Metabolite synthesis by rat liver cells and rat liver mitochondria.

Abstract

The use of rat liver cells prepared by mechanical dispersion techniques has recently been severely criticized on the basis that resulting plasma membrane damage leads to total loss of cytoplasmic constituents, thus leaving only membrane sacs containing mitochondria. In the present study, metabolite synthesis by mechanically prepared cells (M cells), collagenase prepared cells (E cells), and isolated mitochondria is compared.It was found that M cells have high rates of endogenous fatty acid oxidation and incorporate [1-14C]acetate approximately equally into 3-hydroxybutyrate, glutamate, citrate, malate, and 2-oxoglutarate. In contrast, mitochondria synthesize primarily citrate and aspartate but no 3-hydroxybutyrate from [1-14C]acetate. Loss of some cytoplasmic enzymes from M cells during isolation was confirmed. M cells are incapable of gluconeogenesis from lactate, synthesize much less glutamate than E cells, and become more highly oxidized during incubation ([NAD+]/[NADH] ratio of 135 versus 30 for E cells). Metabolite synthesis from [1-14C]acetate by mitochondria can be regulated by adding very low concentrations of organic acids and electron donors to the incubation.From these results it was concluded that M cells carry out metabolic reactions not present in isolated mitochondria. Metabolic reactions in mitochondria will approximate those of M cells if electron donors and mixtures of anions are added to the incubation.