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The aim of this study was to trace D-amphetamine toxicity in isolated rat hepatocytes and to elucidate a possible involvement of CYP3A in the mechanisms of its toxicity. To this end, male Wistar rats were treated with nifedipine (5 mg kg-1 i.p., 5 days), a substrate and inducer of CYP3A. Hepatocytes isolated from nifedipine-treated and control rats were incubated with D-amphetamine at a concentration of 100 μmol L^sup -1^, which was determined to be an average toxic concentration (TC^sub 50^) for the compound. To evaluate the possible toxic effects of D-amphetamine on freshly isolated rat hepatocytes, we assessed the following parameters: cell viability, lactate dehydrogenase (LDH) activity, and glutathione (GSH).
The results showed that nifedipine potentiated amphetamine cytotoxicity in vitro, as follows: cell viability dropped by 65 % (p<0.001), GSH by 80 % (p<0.001), and LDH activity increased by 190 % (p<0.001). To clarify the role of nifedipine in amphetamine cytotoxicity, we used amiodarone, a substrate and an inhibitor of CYP3A. Pre-incubation of nifedipine-treated hepatocytes with amiodarone (14 μmol L^sup -1^) signifi cantly lowered amphetamine cytotoxicity.
Our results confi rmed the toxicity of D-amphetamine in isolated rat hepatocytes and the involvement of CYP3A in its metabolism and hepatotoxicity.
KEY WORDS: amiodarone, amphetamine cytotoxicity, nifedipine
Sazetak
ULOGA CYP3A-ENZIMA U TOKSIENOSTI D-AMFETAMINA U SVJEZE IZOLIRANIMA STAKORSKIM HEPATOCITIMA
Cilj je ovoga istrazivanja bio utvrditi toksienost D-amfetamina u izoliranim stakorskim hepatocitima kako bi se razjasnila eventualna uloga enzima CYP3A u mehanizmu toksienosti ovoga lijeka. U tu su svrhu muzjaci Wistar stakora pet dana primali nifedipin (5 mg kg^sup -1^ ip.), koji je supstrat i induktor CYP3A. Stanice izolirane iz jetre stakora koji su primali nifedipin i kontrolnih stakora inkubirane su s D-amfetaminom u njegovoj srednjoj toksienoj koncentraciji (TC^sub ^50) od 100 μmol L^sup -1^. Za procjenu eventualnoga toksienoga djelovanja D-amfetamina na svjeze izolirane hepatocite uporabili smo sljedeæ e parametre: prezivljenje stanica te razine laktat dehidrogenaze (LDH) i glutationa (GSH). Rezultati su pokazali da je nifedipin pojaeao toksieno djelovanje amfetamina in vitro na sljedeæ i naein: prezivljenje stanica palo je za 65 % (p<0,001), GSH za 80 % (p<0,001), a razine LDH porasle su za 190 % (p<0.001). Radi pojasnjenja uloge nifedipina u citotoksienosti amfetamina uporabili smo amiodaron, koji je supstrat i inhibitor CYP3A. Inkubacija hepatocita stakora koji...