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Use of FRET-Sensor 'Mermaid' to Detect Subtle Changes in Membrane Potential of Primary Mouse PASMCs
Ruth C Dartsch1, Simone Kraut1, Tim Mayer2
1Cardiopulmonary Institute (CPI), Universities of Giessen and Marburg Lung Center (UGMLC), Member of the German Center for Lung Research (DZL), Justus-Liebig-University, 35392 Giessen, Germany.
Cells
|June 26, 2024
Summary
The voltage sensor Mermaid can detect significant (over 10 mV) membrane potential changes in pulmonary arterial smooth muscle cells (PASMCs). However, it struggles with subtle voltage shifts and forms intracellular aggregates, limiting its use for precise measurements.
Area of Science:
- Physiology
- Cell Biology
- Biophysics
Background:
- Pulmonary arterial smooth muscle cells (PASMCs) control vascular tone through membrane potential changes.
- Hypoxic Pulmonary Vasoconstriction relies on these subtle electrical signaling events.
- Accurate voltage sensing is crucial for understanding pulmonary circulation regulation.
Purpose of the Study:
- To evaluate the FRET-based voltage sensor Mermaid for detecting subtle membrane potential changes in mouse PASMCs.
- To assess Mermaid's sensitivity and specificity in response to physiologically relevant stimuli.
- To determine the limitations of Mermaid for studying pulmonary vascular tone regulation.
Main Methods:
- Isolation and lentiviral transduction of mouse PASMCs with Mermaid.
- Live-cell FRET imaging to assess Mermaid's emission ratio.
- Application of potassium chloride (KCl) to induce controlled membrane depolarization.
- Validation of KCl-induced depolarization using patch clamp recordings.
Main Results:
- Mermaid's emission ratio increased dose-dependently with KCl-induced depolarization.
- Unspecific intracellular Mermaid aggregates were observed, limiting sensor utility.
- Mermaid could not resolve subtle membrane potential changes of ≤10 mV, even when analyzing the membrane rim.
- The sensor reliably detected qualitative voltage changes >10 mV.
Conclusions:
- Mermaid is a viable tool for detecting significant (>10 mV) membrane potential changes in PASMCs.
- Limitations include intracellular aggregation and reduced sensitivity to subtle voltage fluctuations.
- Careful consideration of Mermaid's limitations is necessary for accurate interpretation of results in pulmonary circulation research.
