Elevated cortical extracellular fluid glutamate in transgenic mice expressing human mutant (G93A) Cu/Zn superoxide dismutase

Transgenic mice expressing a mutated (G93A) human Cu/Zn superoxide dismutas e (SOD1) develop motor neuron pathology and clinical symptoms similar to th ose seen in patients with amyotrophic lateral sclerosis. Loss of motor neur ons is most prominent in lumbar, followed by cervical cord and then brainst em. No significant cell death has been reported in motor cortex. The integr ity of the cortical glutamate reuptake systems was evaluated using intracer ebral microdialysis and western immunoblot assays for the glutamate transpo rters GLT-1, GLAST, and EAAC1. The basal extracellular fluid revels of aspa rtate, glutamate, glutamine, 3,4-dihydroxyphenylacetic acid, and 5-hydroxyi ndole-3-acetic acid were evaluated by HPLC. The extraction fraction of L-[H -3]glutamate, corrected with [C-14]mannitol, was also evaluated. GLT-1, EAA C1, and GLAST protein levels were determined by semiquantitative chemilumin escence immunoblot of proteins from membrane-enriched fractions. The relati ve optical density of film was translated into relative protein level by co mparison with a standard control mouse. The SOD1 mutant mice demonstrated a significant (p < 0.05) increase in basal levels of extracellular aspartate and glutamate. In addition, when the glutamate extraction fraction was cha llenged with exogenous unlabeled glutamate (500 mu M) by reversed microdial ysis, the glutamate extraction fraction in the mutant SOD1 mice was decreas ed significantly from control levels. The SOD1 mutant mice demonstrated no difference in the cortical protein levels of the glutamate transporter subt ypes. This study demonstrates that in areas of no visible pathology and no loss of glutamate transporter proteins, SOD1 mutant mice have elevated extr acellular fluid aspartate and glutamate levels and a decreased capacity to clear glutamate from the extracellular space.