Damage to the CNS is characterized by activation of microglia and astroglia, a phenomenon termed reactive gliosis. These cellular reactions have been implicated in both regeneration and degeneration of the CNS. Damage-induced production of cytokines and chemokines has been linked to the initiation of gliosis using several injury models, including exposure to known neurotoxicants. The Janus kinase-signal transducers and activators of transcription (JAK-STAT) pathway is a down-stream effector of cytokines and activation of this pathway is associated with gliosis. In particular, the transcription factor STAT3 is critical for astroglial response to CNS damage. Here, we used brain slices from mice treated with the dopaminergic neurotoxicant, 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) to examine astroglial STAT3 activation responses. 12 hours post MPTP (12.5 mg/kg, s.c.) or saline treatment, C57Bl/6 female mice were sacrificed by decapitation and coronal slices of brain were prepared. Striatum was dissected and assayed for activated STAT-3 (Tyr705 phosphorylation). Phospho-STAT3 was present in slices from MPTP treated mice immediately following sacrifice while absent in control mice, recapitulating in vivo observations. These data suggest that activated STAT3 is pre-served in brain slices. Striatal slices were then exposed to phosphate free oxygenated buffer for various times. After 45 minutes both MPTP treated and control striata expressed p-STAT3, indicating that MPTP induced phosphorylation is retained ex vivo and that STAT3 activation can also result from slice injury. Production of cytokines upstream of STAT3 was examined by RT-PCR. Messenger RNA for CNTF and IL-6 was not altered while mRNA for MCP-1 and LIF was increased. Finally we determined that AG490, a JAK inhibitor, and lavendustin A, a tyrosine kinase inhibitor, could diminish STAT3 phosphorylation in slices. These data suggest that activation of STAT3 is an early event in both toxicant and slice-induced glial activation. In addition, this investigation establishes the brain slice preparation method as a reliable model to examine reactive gliosis.
The Toxicologist. Society of Toxicology 43nd Annual Meeting and ToxExpo, March 21-25, 2004, Baltimore, Maryland