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Related Experiment Videos

Protonmotive force in giant mitochondria.

M L Campo, H Tedeschi

    European Journal of Biochemistry
    |May 15, 1984
    PubMed
    Summary

    Researchers used the fluorescent dye pyranine to measure internal mitochondrial pH. Activating metabolism in giant mitochondria with succinate caused an alkaline pH shift, validating pyranine as a reliable pH indicator.

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    Area of Science:

    • Biochemistry
    • Cell Biology
    • Mitochondrial Physiology

    Background:

    • Mitochondria are crucial for cellular energy production through oxidative phosphorylation.
    • Accurate measurement of intramitochondrial pH is essential for understanding mitochondrial function.
    • Previous methods for assessing mitochondrial pH had limitations.

    Purpose of the Study:

    • To validate the use of pyranine (8-hydroxy-1,2,6-pyrenesulfonate) as a quantitative indicator of internal mitochondrial pH.
    • To measure the pH changes in giant mitochondria during metabolic activation.

    Main Methods:

    • Microinjection of the pH-sensitive fluorescent dye pyranine into giant mitochondria or mitoplasts.
    • Inhibition of oxidative phosphorylation using rotenone.
    • Activation of mitochondrial metabolism by the addition of succinate.
    • Monitoring internal mitochondrial pH using fluorescence measurements.

    Main Results:

    • Microinjected pyranine did not impair the oxidative phosphorylation capacity of giant mitochondria or mitoplasts.
    • Activation of metabolism in rotenone-inhibited giant mitochondria by succinate addition induced an internal pH shift.
    • The observed pH shift was approximately 0.3 units towards the alkaline range.
    • This pH shift is comparable to that observed in conventional rat liver mitochondria.

    Conclusions:

    • Pyranine is a suitable and non-disruptive quantitative indicator for internal mitochondrial pH.
    • Metabolic activation of mitochondria leads to a measurable alkaline shift in intramitochondrial pH.
    • Giant mitochondria exhibit similar pH regulatory responses to metabolic stimulation as conventional mitochondria.

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