Cytochrome P450 2B (CYP2B)-mediated activation of methyl-parathion in rat brain extracts.
The role of cytochrome P450 (CYP) and the CYP isoform involved in the activation of the widely used pesticide methyl-parathion (MePA) were investigated in rat brain extracts by measuring the effect of different CYP inhibitors on acetylcholinesterase (AChE) inhibition by MePA. Brain extracts provide a useful tool to study the activation mechanisms of organophosphorus compounds (OP) since they contain both the activating enzyme(s) and the molecular target for OP toxicity. As expected, in incubations of rat brain extract supplemented with NADPH, AChE activity was non-competitively inhibited by the presence of MePA, indicating that MePA was activated to its reactive metabolite methyl-paraoxon (MePO). Indeed, Vmax(app) decreased from 13.4 to 8.7 micromol thionitrobenzoic acid (TNB)/min per mg protein. MePA activation by rat brain extracts, as measured by the AChE inhibition produced by the presence of the pesticide in the incubation, was fully prevented by previously bubbling the incubation mix with CO, by the presence of monoclonal anti-rat CYP2B1/2B2 antibodies and by the addition of phenobarbital (PB), a CYP2B substrate. Interestingly, MePA showed a greater affinity for CYP2B than PB. CYP1A1 antibodies showed no effect on MePA activation. The presence of cytochrome P450 2B (CYP2B) in the rat brain extracts was confirmed by immunoblotting. These results demonstrate indisputably the responsibility of CYP2B in MePA activation in the rat brain in vitro, suggesting that metabolic activation of OP compounds in situ might be crucial for their organ specific toxicity to the central nervous system also in vivo.
Albores
A
A
Sección de Toxicología, Centro de Investigación y de Estudios Avanzados del IPN, Avenida IPN 2508, Col. San Pedro Zacatenco, Mexico City 07360, Mexico. aalbores@mail.cinvestav.mx
Ortega-Mantilla
G
G
Sierra-Santoyo
A
A
Cebrián
M E
ME
Muñoz-Sánchez
J L
JL
Calderón-Salinas
J V
JV
Manno
M
M
D000110:Acetylcholinesterase / Q000378:metabolism; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D001921:Brain / Q000187:drug effects* / Q000201:enzymology; D002800:Cholinesterase Inhibitors / Q000009:adverse effects / Q000494:pharmacology*; D065702:Cytochrome P-450 CYP2B6; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004790:Enzyme Induction; D018846:Excitatory Amino Acids / Q000494:pharmacology; D007527:Isoenzymes; D008297:Male; D008743:Methyl Parathion / Q000009:adverse effects / Q000494:pharmacology*; D009249:NADP / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism*; D010634:Phenobarbital / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar
D002800:Cholinesterase Inhibitors; D018846:Excitatory Amino Acids; D007527:Isoenzymes; D008743:Methyl Parathion; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; D065702:Cytochrome P-450 CYP2B6; D010089:Oxidoreductases, N-Demethylating; D000110:Acetylcholinesterase; D010634:Phenobarbital