiporters / Q000187:drug effects* / Q000378:metabolism; D005452:Fluoresceins; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012965:Sodium Chloride / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology*
chemical_list: D001639:Bicarbonates; D027963:Chloride-Bicarbonate Antiporters; D005452:Fluoresceins; D012965:Sodium Chloride; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D013917:Thrombin