Comparison of the toxicokinetics of the convulsants picrotoxinin and tetramethylenedisulfotetramine (TETS) in mice

Abstract

Acute intoxication with picrotoxin or the rodenticide tetramethylenedisulfotetramine (TETS) can cause seizures that rapidly progress to status epilepticus and death. Both compounds inhibit γ-aminobutyric acid type-A (GABA A) receptors with similar potency. However, TETS is approximately 100 × more lethal than picrotoxin. Here, we directly compared the toxicokinetics of the two compounds following intraperitoneal administration in mice. Using LC/MS analysis we found that picrotoxinin, the active component of picrotoxin, hydrolyses quickly into picrotoxic acid, has a short in vivo half-life, and is moderately brain penetrant (brain/plasma ratio 0.3). TETS, in contrast, is not metabolized by liver microsomes and persists in the body following intoxication. Using both GC/MS and a TETS-selective immunoassay we found that mice administered TETS at the LD 50 of 0.2 mg/kg in the presence of rescue medications exhibited serum levels that remained constant around 1.6 μM for 48 h before falling slowly over the next 10 days. TETS showed a similar persistence in tissues. Whole-cell patch-clamp demonstrated that brain and serum extracts prepared from mice at 2 and 14 days after TETS administration significantly blocked heterologously expressed α 2 β 3 γ 2 GABA A-receptors confirming that TETS remains pharmacodynamically active in vivo. This observed persistence may contribute to the long-lasting and recurrent seizures observed following human exposures. We suggest that countermeasures to neutralize TETS or accelerate its elimination should be explored for this highly dangerous threat agent.

Protective Effects of Crude Extract and Fractions of Newbouldia laevis Leaves in Chemoconvulsant-Induced Seizures

“…Picrotin is known to be inert, and as such, the convulsant activity of picrotoxin is attributable to its picrotoxinin moiety. 28 Some compounds that antagonize picrotoxin effect are known to be effective anticonvulsant agents. For example, benzodiazepines are effective antagonists of convulsions produced by picrotoxin, but they are not effective against convulsions induced by strychnine and other agents that operate independently of GABA system.…”

Identification of the Functional Binding Site for the Convulsant Tetramethylenedisulfotetramine in the Pore of the α2β3γ2 GABAA Receptor

“…Ten millimolar stocks of GABA were made fresh daily using Ringer’s solution (see below for composition). Ten millimolar stocks of picrotoxinin and TETS were prepared in DMSO and diluted down into Ringer’s solution only immediately before application onto the cell to avoid any hydrolysis of picrotoxinin ( Pressly et al, 2020 ). The identity of TETS and picrotoxinin was confirmed by 1 H- and 13 C-NMR; purity of TETS was tested by GC/mass spectrometry or high-pressure liquid chromatography/mass spectrometry and found to be >98% based on total ion fragment analysis.…”

Revision of the Unstable Picrotoxinin Hydrolysis Product

“…Higher pH causes irreversible degradation, purportedly through intramolecular ring opening of the 8,9‐epoxide ( 2 a ) by the C3 alkoxide or by an irreversible second lactone hydrolysis ( 3 a ). Decay of picrotoxinin has taken on new significance with the recent discovery that pH 7.4 buffer supports only a 45‐minute half‐life at room temperature [13–15] . Whereas picrotoxinin is a potent GABA A receptor antagonist, its hydrolysis products 2 a and 3 a lack activity, so care must be taken in the preparation and storage of picrotoxinin solutions [13, 14] .…”

“…Decay of picrotoxinin has taken on new significance with the recent discovery that pH 7.4 buffer supports only a 45‐minute half‐life at room temperature [13–15] . Whereas picrotoxinin is a potent GABA A receptor antagonist, its hydrolysis products 2 a and 3 a lack activity, so care must be taken in the preparation and storage of picrotoxinin solutions [13, 14] . Hydrolysis occurs even faster in vivo (mouse half‐life near 15 min), [14] accounting for large variation in seizure‐induction depending on the route of administration [14] .…”

“…The structures of hydrolysis ( 2 a / 3 a ) and methanolysis products ( 2 b / 3 b ) repeatedly prompted the proposal [11a, 13–15, 20] that the initial, unstable carboxylate corresponds to structure 4 : at pH 7–9, this carboxylate would persist, but further basification would cause transannular cyclization to 2 a . On the face of it, this proposal makes sense since both isolated products lack the C15 lactone.…”