Persistent alterations of calmodulin kinase II activity in chickens after an oral dose of tri-o-cresyl phosphate.
Calmodulin kinase II has been found to be involved in the increased phosphorylation of brain microtubule and spinal cord neurofilament triplet proteins following treatment of animals with organophosphorus compounds that are capable of producing organophosphorus compound-induced delayed neurotoxicity (OPIDN). In this report, chickens were given a single oral neurotoxic dose of 750 mg/kg tri-o-cresyl phosphate (TOCP), and killed after 1 or 21 days of treatment. Crude calmodulin kinase II from brain cytosol as well as phosphocellulose-purified microtubules were prepared from control and treated animals. Phosphorylation reactions were started by adding protein into the phosphorylation buffer in the presence of Mg2+, Ca2+, calmodulin or trifluoperazine, and [gamma-32P]ATP. Proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subjected to autoradiography. The extent of the calmodulin kinase II autophosphorylation as well as the Ca2+/calmodulin-dependent phosphorylation of the purified microtubules was investigated. The enzyme activities isolated from control and treated animals were compared. Autophosphorylation of calmodulin kinase II was found to be higher in both 1-day and 21-day TOCP-treated animals than in control animals. The activity of the kinase to phosphorylate exogenous substrates such as tubulin and microtubule-associated protein-2 (MAP-2) was also higher in the treated hens than in the controls. The increased activity of the kinase was noted at day 1 following treatment when no clinical signs were observed and persisted until day 21 when the animals were paralyzed completely. This finding supports the significance of altered calmodulin kinase II in the pathogenesis of OPIDN.
Lapadula
E S
ES
Department of Pharmacology, Duke University Medical Center, Durham, NC 27710.
Lapadula
D M
DM
Abou-Donia
M B
MB
D000818:Animals; D001259:Ataxia / Q000139:chemically induced / Q000201:enzymology; D001923:Brain Chemistry; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005260:Female; D007381:Intermediate Filament Proteins / Q000032:analysis; D007525:Isoelectric Focusing; D016900:Neurofilament Proteins; D010243:Paralysis / Q000139:chemically induced / Q000201:enzymology; D010750:Phosphoproteins / Q000032:analysis; D010766:Phosphorylation; D011494:Protein Kinases / Q000032:analysis*; D014317:Tritolyl Phosphates / Q000008:administration & dosage / Q000633:toxicity*
D007381:Intermediate Filament Proteins; D016900:Neurofilament Proteins; D010750:Phosphoproteins; D014317:Tritolyl Phosphates; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases