Electrophysiological Evaluation of Paralytic Shellfish Toxins in Bivalves

Paralytic shellfish toxins (PSTs) represented by saxitoxin (STX) are lethal; pre-shipment toxicity testing of shellfish is mandatory. In Japan, screening methods that reliably guarantee toxin concentrations below shipment regulatory limits (0.8 mg STX·2HCl eq./kg) are permitted. In this study, we aimed to develop an electrophysiological method for PSTs by utilizing their property of inhibiting voltage-gated sodium channels (Na_v) expressed in mouse neuroblastoma Neuro 2a cells. In addition to the three reference standards of dcSTX, GTX2/3, and C1/2, extracts from the digestive glands of toxic Patinopecten yessoensis and those from the whole edible parts of P. yessoensis, Mimachlamys nobilis, Perna viridis, and Mytilus galloprovincialis were tested. As a result, the toxicity required to inhibit Na_v by 50% was 1-4 µg STX·2HCl eq./kg, which was well below the shipment regulatory limits and roughly consistent across the three reference standards and all the bivalve extracts. A calibration curve was constructed by globally fitting the dose-response curves obtained from electrophysiological experiments using various bivalve extracts. The electrophysiological toxicity of each extract was then estimated using this calibration curve. The correlation coefficient (r) between the electrophysiological toxicity and the toxicity determined by LC-MS/MS was 0.90. Thus, these findings provide fundamental insights for establishing an electrophysiological screening method for PSTs.