A procedure allowing measurement of cytosolic Ca2+ in rat platelets. Inhibition of a plasma lipoprotein on fura 2-AM loading.
Loading of the fluorescent Ca2+ probe fura 2 in rat platelets is highly inhibited by a plasmatic factor that is removed by gel filtration through a Sepharose C-2B column. Rat plasma also inhibits fura 2 loading in human platelets. The inhibitory effect is abolished by perchloric acid-deproteinization or heat denaturation of plasma suggesting a proteic nature of the inhibitory compound. Indeed the 10,000 x g supernatant of the heat denaturated plasma shows a positive effect on fura 2 accumulation, most likely by partially inhibiting its cellular effux. These effects are only negligibly shown by the corresponding fractions of human plasma. Results obtained by fractionation of rat plasma proteins by means of ion exchange DEAE Sepharose C-6B chromatography and ultracentrifugation through high density saline solutions indicate that the inhibition of cellular fura 2 loading is due to the HDL fraction of rat plasma.
Cavallini
L
L
Department of Biological Chemistry, University of Padova, Italy.
Francesconi
M A
MA
Ruzzene
M
M
Valente
M
M
Deana
R
R
D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D005456:Fluorescent Dyes; D016257:Fura-2 / Q000031:analogs & derivatives; D066298:In Vitro Techniques; D008075:Lipoproteins, HDL / Q000097:blood / Q000302:isolation & purification; D051381:Rats; D011919:Rats, Inbred Strains; D013050:Spectrometry, Fluorescence
D005456:Fluorescent Dyes; D008075:Lipoproteins, HDL; C049925:fura-2-am; D002118:Calcium; D016257:Fura-2