Mitchell 1969 Eur J Biochem

Mitchell P, Moyle J (1969) Estimation of membrane potential and pH difference across the cristae membrane of rat liver mitochondria. Eur J Biochem 7:471-84.

» PMID: 5776240

Mitchell P, Moyle J (1969) Eur J Biochem

Abstract: The cristae membrane or M phase of resting or respiring rat liver mitochondria becomes relatively permeable to K⁺ ions in presence of valinomycin. The equilibrium distribution of K⁺ ions across the membrane can therefore be used to estimate the membrane potential ΔΨ provided that precautions are taken to minimise swelling of the valinomycin-treated mitochondria. The pH difference ΔpH across the M phase of anaerobic mitochondria has been estimated from the buffering powers of the inner and outer phases and from the change of pH observed on lysing the mitochondria with Triton X-100. When the anaerobic mitochondria (State 5), in presence of β-hydroxybutyrate, are brought to a state of steady respiration either in absence (State 4) or in presence (State 3) of phosphate acceptor, the changes of pH and of pK of the medium recorded with H⁺ ion-sensitive and K⁺ ion-sensitive electrodes can be used to estimate the changes of ΔΨ and ΔpH across the M phase. The absolute values of ΔΨ and ΔpH have been estimated from the values determined in State 5 and the change of these values in the transition from State 5 to States 4 and 3.

Continued in Free Text • Keywords: Membrane potential, pH difference, Cristae membrane, Valinomycin, K⁺ ions

Labels: MiParea: Respiration 

Organism: Rat  Tissue;cell: Liver  Preparation: Isolated mitochondria 

Regulation: Coupling efficiency;uncoupling, Ion;substrate transport, mt-Membrane potential, pH  Coupling state: OXPHOS 

Made history 

Abstract Continued

The total protonmotive force Δp =ΔΨ– 59 ΔpH across the M phase of the mitochondria oxidising β-hydroxybutyrate in State 4 at 25° in a 250 mM sucrose medium near pH 7 is estimated to be about 230 mV, of which the major component is ΔΨ when the effect of translocation of K⁺ ions across the M phase is minimised. Under conditions permitting accumulation of a relatively large quantity of cation (State 6), Δp is not significantly different from that in State 4, but the major component is −ZΔpH.

The effects of changing Δp in mitochondrial suspensions in State 4 with uncoupling agent and with pulses of acid, alkali, calcium salt and ADP have been found to be in accord with the chemiosmotic hypothesis. In particular Δp in State 3 (ADP and Pi present) is estimated to be about 30 mV less than in State 4.