Fig. 2. Effect of 10- to 30-min lidocaine exposure followed by 120-min recovery period on neuronal viability. (A –C ) Lidocaine, 37 mm × 30 min; phase contrast, propidium iodide (PI), and annexin V images of same field. Several neurons have small blebs ( arrows in phase image) and annexin V staining without PI staining, indicating apoptosis. Dashed arrows in annexin V image indicate light staining consistent with early apoptosis; solid arrows indicate darker staining consistent with greater externalization of phosphatidylserine and more advanced apoptosis. ( D –E ) Lidocaine, 185 mm × 10 min; phase and PI images of same field. All neurons have large blebs and are PI+, consistent with necrosis. Annexin V image not done because all PI+neurons will be annexin V+, and greater than 92 mm lidocaine inhibited annexin V staining (data not shown). ( F –H ) Tris buffer control, 37 mm (185 mm Tris gave identical results); phase, PI, and annexin V images of same field. (I –J ) Tetrodotoxin as Na+channel–blocking control, 3.5 μm equipotent to 185 mm lidocaine; phase, PI, and annexin V images of same field. Both F and I show normal morphology with no apparent necrosis or apoptosis. Scale bar = 20 μm. Each experiment typical of three experiments; > 7 fields/experiment.