The plasma membrane functions both as a natural insulator and a diffusion barrier to the movement of ions. A wide variety of proteins transport and pump ions to generate concentration gradients that result in voltage differences, while ion channels allow ions to move across the membrane down those gradients. Plasma membrane potential is the difference in voltage between the inside and the outside of a biological cell, and it ranges from ~− 3 to ~− 90 mV. Most of the most significant discoveries in this field have been made in excitable cells, such as nerve and muscle cells. Nevertheless, special attention has been paid to some events controlled by changes in membrane potential in non-excitable cells. The origins of several blood disorders, for instance, are related to disturbances at the level of plasma membrane in erythrocytes, the structurally simplest red blood cells. The high simplicity of erythrocytes, in particular, made them perfect candidates for the electrophysiological studies that laid the foundations for understanding the generation, maintenance, and roles of membrane potential. This article summarizes the methodologies that have been used during the past decades to determine Δψ in red blood cells, from seminal microelectrodes, through the use of nuclear magnetic resonance or lipophilic radioactive ions to quantify intra and extracellular ions, to continuously renewed fluorescent potentiometric dyes. We have attempted to highlight the advantages and disadvantages of each methodology, as well as to provide a description of the technical aspects involved.

Plasma membrane tubulin is an endogenous regulator of P-ATPases and the unusual accumulation of tubulin in the erythrocyte membrane results in a partial inhibition of some their activities, causing hemorheological disorders like reduced cell deformability and osmotic resistance. These disorders are of particular interest in hypertension and diabetes, where the abnormal increase in membrane tubulin may be related to the disease development. Phosphatidylserine is more exposed on the membrane of diabetic erythrocytes than in healthy cells. In most cells, phosphatidylserine is transported from the exoplasmic to the cytoplasmic leaflet of the membrane by lipid flippases. Here we report that phosphatidylserine is more exposed in erythrocytes from both hypertensive and diabetic patients than in healthy erythrocytes, which could be attributed to the inhibition of flippase activity by tubulin. This is supported by: (i)- the translocation rate of a fluorescent phosphatidylserine analog in hypertensive and diabetic erythrocytes was slower than in healthy cells, (ii)- the pharmacological variation of membrane tubulin in erythrocytes and K562 cells was linked to changes in phosphatidylserine translocation, (iii)- the P-ATPase-dependent phosphatidylserine translocation in inside-out vesicles from human erythrocytes was inhibited by tubulin. These results suggest that tubulin regulates flippase activity and hence the membrane phospholipid asymmetry.

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