Regulation of GABA level in rat brain synaptosomes: fluxes through enzymes of the GABA shunt and effects of glutamate, calcium, and ketone bodies.
Stable isotopes were used to measure both the rate of GABA formation by glutamic acid decarboxylase (GAD) and the rate of utilization by GABA-transaminase (GABA-T). The initial rate of GABA accumulation, determined with either [2-15N]glutamine or [2H5]glutamine as precursor, was 0.3-0.4 nmol/min/mg of protein. Addition of the calcium ionophore A23187 enhanced GAD activity, whereas changes in levels of inorganic phosphate and H+ were without influence. Flux through GABA-T (GABA--> glutamate), measured with [15N]GABA as precursor, was 0.82 nmol/min/mg of protein, whereas the reamination of succinic acid semialdehyde (reverse flux through GABA-T) was almost sixfold faster, 4.8 nmol/min/mg of protein. The rate of GABA metabolism via the tricarboxylic acid cycle was very slow, with the upper limit on flux being 0.03 nmol/min/mg of protein. Addition of either acetoacetate or beta-hydroxybutyrate raised the internal content of glutamate and reduced that of aspartate; the GABA concentration and the rate of its formation increased. It is concluded that in synaptosomes (a) GABA-T is a primary factor in regulating the turnover of GABA, (b) a major regulator of GAD activity is the concentration of internal calcium, (c) GAD in nerve endings may not be saturated with its substrate, glutamate, and the concentration of the latter is a determinant of flux through this pathway, and (d) levels of ketone bodies increase, and maintain at a higher value, the synaptosomal content of GABA, a phenomenon that may contribute to the beneficial effect of a ketogenic diet in the treatment of epilepsy.
Erecińska
M
M
Department of Pharmacology, University of Pennsylvania, School of Medicine, Philadelphia, USA.
Nelson
D
D
Daikhin
Y
Y
Yudkoff
M
M
D000612:4-Aminobutyrate Transaminase / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology*; D018698:Glutamic Acid / Q000494:pharmacology*; D005973:Glutamine / Q000378:metabolism; D007657:Ketone Bodies / Q000494:pharmacology*; D008297:Male; D010710:Phosphates / Q000494:pharmacology; D011522:Protons; D051381:Rats; D017207:Rats, Sprague-Dawley; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000096:biosynthesis / Q000378:metabolism*
D007657:Ketone Bodies; D010710:Phosphates; D011522:Protons; D005973:Glutamine; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; D000612:4-Aminobutyrate Transaminase; D002118:Calcium