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Background:  HIV infection of the central nervous system results in neuronal death and damage in the cerebral cortex, basal ganglia, and hippocampus. Neurons in these regions are also relatively sensitive to excitotoxic insults that occur through over-activation of NMDA receptors. During HIV infection of the central nervous system, HIV-infected macrophages/microglia in the brain release neurotoxins, including glutamate or quinolinic acid that activate NMDA receptors. To define the role of NMDA receptors in HIV-induced neurodegeneration, we examined effects of HIV-infected macrophages on survival of primary rodent hippocampal neurons, whose developmental expression of NMDA receptor subtypes is well established both in vivo and in vitro. Through the use of NMDA receptors subtype-specific inhibitors, we analyzed the contribution of NMDA receptor subtypes in mediating HIV-induced neurotoxicity.

Methods:  Primary rodent cultures of hippocampal neurons were aged for 7, 14, or 21 days in vitro. Cultures were pre-incubated with NMDA receptor antagonists for 1 hour and exposed for 24 hours to supernatants from macrophages infected with the HIV CNS-isolate, Jago. Cultures were then fixed and stained for MAP-2, a neuronal marker. The number of surviving neurons was determined for each condition and statistical comparisons were made by a paired t-test.

Results:  Immature hippocampal neurons (7 days in vitro) were resistant to HIV-induced neurotoxicity, whereas mature neurons (14 days in vitro and 21 days in vitro) were susceptible. Neuronal loss in mature hippocampal cultures was blocked by competitive and non-competitive NMDA receptor antagonists. In addition, NMDA receptor antagonists specific for the NMDA receptor-2B subtype completely protected hippocampal neurons at 14 days in vitro, but only partially at 21 days in vitro.

Conclusions:  NMDA receptors play a significant role in HIV-induced neurodegeneration in our model, and susceptibility follows known NMDA receptor subunit expression patterns. Immature neurons (with lower NMDA receptor expression) are resistant, while mature neurons are susceptible to HIV-induced neurodegeneration. Antagonists of the NMDA receptor-2B subunit, which predominates in developing neurons, completely protect neurons at 14 days in vitro, but only partially protect at 21 days in vitro. This change in neuronal susceptibility may reflect increasing expression of NMDA receptor-2A in maturing hippocampal neurons. We are currently dissecting the role of NMDA receptor-2A in this process. Understanding the role of NMDA receptors subtypes in neurodegeneration may reveal new therapeutic targets.

Keywords: Neurodegeneration; NMDA receptor; hippocampal neurons