Neurobehavioral Toxicity And Mechanisms Of Prenatal And Postnatal PFOS Exposure In Wistar Rat

Perfluorooctane sulfonate (PFOS) has been widely used in fire extinguishing agent, textile products, pesticides and other industrial and consumer applications because of its surface activity and stability. Toxicological studies indicate that PFOS is one kind of environmental pollutants with systemic multiple organ toxicities. The Stockholm Convention has officially listed PFOS as a persistent organic pollutant in May,2009. PFOS could enter the fetus and infants through the placenta barrier and breast milk, then accumulate in the infants'brain through the blood-brain barrier, and impact on the early development of the nervous system. However, limited information is available on the mechanisms of the developmental neurotoxicity of PFOS.In this study, the perinatal Wistar rats were exposed to 0,5,15 mg/L PFOS by drinking water, and the rat offspring were cross-fostered on postnatal day 1 (PNDl). The aims were to compare the neurotoxicity effects of prenatal and the postnatal PFOS exposure, and to determine the critical period of the developmental neurotoxicity effects induced by PFOS. Moreover, the mechanisms underlying the developmental neurotoxicity of PFOS in rat offspring were elucidated in relation to the nerve development, the apoptosis of neurons in hippocampus, and the calcium signaling pathways. The main contents include,(1) PFOS accumulation in hippocampus of rat offspring were detected by using liquid chromatography and mass spectrometry. The results showed that the PFOS concentrations in the hippocampus were higher than those in the serum on PND1, which maybe due to the higher cell membrane permeability before the establishment of blood brain barrier, suggestsing that relatively high PFOS accumulation level in the target organ during the developmental stage and the potential neurotoxic effects. The PFOS concentrations in hippocampus were decreased after birth in the sustained exposed groups and the prenatal exposed groups, while they were gradually increased in both serum and hippocampus in the postnatal exposed groups. On PND35, serum and hippocampal PFOS concentrations in prenatal exposure groups were significantly lower than that in the postnatal exposure groups.(2) Neurobehavioral toxicity of prenatal and postnatal PFOS exposure was investigated by evaluating the spatial learning and memory abilities of rat offspring. The grip strength tests showed that the muscle strength of the rat offspring were not significantly affected. Besides, the visible platform tests showed that the swimming ability of rat offspring was not significantly affected. However, the hidden platform test and probe trial test showed that the prenatal and/or postnatal exposure to PFOS resulted in a decrease of the spatial learning and memory ability of the rat offspring. Although the PFOS concentration in the hippocampus of rat offspring in prenatal exposure groups was significantly lower than that in postnatal exposure groups on PND35, the results showed that the degree of the neurobehavioral injury in the prenatal exposed groups was comparable, or even higher than that in the postnatal exposure groups. Neurobehavioral toxicity results suggested that the prenatal PFOS exposure caused long-term effects on rat nervous system function, and the embryonic period is the key period of developmental neurotoxicity induced by PFOS.(3) The relationship between neurobehavioral toxicity of PFOS and synaptic development was elucidated, by analyzing the differential expression of key proteins and genes in neural development. Results showed that the neural development key proteins, including growth-associated protein-43 (GAP-43), neural cell adhesion molecule 1 (NCAM1), nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF), were decreased on PND35, and the corresponding genes (gap-43, ncaml, ngf, bdnf) were significant increased on the mRNA level, which indicated that PFOS influenced the process of transcriptional and translational processes, interfered the expression of neural developmental key proteins, then disturbed the nervous system development, and affected the learning and memory ability of rat offspring. The change degree of the nerve developmental key factors in the prenatal exposure groups were similar to that in the postnatal exposure groups, or even more serious, which also indicated the high risk of PFOS exposure in the embryonic stage.(4) The effects of prenatal and postnatal PFOS exposure on apoptosis in hippocampal neuron of rat were detected, to elucidate the neurobehavioral toxicity mechanism of PFOS on cellular level. The results showed that PFOS caused varying degrees of increased apoptosis on PNDs 1,7,35. Meantime, only prenatal exposure to PFOS could also result in the increased apoptosis in hippocampus. Moreover, the intracellular free calcium concentration ([Ca2+]i) increased in a similar trend with apoptosis. Calcium-related apoptotic genes, apotosis-linked gene-2 (alg-2) and death associated protein kinase (dapk2) genes, were up-regulated on PND35, and had significant difference in both continued exposure groups and prenatal exposure groups, indicating that [Ca2+]i overload acted as a potential mechanism of the hippocampus apopotosis induced by PFOS. The anti apoptosis gene (bcl-2) was significantly up-regulated on both PND7 and PND35, in comparable levels in both prenatal and postnantal exposure groups. At the same time, the expression of Bcl-2 protein in the TT15 exposed group was significantly decreased on PND35. The inconsistent change trend of bcl-2 gene and protein indicated that the feedback regulation mechanism in transcription and translation level was damaged, and ultimately anti apoptosis mechanism is destroyed. PFOS exposure caused the increase of [Ca2+]i, then activated the Ca2+associated apoptosis related genes, intefered the expression of anti apoptotic protein, and resulted in increased apoptosis of hippocampal cells, which might be one of mechanisms of the declined learning and memory ability.(5) The interference of pernatal and postnatal PFOS exposure on key calcium signaling pathways in hippocampal neuron of rat were evaluate, to clarify the association between the neurobehavioral toxicity of PFOS and calcium signaling pathways. The results showed that PFOS exposure can inhibit the expression of calpain ? and activate the expression of calpain II, then lead to the disorder of calcium signaling pathways. The expression of protein kinase A (PKA), cyclic adenosine monophosphate (cAMP), and phosphorylated cAMP response element-binding protein (p-CREB) was inhibited, and the expression of calmodulin-dependent protein kinases ? (CaMKII) was over activated. The trend of calmodulin (CaM) and mitogen-activated protein kinase (MAPK) was not significantly changed on PND35. The changes level of each protein in prenatal exposure groups were comparable with those in the postnatal exposure groups. The over activation of calpain II may interfere the phosphorylation of CREB by inhibiting the activity of cAMP-PKA pathway, then led to the developmental neurotoxicity.