The study aimed at determining mechanism(s) by which amobarbital (amytal) suppresses glucose oxidation in cerebellar granule neurons in primary cultures, a glutamatergic preparation. When challenged with a depolarizing K + concentration (55 mM), the cells doubled their rate of glucose oxidation (production of 1 4 CO 2 from U-[ 1 4 C]glucose) and glycolysis (lactate accumulation). At normal K + concentration, amobarbital reduced 1 4 CO 2 production with half-maximum effect at 0.5-1 mM; at 55 mM K + , the inhibition was more potent, with more than half of the K + -induced stimulation abolished at 50 μM. Dixon plot analysis showed a single inhibitory mechanism at 5.4 mM K + , but at 55 mM K + , two kinetically different mechanisms could be distinguished. A more pronounced compensatory amobarbital-induced increase in glycolysis at 5.4 than at 55 mM K + suggested that amobarbital in addition to its inhibition of mitochondrial respiration inhibited K + -induced increase in energy demand, probably by its known suppression of stimulated glutamate release.