Metabolic Effects of Sodium Fluorocitrate and Potential Treatments on Saccharomyces Cerevisiae: Changes in Oxygen Consumption

Abstract

Sodium fluoroacetate (1080) is a colorless, odorless, tasteless, water‐soluble metabolic poison. In the body, fluoroacetate is converted to fluorocitrate (FC) which blocks the citric acid cycle and drastically decreases ATP production. Symptoms of exposure include nausea, vomiting, abdominal pains, salivation, irrational fear, weakness, tachypnea, cyanosis, sweating, increased temperature, and death. Signs and symptoms of 1080 poisoning are nonspecific, further complicating the diagnosis. 1080 is tightly regulated within the United States, but it is commonly used in Australia, New Zealand, Mexico, Japan, South Korea, and Israel as a rodenticide to control invasive and predatory species. An eco‐terrorist scare in New Zealand (2015) and findings reported by the CIA (2007) have led 1080 to being identified as a potential terrorist weapon.To identify potential countermeasures against 1080 poisoning, a Saccharomyces cerevisiae (yeast) model was selected to assess the impact of FC on cellular respiration and to screen candidate therapeutics. In response to FC, yeast showed depressed mitochondrial activity as measured by oxygen consumption rate (OCR). Potential treatments, thonzonium bromide, nifuroxazide, pyrvinium pamoate, menadione, mefloquine, clemastine fumarate, bisacodyl, L‐ascorbic acid, thiethylperazine and vinpocitine, were examined in this study. All treatments are approved by the Food and Drug Administration. Toxicity of each treatment was also assessed.Assays were performed in yeast extract peptone galactose media, a metabolically restrictive media. To assess the effect of FC on OCR, cells were exposed to 50 µM, 100 µM, and 200 µM FC in liquid culture for 1, 4, or 24 hours. After incubation, mitochondrial respiration was assessed via the MitoXpress assay (Agilent), a fluorescence‐based assay that measures OCR. Once established as a viable model, yeasts were then used to screen therapeutics to improve cellular respiration. Yeast was co‐incubated with fluorocitrate (200 µM), and potential treatments at three increasing concentrations for 4 hours and the effects on mitochondrial respiration were assessed via the MitoXpress assay.Following the decreased OCR in yeast exposed to FC, 6 of the 10 treatments successfully restored the mitochondrial function of yeast as demonstrated by an increase in OCR. As a result, the compounds nifuroxazide, pyrvinium pamoate, vinpocitine, clemastine fumarate, bisacodyl, and L‐ascorbic acid are moving forward for further testing in cardiomyocytes and hepatic spheroids, with potential in vivo assessment, to characterize their effectiveness against 1080 poisoning. Future studies will continue to use yeast as a valuable screening method to identify other potential treatments for sodium fluoroacetate.