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Assessment of Open Probability of the Mitochondrial Permeability Transition Pore in the Setting of Coenzyme Q Excess
Published on: June 1, 2022
Individual Cardiac Mitochondria Undergo Rare Transient Permeability Transition Pore Openings
Xiyuan Lu1, Jennifer Q Kwong1, Jeffery D Molkentin1
1From the Department of Pharmacology, University of California, Davis (X.L., D.M.B.); Department of Pediatrics, Cincinnati Children's Hospital Medical Center, University of Cincinnati, OH (J.Q.K., J.D.M.).
Rationale:
Mitochondria produce ATP, especially critical for survival of highly aerobic cells, such as cardiac myocytes. Conversely, opening of mitochondrial high-conductance and long-lasting permeability transition pores (mPTP) causes respiratory uncoupling, mitochondrial injury, and cell death. However, low conductance and transient mPTP openings (tPTP) might limit mitochondrial Ca(2+) load and be cardioprotective, but direct evidence for tPTP in cells is limited.
Objective:
To directly characterize tPTP occurrence during sarcoplasmic reticulum Ca(2+) release in adult cardiac myocytes.
Methods And Results:
Here, we measured tPTP directly as transient drops in mitochondrial [Ca(2+)] ([Ca(2+)]mito) and membrane potential (ΔΨm) in adult cardiac myocytes during cyclic sarcoplasmic reticulum Ca release, by simultaneous live imaging of 500 to 1000 individual mitochondria. The frequency of tPTPs rose at higher [Ca(2+)]mito, [Ca(2+)]i, with 1 μmol/L peroxide exposure and in myocyte from failing hearts. The tPTPs were suppressed by preventing mitochondrial Ca(2+) influx, by mPTP inhibitor cyclosporine A, sanglifehrin, and in cyclophilin D knockout mice. These tPTP events were 57±5 s in duration, but were rare (occurring in <0.1% of myocyte mitochondria at any moment) such that the overall energetic cost to the cell is minimal. The tPTP pore size is much smaller than for permanent mPTP, as neither Rhod-2 nor calcein (600 Da) were lost. Thus, proteins and even molecules the size of NADH (663 Da) will be retained during these tPTP.
Conclusions:
We conclude that tPTP openings (MitoWinks) may be molecularly related to pathological mPTP, but are likely to be normal physiological manifestation that benefits mitochondrial (and cell) survival by allowing individual mitochondria to reset themselves with little overall energetic cost.
Insights
Transient mitochondrial permeability transition pore openings (tPTP) are identified in cardiac cells. These brief events help mitochondria survive by managing calcium levels with minimal energy cost.
Area of Science:
- Mitochondrial physiology
- Cardiovascular research
- Cellular bioenergetics
Background:
- Mitochondria are vital for ATP production, particularly in high-energy demand cardiac myocytes.
- Opening of mitochondrial permeability transition pores (mPTP) can lead to cell death.
- Transient mPTP openings (tPTP) may offer cardioprotection by limiting mitochondrial calcium overload, but direct cellular evidence was lacking.
Purpose of the Study:
- To directly characterize the occurrence and nature of transient mitochondrial permeability transition pore (tPTP) openings in adult cardiac myocytes.
- To investigate the conditions influencing tPTP frequency during sarcoplasmic reticulum calcium release.
Main Methods:
- Simultaneous live imaging of mitochondrial calcium ([Ca(2+)]mito) and membrane potential (ΔΨm) in adult cardiac myocytes.
- Quantification of tPTP events during cyclic sarcoplasmic reticulum calcium release.
- Assessment of tPTP frequency under varying calcium loads, oxidative stress, and in failing heart models.
- Pharmacological and genetic inhibition of mPTP to evaluate tPTP suppression.
Main Results:
- Transient drops in [Ca(2+)]mito and ΔΨm directly measured as tPTPs in cardiac myocytes.
- tPTP frequency increased with higher cytosolic and mitochondrial calcium, peroxide exposure, and in myocytes from failing hearts.
- tPTPs were suppressed by inhibiting mitochondrial calcium influx and by mPTP inhibitors (cyclosporine A, sanglifehrin) and in cyclophilin D knockout mice.
- tPTP events were short-lived (57±5 s), rare (<0.1% of mitochondria), retained molecules up to NADH (663 Da), and had minimal energetic cost.
Conclusions:
- Transient mitochondrial permeability transition pore openings (MitoWinks) are identified as a physiological event in cardiac myocytes.
- tPTPs, though related to pathological mPTP, likely contribute to mitochondrial and cell survival by allowing self-resetting with low energetic impact.
- These findings provide direct evidence for a potentially cardioprotective role of transient mPTP activity.
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