Nnitric oxide function in regulation of Ca2+-dependent K+-permeability of human erythrocyte

Nitric oxide performs multiplefunctions in the human body. Specifically, NO affects the stability of hypotonic red blood cells, regulates the transfer of oxygen, affectsthe deformability of red blood cells, as well as programmed death of erythrocytes. Erythrocyte membrane contains Ca2 +-activated K+-channels of intermediate conductance, or the Gardos-channels, which play a role in death of erythrocytes. They may participate in deformabilityof cells: Ca2+-induced decrease in the deformability of red blood cells eliminated by aligning the gradient of potassium ions.Question of the participation of nitric oxide in the regulation of Ca2+-activated K+-channels of erythrocytes remains unsolved. In thepresent study, we investigated the effect of nitric oxide on Ca2+-dependent K+-permeability of the membrane of human erythrocytes. Thestudies were conducted using the method of recording the membrane potential in erythrocyte suspensions by changing the pH of theincubation medium in the presence of the protonophore. Adding the calcium ionophore A23187 to a suspension of cells containing calciumchloride led to potassium ions issue and the development of hyperpolarizing response of the membrane of red blood cells, which isreflected in the change of pH of the suspension. The activity of Ca2+-activated potassium channels was estimated by the amplitude of thehyperpolarizing response and the speed of its development. Data analysis was performed using the program Statistica 6.0 for Windowsof Statsoft Company. The formed sample does not obey the normal distribution; so nonparametric tests were used for testing statisticalhypotheses. Adding of red blood cells 10-8 to 10-6 M sodium nitroprusside (NP) to the incubation medium did not cause any changes inthe amplitude of hyperpolarizing response of red blood cells. Increasing concentrations of NP in the incubation medium of erythrocytesto 10-5 M resulted in a significant decrease in the amplitude of hyperpolarizing response. Further increase in the concentration of sodiumnitroprusside in the incubation medium to 10-4 10-3 M led to complete suppression of hyperpolarizing response of red blood cells. Perhapsthe effects obtained are due to decomposition of NP with fragments of electron-transport chain present in the membrane of redblood cells and the inhibitory effect of cyanide ion. Incubation of erythrocyte with L-arginine (10-5 M) caused a significant increase inthe amplitude and velocity of hyperpolarizing response of red blood cells. Inhibition of NO-synthase with L-NMMA (Sigma) (24 10-5M) significantly reduced the amplitude of the hyperpolarizing response. Thus, the experiments revealed that intracellular NO productionby L-arginine increased the Ca2+-dependent potassium permeability of erythrocyte membranes. The same effect is obtained by increasingthe intracellular concentration of cGMP by dibutyryl-cGMP or phosphodiesterase inhibitors.

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