PHARMACOLOGY OF NEUROTOXINS

National Institute Of Mental Health

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Abstract

The release of soluble neurotoxins and growth promoting factors by activated macrophages/microglia has been reported following chronic HIV infection and various types of brain injury. A neuronal cultured cell bioassay using neonatal rat has been refined for quantitative measurements. The isolation and characterization of soluble factors isolated from conditioned media from activated human peripheral blood monocytes or HIV-infected macrophages has been monitored with this bioassay. Extracts of conditioned media have been separated into component classes (lipids vs. non-lipids, low versus high molecular weight). Neuronal toxicity has been demonstrated in six of eleven batches of conditioned media from HIV-infected-LPS-stimulated macrophages. Activity was highest in a lipid-free fraction containing substances less than 3000 Da. The neurotoxicity was trypsin insensitive. Glutamate concentrations were elevated in neurotoxic media. Three tests demonstrated that glutamate concentrations are alone sufficient to account for the observed neurotoxicity. Glutamate concentrations were lowered to control levels and neurotoxicity was abolished by treatment of the conditioned media with glutamate pyruvate transaminase. Pretreatment of neurotoxic media by pre-incubation with astrocytes also reduced glutamate concentrations and abolished neurotoxicity. Finally, addition of glutamate to control conditioned media in concentrations matching those in the neurotoxic HIV/LPS media replicated the observed extent and morphology of the neurotoxic effect. In a separate project, the CNS fraction of the excitoxic amino acid quinolinate that derives from kynurenine metabolism has been determined in pharmacokinetic studies. Stable isotope labeled L-kynurenine was prepared and infused into gerbils. The enrichment of peripheral vs. central circulating kynurenine was measured in control and immune stimulated gerbil brains in order to determine the origin of the enhanced quinolinate concentration in animals with CNS inflammation.