Glial Alkalinization Detected In Vivo by 1H‐15N Heteronuclear Multiple‐Quantum Coherence‐Transfer NMR in Severely Hyperammonemic Rat

Brain [5- 15 N]glutamine amide protons were selectively observed in vivo by 1 H- 15 N heteronuclear multiple-quantum coherence-transfer NMR in spontaneously breathing, severely hyperammonemic rats during intravenous [ 15 N]ammonium acetate infusion and the subsequent recovery period. The linewidth of brain [5- 15 N]-glutamine amide proton H z increased from 36 ± 2 Hz at 3.4 h to 58 ± 6 Hz after 5.7 h of infusion, a net increase of 22 ± 6 Hz. Concomitantly, brain ammonia concentration increased from 1.7 to 3.5 ± 0.2 μmol/g and the rat progressed from grade III to grade IV encephalopathy. On recovery to grade III and decrease of brain ammonia concentration to 1.3 μmol/g, the linewidth returned to 37 ± 2 Hz. In aqueous solution, [5- 15 N]glutamine amide proton H z underwent a 17-Hz linebroadening when pH was raised from 7.1 to 7.5 at 37°C, due to the increased rate of base-catalyzed exchange with water proton. Hence, linebroadening is a sensitive measure of changing intracellular pH. The 22-Hz linebroadening observed in vivo in severely hyperammonemic grade IV rats strongly suggests that the intracellular pH increases from 7.1 to about 7.4-7.5 in astrocytes where glutamine is synthesized and mainly stored. Probable mechanisms for the ammonia-induced alkalinization and decreased intraglial buffering capacity, as well as implications of the result for pathogenesis of hepatic encephalopathy, are discussed.