Enhanced Ca influx in mechanically distorted erythrocytes measured with F nuclear magnetic resonance spectroscopy.

We present the first direct nuclear magnetic resonance (NMR) evidence of enhanced entry of Ca ions into human erythrocytes (red blood cells; RBCs), when these cells are mechanically distorted. For this we loaded the RBCs with the fluorinated Ca chelator, 1,2-bis(2-amino-5-fluorophenoxy)ethane-N,N,N',N'-tetraacetic acid (5FBAPTA), and recorded F NMR spectra. The RBCs were suspended in gelatin gel in a special stretching/compression apparatus. The 5FBAPTA was loaded into the cells as the tetraacetoxymethyl ester; and C NMR spectroscopy with [1,6-C]D-glucose as substrate showed active glycolysis albeit at a reduced rate in cell suspensions and gels. The enhancement of Ca influx is concluded to be via the mechanosensitive cation channel Piezo1. The increased rate of influx brought about by the activator of Piezo1, 2-[5-[[(2,6-dichlorophenyl)methyl]thio]-1,3,4-thiadiazol-2-yl]-pyrazine (Yoda1) supported this conclusion; while the specificity of the cation-sensing by 5FBAPTA was confirmed by using the Ca ionophore, A23187.