Strain differences in diazepam metabolism at its three metabolic sites in sprague-dawley, brown norway, dark agouti, and wistar strain rats.
Knowledge of strain differences in drug metabolism is important for the selection of animals for pharmacokinetic, pharmacodynamic, and toxicological studies. Hepatic microsomes from Sprague-Dawley (SD) and Brown Norway (BN) rats had 300-fold higher diazepam p-hydroxylation activity than Dark Agouti (DA) and Wistar (W) rats at a low diazepam concentration (3 microM). Kinetic studies indicated that diazepam p-hydroxylation in SD and BN rats proceeded with lower K(m) and higher V(max) values than it did in DA and W rats. However, the expression levels of cytochrome P450 CYP2D1, the reported enzyme for diazepam p-hydroxylation, did not cosegregate with the activity. These results suggest the presence of a new high-affinity diazepam p-hydroxylation enzyme other than CYP2D1 in SD and BN rats. DA rats showed 3- and 2-fold higher diazepam 3-hydroxylation and N-desmethylation activities, respectively, than the other rat strains. In agreement with this, DA rat liver microsomes had a higher expression of CYP3A2, which is responsible for diazepam 3-hydroxylation and partly responsible for N-desmethylation. Values of CL(int) (V(max)/K(m)) indicated that p-hydroxy-diazepam is the major metabolite in SD and BN rats, whereas 3-hydroxy-diazepam is the major metabolite in DA and W rats. The sum of the CL(int) in each strain was in the order of DA > SD = BN >> W. Strain differences in the pharmacodynamics of diazepam between SD and DA rats may be due to these differences in diazepam metabolism. We found that both the rate of elimination of diazepam and the major metabolic pathways in diazepam metabolism differed among the different rat strains due to polymorphic expression of the two enzymes involved in diazepam metabolism.
Saito
Konomu
K
Laboratory of Toxicology, Department of Environmental Veterinary Sciences, Graduate School of Veterinary Medicine, Hokkaido University, N18W9 North Ward, Sapporo 060-0818, Japan. fujita@vetmed.hokudai.ac.jp
Sakai
Noriaki
N
Kim
Hyung-Sub
HS
Ishizuka
Mayumi
M
Kazusaka
Akio
A
Fujita
Shoichi
S
D000429:Alcohol Oxidoreductases; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015153:Blotting, Western / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D051544:Cytochrome P-450 CYP3A; D000072467:Cytochrome P450 Family 2; D003975:Diazepam / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000494:pharmacology / Q000031:analogs & derivatives; D006900:Hydroxylation / Q000187:drug effects; D007106:Immune Sera / Q000276:immunology / Q000378:metabolism; D007700:Kinetics; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D009249:NADP / Q000378:metabolism; D011110:Polymorphism, Genetic / Q000235:genetics*; D051381:Rats; D011914:Rats, Inbred BN; D011919:Rats, Inbred Strains; D011922:Rats, Mutant Strains; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D013045:Species Specificity; D039181:Steroid 16-alpha-Hydroxylase / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013693:Temazepam / Q000031:analogs & derivatives* / Q000378:metabolism
D007106:Immune Sera; D008565:Membrane Proteins; C049480:4'-hydroxydiazepam; D009249:NADP; D013693:Temazepam; D000429:Alcohol Oxidoreductases; D001189:Aryl Hydrocarbon Hydroxylases; C450279:CYP2C11 protein, rat; C504568:Cyp2d1 protein, rat; C119985:Cyp2d2 protein, rat; C465260:Cyp3a2 protein, rat; D051544:Cytochrome P-450 CYP3A; D000072467:Cytochrome P450 Family 2; D039181:Steroid 16-alpha-Hydroxylase; D003975:Diazepam