No study to date has addressed biochemical, histopathological and clinical differences in hens given the same dose of the three isoforms of methamidophos, nor have they addressed potential treatments. The aim of this study was to evaluate the acute and delayed effects of methamidophos enantiomers and TOCP (a positive inducer of delayed neuropathy) measuring AChE, NTE and calpain activities in brain, neuropathological damages in spinal

cord and signs of ataxia as biochemical, histopathological and clinical indicators, respectively. In addition, the benefit of nimodipine and Ca-glu on OPIDN in methamidophos-treated hens was also investigated. Racemic methamidophos, nimodipine, sodium dodecyl sulfate (SDS), paraoxon, bovine serum albumin (BSA), dl-dithiothreitol (DTT), ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetra-acetic acid (EGTA), Selleck PLX4720 2-mercaptoethanol, casein, DEAE cellulose, coomassie brilliant blue G-250, tris(hydroxymethyl) aminomethane, ethylenediaminetetraacetic HDAC inhibitor acid (EDTA), phosphoric acid 85%, acetylthiocholine (ACTh) and 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) were purchased from Sigma, St Louis, MO, USA; TOCP was purchased from Acros Organics, Pittsburg, PA, USA; mipafox and phenyl valerate were obtained from Oryza Laboratories, Inc., Chelmsford, MA, USA; sodium citrate and triton X-100 were purchased from Rhiedel-de Haën, Hannover, Germany; 4-aminoantipyrine, potassium ferricyanide, and

dimethylformamide were purchased from Merck, Darmstadt, Germany; calcium gluconate (Ca-glu) 10% (w/v) injectable ampoules from Hypofarma, Ribeirão das Neves, MG, Brazil; deltametrin (K-otrine®) was obtained from Bayer Cropscience Ltda, Rio de Janeiro, RJ, Brazil; and piperazine citrate (Proverme®) was purchased from Tortuga Agrarian Zootechnical Company, São Paulo, Brazil; ampoule of ketamine 10% was purchased from Agener União, Embu Guaçu, SP, Brazil. The enantiomeric separation of (±)-methamidophos G protein-coupled receptor kinase was conducted according to the method described by Emerick et al. (2012b). The enantiomers of methamidophos were obtained with 99.5% of optical purity for the (+)-methamidophos and 98.3% of optical purity for the (−)-methamidophos.

All other chemicals employed in this study were of analytical grade. Thirty-nine isabrown leghorn hens (aged 70–90 weeks, weighing 1.5–2.0 kg) were obtained from the Hayashi farm cooperative of Guatapará, SP, Brazil. Before the experiments were initiated, the hens were treated to eliminate ecto-parasites (using deltametrin) and endo-parasites (using piperazine citrate), as described elsewhere ( DeOliveira et al., 2002 and Emerick et al., 2010). After this treatment (1 month), the hens were housed at a density of 3 per cage in a temperature- and humidity-controlled room (24 ± 2 °C and 55% ± 10 RH) on an automatic 12:12 light–dark photocycle with lights activated at 8 a.m. Purina® feed and filtered tap water were provided ad libitum.