A rapid assay for the brevetoxin group of sodium channel activators based on fluorescence monitoring of synaptoneurosomal membrane potential.
A functional pharmacologically-based assay for the brevetoxin group of sodium channel activators was developed using synaptoneurosomes isolated from the brains of CD1 mice. The assay can detect the depolarizing effect of brevetoxin congeners PbTx-2 and PbTx-3 as enhancements of the veratridine-dependent increase in fluorescence of the voltage-sensitive fluorescent probe rhodamine 6G. The assay is relatively rapid and can detect brevetoxin activity in the nanomolar range. The synaptoneurosomal assay has been used to analyse mussel tissue extracts spiked with PbTx-2, and composite toxicity, expressed as PbTx-3 equivalents in extracts of oysters naturally exposed to brevetoxins. In this latter context, the synaptoneurosomal technique was shown to compare favorably with the cytotoxicity assay, the receptor binding assay and HPLC/MS. Our results support the concept that this membrane potential assay detects brevetoxins based on their interaction with sodium channels.
David
Laurence S
LS
Department of Biological Sciences, Simon Fraser University, 8888 University Drive, Burnaby, BC, Canada V5A 1S6.
Plakas
Steven M
SM
El Said
Kathleen R
KR
Jester
Edward L E
EL
Dickey
Robert W
RW
Nicholson
Russell A
RA
D000818:Animals; D049872:Bivalvia / Q000737:chemistry; D004141:Dinoflagellida / Q000737:chemistry; D005453:Fluorescence; D005456:Fluorescent Dyes; D008297:Male; D008387:Marine Toxins / Q000032:analysis* / Q000633:toxicity*; D008564:Membrane Potentials / Q000187:drug effects*; D051379:Mice; D008815:Mice, Inbred Strains; D010125:Ostreidae / Q000737:chemistry; D035964:Oxocins / Q000032:analysis* / Q000633:toxicity*; D012016:Reference Values; D012235:Rhodamines; D012680:Sensitivity and Specificity; D062687:Sodium Channel Agonists; D015222:Sodium Channels / Q000378:metabolism; D013574:Synaptosomes / Q000187:drug effects*; D013997:Time Factors; D014701:Veratridine / Q000494:pharmacology; D000095702:Polyether Toxins
D005456:Fluorescent Dyes; D008387:Marine Toxins; D035964:Oxocins; D012235:Rhodamines; D062687:Sodium Channel Agonists; D015222:Sodium Channels; D014701:Veratridine; C051732:Ptychodiscus brevis T2 toxin; C026188:rhodamine 6G; C053342:brevetoxin; D000095702:Polyether Toxins