Osmotically-Induced Alterations in Volume and Ultrastructure of Mitochondria Isolated From Rat Liver and Bovine Heart

Abstract

Detailed studies correlating changes in mitochondrial optical density, packed volume, and ultrastructure associated with osmotically-induced swelling were performed . Various swelling states were established by incubating mitochondria (isolated in 0 .25 M sucrose) at 0°C for 5 min in series of KCI and sucrose solutions ranging in tonicity from 250 to 3 milliosmols . Reversibility of swelling was determined by examining mitochondria exposed to 250 milliosmols media after they had been induced to swell . Swelling induced by lowering the ambient tonicity to approximately 130 (liver mitochondria) and 90 (heart mitochondria) milliosmols involves primarily swelling of the inner compartment within the intact outer membrane . Decreasing the ambient tonicity beyond this level results in rupture of the outer membrane and expansion of the inner compartment through the break . The maximum extent of swelling, corresponding with complete unfolding of the cristae and an increase in over-all mitochondrial volume of approximately 6-fold (liver mitochondria) and 11-fold (heart mitochondria), is reached at approximately 15 (liver mitochondria) and 3 (heart mitochondria) milliosmols . Exposure of liver mitochondria to media of lower tonicity results in irreversibility of inner compartment swelling and escape of matrix material . These changes appear to result from increased inner membrane permeability, possibly due to stretching .

Energy-Linked Ultrastructural Transformations in Isolated Liver Mitochondria and Mitoplasts

“…Empirically, however, an ever increasing number of investigators have confirmed the condensed configuration in freshly isolated liver mitochondria, heart mitochondria, kidney mitochondria, skeletal muscle mitochondria, and brain mitochondria after fixation in osmium tetroxide or glutaraldehyde followed by osmium tetroxide (1-3, 6-10, 12, 14, 15, 17-19, 26-30, 32-37) . A condensed configuration also has been observed in isolated mitochondria by the use of negative staining (13,31) and in isolated mitochondria exposed to osmium tetroxide vapors (30) . In addition, the condensed configuration has been shown in mitochondria in the intact cell to be linked to induced change in physiological state as observed after either osmium tetroxide or glutaraldehyde fixation (4, 11, 13) .…”

“…10) . In another investigation Stoner and Sirak (30) found it necessary to use low osmolar concentra- …”

“…Lastly, only the condensed configuration of freshly isolated mitochondria, as observed after fixation in osmium tetroxide or glutaraldehyde, is compatible with the fact that freshly isolated mitochondria contain a compartment of sucrose-inaccessible water which is equal in volume to the inner compartment of the condensed configuration but is approximately one-half the volume of the inner compartment of the orthodox configuration (18,26,(46)(47)(48) . Passive osmotic volume changes also show conversion of the condensed to the orthodox configuration and its reversal by the controlled use of sucrose (30) . Consideration of the above evidence taken together with the freeze-cleave results presented in this report leaves little doubt that potassium permanganate, at least as used by Butler and Judah (16,40), fails to preserve the natural configuration of mitochondria related to freshly prepared preparations or to specific metabolic states, but rather causes a severe swelling during fixation .…”

Dynamics, structure, and function are coupled in the mitochondrial matrix.

“…In rat liver, the matrix of the condensed mitochondrion occupies about 50%o of the total mitochondrial volume and contains about 56% (wt/vol) protein, whereas the matrix of the orthodox mitochondrion occupies nearly the entire mitochondrial volume and contains about 26% protein (7). These mitochondrial configurations are dependent on the osmolarity of the external medium and on respiratory state-i.e., whether or not the mitochondria are active in electron transport and ATP synthesis (5,6,8).…”

Ultrastructural Studies of Beef Heart Mitochondria

“…The first is large amplitude swelling brought about by passive osmotic manipulation as first quantitated by Tedeschi and Harris (43) . This phenomenon has been studied recently with the electron microscope by Stoner and Sirak (44) . The second volume change is a low amplitude contraction brought about by addition of ADP.…”