Distribution Of Cytochrome Oxidase Activity Research Articles

A periodic pattern in the distribution of cytochrome oxidase activity in the visual cortex in kittens

The aim of the present work was to study the distribution of cytochrome oxidase activity in the visual cortex of kittens in normal conditions. Studies were performed using computer filtration of digitized images of frontal sections. The occurrence of regularly alternating levels of enzyme activity was demonstrated in layers III, IV, and VI of field 17 and layers IV and VI in field 18.

Comparative Histochemical Study of Cytochrome Oxidase Reactivity in the Rat Superior Colliculus in Norm and after Unilateral Destruction of the Inner Ear Cochlea

A comparative histochemical study of cytochrome oxidase (CO) activity in the superior colliculus (SC) of intact laboratory rats and animals with unilaterally lesioned inner ear cochlea was carried out. It was shown that distribution of CO activity in the SC of intact rats corresponded to its cytoarchitectonic laminar organization had a heterogeneous distribution between layers and homogeneous distribution inside the layers. Unilateral lesion of the inner ear cochlea results in the appearance of alternating columns with low and high CO activity in the medial complex layers and deep gray layer; these columns are directed mediolaterally and differentiated bilaterally. The appearance of the CO columns seems to be due to the spatially arranged organization of afferents coming onto SC from auditory structures.

Hierarchy within human SI: supporting data from cytochrome oxidase, acetylcholinesterase and NADPH-diaphorase staining patterns

The human primary somatosensory cortex consists of four cytoarchitectonic subdivisions (3a, 3b, 1 and 2) that are likely to contain distinct somatosensory representations. The intraareal organization of these areas as well as that of the primary motor cortex (area 4) has been analyzed using histochemical stains of cytochrome oxidase, acetylcholinesterase and NADPH-diaphorase activity in normal human brains. Cytochrome oxidase activity was revealed in individual cortical neurons and neuropil. Areas 4, 3a and 3b were on average darker than areas 1 and 2. The laminar distribution of cytochrome oxidase activity varied in different areas. A prominent dark band was present in layers IV and lower III in areas 3a and 3b and in layer III in areas 1, 2 and 4. Acetylcholinesterase staining revealed fibers and pyramidal cells in layers III and V; stained layer III pyramids were rare in areas 3a and 3b and numerous in areas 1, 2 and 4. NADPH-diaphorase positive elements included Golgi-like stained non-pyramidal neurons and Nissl-like stained pyramidal neurons; the former were found, in small numbers, in layer II of areas 4, 3a, 3b and 1, and the latter in layers III and V of areas 4 and 3a and in layer V of areas 1 and 2. The dark cytochrome oxidase staining of layer IV and the paucity of acetylcholinesterase positive pyramids in areas 3a and 3b resemble the pattern found in primary visual and auditory areas, whereas the dark cytochrome oxidase staining in layer III and abundance of acetylcholinesterase positive pyramids in areas 1 and 2 that of association areas. These results suggest that the four areas included in human SI constitute hierarchical stages of cortical processing, with 3a and 3b corresponding to primary and 1 and 2 to secondary areas.

Cytochrome Oxidase, Acetylcholinesterase, and NADPH-Diaphorase Staining in Human Supratemporal and Insular Cortex: Evidence for Multiple Auditory Areas

The pattern of cytochrome oxidase, acetylcholinesterase, and NADPH-diaphorase activity was studied in the supratemporal plane, the posterior part of the superior temporal gyrus, and the insula of normal human brains. Five dark cytochrome oxidase regions were found: (i) on Heschl's gyrus (area TC of von Economo and Koskinas); (ii) on the planum polare (area TC/TG); (iii) posterior to Heschl's gyrus (within area TA); (iv) on the posterior convexity of the superior temporal gyrus (within area TA); and (v) on the posterosuperior insula (area IB). More lightly stained cortex separated these regions (areas IA, TD, and part of TB). The laminar distribution of cytochrome oxidase activity varied in different areas. Acetylcholinesterase-positive fibers predominated in area TC and pyramidal neurons in areas TA and IA and in parts of TB; a mixture of fiber and neuronal staining was found in TC/TG, TD, and IB. NADPH-diaphorase positive profiles included large darkly stained nonpyramidal neurons, mostly in infragranular layers and in subcortical white matter, small faintly stained cells, and a dense array of fibers. The NADPH-diaphorase staining pattern did not vary between areas. The present results suggest that the supratemporal plane, the posterior part of the superior temporal gyrus, and the insula contain at least eight putative cortical areas. Comparison with activation studies by others suggest that, apart from the primary auditory area, six other putative areas may be auditory whereas one putative area, on posterior insula, may be vestibular.

Modular organization of human extrastriate visual cortex: evidence from cytochrome oxidase pattern in normal and macular degeneration cases.

The human extrastriate occipital cortex contains several visual areas that are probably analogues of macaque areas V2, V3, VP, V4 and V5. Tracing of callosal connections has led to the anatomical identification of these areas and to the characterization of some of them by cyto- and myeloarchitecture (Clarke and Miklossy, 1990). The pattern of cytochrome oxidase activity in these visual areas is now described in a normal case and in a case of age-related bilateral macular degeneration. In normal cortex, the laminar distribution of cytochrome oxidase activity was similar in V2, V3, VP, V4 and V5; a prominent dark band covered most of layers III and IV, and its upper and lower limits were gradual. In V2, V3, V4 and V5 but not VP, layer II tended to be darker than the infragranular layers. The overall intensity of the staining varied between areas: VP was very light, V2, V3 and V4 were darker, and V5 was very dark. A different, two-band pattern of cytochrome oxidase activity was found in a restricted region of the posterosuperior precuneus. The bilateral age-related macular degeneration had led to a great loss of ganglion cells in the central, but not in the peripheral retinae. The central representation in the lateral geniculate nuclei showed abnormally weak staining for cytochrome oxidase, particularly in the parvocellular layers. In the cortex, the contrast between lightly and darkly stained regions was greater than in the normal case. In particular, V5 was very heavily stained, and in V1 and V2 there were two different types of dark stripes that may represent compartments driven predominantly by the magnocellular system.

Parvalbumin Immunoreactivity and Cytochrome Oxidase Activity in the Brain of the Mongolian Gerbil.

Parvalbumin was purified from the skeletal muscle of the Mongolian gerbil (Meriones unguiculatus) and used to raise a polyclonal antiserum in a rabbit. The affinity-purified antibodies were used to study the distribution of parvalbumin in the brain of adult and developing gerbil and compare it with the distribution of cytochrome oxidase activity. In the adult brain, immunoreactivity was found in distinct subpopulations of neurons. There were densely stained areas, most of which contained immunopositive cell somata and their processes in the diffusely stained neuropil. Some highly stained areas were filled with immunopositive dot-like structures or fiber segments. Comparison of these parvalbumin-rich areas with cytochrome oxidase-rich areas revealed a close spatial correlation. In the developing brain, the spatial coincidence was still observed. In the developing cerebral cortex, parvalbumin-positive cells first appeared preferentially in the area which had had high cytochrome oxidase activity in the neuropil. At the cellular level, cytochrome oxidase staining often condensed in a part of the cell body before whole cell bodies became stained. Thus, the adult pattern of spatial coincidence appears, in at least some areas, to be generated in several stages.

Calbindin and parvalbumin cells in monkey VPL thalamic nucleus: distribution, laminar cortical projections, and relations to spinothalamic terminations

The ventral posterior lateral nucleus (VPL) of the monkey thalamus was investigated by histochemical staining for cytochrome oxidase (CO) activity and by immunocytochemical staining for the calcium-binding proteins parvalbumin and 28 kDa calbindin. Anterograde and retrograde tracing experiments were used to correlate patterns of differential distribution of CO activity and of parvalbumin and calbindin cells with the terminations of spinothalamic tract fibers and with the types of cells projecting differentially to superficial and deeper layers of primary somatosensory cortex (SI). VPL is composed of CO-rich and CO-weak compartments. Cells are generally smaller in the CO-weak compartment. Parvalbumin-immunoreactive cells and parvalbumin-immunoreactive medial lemniscal fiber terminations are confined to the CO-rich compartment. Calbindin-immunoreactive cells are found in both the CO-rich and CO-weak compartments. The CO-weak compartment, containing only calbindin cells, forms isolated zones throughout VPL and expands as a cap covering the posterior surface of the ventral posterior medial nucleus (VPM). Spinothalamic tract terminations tend to be concentrated in the CO-weak compartment, especially in the posterior cap. Other CO-weak, parvalbumin-negative, calbindin-positive nuclei, including the posterior, ventral posterior inferior, and anterior pulvinar and the small-celled matrix of VPM are also associated with concentrations of spinothalamic and caudal trigeminothalamic terminations. Parvalbumin cells are consistently larger than calbindin cells and are retrogradely labeled only after injections of tracers in middle and deep layers of SI. The smaller calbindin cells are the only cells retrogradely labeled after placement of retrograde tracers that primarily involve layer I of SI. The compartmental organization of VPL is similar to but less rigid than that previously reported in VPM. VPL and VPM relay cells projecting to different layers of SI cortex can be distinguished by differential immunoreactivity for the two calcium-binding proteins. The small-celled, CO-weak, calbindin-positive zones of VPL and VPM appear to form part of a wider system of smaller thalamic neurons unconstrained by traditional nuclear boundaries that are preferentially the targets of spinothalamic and caudal trigeminal inputs, and that may have preferential access to layer I of SI.

Differential expression of cytochrome oxidase (COX) genes in different regions of monkey brain.

A frontal pole cDNA library from monkey (Macaca mulatta) brain was screened to identify mRNAs that are expressed more in frontal pole as compared to primary visual cortex. Three cDNA clones, whose greater expression was confirmed by Northern blot analysis, were identified as cytochrome oxidase (COX) subunits I, II, and III (COX I, II, and III). Each clone showed higher levels of mRNA in the frontal pole, dorsal lateral prefrontal cortex, and hippocampus than in the primary visual or somatosensory cortices. COX histochemistry of prefrontal, visual, and somatosensory cortical regions demonstrated heterogeneous distributions, with highest activity in dendrite-rich neuropil of the cortex. A laminar distribution of COX mRNA expression also was demonstrated with in situ hybridization. mRNA was detected in cell bodies and in apical dendrites. These results indicate region specific differences in the distribution of COX activity and in the corresponding mRNA for three of its subunits within the monkey brain. Such differences may be related to differences in the distribution of neuropil as compared with cell bodies among the brain regions studied, and may be relevant to selective vulnerability in Alzheimer's disease.

Some observations upon the distribution of cytochrome oxidase activity in the globus pallidus of the rat.

The histochemical distribution of the enzyme cytochrome oxidase (CO) was explored in the globus pallidus (GP) of the rat, and it was compared with the distribution of acetylcholinesterase (AChE), another well-known enzyme of the basal ganglia in mammals. This neurochemical research illustrates two prominent findings. In the first place, a regional compartmentalization was detected in the pallidal distribution of CO, in which dorsal territories of the GP exhibited a more abundant presence of this enzyme. In addition, the distribution of this mitochondrial enzyme was not uniform all over those dorsal pallidal territories. Thus, the pallidal concentration of CO was gradually decreasing dorsoventrally and lateromedially, and, sometimes, areas highly stained for CO were intermingled with zones in which this enzyme was less conspicuous. In the second place, the pallidal distribution of AChE was the opposite of that found for CO (i.e. more abundant in ventral and medial territories of the GP). The functional significance of these findings is discussed in the light of the heterogeneous hodological neuroanatomy of the GP of rodents.

A Note upon the Striatal Distribution of Cytochrome Oxidase Activity in the Rat and in the Cat

The distribution of cytochrome oxidase (CO) activity was explored in the striatum of two mammalian species, the rat and the cat. Regional differences in the striatal distribution of CO were detected in both species. Thus, in most of our experimental material for rodents, an extensive band richer in CO was present in peripheral regions of the rostral and outer part of the corpus striatum. This striatal band was running in coronal sections, from medial and dorsal to lateral and ventral. In the cat's striatum, rostral and, above all, dorsal territories of the caudate nucleus were prominently stained for CO. In addition, in both species, although more sharply shown in the cat, it was possible to delineate a heterogeneous distribution of this mitochondrial enzyme following the local compartmental design of acetylcholinesterase (AChE) within the mammalian striatum. Zones with low concentration of AChE were in register with areas in which CO was low as well. This regional and local striatal heterogeneity might be a consequence of differences in the mitochondrial activity of cells located in diverse histochemical striatal parcels and, therefore, with different functional targets outside the striatum.

Laminar distribution of cytochrome oxidase staining in cetacean isocortex.

The distribution of cytochrome oxidase activity was studied in the cerebral cortex of two species of cetaceans, the harbour porpoise Phocoena phocoena, and the bottlenose dolphin Tursiops truncatus. Two main patterns of distribution of cytochrome oxidase were detected. The first, characteristic of the visual and auditory cortices of the lateral and suprasylvian gyri, is typified by a peak density in layer III, contrasting with low levels in layers II, V and VI. The second is found in wide areas of the limbic lobe, the insular cortex, the temporal operculum and the occipital cortex. In these regions, distribution of cytochrome oxidase is more uniform, with little difference between layers III, V and VI. A transitional pattern is found in the most dorsal parts of the limbic lobe, the parietal operculum, the ectosylvian gyrus and in orbitofrontal cortex. As areas of high cytochrome oxidase activity have been described in various land mammals to correspond to zones of major excitatory input and, in particular, to characterise the cortical layers that receive thalamocortical afferents, we propose that the thalamocortical input to cetacean sensory cortex, in which a typical layer IV is absent, may be mainly to layer III. This view is supported by the high density of neurons positive for the inhibitory transmitter gamma-aminobutyric acid that is also found in layer III of cetacean cortex, another typical feature of thalamocortical recipient zones.

Heterogeneous distribution of cytochrome oxidase activity in the rat substantia nigra: correlation with tyrosine hydroxylase and dynorphin immunoreactivities

Cytochrome oxidase (COase) activity, an endogenous marker of neuronal activity, was examined in the substantia nigra of the adult rat at the light-microscopic level. In addition, the pattern of histochemical staining observed for COase activity was correlated with immunohistochemistry for tyrosine hydroxylase (a marker of dopaminergic neurons) and for dynorphin (a peptide present in afferents from the striatum). Differential oxidative metabolic activity was revealed in subregions of the substantia nigra by COase histochemistry. Neurons of the substantia nigra pars compacta (SNc), ventral tegmental area, and ventrally displaced dopaminergic neurons were characterized by little or no staining for COase. n contrast, the substantia nigra pars reticulata (SNr) possessed a heterogeneous distribution of COase activity that was characterized by denzer staining in the ventrolateral than the dorsomedial part of the nucleus throughout its rostrocaudal extent, with the exception of the most rostral levels. This pattern of COase activity was inversely correlated with the density of ventrally descending tyrosine hydroxylase-positive dendrites arising from the medial portion of the SNc, as well as with the density of dynorphin immunoreactivity. The results suggest that the SNc and SNr possess distinct levels of oxidative metabolic activity. Furthermore, within the SNr itself, different levels of COase activity characterize subpopulations of neurons which may be differentially regulated by both striatal and dopaminergic influences.

Brain cytochrome oxidase: purification, antibody production, and immunohistochemical/histochemical correlations in the CNS

Cytochrome oxidase (CO) is a mitochondrial energy-generating enzyme used in brain studies as a marker of neural functional activity. The activity of CO in different brain regions, revealed histochemically, is distributed nonhomogeneously but in distinct patterns. Localized differences in CO activity could arise from localized differences in enzyme amount or from localized regulation of enzyme turnover number (molecular activity). To distinguish between these alternatives, we used antibodies against purified calf brain CO to assess the immunohistochemical distribution of CO amount (protein immunoreactivity) in several brain regions. Calf brain mitochondria (synaptic and nonsynaptic populations) were isolated from gray matter homogenates by differential centrifugation. CO was purified from detergent extracts of the mitochondria by cytochrome c-Sepharose 4B affinity chromatography. Antisera against the purified CO were raised in rabbits. The antibodies reacted specifically with CO, predominantly subunit IV, in SDS immunoblots. The antibodies did not react in SDS immunoblots with any other proteins solubilized from mitochondria or caudate nucleus but did cross-react with brain CO from other mammalian species and with bovine heart CO. The immunohistochemical distribution of CO amount matched the histochemical distribution of CO activity in all regions tested, including the monkey hippocampus and the mouse olfactory bulb, somatosensory (barrel) cortex, and cerebellum. Thus, the amount of CO in neural tissue is distributed in the same nonhomogeneous pattern as the histochemical activity of CO. The results suggest that mechanisms exist by which CO molecules are selectively distributed within neurons to meet local metabolic demands posed by neural functional activity.

Cytochrome oxidase activity in the rat caudate nucleus: light and electron microscopic observations.

Cytochrome oxidase (CO) activity was examined in the neostriatum of normal adult rats at the light and electron microscopic level. At the light microscopic level a heterogeneous distribution of CO activity was observed and was characterized by patches of high activity ranging in size from 200 to 800 microns surrounded or adjacent to regions of lower activity. The most dorsomedial and ventromedial regions of the caudate nucleus appeared to be consistently high in activity in all animals. At the ultrastructural level CO reaction product was localized to the membranes and intracristal spaces of mitochondria. The most reactive mitochondria (those containing the denest precipitates of reaction product) were found within the dendrites of spiny neurons in all caudate regions. In areas of high CO activity the mitochondria within bundles of myelinated fibers and in many axon terminals were also highly reactive whereas those in neuronal somata, primary dendrites, and glial cells and processes exhibited relatively little activity. Quantitative study showed that mitochondria within dendrites accounted for most of the CO activity in caudate neuropil. The mitochondria within dendrites and axon terminals were more reactive in regions of high CO activity than in regions of low CO activity. No differences in the density of synapses or in the proportions of axospinous and axodendritic synapses were observed between CO-rich and CO-poor areas. Heterogeneity in the distribution of CO activity in the caudate nucleus may be related to the "patchy" pattern of localization previously observed for some neostriatal afferents, enzymes, transmitters, peptides, and receptor ligands.(ABSTRACT TRUNCATED AT 250 WORDS)

Cytochrome oxidase activity in the striate cortex and lateral geniculate nucleus of the newborn and adult macaque monkey.

The laminar location of cytochrome oxidase staining has been compared in the lateral geniculate nucleus and area 17 in newborn and adult macaque monkeys. In area 17 of the adult, the distribution of cytochrome oxidase activity confirmed published findings. In the newborn animals, the tissue reacted as strongly for cytochrome oxidase as in the adult but the pattern of labelling was different in two respects. Firstly in layer 1 activity was stronger and occupied a wider portion of this layer. Secondly, cytochrome oxidase staining in layer 4C occupied two separate bands, a small narrow band at the bottom of 4C beta and a wider one occupying the full width of 4C alpha and spilling over into 4B. The pattern of cytochrome oxidase activity did not appear to be influenced by eccentricity in the newborn whereas, in the adult, label in 4C was more intense in cortex subserving central vision. In the lateral geniculate nucleus of the adult, the magnocellular layers and the most dorsal parvocellular layer reacted most strongly for cytochrome oxidase. In the newborn, parvocellular layers were more uniformly labelled and the difference between parvo- and magnocellular layers more pronounced. These results are discussed in relationship to the development of thalamo-cortical projections in the monkey.

A cytochemical investigation into the distribution of cytochrome oxidase activity within the mitochondria of brown adipose tissue from the prenatal rat

Using 3,3′-diaminobenzidine as substrate, the localization of cytochrome oxidase activity has been studied in mitochondria of brown adipose tissue from late prenatal rats. The reaction demonstrated cytochrome c oxidase since it was stimulated by exogenous cytochrome c, inhibited by KCN, and was insensitive to antimycin. The reaction product was found in the cristal and intermembrane spaces of unfixed, isolated mitochondria after as little as 2 minutes of incubation. Even after 45 minutes of incubation, reaction product was not clearly localized to the type II inclusions. This negative result was not altered by the presence of exogenous cytochrome c or by 2,4-dinitrophenol in the incubation medium. In fixed tissue, 30–40% of the mitochondrial profiles contained cristae oriented longitudinally. These had no observably different cytochrome oxidase activity compared to cristae oriented transversely. The results are discussed. It is concluded that they support the hypothesis that type II inclusions are composed of incomplete inner membrane lacking some membrane-bound respiratory enzymes.

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The distribution of cytochrome-oxidase activity in rabbit brain.

1. The cytochrome-oxidase activity in eleven structures of the rabbit brain has been investigated. 2. Kinetic data suggest that the same enzyme occurs in all brain structures but that the activity on a tissue-weight basis differs from structure to structure, being in general higher in the phylogenetically newer structures. 3. When the kinetic data are related to cell density (as measured by DNA concentration) all the neuron-containing structures of the cerebrum and brain stem show the same activity but the corpus callosum and the cerebellum show much less. 4. A special study was made of the distribution of activity and cell density within the diencephalon. 5. The results suggest that all neurons in the cerebrum and brain stem have a similar cytochrome-oxidase activity, which is about 80 times that of glia, and that the cerebellum is relatively richer in glia or that it contains neurons with a much lower cytochrome-oxidase activity, or both.

The distribution of cytochrome oxidase activity in Ehrlich ascites cell particulates obtained from glycerol homogenates

A method of fractionation of Ehrlich ascites cells has been developed for the separation of particulates derived by the previously developed glycerol technique for homogenization of this type of cell suspension. The success of the fractionation procedure which yields mitochondrial fractions containing much larger total cellular cytochrome oxidase than obtained by other methods was due to the effectiveness of glycerol in the cell disruption process and to the dilution of the glycerol homogenates of the Ehrlich ascites cells with saline solution containing sucrose. Fairly pure mitochondria, microsomes, and nonparticulate supernatants as judged by cytochrome oxidase activities were easily obtained.