Abstract
Microcystins (MCs) are produced during the growth and proliferation of some species of cyanobacteria, mainly Microcystis aeruginosa, which has massive growth in eutrophic water bodies. Microcystins are highly toxic metabolites derived from some cyanobacteria species that exert its main effect in the liver through the inhibition of protein phosphatase (PP1 and PP2A). However, other damages in fish species are less documented and could be unexpected. The aim of the current study was to evaluate the effects of Microcystis aeruginosa extract (MaE) into the central nervous system (CNS) of the Nile tilapia. The MaE was normalized by MCs content (MC-LR). We include a positive control for protein phosphatase inhibition, norcantharidin intraperitoneally dosed at sublethal levels. On the eighth day, measurement of neurotransmission biomarkers (AChE, BChE, CbE, and GABA) were measured, as well as levels of mitochondrial calcium and the mitochondrial membrane potential by flow cytometry in the brain and spinal cord were assessed, in addition to the PP1/PP2A activity in the liver. The MCs elicited mortality at 5 µg/L. The positive control and MCs at sublethal levels inhibited the PP1/PP2A activity in the liver and induced alterations in the neurotoxicity biomarkers evaluated in the CNS. This response is probably due to the disruption of transport ions, dependent and independent of ATP because of alterations in the mitochondrial membrane potential and mitochondrial calcium. The findings of this study suggest that pollutants capable of inducing cyanobacterial blooms are able, in an indirect way, to exert neurotoxic effects in fish species through MC levels.
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The authors declare that all other data supporting the findings of this study are available within the article and its supplementary information files.
References
Baganz, D., Staaks, G., & Steinberg, C. (1998). Impact of the cyanobacteria toxin, microcystin-lr on behaviour of zebrafish Danio Rerio. Water Res., 32(3), 948–952. https://doi.org/10.1016/S0043-1354(97)00207-8
Baganz, D., Staaks, G., Pflugmacher, S., & Steinberg, C. E. W. (2004). A comparative study on microcystin-LR induced behavioural changes of two fish species (Danio rerio and Leucaspius delineatus). Environmental Toxicology, 19, 564–570. https://doi.org/10.1002/tox.20063
Bai, T., Dong, D. S., & Pei, L. (2013). Resveratrol mitigates isoflurane-induced neuroapoptosis by inhibiting the activation of the Akt-regulated mitochondrial apoptotic signaling pathway. International Journal of Molecular Medicine, 32(4), 819–826. https://doi.org/10.3892/ijmm.2013.1464
Bak, L. K., Schousboe, A., & Waagepetersen, H. S. (2006). The glutamate/GABA-glutamine cycle: Aspects of transport, neurotransmitter homeostasis and ammonia transfer. Journal of Neurochemistry, 98(3), 641–653. https://doi.org/10.1111/j.1471-4159.2006.03913.x
Carbis, C. R., Rawlin, G. T., Grant, P., Mitchell, G. F., Anderson, J. W., & McCauley, I. (1997). A study of fetal carp, Cyprinus carpio L., exposed to Microcystis aeruginosa at Lake Mokoan, Australia, and possible implications for fish health. Journal of Fish Diseases, 20, 81–91. https://doi.org/10.1046/j.1365-2761.1997.d01-111.x
Carmichael, W. W., Azevedo, S. M. F. O., An, J. S., Molica, R. J. R., Jochimsen, E. M., Lau, S., Rinehart, K. L., Shaw, G. R., & Eaglesham, G. K. (2001). Human fatalities from cyanobacteria: Chemical and biological evidence for cyanotoxins. Environmental Health Perspectives, 109(7), 663–668. https://doi.org/10.1289/ehp.01109663
Cazenave, J., Bistoni, M. A., Pesce, S. B., & Wunderlin, D. A. (2006). Differential detoxification and antioxidant response in diverse organs of Corydoras paleatus experimentally exposed to microcystin-RR. Aquatic Toxicology, 76(1), 1–12. https://doi.org/10.1016/j.aquatox.2005.08.011
Cazenave, J., Nores, M. L., Miceli, M., Díaz, M. P., Wunderlin, D. A., & Bistoni, M. A. (2008). Changes in the swimming activity and the glutathione S-transferase activity of Jenynsia multidentata fed with microcystin-RR. Water Research, 42(4–5), 1299–1307. https://doi.org/10.1016/j.watres.2007.09.025
Chang H, Huang H, Huang T, Yang P, Wang Y, Juan H (2013) Flow cytometric detection of mitochondrial membrane potential. Bio-protocol 3(8):e430. https://doi.org/10.21769/BioProtoc.430
Chen, J., Xie, P., Li, L., & Xu, J. (2009). First identification of the hepatotoxic microcystins in the serum of a chronically exposed human population together with indication of hepatocellular damage. Toxicological Sciences, 108(1), 81–89. https://doi.org/10.1093/toxsci/kfp009
Chorus I, Bartram J (1999) Toxic cyanobacteria in water: A guide to their public health consequences, monitoring and management/edited by Ingrid Chorus and Jamie Bertram. World Health Organization. https://apps.who.int/iris/handle/10665/42827
Deng, L., Dong, J., & Wang, W. (2013). Exploiting protein phosphatase inhibitors based on cantharidin analogues for cancer drug discovery. Mini Reviews in Medicinal Chemistry., 13(8), 1166–1176. https://doi.org/10.2174/1389557511313080005
Dzul-Caamal, R., Salazar-Coria, L., Olivares-Rubio, H. F., Rocha-Gómez, M. A., Girón-Pérez, M. I., & Vega-López, A. (2016). Oxidative stress response in the skin mucus layer of Goodea gracilis (Hubbs and Turner, 1939) exposed to crude oil: A non-invasive approach. Comparative Biochemistry and Physiology Part a: Molecular & Integrative Physiology, 200, 9–20. https://doi.org/10.1016/j.cbpa.2016.05.008
Finkel, T., Menazza, S., Holmström, K. M., Parks, R. J., Liu, J., Sun, J., Liu, J., Pan, X., & Murphy, E. (2015). The ins and outs of mitochondrial calcium. Circulation Research, 116(11), 1810–1819. https://doi.org/10.1161/CIRCRESAHA.116.305484
Fontanillo, M., & Köhn, M. (2018). Microcystins: Synthesis and structure-activity relationship studies toward PP1 and PP2A. Bioorganic & Medicinal Chemistry, 26(6), 1118–1126. https://doi.org/10.1016/j.bmc.2017.08.040
Frezza, C., Cipolat, S., & Scorrano, L. (2007). Organelle isolation: Functional mitochondria from mouse liver, muscle and cultured filroblasts. Nature Protocols, 2(2), 287–295. https://doi.org/10.1038/nprot.2006.478
Gélinas, M., Juneau, P., & Gagné, F. (2012). Early biochemical effects of Microcystis aeruginosa extracts on juvenile rainbow trout (Oncorhynchus mykiss). Comparative Biochemistry and Physiology Part b: Biochemistry and Molecular Biology, 161(3), 261–267. https://doi.org/10.1016/j.cbpb.2011.12.002
Guillard, R. R. L. (1975). Culture of phytoplankton for feeding marine invertebrates. In W. L. Smith & M. H. Chanley (Eds.), Culture of Marine Invertebrate Animals (pp. 26–60). Plenum Press.
Guiry in Guiry MD, Guiry GM (2020) AlgaeBase. World-wide electronic publication, National University of Ireland, Galway. Available on http://www.algaebase.org/search/species/detail/?species_id=30050
Gupta, N., Pant, S. C., Vijayaraghavan, R., & Rao, P. V. L. (2003). Comparative toxicity evaluation of cyanobacterial cyclic peptide toxin microcystin variants (LR, RR, YR) in mice. Toxicology, 188, 285–296. https://doi.org/10.1016/s0300-483x(03)00112-4
Harada, K. I., & Tsuji, K. (1998). Persistence and decomposition of hepatotoxic microcystins produced by cyanobacteria in natural environment. Journal of Toxicology: Toxin Reviews, 17(3), 385–403. https://doi.org/10.3109/15569549809040400
Hestrin, N. (1949). The reaction of acetylcholine and other carboxylic acid derivatives with hydroxylamine, and its analytical application. Journal of Biological Chemistry, 180(1), 249–261.
Hill, T. A., Stewart, S. G., Gordon, C. P., Ackland, S. P., Gilbert, J., Sauer, B., Sakoff, J. A., & McCluskey, A. (2008). Norcantharidin analogues: Synthesis, anticancer activity and protein phosphatase 1 and 2A inhibition. Chem Med Chem, 3(12), 1878–1892. https://doi.org/10.1002/cmdc.200800192
Hinojosa MG, Gutiérrez-Praena D, Prieto AI, Guzmán-Guillén R, Jos A, Cameán AM (2019) Neurotoxicity induced by microcystins and cylindrospermopsin: A review. Science Total Environ 10 (668):547–565. /https://doi.org/10.1016/j.scitotenv.2019.02.426
Hotta, Y., Ezaki, S., Atomi, H., & Imanaka, T. (2002). Extremely stable and versatile carboxylesterase from a hyperthermophilic archaeon. Applied and Environment Microbiology, 68(8), 3925–3931. https://doi.org/10.1128/AEM.68.8.3925-3931.2002
Hubbard, M. J., & McHugh, N. J. (1996). Mitochondrial ATP synthase F1-β-subunit is a calcium-binding protein. FEBS Letters, 391(3), 323–329. https://doi.org/10.1016/0014-5793(96)00767-3
Jonas, A., Scholz, S., Fetter, E., Sychrova, E., Novakova, K., Ortmann, J., Benisek, M., Adamovsky, O., Giesy, J. P., & Hilscherova, K. (2015). Endocrine, teratogenic and neurotoxic effects of cyanobacteria detected by cellular in vitro and zebrafish embryos assays. Chemosphere, 120, 321–327. https://doi.org/10.1016/j.chemosphere.2014.07.074
Kalueff, A. V., & Nutt, D. J. (2007). Role of GABA in anxiety and depression. Depression and Anxiety, 24(7), 495–517. https://doi.org/10.1002/da.20262
Kataoka, M., Fukura, Y., Shinohara, Y., & Baba, Y. (2005). Analysis of mitochondrial membrane potential in the cells by microchip flow cytometry. Electrophoresis, 26(15), 3025–3031. https://doi.org/10.1002/elps.200410402
Kawamoto EM, Vivar C, Camandola S (2012) Physiology and pathology of calcium signaling in the brain. Front Pharmacol 3(61). https://doi.org/10.3389/fphar.2012.00061
Kist, L. W., Rosemberg, D. B., Pereira, T. C., de Azevedo, M. B., Richetti, S. K., de Castro, L. J., Yunes, J. S., Bonan, C. D., & Bogo, M. R. (2012). Microcystin-LR acute exposure increases AChE activity via transcriptional ache activation in zebrafish (Danio rerio) brain. Comparative Biochemistry and Physiology Part c: Toxicology & Pharmacology, 155(2), 247–252. https://doi.org/10.1016/j.cbpc.2011.09.002
Khuhawar, M. Y., & Rajper, A. D. (2003). Liquid chromatographic determination of gamma-aminobutyric acid in cerebrospinal fluid using 2-hydroxynaphthaldehyde as derivatizing reagent. Journal of Chromatography. b, Analytical Technologies in the Biomedical and Life Sciences, 788(2), 413–418. https://doi.org/10.1016/s1570-0232(03)00062-x
Knedel M, Böttger R (1967) [A kinetic method for determination of the activity of pseudocholinesterase (acylcholine acyl-hydrolase 3.1.1.8.)] Klin Wochenschr 45(6):325–7. https://doi.org/10.1007/bf01747115
Komárek J, Anagnostidis K (2005) Süsswasserflora von Mitteleuropa. Cyanoprokaryota: 2. Teil/2nd Part: Oscillatoriales. Vol. 19. München: Elsevier Spektrum Akademischer Verlag.
Landsberg, J. H. (2002). The effects of harmful algal blooms on aquatic organisms. Fisheries Science, 10, 113–390. https://doi.org/10.1080/20026491051695
Lewis, R. S. (2011). Store-operated calcium channels: New perspectives on mechanism and function. Cold Spring Harbor Perspectives in Biology, 3, a0039701. https://doi.org/10.1101/cshperspect.a003970
Li, B., Liu, Y., Zhang, H., Liu, Y., Liu, Y., & Ping Xie, P. (2021). Research progress in the functionalization of microcystin-LR based on interdisciplinary technologies. Coordination Chemistry Reviews, 443, 214041. https://doi.org/10.1016/j.ccr.2021.214041
Liang, J., Li, T., Zhang, Y. L., Guo, Z. L., & Xu, L. H. (2011). Effect of microcystin-LR on protein phosphatase 2A and its function in human amniotic epithelial cells. Journal of Zhejiang University. Science. B, 12(12), 951–960. https://doi.org/10.1631/jzus.B1100121
Lires-Deán, M., Caramés, B., Cillero-Pastor, B., Galdo, F., López-Armada, M., & Blanco, F. J. (2008). Anti-apoptotic effect of transforming growth factor-β1 on human articular chondrocytes: Role of protein phosphatase 2A. Osteoarthritis Cartilage, 16(11), 1370–1378. https://doi.org/10.1016/j.joca.2008.04.001
Liu, J., & Sun, Y. (2015). The role of PP2A-associated proteins and signal pathways in microcystin-LR toxicity. Toxicology Letters, 236(1), 1–7. https://doi.org/10.1016/j.toxlet.2015.04.010
Malbrouck, C., & Kestemont, P. (2006). Effects of microcystins on fish. Environmental and Chemistry, 25(1), 72–86. https://doi.org/10.1897/05-029r.1
Malécot, M., Mezhoud, K., Marie, A., Praseuth, D., Puiseux-Dao, S., & Edery, M. (2009). Proteomic study of the effects of microcystin-LR on organelle and membrane proteins in Medaka fish liver. Aquatic Toxicology, 94(2), 153–161. https://doi.org/10.1016/s0009-2797(02)00075-3
Mezhoud, K., Praseuth, D., Puiseux-Dao, S., Francois, J.-C., Bernard, C., & Edery, M. (2008). Global quantitative analysis of protein expression and phosphorylation status in the liver of the medaka fish (Oryzias latipes) exposed to microcystin-LR I. Balneation Study. Aquat Toxicol, 86(2), 166–175. https://doi.org/10.1016/j.aquatox.2007.10.010
Mikhailov, A., Härmälä-Braskén, A. S., Hellman, J., Meriluoto, J., & Eriksson, J. E. (2003). Identification of ATP-synthase as a novel intracellular target for microcystin-LR. Chemico-Biological Interactions, 142(3), 223–237. https://doi.org/10.1016/s0009-2797(02)00075-3
Olivares-Rubio, H. F., Martínez-Torres, M. L., Domínguez-López, M. L., García-Latorre, E., & Vega-López, A. (2013). Pro-oxidant and antioxidant responses in the liver and kidney of wild Goodea gracilis and their relation with halomethanes bioactivation. Fish Physiology and Biochemistry, 39(6), 1603–1617. https://doi.org/10.1007/s10695-013-9812-8
Olivares-Rubio, H. F., Martínez-Torres, M. L., Nájera-Martínez, M., Dzul-Caamal, R., Domínguez-López, M. L., García-Latorre, E., & Vega-López, A. (2015). Biomarkers involved in energy metabolism and oxidative stress response in the liver of Goodea gracilis Hubbs and Turner, 1939 exposed to the microcystin-producing Microcystis aeruginosa LB85 strain. Environmental Toxicology, 30(10), 1113–1124. https://doi.org/10.1002/tox.21984
Qian, H., Liu, G., Lu, T., & Sun, L. (2018). Developmental neurotoxicity of Microcystis aeruginosa in the early life stages of zebrafish. Ecotoxicology and Environmental Safety, 151, 35–41. https://doi.org/10.1016/j.ecoenv.2017.12.059
Parekh, A. B., & Putney, J. W. (2005). Store-operated calcium chan-nels. Physiological Reviews, 85, 757–810. https://doi.org/10.1152/physrev.00057.2003
Perry, S. W., Norman, J. P., Barbieri, J., Brown, E. B., & Gelbard, H. A. (2011). Mitochondrial membrane potential probes and the proton gradient: A practical usage guide. BioTechniques, 50(2), 98–115. https://doi.org/10.2144/000113610
Prakriya, M., & Lewis, R. S. (2015). Store-operated calcium channels. Physiological Reviews, 95(4), 1383–1436. https://doi.org/10.1152/physrev.00020.2014
Sanchez, W., Burgeot, T., & Porcher, J. (2013). A novel “Integrated biomarker response” calculation based on reference deviation concept. Environmental Science and Pollution Research, 20, 2721–2725. https://doi.org/10.1007/s11356-012-1359-1
Silberman, S. R., Speth, M., Nemani, R., Ganapathi, M. K., Dombradi, V., Paris, H., & Lee, E. Y. (1984). Isolation and characterization of rabbit skeletal muscle protein phosphatases C-I and C-II. Journal of Biological Chemistry, 259(5), 2913–2922.
Stathopulos, P. B., & Ikura, M. (2017). Store operated calcium entry: From concept to structural mechanisms. Cell Calcium, 63, 3–7. https://doi.org/10.1016/j.ceca.2016.11.005
Tang, Z. B., Chen, Y. Z., Zhao, J., Guan, X. W., Bo, Y. X., Chen, S. W., & Hui, L. (2016). Conjugates of podophyllotoxin and norcantharidin as dual inhibitors of topoisomerase II and protein phosphatase 2A. European Journal of Medicinal Chemistry, 123, 568–576. https://doi.org/10.1016/j.ejmech.2016.07.031
Toivola, D. M., Erikssonb, J. E., & Brautigan, D. L. (1994). Identification of protein phosphatase 2A as the primary target for microcystin-LR in rat liver homogenates. FEBS Letters, 344, 175–180. https://doi.org/10.1016/0014-5793(94)00382-3
Vega-López, A., Pagadala, N. S., López-Tapia, B. P., Madera-Sandoval, R. L., Rosales-Cruz, E., Nájera-Martínez, M., & Reyes-Maldonado, E. (2019). Is related the hematopoietic stem cells differentiation in the Nile tilapia with GABA exposure? Fish & Shellfish Immunology, 93, 801–814. https://doi.org/10.1016/j.fsi.2019.08.032
Wang, G., Dong, J., & Deng, L. (2018). Overview of cantharidin and its analogues. Current Medicinal Chemistry, 25(17), 2034–2044. https://doi.org/10.2174/0929867324666170414165253
Wang H, Xu C, Liu Y, Jeppesen E, Svenning JC, Wu J, Zhang W, Zhou T, Wang P, Nangombe S, Ma J, Duan H, Fang J, Xie P. (2021) From unusual suspect to serial killer: Cyanotoxins boosted by climate change may jeopardize megafauna. Innovation (N Y). 2(2):100092. https://doi.org/10.1016/j.xinn.2021.100092
Wiegand, C., & Pflugmacher, S. (2005). Ecotoxicological effects of selected cyanobacterial secondary metabolites a short review. Toxicology and Applied Pharmacology, 203(3), 201–218. https://doi.org/10.1016/j.taap.2004.11.002
Wu, Q., Yan, W., Liu, C., Li, L., Yu, L., Zhao, S., & Li, G. (2016). Microcystin-LR exposure induces developmental neurotoxicity in zebrafish embryo. Environmental Pollution, 213, 793–800. https://doi.org/10.1016/j.envpol.2016.03.048
Wu, Q., Yan, W., Cheng, H., Liu, C., Hung, T. C., Guo, X., & Li, G. (2017). Parental transfer of microcystin-LR induced transgenerational effects of developmental neurotoxicity in zebrafish offspring. Environmental Pollution, 231(Pt 1), 471–478. https://doi.org/10.1016/j.envpol.2017.08.038
Zhang, Y., Zhang, J., Wang, E., Qian, W., Fan, Y., Feng, Y., Yin, H., Li, Y., Wang, Y., & Yuan, T. (2018). Microcystin-leucine-arginine induces tau pathology through Bα degradation via protein phosphatase 2A demethylation and associated glycogen synthase kinase-3β phosphorylation. Toxicological Sciences, 162(2), 475–487. https://doi.org/10.1093/toxsci/kfx271
Zhang, H., Li, B., Liu, Y., Chuan, H., Liu, Y., & Xie, P. (2022). Immunoassay technology: Research progress in microcystin-LR detection in water samples. Journal of Hazardous Materials, 424(Pt B), 127406. https://doi.org/10.1016/j.jhazmat.2021.127406
Zsombok, A., Schrofner, S., Hermann, A., & Kerschbaum, H. H. (2005). A cGMP-dependent cascade enhances an L-type-like Ca2+ current in identified snail neurons. Brain Research, 1032(1–2), 70–76. https://doi.org/10.1016/j.brainres.2004.11.003
Funding
This study was supported by Instituto Politécnico Nacional SIP code 20180917 and SIP code 20201044. M. Najera-Martínez is a graduate DSc in Chemobiological Sciences which received financial support by a postdoctoral fellowship granted by CONACyT, México. G.G. Landon-Hernández is a graduate BSc student. J.P. Romero-López is a graduate DSc in Chemobiological Sciences who received scholarship from CONACyT and BEIFI-IPN. M.L. Domínguez-López and A. Vega-López are fellow of Estímulos al Desempeño en Investigación and Comisión y Fomento de Actividades Académicas (Instituto Politécnico Nacional) and Sistema Nacional de Investigadores (SNI, CONACyT, México).
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MNM, GGLH, and JPRL conducted the experiments; MNM, MLDL, and AVL designed the experiments and interpreted the results; and AVL wrote the manuscript.
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Nájera-Martínez, M., Landon-Hernández, G.G., Romero-López, J.P. et al. Disruption of Neurotransmission, Membrane Potential, and Mitochondrial Calcium in the Brain and Spinal Cord of Nile Tilapia Elicited by Microcystis aeruginosa Extract: an Uncommon Consequence of the Eutrophication Process. Water Air Soil Pollut 233, 6 (2022). https://doi.org/10.1007/s11270-021-05480-x
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DOI: https://doi.org/10.1007/s11270-021-05480-x