Ammonia and amino acid metabolism and transport in brain in vitro
Creators: name: Benjamin, Abraham Moses
Publisher: name: University of British Columbia
Year: 2011

description: Studies have been made of the factors controlling the formation, transport and utilization of ammonia in the brain and its effects on brain metabolism, of the processes promoting amino acid fluxes in brain under a variety of conditions, especially those leading to increased nerve activity, and on the specific locations, and sites of formation, of amino acids in the brain. By using tetrodotoxin (2 µM) to suppress partly the neuronal efflux of amino acids brought about by the joint action of protoveratrine (5 µM) and ouabain (0.1 mM), the former drug being used to promote neuronal efflux of amino acids, and the latter being added to diminish re-uptake of amino acids, it has been shown that the major pools of glutamate, aspartate, glycine, serine and probably γ-aminobutyrate, are in the neurons. However, the major pool of glutamine appears to be in the glia. Glutamine formation takes place in the glia and is a process partly controlled by the concentrations of cations (K⁺, Na⁺, Ca⁺⁺) within these cells. Fluoroacetate (3 mM) acts mainly in the glia as it suppresses glutamine synthesis, but not the proto-veratrine-stimulated brain respiration. Malonate (2 mM) acts mainly in the neurons since it suppresses the protoveratrine - stimulated respiration but not the synthesis of glutamine. The amino acids, particularly glutamate, γ-aminobutyrate, aspartate and glycine, are released from brain cortex slices under conditions associated with brain cell excitation. The release processes are partly or wholly blocked by tetrodotoxin (2 µM). Tetrodotoxin does not affect the release of glutamine nor does protoveratrine accelerate it. This result is in accord with the conclusion that the main depot of glutamine lies not in the neurons but in the glia. Protoveratrine brings about an increased rate of formation of glutamine in incubated brain slices, suggesting that glutamate released from the neurons is taken up by the glia and there converted to glutamine. L-Glutamine is more effective than L-glutamate as a precursor of γ-aminobutyrate in brain slices. As glutamic acid decarboxylase is localized in the neurons, it is concluded that glutamine released from the glia is taken up by the neurons and there converted to glutamate and γ-aminobutyrate. Changes in the contents of NH₄⁺ in incubated brain slices are accompanied by quantitatively equivalent changes in the amino acid contents of the tissue. Amytal (1 mM) suppresses endogenous glutamate oxidation and enhances the neuronal contents of glutamate and γ -aminobutyrate. It diminishes ammonia liberation. Ammonia is formed aerobically by brain cortex slices in a glucose-free medium largely by endogenous glutamate oxidation within the neurons, and also by glutamine hydrolysis. External L-glutamate is taken up against a concentration gradient largely by the glia and is less effective than endogenous glutamate as a source of ammonia in brain. Ammonium ions are not accumulated in brain slices against a concentration gradient. They are presumably formed, up to a limiting concentration, in the neurons independently of the external NH₄⁺ concentration. Ammonium ions affect both neuronal and glial metabolism and the brain cell transport of Na⁺ and K⁺ in the incubated brain slices. The decrease of is partly due to exchange with NH₄⁺. The exchange process is most marked in infant rat brain. The effects of NH₄⁺ in inhibiting respiration, diminishing ATP concentrations, and changing the cationic concentrations at the brain cell membrane are more pronounced in the stimulated than in the unstimulated brain tissue. It is concluded that the effects of NH₄⁺ on brain metabolism and cation transport may be explained by its inhibitory effect on ATP formation in the neurons, by removal of α-ketoglutarate and hence by partly blocking the operation of the citric acid cycle. This may be one of the reasons for ammonia toxicity in the central nervous system.