Mouse (Mus Musculus) Embryonic Cerebral Cortex Cell Death Caused by Carbofuran Insecticide Exposure

Abstract

Introduction: The aim of the study was to describe the process of neuron death in the cerebral cortex caused by embryonic carbofuran exposure.

Material and methods: 81 mouse foetuses from 27 breeding mice were used in the study. Carbofuran was administered by gavage from the 6^th to the 15^th day of gestation to two groups: one at 0.0208 and the other at 0.0417 mg/kg b.w. On the 17^th day, the mice were sacrificed and the foetuses were taken to measure the ROS (malondialdehyde/MDA and superoxide dismutase/SOD) activity in brain tissue, the number of apoptotic embryonic cerebral cortex neurons using a TUNEL assay, and necrotic cells using HE staining. Examination of p53 and caspase 3 expression was done by immunohistochemistry. Data were analysed using analysis of variance (ANOVA) and Duncan's test.

Results: Increased activity of cerebral ROS characterised by significant elevation of the MDA level (P < 0.05), decreased SOD (P < 0.01), increased p53 and caspase 3 expression, and cerebral cortical neuron death either by necrosis or apoptosis (P < 0.05) were found. At the low dose carbofuran increased expression of p53, caspase 3, and apoptosis. At the high dose it increased levels of MDA and necrosis.

Conclusion: Increased expression of p53 and caspase 3 and apoptosis indicated that carbofuran may cause apoptosis through the intrinsic pathway. The increased apoptosis grants an opportunity to prevent and treat the effect of ROS due to gestational carbofuran exposure.

Keywords: ROS; apoptosis; carbofuran; mice; necrosis.

Fig. 1

Embryonic cerebral cortex neuron p53 expression in 6–15-day-old foetuses gestated by carbofuran-exposed mice (400×). Brown cells (marked by arrows) indicate p53-expressing neuronal cells a – p53 expression in the embryonic cerebral cortex (yellow box), b – control group, c – lower-dose group (0.0208 mg/kg b.w.), d – higher-dose group (0.0417 mg/kg b.w.)

Fig. 2

Embryonic cerebral cortex neuronal cell caspase 3 expression examined in 6–15-day-old foetuses gestated by CF-exposed mice. Arrows mark neuronal cells expressing caspase 3 a – caspase 3 expression located in the embryonic cerebral cortex (yellow box), b– control group, c – lower-dose group (0.0208 mg/kg b.w.), d – higher-dose group (0.0417 mg/kg b.w.)

Fig. 3

Embryonic cerebral cortex neuron apoptosis examined in 6–15-day-old foetuses gestated by CF-exposed mice. Arrows mark apoptotic neurons (dark brownish cells) a – apoptotic neurons located in the embryonic cerebral cortex (yellow box), b – control group, c – lower-dose group (0.0208 mg/kg b.w.), d – higher-dose group (0.0417 mg/kg b.w.)

Fig. 4

Embryonic neuron necrosis in 6–15-day-old foetuses gestated by CF-exposed mice. Neurons undergoing necrosis (marked by arrows) are shrunken and have solid pyknotic nuclei and eosinophilic cytoplasm a – necrotic neurons located in the embryonic cerebral cortex (yellow box), b – control group, c – lower-dose group (0.0208 mg/kg b.w.), d – higher-dose group (0.0417 mg/kg b.w.)