The synergism of n-hexane-induced neurotoxicity by methyl isobutyl ketone following subchronic (90 days) inhalation in hens: induction of hepatic microsomal cytochrome P-450

Abstract

The effect of methyl isobutyl ketone (MiBK) on n-hexane-induced neurotoxicity was investigated via inhalation in seven groups of five hens each for 90 days followed by a 30-day observation period. One group was exposed to vapors containing 1000 ppm n-hexane and another group to vapors having 1000 ppm MiBK. Four groups were exposed simultaneously to 1000 ppm of n-hexane and 100, 250, 500, or 1000 ppm MiBK. Another group was exposed similarly to ambient air in an exposure chamber and used as a control. Hens continuously exposed to 1000 ppm MiBK developed leg weakness with subsequent recovery, while inhalation of the same concentration of n-hexane produced mild ataxia. Hens exposed to mixtures of n-hexane and MiBK developed clinical signs of neurotoxicity, the severity of which depended on the MiBK concentration. Thus, all hens exposed to 1000 ppm n-hexane in combination with 250, 500, or 1000 ppm MiBK progressed to paralysis. Hens continuously exposed to 1000/100 n-hexane/MiBK showed severe ataxia which did not change during the observation period. The neurologic dysfunction in hens exposed simultaneously to n-hexane and MiBK was accompanied by large swollen axons and degeneration of the axon and myelin of the spinal cord and peripheral nerves. The results indicate that the nonneurotoxic chemical MiBK synergized the neurotoxic action of the weak neurotoxicant n-hexane since the coneurotoxicity coefficient for joint exposure was more than two times the additive effect of each treatment alone. In another experiment, to investigate the mechanism of MiBK synergism of n-hexane neurotoxicity, continuous inhalation for 50 days of 1000 ppm n-hexane had no effect on hen hepatic microsomal enzymes, whereas inhalation of 1000 ppm MiBK for 50 days or a mixture of 1000 ppm of each of n-hexane and MiBK for 30 days significantly induced aniline hydroxylase activity and cytochrome P-450 contents in hen liver microsomes. Liver microsomal proteins from these hens and from hens treated with beta-naphthoflavone (beta-NF) and phenobarbital (PB) were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. While beta-NF increased the 55-kDa band (1408%), PB, MiBK, and MiBK/n-hexane increased the protein band (49 kDa) (258, 335, and 253%, respectively), indicating that MiBK induces chicken hepatic cytochrome P-450. The results suggest that the synergistic action of MiBK on n-hexane neurotoxicity may be related to its ability to induce liver microsomal cytochrome P-450, resulting in increased metabolic activation of n-hexane to more potent neurotoxic metabolites.