Chronic In Vivo Effects of Repeated Exposure to Low Oral Doses of Tetrodotoxin: Preliminary Evidence of Nephrotoxicity and Cardiotoxicity.

Tetrodotoxin (TTX) is one of the most potent naturally occurring neurotoxins. InitiallyTTX was associated with human food intoxications in Japan, but nowadays, concerns about thehuman health risks posed by TTX have increased in Europe after the identification of the toxin infish, marine gastropods, and bivalves captured in European waters. Even when TTX monitoring isnot currently performed in Europe, an acute oral no observable effect level (NOAEL) of 75 µg/kghas been recently established but, to date, no studies evaluating the chronic oral toxicity of TTXhave been released, even when EFSA has highlighted the need for them. Thus, in this work, thechronic effects of low oral TTX doses (below the acute lethal dose 50) were evaluated followinginternationally adopted guidelines. The results presented here demonstrate that low oral doses ofTTX have deleterious effects on renal and cardiac tissues. Moreover, alterations in bloodbiochemistry parameters, urine production, and urinalysis data were already detected at the oraldose of 75 µg/kg after the 28 days exposure. Thus, the data presented here constitute an initialapproach for the chronic evaluation of the in vivo toxicity of tetrodotoxin after its ingestion throughcontaminated fishery products.

A fluorimetric microplate assay for detection and quantitation of toxins causing paralytic shellfish poisoning.

Paralytic shellfish poisoning is one of the most severe forms of food poisoning. The toxins responsible for this type of poisoning are metabolic products of dinoflagellates, which block neuronal transmission by binding to the voltage-gated Na(+) channel. Accumulation of paralytic toxins in shellfish is an unpredictable phenomenon that necessitates the implementation of a widespread and thorough monitoring program for mollusk toxicity. All of these programs require periodical collection and analysis of a wide range of shellfish. Therefore, development of accurate analytical protocols for the rapid determination of toxicity levels would streamline this process. Our laboratory has developed a fluorimetric microplate bioassay that rapidly and specifically determines the presence of paralytic shellfish toxins in many seafood samples. This method is based on the pharmacological activity of toxins and involves several steps: (i) Incubation of excitable cells in 96 well microtiter plates with the fluorescent dye, bis-oxonol, the distribution of which across the membrane is potential-dependent. (ii) Cell depolarization with veratridine, a sodium channel-activating toxin. (iii) Dose-dependent inhibition of depolarization with saxitoxin or natural samples containing paralytic shellfish toxins. Measuring toxin-induced changes in membrane potential allowed for quantification and estimation of the toxic potency of the samples. This new approach offers significant advantages over classical methods and can be easily automated.