Visualizing subcellular changes in the NAD(H) pool size versus redox state using fluorescence lifetime imaging...

Area of Science:

• Biophysics
• Cellular Metabolism
• Microscopy

Background:

• NADH autofluorescence imaging visualizes cellular energy metabolism.
• It is sensitive to both redox ratio and NAD(H) pool size, which can change independently.
• Distinguishing these factors is crucial for understanding cellular processes like aging.

Purpose of the Study:

• To evaluate the potential of NADH fluorescence lifetime imaging microscopy (FLIM) to differentiate between redox ratio and NAD(H) pool size changes.
• To assess the impact of modified NAD(H) pools on NADH FLIM.
• To establish NADH FLIM as a tool for analyzing cellular metabolism.

Main Methods:

• Targeted modifications of NAD(H) pool size and ratio in cells and mice.
• Analysis of NADH FLIM data to assess sensitivity to pool size and redox alterations.
• Comparison of lifetime components' response to different metabolic changes.

Main Results:

• NADH FLIM is sensitive to NAD(H) pool size, mimicking redox alteration effects.
• Specific components of NADH fluorescence lifetime are differentially affected by redox versus pool size changes.
• FLIM can distinguish between these two modalities independently.

Conclusions:

• NADH FLIM can differentiate between changes in cellular redox state and NAD(H) pool size.
• This technique provides high spatial resolution for evaluating cellular metabolism and relative NAD(H) levels.
• NADH FLIM is a valuable tool for studying aging and metabolic disorders.