Visualization of mitochondrial membrane potential in mammalian cells

Area of Science:

• Cell Biology
• Biochemistry
• Mitochondrial Research

Background:

• Mitochondrial membrane potential (Δψ) is crucial for cellular energy production.
• Assessing Δψ in living cells provides insights into mitochondrial function and bioenergetic state.
• Fluorescent lipophilic cations are commonly used to measure Δψ due to their accumulation in mitochondria.

Purpose of the Study:

• To describe the application of tetramethylrhodamine methyl ester (TMRM) and rhodamine 123 (Rhod123) for semi-quantitative Δψ analysis.
• To present strategies for both steady-state and dynamic Δψ measurements in living mammalian cells.
• To provide practical protocols for assessing Δψ in different cell types.

Main Methods:

• Utilized fluorescent lipophilic cations TMRM and Rhod123.
• Employed two measurement strategies: steady-state and dynamic Δψ analysis.
• Applied live-cell fluorescence microscopy to primary human skin fibroblasts and neuron/astrocyte co-cultures.

Main Results:

• Demonstrated the utility of TMRM and Rhod123 for semi-quantitative Δψ assessment.
• Showcased steady-state measurements for comparing Δψ across conditions (e.g., disease, treatments).
• Illustrated dynamic measurements for monitoring temporal Δψ changes during acute perturbations.

Conclusions:

• TMRM and Rhod123 are effective tools for evaluating mitochondrial membrane potential in living cells.
• The presented methods allow for both comparative and dynamic analyses of Δψ.
• These protocols facilitate the study of mitochondrial bioenergetics in various cellular models.