Quantitative cytochemical mapping of mitochondrial enzymes in rat cerebella

Mitochondrial metabolic competence, defined as the organelle's capacity to provide adequate amounts of ATP in due time, appears to constitute an impor tant determinant in several biological processes and pathological condition s. Thus, the assessment of the metabolic efficiency of the mitochondrial po pulation in a given tissue area or cellular compartment may provide clues t o identifying alterations of the cellular bioenergetic machinery, which may constitute a predisposing condition leading to impaired organ and system f unctions. In the cerebellar cortex of adult rats, the activities of the enzymes cytoc hrome oxidase (COX) and succinic dehydrogenase (SDH) were, respectively, ev idenced by means of the diaminobenzidine and copper ferrocyanide preferenti al cytochemical techniques. At the electron microscope, the activities of t hese two key molecules of the respiratory chain were clearly visualised as dark precipitates at the inner mitochondrial membrane sites where COX and S DH are located. By means of the disector method, unbiased mitochondrial sam plings were carried out to measure: the number of mitochondria/mum(3) of ti ssue (numeric density: Nv); the mitochondrial volume fraction/mum(3) of tis sue (volume density: Vv) and the average mitochondrial volume (V) both on C OX- and SDH-positive organelles in the cerebellar glomeruli and Purkinje ce lls, respectively. The ratio R (total area of the precipitates due either t o COX or SDH activity within the single mitochondrion/area of the same orga nelle) was also evaluated to get information on the enzyme activity related to mitochondrial size. The documented accumulation of mutant mitochondrial DNA particularly in pos tmitotic cells results in a marked heteroplasmy (mixtures of normal and mut ated genomes) at mitochondrial and cellular levels, thus the cellular poten tial for energy production is demanded to a mosaic of organelles with diffe rent functional capabilities. Assessment of the mitochondrial mosaic outlin e by means of quantitative cytochemistry of key enzymes of the respiratory chain, such as COX and SDH, may allow for the morphofunctional metabolic ma pping of mitochondrial efficiency in discrete cellular or tissue compartmen ts. (C) 2001 Elsevier Science Ltd. All rights reserved.