Thesis (M.Sc.in Pharm)--Chulalongkorn University, 2003

Geldanamycin (GA) is a natural benzoquinone ansamycin class antibiotic which possesses antitumor activity. This study was designed to investigate a suspected and potential neuroprotective effect of GA against K+ deprivation-induced cell injury and death in primary cultured rat cerebellar granule cells. Cell metabolic activity and viability (assessed by MTT reduction and LDH release), content of glutathione and lipid peroxidation, were used as the measuring endpoints. Exposure of cultured cerebellar granule neurons for 12-72 hr to geldanamycin at different concentrations (0.01-1.0 micromolar) showed concentration- and exposure time-dependent neuroprotective effects. At low concentrations of GA with short incubation periods, GA showed a little effect on neuronal metabolic activity. After 24-48 hr pre-exposure to GA at concentrations higher than 0.05 micromolar, GA had a marked effect on neuronal metabolic activity whereas an exposure to very high concentrations and prolonged duration caused reduction in cell survival. With 24 hr pre-exposure to GA at concentrations higher than 0.025 micromolar, cell death was significantly prevented. Apparently, the maximal magnitude of neuroprotection may be obtained from 24 hr pre-exposure to 0.10 micromolar GA. After switching cultured cerebellar granule neurons from a serum medium containing 25 millimolar K+ (high K+) to a serum-free medium containing 5 millimolar low K+ (low K+) for 24 hr, approximately 50% of neurons had loss their viability. Simultaneous exposure with GA and low K+ for 24 hr did not protect neuronal cell death while pretreatment with 0.025-0.10 micromolar GA for 24 hr before shifting to low K+ medium significantly boosted up mitochondrial metabolic activity and prevented neuronal cell death in low K+ condition. Furthermore, GSH assay revealed that total GSH content was reduced by nearly 40% after switching cultured neurons to low K+ for 24 hr. Pretreatment with 0.1 micromolar GA for 24 hr effectively inhibited low K+-induced GSH diminution. Moreover, the same pretreatment with GA also prevented an increase in cellular lipid peroxidation after 24-hr exposure to low K+. These findings suggest that GA pretreatment may prevent granule cell death by induction of proteins that regulate GSH metabolism and lipid peroxidation