Acetylcholinesterase inhibitors anatoxin

An enzymatic assay can also be used for detecting anatoxin-a(s). " This toxin inhibits acetylcholinesterase, which can be measured by a colorimetric reaction, i.e. reaction of the acetyl group, liberated enzymatically from acetylcholine, with dithiobisnitrobenzoic acid. The assay is performed in microtitre plates, and the presence of toxin detected by a reduction in absorbance at 410 nm when read in a plate reader in kinetic mode over a 5 minute period. The assay is not specific for anatoxin-a(s) since it responds to other acetylcholinesterase inhibitors, e.g. organophosphoriis pesticides, and would need to be followed by confirmatory tests for the cyanobacterial toxin. 

Anatoxin-a(s) inhibits acetylcholinesterase by acting as an irreversible active-site-directed inhibitor [61]. This prevents degradation of ACh and leads to over-stimulation of the muscle cells (Figure 6.1) [56,62]. Thus, although the mechanism of action of anatoxin-a(s) is quite different from that of anatoxin-a, the observed toxicity is similar. In addition, it was the first irreversible acetylcholinesterase inhibitor to be found in a cyanobacterium. 

It is also important to mention the use of the reactivation of the acetylcholinesterase by pyridine-2-aldoxime methochloride to discriminate between the toxin and potential insecticides [96]. Once phos-phorylated, the active site serine of the enzyme can be reactivated by powerful nucleophilic agents such as oximes. However, this reactivation is not possible if attempted too late due to the stable adduct formed by the dealkylation (aging) of the inhibitor s remaining group. When acetylcholinesterase is inhibited by anatoxin-a(s), it shows immediately the characteristics of an aged enzyme and cannot be reactivated. In this way, it is possible to distinguish between the inhibition caused by anatoxin-a(s) and the one provoked by other insecticides. 

Hyde, E.G., and Carmicheal, W.W. 1991. Anatoxin-a(s), a naturally occorring organophosphate, is an irreversible active site directed inhibitor of acetylcholinesterase (EC 3.1.1.T). JBiochem Toxicol 6, 195-201. 

Anatoxin-a(s) acts as an irreversible inhibitor of acetylcholinesterase, reducing blood pressure and heart rate in rats dramatically. " It has been identified as the active agent in a number of animal poisonings, including the deaths of numerous birds in 1993, which coincided with huge cyanobacterial blooms in freshwater lakes. "" As a result, a biosensor has recently been developed for the detection of 61 in environmental samples. " ... 

To date, anatoxin-a(S) is the only known natural organophosphate inhibitor of the neuronal enzyme acetylcholinesterase. The freshwater environment... 

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