Kinetic characteristics of acetylcholinesterase and structural-functional state of rat erythrocyte membranes at moderate hypothermia

Abstract

Artificial hypothermic state of homeothermic animals contributes to the stimulation of free radical processes in red blood cells. In order to understand what are the consequences of oxidative damage of erythrocyte membrane, we examined the dependence of the kinetic characteristics of integral membrane enzyme acetylcholinesterase (AChE) and structural and functional state of the membrane on the duration of mild hypothermia. For this purpose we reduced body temperature of adult Wistar rats by external cooling to 30°C (short-term moderate hypothermia) and then prolonged hypothermia up to 1.5 and 3 h. A short-term hypothermia was followed with an increase in V_max and a decrease in K_m, promoting an increase in the catalysis effectiveness.The optimum point on the graph of the concentration dependence was shifted to the area of lower concentrations, and the character of enzyme–substrate interactions at high concentrations of the enzyme changed. Upon prolongation of hypothermia, changes in the AChE kinetic characteristics favored normalization of the enzyme activity and concentration dependence. To test the hypothesis of a possible influence of the lipid matrix on the kinetic characteristics of AchE, we studied structural properties of the erythrocyte membranes using fluorescent probe pyrene. The observed changes in the structural and dynamic characteristics of erythrocyte membranes after a 1.5-h hypothermia suggested a reduction in microviscosity of both total and annular lipids. Prolongation of hypothermia up to 3 h favored normalization of this parameter. It was found that the indicators of the structural state of erythrocyte membranes at different durations of hypothermia correlate with certain kinetic characteristics of AChE. The data indicate that the prolongation of mild hypothermia up to 3 h triggers adaptive mechanisms directed to normalization of the erythrocytes membrane functioning.