Research On The Molecular Mechanisms About The Neurotoxicity Of Microcystins

Microcystins(MCs), a group of cyclic heptapeptide compounds with multiorgan toxicity produced by cyanobacterial species. In decades, because of the improvement of social productivity and the pressure of the huge number of population, the increasing eutrophication was happened in many fresh waters world wide. Therefore bring about the frequent occurrence of cyanobacterial blooms in world wide. And concequently, the cyanotoxins produced by the cyanobacteria have become a great threat to aquatic animals, livestock, and human health, and more and more researchers pay theie attention on the cyanotoxins. Among them, microcystins (MCs) are the most common and frequent all over the world. So far, more than 100 different structural analogues of MCs have been identified, among which, microcystin-LR (MC-LR) being the most common and toxic, then is microcystn-YR (MC-YR) and microcystin-RR (MC-RR). A number of studies have described the specific hepatotoxicity of the MCs, hoever, the tragic event occurring in Caruaru.Brazil in February 1996, suggests the neurotoxicity of the microcystins. As we know, because of the brain-blood barrier, so many Macromolecular materials can not through it and come into the brain, however, the exsist of the organic anion transporting polypeptides (OATP) which is pecifically required for the active uptake of MCs into brain, make it possible that microcystins can through the brain-blood barrier and into the brain, and may induce serious neurotoxicity. More and more studies have found the effect of neuro txoxicity, but the molecular mechanisms by which it express its toxicity in the nervous system still largely unknown. To further our understanding of mechanisms, in this study, we use the proteomics combind with histopathological changes and biochemical analysis and other technologies, to give a complete research of the neurotoxicity of microcystins.We give the mature zebrafish the microcystin injection by the method of intraperitoneal injection of MC-RR and MC-LR at 0.5LD50 dose, after 24 h, we got the brain of zebrafish and treatment them properly. First, we detection the microcystin content in the brain, and found that the contents of microcysin were 0.109 and 0.036 μg/mg DW in each toxin treatment. As microcystins can potently inhibit the activity of protein phosphatase 1 and 2A, we tested the activity of PP but to found that there has a slightly increase of the protein phosphatase, but has no significant differences compared with control. Histopathological study indicates that with the treatment of microcystins, the number of neurocyte was significantly reduced and we can see the sparce density of the neurocytes. Ultrastructure pathology study shows the neuritis in the neuropilus were swollen and show the vacuoles, the neurofilaments are disappeared, and the density of mitochondria was increased. As time extended, the injury of the neuropilus become more serious,as we can see the highly empty of the neuritis and the diappearence of the organelles in the cells, and also we can see the capillary for which has a slightly swollen endothelial cell cytoplasm. During our experiment, the most serious injuries happened at the 24 h treatment, the injuries were so serious that we can not distinguish the structure of the tissue, the mean time, we can see the neuritis were very serious swallowed, the neurofi laments were disappeared and the rarefaction and vacuoles of the mitochondrial structure, and the break and disappearance of the mitochondrial cristae. To study the mechanisms of microcystins induced neurotoxicity, a proteomic analysis was performed on zebrafish brain injected with 0.5LD50 MC-RR and MC-LR respectively until 24 h treament. Two-dimentional electrophoresis coupled with mass spectrometry was employed to detect and identify the protein profiles. Results showed that compared with control, the MC-RR and the MC-LR treaments have 77 and 90 spots respectively displayed a significant increase or decrease of abundance. Also we compared the treament between MC-RR and MC-LR, and found that there have 45 spots show the differencially expressed. In total,81 proteins were identified. The results suggest that microcystins induced mechanisms against oxidative stress, the cytoskelton assembly, and the adaptative changes in the energetic metabolism. It also affects the expression of the subunits of protein phosphatases 1 and the signal transduction and many other biochemical processes. We can see the complicated regulatory mechanisms of organisms in the response to microcystins treatments.