(B) OPA1 FI, but not mtDsRed FI, is reduced in mitochondria undergoing autophagy. INS1 cells expressing mtDsRed and LC3:GFP were treated with 0.2 μM bafilomycin (45 min) or a cocktail of pepstatin A and E64d (90 min) and fixed. Mitochondria inside APs were defined by colocalization of mtDsRed and LC3:GFP. OPA1 FI in APs was normalized to its FI in mitochondria that were located outside APs (organelles positive for mtDsRed with subthreshold GFP FI). OPA1 FI within APs was 50% (*P0.001) and 54% (**P=0.003) upon treatment with bafilomycin or a cocktail of pepstatin A and E64d, respectively. Mitochondria inside APs had similar (96%) mtDsRed FI to those located outside APs (P=0.47). Scale bar, 2 μm. 