Spatial distribution and temporal change in cytosolic pH and [Ca2+] in resting and activated single human platelets.
In human platelets, a rapid rise in cytoplasmic Ca2+ and slower rise in cytoplasmic pH follow stimulation by thrombin. With the Ca2+ probe Fura-2 and the pH probe SNARF-1 for digitized fluorescence microscopy, we studied simultaneously the distribution and changes with time of [pH]i and [Ca2+]i in individual human platelets. In platelets coloaded with both probes, the probes had no detectable fluorescence at each other's excitation wavelength. The monovalent cation ionophore, nigericin (2 microM), produced a homogeneous rise in pH but no change in [Ca2+]. Platelets, in contact with glass, spread and developed an irregular, apparently mutually independent rise in both [Ca2+] and pH. Stimulation of platelets by thrombin 1.0 U/ml elevated [Ca2+]i and produced slow alkalinization without initial acidification. Replacement of extracellular Na+ by choline abolished thrombin-induced alkalinization, but had no effect on thrombin-induced [Ca2+]i elevation. ADP 10 microM caused a rapid rise of [Ca2+]i and transient alkalinization. Most stimulated platelets developed a gradient in pH, that was highest in the center. ADP and thrombin caused oscillation of [Ca2+]i but not of [pH]i. We conclude that alkalinization in stimulated platelets, presumably involving Na+/H+ antiport, is not essential for the rise of [Ca2+]i that may accompany it.
Ariyoshi
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Department of Surgery, Beth Israel Hospital, Boston, Massachusetts, USA.
Salzman
E W
EW
D000244:Adenosine Diphosphate / Q000494:pharmacology; D001578:Benzopyrans; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002412:Cations; D003600:Cytosol / Q000378:metabolism; D005109:Extracellular Matrix / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes; D016257:Fura-2; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009284:Naphthols; D009550:Nigericin / Q000494:pharmacology; D015539:Platelet Activation; D012235:Rhodamines; D012964:Sodium / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D013917:Thrombin / Q000494:pharmacology
D001578:Benzopyrans; D002412:Cations; D005456:Fluorescent Dyes; D009284:Naphthols; D012235:Rhodamines; C091666:seminaphthorhodaminefluoride; D000244:Adenosine Diphosphate; D012964:Sodium; D013917:Thrombin; D009550:Nigericin; D002118:Calcium; D016257:Fura-2