Heterogeneous distribution of drug metabolism in elutriated rat hepatocytes

Centrifugal elutriation was used to separate isolated rat hepatocytes into five fractions containing cells of different sizes. These fractions were compared with regard to cell size, morphology and function. Analyzed by flow cytometry, the small cells were found to be enriched in fraction 1 and the large cells in fraction 5. Evaluation by light and electron microscopy indicated that the fractions contained single hepatocytes of normal structure. The cytochrome P-450 content and the 7-ethoxycoumarin O-deethylase activity were assessed in hepatocytes from untreated animals, those treated with phenobarbital, and those treated with phenobarbital plus allylisopropylacetamide. In both untreated and phenobarbital-treated animals, cytochrome P-450 content and 7-ethoxycoumarin O-deethylase activity rose significantly from fraction 1 to fraction 5. The P-450 content gradually rose up to 2-fold. The enzyme activity rose 5-fold, and it increased steeply between fractions 2 and 3. The cytochrome P-450 content in phenobarbital-plus-allylisopropylacetamide-treated animals was decreased in all fractions but more extensively in fraction 5 than in fraction 1.