Differential changes in rat brain nitric oxide synthase in vivo and in vitro by methylmercury.
Alterations in mRNA level, protein content and enzyme activity for nitric oxide synthase (NOS) in the cerebrum and cerebellum during a continuous exposure of neurotoxic metal, methylmercury, were examined in Wistar rats. Subcutaneous (s.c.) administration of methylmercuric chloride (MMC, 10 mg kg-1 day-1, 8 days) resulted in significant increases with time of NOS activities in the cerebrum (1. 6-1.9-fold, 5-8 days) and cerebellum (1.4-fold, 8 days). RT-PCR and immunoblot analyses indicated that the increase in the enzyme activity caused by this metal appears to be due to increase in protein levels of neuronal NOS (nNOS), but not inducible NOS (iNOS) because little appreciable mRNA and protein for iNOS were seen during MMC exposure. The direct effect of mercuric compounds on nNOS activity in vitro was evaluated using 20,000xg supernatant from rat cerebellum homogenate. In contrast to the in vivo observation, inorganic-, alkyl-, and aryl-mercuric compound showed potent inhibition of nNOS activity with IC50 values of 11-43 microM, whereas dimethylmercury (DMM) was without effect on the enzyme activity. Further experiments indicated that the inhibition of nNOS by organomercurial occurred via thiol modification.
Graduate School Doctoral Program in Medical Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305, Japan.
M
M
Shinyashiki
Y
Y
Kumagai
H
H
Nakajima
J
J
Nagafune
S
S
Homma-Takeda
M
M
Sagai
N
N
Shimojo
D000818:Animals; D001921:Brain / Q000187:drug effects* / Q000201:enzymology*; D002531:Cerebellum / Q000187:drug effects / Q000201:enzymology; D015151:Immunoblotting; D066298:In Vitro Techniques; D008297:Male; D008767:Methylmercury Compounds / Q000494:pharmacology*; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D014158:Transcription, Genetic
D008767:Methylmercury Compounds; D012333:RNA, Messenger; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; C496322:Nos1 protein, rat; C496318:Nos2 protein, rat; C004925:methylmercuric chloride