Fig. 4. Geldanamycin (GA) blocks astrocyte death induced by glucose deprivation (GD). Sister early cultures of astrocytes subjected to 24 h GD without (A,B) or after pretreatment with GA (C,D). The same microscope field of each culture after 24 h GD was photographed using phase contrast optics (A,C) or by fluorescence microscopy (B,D) after staining with propidium iodide (PI) and Hoechst dye. The insert in B shows two of the apoptotic PI-staining nuclei from the lower part of the field, at higher magnification so the apoptotic bodies can be easily seen. (E) The extent of apoptotic and necrotic cell death in early cultures was determined by nuclear morphology after GD and PI plus Hoechst staining. At least 100 cells per culture were counted, with 4–8 cultures per condition. Values are means ± S.E.M. P<0.05 compared to GD without GA treatment (Control). (F) Quantitation of cell death in mature cultures after 24 h GD by lactate dehydrogenase (LDH) release. GA was present beginning 8 or 4 h prior to GD or first added at the beginning of GD (During); No Tx indicates GD without GA treatment. Pretreatment for 8 or 4 h was most effective, but treatment only during GD was still able to reduce injury in older cultures. ^*P<0.05 compared to GD with GA (No Tx) condition. N=8–12 cultures per condition.