Apparent unmasking of [3H]GDP binding in rat brown‐fat mitochondria is due to mitochondrial swelling

Abstract

1. The presence of and biochemical background for the so‐called ‘unmasking’ phenomenon in rat brown‐fat mitochondria was investigated (i.e. the apparent increase in [3H]GDP binding to the ‘uncoupling’ protein thermogenin, without a concomitant increase in the amount of the protein). 2. It was found that an unmasking could be observed both 1 h after norepinephrine injection and after 1 h cold stress, provided that the rats were preacclimated to 28°C. The unmasking could be observed both when a filtration method and when a centrifugation method for determination of [3H]GDP‐binding capacity were used; however, the absolute values were higher with the filtration method. 3. Based on observations of slower cytochrome‐c oxidase sedimentation during centrifugation, the possibility that the matrix volume of brown‐fat mitochondria isolated from warm‐acclimated animals was smaller than that of cold‐stressed animals was investigated with 3H2O. The cold stress increased the matrix volume from being nearly non‐existent to about 1 μl/mg. A preswelling procedure in an ionic medium could similarly increase the matrix volume in mitochondria from warm‐acclimated animals but was without significant effect in the already swollen mitochondria from cold‐stressed animals or from animals adapted to a lower temperature. 4. In mitochondria from warm‐acclimated animals, the ionic preswelling procedure was fully able to increase the apparent amount of GDP binding to that observed in mitochondria from cold‐stressed animals, but it was practically without effect on GDP binding in mitochondria from cold‐stressed animals or from animals acclimated to a lower temperature. 5. It is concluded that the apparent ‘unmasking’ phenomenon, observed when the tissue is less activated than in normal control situations, is not (as hitherto anticipated) due to a specific change in thermogenin as such, but is a reflection of a general mitochondrial phenomenon. Copyright © 1987, Wiley Blackwell. All rights reserved