Avermectin exposure induces apoptosis in King pigeon brain neurons

Abstract

The effect of avermectin was studied on King pigeon brain nerve cells by cytotoxicity [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide, MTT] and apoptosis [acridine orange/ethidium bromide (AO/EB) assay, transmission electron microscope (TEM) evaluation, measurement of mitochondrial membrane potential (Δψm), phosphatidylserine (PS) exposure, caspases activities, DNA fragmentation, reactive oxygen species (ROS) and caspase-3 mRNA expression] within the 2.5–10μgL−1 concentration-range. The results revealed that within the concentrations of 2.5–10μgL−1, avermectin showed obvious cytotoxicity and induced apoptosis in a dose-dependent manner to neurons of King pigeon in vitro. Cell viability were 99.93±8.52%, 82.02±4.99% and 78.23±5.67% after 24h of treatment with avermectin at the concentrations of 0, 2.5 and 5μgL−1, which decreased to 56.36±2.17% of 10μgL−1. Treated cells showed typical apoptosis morphological changes including cytoplasmic vacuolation, chromatin condensation, unclear nuclear membrane and decreased/swollen mitochondria. Typical biochemical hallmarks of apoptosis including Δψm loss, PS exposure, activations of caspase-3, caspase-8 and caspase-9, DNA fragmentation were observed too. Moreover, the levels of ROS in the avermectin treatment groups increased significantly compared to control group. Furthermore, the caspase-3 mRNA levels increased significantly following AVM treatment. In conclusion, our experimental results show that avermectin has cytotoxicity to brain neurons of King pigeon in vitro and the mechanism of neurotoxicity induced by avermectin is closely related to apoptosis.