Academic journal article Environmental Health Perspectives

Investigation of Microcystin Congener-Dependent Uptake into Primary Murine Neurons

Academic journal article Environmental Health Perspectives

Investigation of Microcystin Congener-Dependent Uptake into Primary Murine Neurons

Article excerpt

BACKGROUND: Contamination of natural waters by toxic cyanobacteria is a growing problem worldwide, resulting in serious water pollution and human health hazards. Microcystins (MCs) represent a group of > 80 cyclic heptapeptides, mediating cytotoxicity via specific protein phosphatase (PP) inhibition at equimolar concentrations (comparable toxicodynamics). Because of the structure and size of MCs, active uptake into cells occurs via organic anion-transporting polypeptides (OATP/Oatp), as confirmed for liver-specific human OATP1B1 and OATP1B3, mouse Oatp1b2 (mOatp 1b2), skate Oatp 1d1, and the more widely distributed OATP1A2 expressed, for example, at the blood--brain barrier. Tissue-specific and cell-type--specific expression of OATP/Oatp transporters and specific transport of MC congeners (toxicokinetics) therefore appear prerequisite for the reported toxic effects in humans and other species upon MC exposure. Beyond hepatotoxicity induced by the MC-LR congener, the effects of other MC congeners, especially neuronal uptake and toxicity, are unknown.

OBJECTIVES: In this study we examined the expression of mOatps and the uptake of congeners MC-LR, MC-LW, and MC-LF in primary murine neurons.

METHODS: Intracellular MC accumulation was indicated indirectly via uptake inhibition experiments and directly confirmed by Western blot analysis and a PP inhibition assay. Neuronal mOatp expression was verified at the mRNA and protein level.

RESULTS: MCs can cross neuronal cell membranes, with a subsequent decrease of PP activity. Of 15 mOatps, 12 were expressed at the mRNA level, but we found detectable protein levels for only two: mOatp 1a5 (Slco 1a5) and the known MC-LR transporter mOatp 1b2 (Slco 1b2).

CONCLUSIONS: These data suggest mOatp-mediated uptake of MC congeners into neurons, thus corroborating earlier assumptions of the neurotoxic potential of MCs.

KEY WORDS: cyanobacteria, microcystin congeners, neurotoxicity, organic anion-transporting polypeptides. Environ Health Perspect 118:1370-1375 (2010). doi:10.1289/ehp.0901289 [Online 14 May 2010]

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Cyanobacteria are abundant in marine, brackish, and fresh waters. During the past decades, cyanobacterial poisonings of aquatic and terrestrial animals as well as humans (Kuiper-Goodman et al. 1999) have been reported worldwide, especially in conjunction with cyanobacterial blooms in water reservoirs, rivers, lakes, and ponds used for drinking water or recreational purposes or as water resources for livestock. Current research suggests that climate change, specifically global warming, will promote the incidence and severity of cyanobacterial mass occurrences (Carmichael 2008; Paerl and Huisman 2008; Paul 2008), thereby increasing the potential threat posed by cyanobacterial toxins to human health and livestock (World Health Organization 1999).

Microcystin (MC), the most common freshwater cyanotoxin, represents a group of cyclic heptapeptides encompassing > 80 structural variants (Meriluoto and Spoof 2008), with molecular weights ranging between 900 and 1,100 Da (Zurawell et al. 2005). Their inherent capability for inhibiting serine/threonine-specific protein phosphatases (PPs; e.g., PP1, PP2A, PP4, PP5) via covalent binding to the PP catalytic subunit (Hastie et al. 2005) results, once taken up into the cell, in a disruption of cellular phosphorylation/dephosphorylation homeostasis. This in turn leads to several downstream responses, such as disintegration of cytoskeletal structure, inhibition of gluconeogenesis, and enhanced glycolysis (Batista et al. 2003; Falconer and Yeung 1992), frequently with later onset of apoptosis and necrosis (Fiadmark et al. 1999; Komatsu et al. 2007). However, to exert the MC-specific PP inhibition, sufficient concentrations of MC must enter the cell. Cellular uptake of MCs has been demonstrated to occur exclusively via an active transport, whereas passive transmembrane diffusion can be excluded (Eriksson et al. …

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