Effect of oxidative stress on the expression of serotonin transporter, 5-HT2A and 5-HT1B receptors in horse leukocytes

Introduction: Serotonin (5-HT) is involved in different pathways of the immune regulation. A dysmetabolism of the 5-HT system may be related to different pathologies, such as infectious and autoimmune diseases. In this preliminary study horse mixed-leukocytes were exposed to H2O2, responsible of a moderate mortality and growth inhibition, in order to evaluate the possible influence of experimental oxidative stress on 5-HT transporter (SERT), 5-HT2A and 5-HT1B receptors. Methodology: Mixed-leukocytes were obtained from blood samples of healthy horses by separation with ficoll. Total RNA was extracted through Trizol kit and 1 microgram was retro transcribed into cDNA for RTq PCR evaluation of SERT, 5-HT2A and 5-HT1B. Beta-actin was used as endogenous control. Results were expressed according to ΔΔCt calculation and by a relative quantization software. Results: RTq PCR results showed a SERT mRNA increase after 3 h exposition to H2O2 (2 micromol) and a decrease after 18 h or to H2O2 (10 micromol) vs control. Both the expressions of 5-HT2A and 5-HT1B receptors increased after 3 h exposition to H2O2 (2 micromol) and fell down under control values after 18 h or to H2O2 (10 micromol). Conclusions: Exposition to a lower concentration of H2O2 may be responsible for cell apoptosis. It is well known that H2O2 generation requires 5-HT internalization through an active transport mechanism into the cell and its degradation by monoamine oxidase A. Exposition to H2O2 at higher concentrations or for a longer time could have produced inhibition of transport mechanisms and peroxidation of membrane lipids leading to a lowered receptor density. These results pave the way for further investigations on the effect of both physiological and pathological stress in leukocytes.