FATTY-ACID OXIDATION AND KETOGENESIS BY ASTROCYTES IN PRIMARY CULTURE

The oxidation of the fatty acids octanoate and palmitate to CO2 and the ketone bodies acetoacetate and D-(-)-3-hydroxybutyrate was examined in astrocytes that were prepared from cortex of 2-day-old rat brain and grown in primary culture to confluence. Accumulation of acetoacetate (by mass) in the culture medium of astrocytes incubated with octanoate (0.3-0.5 mM) was 50-90 nmol C2 units h-1 mg of protein-1. A similar rate was obtained using radiolabeled tracer methodology with [1-C-14]octanoate as labeled substrate. The results from the radiolabeled tracer studies using [1-C-14]- and [7-C-14]octanoate and [1-C-14]-, [13-C-14]-, and [15-C-14]palmitate indicated that a substantial proportion of the omega-terminal four-carbon unit of these fatty acids bypassed the beta-ketothiolase step of the beta-oxidation pathway and the 3-hydroxy-3-methylglutaryl (HMG)-CoA cycle of the classic ketogenic pathway. The [C-14]acetoacetate formed from the 1-C-14-labeled fatty acids, obligated to pass through the acetyl-CoA pool, contained 50% of the label at carbon 3 and 50% at carbon 1. By contrast, the [C-14]acetoacetate formed from (omega-1)-labeled fatty acids contained 90% of the label at carbon 3 and 10% at acid contained 20% of the label at carbon 3 and 80% at carbon 1. These results indicate that acetoacetate is primarily formed either by the action of 3-oxo-acid-CoA transferase (EC 2.8.3.5) or acetoacetyl-CoA deacylase (EC 3.1.2.11) or both on acetoacetyl-CoA and not by the action of the mitochondrial HMG-CoA cycle involving HMG-CoA lyase (EC 4.1.3.4), which was readily detected, and HMG-CoA synthase (EC 4.1.3.5), which was barely measurable.