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PubMed2005

Cerebral metabolic response to low blood flow: possible role of cytochrome oxidase inhibition.

The reactions of cerebral metabolism to imposed changes of cerebral blood flow (CBF) are poorly understood. A common explanation of the mismatched CBF and oxygen consumption (CMR(O(2))) during neuronal excitation holds that blood flow rises more than oxygen consumption to compensate for an absent oxygen reserve in brain mitochondria. The claim conversely implies that oxygen consumption must decline when blood flow declines. As the prevailing rate of reaction of oxygen with cytochrome c oxidase is linked to the tension of oxygen, the claim fails to explain how oxygen consumption is maintained during moderate reductions of CBF imposed by hyperventilation (hypocapnia) or cyclooxygenase (COX) inhibition. To resolve this contradiction, we extended the previously published oxygen delivery model with a term allowing for the adjustment of the affinity of cytochrome c oxidase to a prevailing oxygen tension. The extended model predicted constant oxygen consumption at moderately reduced blood flow. We determined the change of affinity of cytochrome c oxidase in the extended model by measuring CBF in seven, and CMR(O(2)) in five, young healthy volunteers before and during COX inhibition with indomethacin. The average CBF declined 35%, while neither regional nor average CMR(O(2)) changed significantly. The adjustment of cytochrome c oxidase affinity to the declining oxygen delivery could be ascribed to a hypothetical factor with several properties in common with nitric oxide.

Adult↗
PubMed2003

Basal levels of metabolic activity are elevated in Genetic Absence Epilepsy Rats from Strasbourg (GAERS): measurement of regional activity of cytochrome oxidase and lactate dehydrogenase by histochemistry.

The Genetic Absence Epilepsy Rats from Strasbourg (GAERS) are considered an isomorphic, predictive, and homologous model of human generalized absence epilepsy. It is characterized by the expression of spike-and-wave discharges in the thalamus and cortex. In this strain, basal regional rates of cerebral glucose utilization measured by the quantitative autoradiographic [(14)C]2-deoxyglucose technique display a widespread consistent increase compared to a selected strain of genetically nonepileptic rats (NE). In order to verify whether these high rates of glucose metabolism are paralleled by elevated activities of the enzymes of the glycolytic and tricarboxylic acid cycle pathways, we measured by histochemistry the regional activity of the two key enzymes of glucose metabolism, lactate dehydrogenase (LDH) for the anaerobic pathway and cytochrome oxidase (CO) for the aerobic pathway coupled to oxidative phosphorylation. CO and LDH activities were significantly higher in GAERS than in NE rats in 24 and 28 of the 30 brain regions studied, respectively. The differences in CO and LDH activity between both strains were widespread, affected all brain systems studied, and ranged from 12 to 63%. The data of the present study confirm the generalized increase in cerebral glucose metabolism in GAERS, occurring both at the glycolytic and at the oxidative step. However, they still do not allow us to understand why the ubiquitous mutation(s) generates spike-and-wave discharges only in the thalamocortical circuit.

Animals↗
PubMed1988

Isolation from porcine intestinal extracts of a cholecystokinin-like peptide and a peptide with homology to cytochrome oxidase polypeptide VII and chymodenin.

A number of different forms of cholecystokinin (CCK) exist in the brain and intestine. Gel permeation and ion exchange chromatography and high performance liquid chromatography have been used to isolate a peptide from partially purified porcine intestinal extracts with N-terminal homology to porcine brain CCK-58. This peptide contracted both the guinea pig ileum longitudinal muscle and gallbladder muscle and these responses were inhibited by dibutyryl cyclic GMP or proglumide. The potency was approximately 1/100 of that of CCK-8. The reason for this low potency is unclear, but it is possible that a critical part of the biologically active region is modified or that it is a truncated form of CCK-58. A further peptide was isolated with a sequence homologous to cytochrome oxidase polypeptide VII and chymodenin.

Amino Acid Sequence↗
PubMed2006

Total sleep deprivation inhibits the neuronal nitric oxide synthase and cytochrome oxidase reactivities in the nodose ganglion of adult rats.

Sleep disorders are a form of stress associated with increased sympathetic activity, and they are a risk factor for the occurrence of cardiovascular disease. Given that nitric oxide (NO) may play an inhibitory role in the regulation of sympathetic tone, this study set out to determine the NO synthase (NOS) reactivity in the primary cardiovascular afferent neurons (i.e. nodose neurons) following total sleep deprivation (TSD). TSD was performed by the disc-on-water method. Following 5 days of TSD, all experimental animals were investigated for quantitative nicotinamine adenine dinucleotide phosphate-diaphorase (NADPH-d, a co-factor of NOS) histochemistry, neuronal NOS immunohistochemistry and neuronal NOS activity assay. In order to evaluate the endogenous metabolic activity of nodose neurons, cytochrome oxidase (COX) reactivity was further tested. All the above-mentioned reactivities were objectively assessed by computerized image analysis. The clinical significance of the reported changes was demonstrated by alterations of mean arterial blood pressure (MAP). The results indicated that in normal untreated rats, numerous NADPH-d/NOS- and COX-reactive neurons were found in the nodose ganglion (NG). Following TSD, however, both the labelling and staining intensity of NADPH-d/NOS as well as COX reactivity were drastically reduced in the NG compared with normal untreated ganglions. MAP was significantly higher in TSD rats (136+/-4 mmHg) than in normal untreated rats (123+/-2 mmHg). NO may serve as an important sympathoinhibition messenger released by the NG neurons, and decrease of NOS immunoexpression following TSD may account for the decrease in NOS content. In association with the reduction of NOS activity, a defect in NOS expression in the primary cardiovascular afferent neurons would enhance clinical hypertension, which might serve as a potential risk factor in the development of TSD-relevant cardiovascular disturbances.

Animals↗
PubMed2002

A mutation in subunit I of cytochrome oxidase from Rhodobacter sphaeroides results in an increase in steady-state activity but completely eliminates proton pumping.

The heme-copper oxidases convert the free energy liberated in the reduction of O(2) to water into a transmembrane proton electrochemical potential (protonmotive force). One of the essential structural elements of the enzyme is the D-channel, which is thought to be the input pathway, both for protons which go to form H(2)O ("chemical protons") and for protons that get translocated across the lipid membrane ("pumped protons"). The D-channel contains a chain of water molecules extending about 25 A from an aspartic acid (D132 in the Rhodobacter sphaeroides oxidase) near the cytoplasmic ("inside") enzyme surface to a glutamic acid (E286) in the protein interior. Mutations in which either of these acidic residues is replaced by their corresponding amides (D132N or E286Q) result in severe inhibition of enzyme activity. In the current work, an asparagine located in the D-channel has been replaced by the corresponding acid (N139 to D; N98 in bovine enzyme) with dramatic consequences. The N139D mutation not only completely eliminates proton pumping but, at the same time, confers a substantial increase (150-300%) in the steady-state cytochrome oxidase activity. The N139D mutant of the R. sphaeroides oxidase was further characterized by examining the rates of individual steps in the catalytic cycle. Under anaerobic conditions, the rate of reduction of heme a(3) in the fully oxidized enzyme, prior to the reaction with O(2), is identical to that of the wild-type oxidase and is not accelerated. However, the rate of reaction of the fully reduced enzyme with O(2) is accelerated by the N139D mutation, as shown by a more rapid F --> O transition. Whereas the rates of formation and decay of the oxygenated intermediates are altered, the nature of the oxygenated intermediates is not perturbed by the N139D mutation.

Amino Acid Substitution↗
PubMed1987

Glial glutamate dehydrogenase: ultrastructural localization and regional distribution in relation to the mitochondrial enzyme, cytochrome oxidase.

Glutamate dehydrogenase (GDH) is primarily a mitochondrial enzyme involved in the metabolism of glutamate. We have recently shown by light microscopic immunocytochemistry that, within detergent-permeabilized brain tissue, GDH is enriched in glial cells, particularly in regions utilizing L-glutamate as a neurotransmitter. In this study, we used immunogold labeling to quantitatively establish that the form of the enzyme recognized by the presently used GDH antiserum is associated primarily with a subpopulation of mitochondria in ultrathin, plastic-embedded sections of the rat cortex and striatum. Permeabilization with detergents was omitted in these studies, so as to preserve the ultrastructure. As expected, labeled mitochondria occurred both in neurons and glia. Furthermore, light microscopic comparisons of the regional distributions of peroxidase immunoreactivity for GDH and a histochemical reaction product for a second mitochondrial enzyme, cytochrome oxidase (CO), were used to demonstrate that high levels of GDH in glia of glutamate-receptive areas do not necessarily reflect the areas' demand for elevated oxidative metabolism. While all regions showing intense labeling for glial GDH also exhibited high levels of CO activity, many additional regions showing high levels of CO activity contained no detectable immunoreactivity for glial GDH. These light-microscopic comparisons reveal that the energy requirements are not the only factors accounting for the regional heterogeneity of the enzyme. We conclude that glial mitochondria are heterogeneous with respect to their GDH content and that GDH is enriched in areas exhibiting chronically active glutamatergic transmission.

Animals↗
PubMed2001

Early vacuolization and mitochondrial damage in motor neurons of FALS mice are not associated with apoptosis or with changes in cytochrome oxidase histochemical reactivity.

Overexpression of mutated superoxide dismutase (SOD1) in transgenic mice causes a progressive motor neuron degeneration in the spinal cord similar to that in human amyotrophic lateral sclerosis (ALS). Ultrastructural analysis of motor neurons at different stages of the disease in transgenic C57BL/6 mice carrying the G93A mutation of SOD1 showed, at about 2 weeks of age, much earlier than the initial symptoms of the disease, microvacuoles in the cytoplasm, with marked swelling of the mitochondria. Nuclei with an apoptotic morphology were never observed in these motor neurons. Swollen mitochondria were also seen in the distal part of motor axons of phrenic nerves and in the large axons of sciatic nerves before the onset of the disease, but no mitochondrial alterations were seen in skeletal muscles or in the small sciatic nerve axons. Moreover, we found no apparent changes in the histochemical reactivity of cytochrome oxidase in motor neurons of transgenic mice even at the advanced stage of the disease, suggesting that partial neuronal activity in these cells may be maintained despite the altered mitochondria. Immunoreactivity for human SOD1 was high around vacuoles in the motor neurons of transgenic mice but no cytoplasmic intracellular SOD1 aggregates were observed. Our data indicate that mitochondrial swelling may be an important factor triggering the cascade leading to progressive motor neuron death. Activation of the mitochondrial permeability transition pore may be involved in this process, through excitotoxicity or other neurotoxic stimuli.

Amyotrophic Lateral Sclerosis↗
PubMed1992

Direct W-like geniculate projections to the cytochrome oxidase (CO) blobs in primate visual cortex: axon morphology.

The primate lateral geniculate nucleus (LGN) is composed of large, medium, and small cells located, respectively, in magnocellular (M), parvocellular (P), and specialized layers (intercalated and S-layers in simians, koniocellular (K) layers in prosimians). Several studies have examined the physiology and connections of M and P LGN cells and have concluded that they provide separate contributions to visual perception via separate pathways. Less is known about the structure and contributions of the small LGN cells. This study examined the distribution and structure of K LGN cell axons in the cortex of the prosimian, Galago crassicaudatus. Wheat germ agglutinin conjugated to horseradish peroxidase, or Phaseonlus vulgaris leucoaglutinin, was injected into the LGN K layers to demonstrate the overall axon projection pattern and the details of individual axons, respectively. Location of axons within striate cortex was specified relative to boundaries determined by Nissl or cytochrome oxidase (CO) stains on the same or adjacent sections. Our results show that K LGN axons end as single complex arbors within one CO blob zone in layer III; they never terminate in interblob zones. These axons also emit a collateral in layer I that arborizes more broadly and spans both CO blob and interblob zones. These data, together with data on K cell physiology and intralaminar cortical connections, suggest that the LGN small cell pathway could modulate the activity of the other two pathways in striate cortex and contribute directly to visual perception.

Animals↗
PubMed1983

Inhibition of cytochrome oxidase activity by local anaesthetics.

Using a polarographic method, we studied the inhibition of mitochondrial electron transport at the cytochrome c oxidase site caused by eight local anaesthetics. The diversity of the types of inhibition observed indicate the importance of electrostatic interactions between the anaesthetic molecules and the membrane protein. A linear relationship was recognized between the anaesthetic activity of infiltration and the affinity for the enzyme. We also observed a significant relationship between this affinity and the octanol-water partition coefficient. This result suggests that lipophilic interactions are involved in cytochrome oxidase-anaesthetic binding. We tried to establish a parallel between this binding and the mechanism of anaesthesia involving the nerve membrane proteins.

Anesthetics, Local↗
PubMed2006

Alteration of cytochrome oxidase subunit I labeling is associated with severe mitochondriopathy in NRTI-related hepatotoxicity in HIV patients.

Liver mitochondrial toxicity induced by nucleoside reverse transcriptase inhibitors (NRTI) in human immunodeficiency virus (HIV) patients has been associated with a wide range of liver involvement ranging from low-grade hepatotoxicity, asymptomatic lactacidemia to severe liver insufficiency, with massive steatosis and life-threatening lactic acidosis. Considerable efforts have been made in the last few years to establish clinical guidelines to avoid life-threatening NRTI-associated lactic acidosis. However, the important issue of low-grade NRTI-associated hepatotoxicity still needs to be unravelled since its natural history is largely unknown. We have recently reported a series of 13 monoinfected HIV patients with low-grade NRTI-associated toxicity. Our results outlined the heterogeneity of NRTI-induced hepatotoxicity and raised the question of its diagnosis. The present study evaluates the expression of cytochrome oxidase (COX) subunits I and IV, encoded by mitochondrial and nuclear DNA, respectively, in NRTI hepatotoxicity. The aim of our study was to compare the detection rate of mitochondrial abnormalities of immunohistochemistry for COX subunit I with electron microscopy. COX subunit I and IV labeling was performed together with light microscopy and ultrastructural analysis in a series of 55 liver biopsies from HIV monoinfected and HIV-hepatitis C virus coinfected patients. Clinical data were also recorded. Our major findings were: (i) decreased COX subunit I labeling is associated with severe ultrastructural mitochondrial alterations and may represent overt NRTI-induced mitochondrial cytopathy; (ii) mild ultrastructural damage associated with normal COX subunit I labeling is of unknown clinical significance. The results of the study suggest that COX subunit I labeling may be a valuable tool for the diagnosis of mitochondrial liver disease in HIV patients.

Adult↗
PubMed1991

Cytochrome oxidase activity as a measure of synaptogenesis by multifunctional neuron L7 of Aplysia.

Neuron L7 of the marine mollusc, Aplysia californica, is unique in that it innervates five different target tissues in the animal. We show that when L7 is grown in vitro with two of these targets, that is, muscle cells isolated from the auricle or the gill vein, newly formed L7 neurites contact the muscle cells. Chemical synapses are formed since intracellular stimulation of L7 elicits contraction of individual muscle cells. Interestingly, auricle muscles are also innervated by neuron RBhe and co-cultures of RBhe and auricle muscle cells also exhibit synapse formation. To explore the molecular basis for synaptogenesis between L7 and its targets, it would be useful to quantify the extent of synapse formation in vitro, that is, to determine how many muscle cells can be innervated by a single L7. We show that this can be attained by staining for cytochrome oxidase activity. Cultures of auricle and gill vein muscles were exposed to the appropriate neurotransmitter in order to elicit contraction. The cells were then fixed and stained. In both cases, only cells that contracted were stained and electron microscopy showed reaction product associated with the cristae of mitochondria. When this procedure was applied to cultures of L7 and muscle cells, 38 +/- 2.8% (S.E.M.; n = 7) of the cells on the neurites were stained and therefore responded to L7 stimulation. Thus, part of the L7-RBhe circuit can be assembled in vitro and the extent of synaptogenesis can be accurately quantitated.

Animals↗
PubMed1991

Acidic cytochrome c6 of unicellular cyanobacteria is an indispensable and kinetically competent electron donor to cytochrome oxidase in plasma and thylakoid membranes.

Cytochromes c6 from three cyanobacteria were tested as substrates for membranous cyt. c oxidase(aa3) of Anacystis and Synechocystis using intact spheroplasts or isolated plasma(CM) and thylakoid(ICM) membranes. Neither spheroplasts nor CM/ICM gave significant O2 uptake rates with NADH without added cyt. c. Horse cyt. c (at low ionic strength) or cyt. c6 from Anacystis, Synechocystis or Microcystis (at high ionic strength) supported substantial HCN- & CO-sensitive NADH oxidase activity, consistent with in vivo O2 uptake. Cyanobacterial respiratory electron transport involves NADH dehydrogenase(fpN), plastoquinone, cyt. b/c(f), cyt. c6 & cyt. aa3, in both CM & ICM. In ICM, fpN and cyt. aa3 are functionally replaced in the light by PS II and PS I, respectively. In both membranes, cyt. c6 is an obligatory electron donor to cyt. aa3 &/or to P700. Respiratory action of acidic cyt. c6 (in unicellular species) may be unmasked only under conditions of elevated ionic strength.

Animals↗
PubMed1992

Competition between an aphakic and an occluded eye for territory in striate cortex of developing rhesus monkeys: cytochrome oxidase histochemistry in layer 4C.

Monkey models were used to examine the effects of competition for cortical territory between two eyes which were deprived simultaneously, but each eye experienced a different type of deprivation. We wanted to determine whether, under this condition of binocular unequal deprivation, the postnatal process of segregation into ocular dominance columns proceeds according to the same rules as those that apply to competition between a deprived and an undeprived fellow eye. Our models involved surgical removal of the natural lens from one eye in newborn rhesus monkeys. The resulting aphakia was corrected optically to a near point with extended-wear contact lenses. The fellow eyes were either left unmanipulated or occluded with opaque contact lenses for varying periods during the day. At the end of the rearing period, some monkeys from each experimental group had either one eye enucleated or sustained injury to the retinal ganglion cells of one eye. The histochemical reaction for cytochrome oxidase was used to reveal the widths of ocular dominance columns in layer 4C of striate cortex in these monkeys. Under all experimental conditions, the axons related to the two eyes occupied segregated fields. The amount of cortical territory related to the aphakic, optically corrected eye depended on the manipulations of the fellow eye. In competition with an unmanipulated fellow eye, the aphakic eye's territory was greatly reduced. In competition with a part-time occluded eye, its territory was reduced to a lesser degree, depending on the duration of the occlusion. In competition with a continuously occluded eye, however, the space related to the aphakic, optically corrected eye was slightly greater than that related to the occluded eye. Since neither the aphakic nor the continuously occluded eye receives normal visual input, they are both impaired. Therefore, they may compete on an almost equal basis for synaptic territory in layer 4C of striate cortex. Moreover, it is likely that activities originating in the aphakic and the continuously occluded eye are asynchronous, and that this condition is sufficient to drive the postnatal segregation of inputs from the two deprived eyes.

Aging↗
PubMed1991

Hormonal regulation of cytochrome oxidase subunit messenger RNAs in rat Sertoli cells.

Using differential hybridization to screen a rat Sertoli cell cDNA library for hormonally regulated gene products, we isolated a clone, designated 13-10, which contained a 1.0-kilobase insert and hybridized to a 1.7-kilobase message in total testis, Sertoli cells, and peritubular cells. This mRNA was decreased relative to untreated control levels in total testicular RNA from hypophysectomized rats, but was increased by FSH treatment begun on the day of hypophysectomy. FSH caused a transient rise in 13-10 mRNA at 24 h in cultured Sertoli cells. There was no comparable rise in beta-actin RNA or the RNA/DNA ratio at this time, suggesting that the effect on 13-10 was specific. Testosterone had no effect at any time interval studied. The 13-10 mRNA was not increased in peritubular cells treated in vitro with FSH or testosterone. Sequence analysis of 13-10 revealed more than 99% homology with a portion of the sequence of rat liver cytochrome oxidase subunit I (COX I). The clone included 58% of the open reading frame of COX I as well as that for the adjacent Ser-tRNA. COX I is a mitochondrial gene, and Southern analysis confirmed 13-10 sequence in testicular mitochondrial DNA. In addition to FSH, forskolin and (Bu)2cAMP also increased COX I steady state mRNA in Sertoli cells (3.8-, 4.1-, and 9.2-fold, respectively). (Bu)2cAMP increased mRNA for other mitochondrial gene products, COX subunit II and 16S rRNA (6.9- and 5.4-fold, respectively), whereas the smaller effects elicited by forskolin and FSH were not statistically significant.(ABSTRACT TRUNCATED AT 250 WORDS)

Actins↗
PubMed1995

Sex differences in cytochromes oxidase and P-45011 beta in the rat adrenal cortex.

A comparative study of cytochrome c oxidase (COX) activity and expression as well as cytochrome P-45011 beta expression has been carried out on the adrenal cortex of male and female rats. COX has also been examined in rat liver. In addition, the effect of testosterone replacement in orchiectomized male rats on adrenal COX has also been investigated. Adult male rats had higher COX activity in adrenal (255%) and liver (144%) mitochondria compared to adult female rats. Male rat adrenals and liver also had increased levels of COX II, a mitochondria-encoded COX subunit, and of COX IV, a nucleus-encoded COX subunit, as measured by Western analysis. In contrast, cytochrome P-45011 beta levels were lower (48%) in adrenal mitochondria from male rats than those of female rats. There was no significant sex difference in the level COX II and COX IV mRNAs in adrenal or liver, whereas the cytochrome P-45011 beta mRNA was 4-fold higher in female adrenals than in males. In male rats, orchiectomy caused a 23% decrease and testosterone replacement a 66% increase in adrenal COX activity. There were no corresponding changes in the levels of mRNAs encoding for COX subunits, suggesting post-transcriptional effects of testosterone on COX. These results are consistent with a regulatory role of testosterone on the expression of components of the respiratory and steroidogenic electron transport chains.

Adrenal Cortex↗
PubMed1986

Proton translocation by cytochrome oxidase in (antimycin + myxothiazol)-treated rat liver mitochondria using ferrocyanide or hexammineruthenium as electron donor.

When O2 was injected into an anaerobic suspension of valinomycin-treated rat liver mitochondria inhibited with rotenone, antimycin, and myxothiazol, a small amount of O2 (0.23-0.33 ng-atom of O/mg of protein) was reduced extremely rapidly (within the 2 s time-resolution of the oxygen electrode). The subsequent steady-state rate of flow of electrons to oxygen was very low [less than 3 nequiv. X s-1 X (g of mitochondrial protein)-1]. In the presence of valinomycin there was a rapid ejection of protons synchronous with the rapid phase of O2 consumption corresponding to 0.38-0.61 nequiv. of H+ X (mg of mitochondrial protein)-1. When valinomycin was replaced by carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) there was a rapid alkalification of the medium corresponding to 0.20-0.42 nequiv. of H+ X (mg of mitochondrial protein)-1. When 2 mM-Fe(CN)6(4-) was present to re-reduce endogenous cytochrome c, O2 consumption was still biphasic but the second phase of O2 consumption was very much more rapid [600 nequiv. X s-1 X (g of protein)-1], and resulted in the virtually complete consumption of the O2 in the pulse within 4 s. With 60 microM-Ru(NH3)6(2+) as reductant, O2 consumption was even faster [1200 nequiv. X s-1 X (g of protein)-1]. In a medium containing 150 mM-choline chloride with Ru(NH3)6(2+) as reductant, the proton per reducing equivalent stoichiometry (delta H+O/e-) was +0.95 in the presence of valinomycin and -0.94 in the presence of FCCP. In choline chloride medium containing Ru(NH3)6(2+) and valinomycin, there was an uptake of K+ ions corresponding to 1.86 K+/e-. It is concluded that nearly 1 proton is translocated outwards through cytochrome oxidase per oxidizing equivalent injected in this medium. In low ionic strength sucrose-based medium, with Ru(NH3)6(2+) as reductant, delta H+O/e- was 1.05 in the presence of valinomycin, and -0.71 in the presence of FCCP. It is concluded that the translocation of protons is accompanied by net acid production in this medium.

Animals↗
PubMed1998

mtDNA replicative potential remains constant during ageing: polymerase gamma activity does not correlate with age related cytochrome oxidase activity decline in platelets.

Progressive age-related oxidative phosphorylation (OxPhos) decline is well known in human tissues. Depletion of mitochondrial DNA (mtDNA) causes OxPhos defects in patients with myopathic syndromes and deficient mtDNA replication has been observed in cells cultured from patients with mitochondrial disease. Patients undergoing treatment for AIDS develop OxPhos defects via mtDNA depletion resulting from inhibition of mtDNA polymerase gamma (Polgamma) by 2'-deoxy 3'-azido thymidine. These findings by others give rise to a possible link between mtDNA replication and bioenergetic decline in disease and during ageing. We have designed an in vitro assay for Polgamma function in small tissue samples to explore this possible link. Platelet homogenate Polgamma showed an activity with a K m of 150 microM (dTTP), a V max of 11.8 pmol/min/mg, inhibited (41% inhibition; 50 microM) by ethidium bromide. Determination of several storage characteristics showed that platelets were a convenient source of Polgamma for assay. Polgamma activity in 45 subjects did not coincide with significant age-related decline (P<0.002; P) observed in cytochrome oxidase (CytOx) activity or with citrate synthase activity. Of the activities studied, the only significant age-wise variation was a 24% CytOx deficiency in elderly (>50; n = 19) compared to young (<51; n = 24) individuals (P<0.01; t). These results suggest a maintenance of total cellular mtDNA Polgamma processive levels during ageing, largely independent of total cellular bioenergetic status or mitochondrial number/density. The processive component of Polgamma is therefore unlikely to make a major contribution to age-related bioenergetic activity decline. This does not, however, preclude the possibility that transient periods of inhibition at crucial points of the cell cycle or development may augment existing intracellular deficiencies. The assay described here greatly facilitates study of Polgamma activity in patients with conditions involving mtDNA depletion or rearrangement.

Adult↗
PubMed1996

Bicuculline microinjections into the ventral tegmental area of the rat: alteration of self-stimulation thresholds and of cytochrome oxidase activity in the brain.

Abuse of drugs that potentiate GABAergic neurotransmission, namely benzodiazepines, is difficult to understand because this potentiation should elicit, among other effects, a decrease in activity within the mesolimbic system. Abuse of benzodiazepines is difficult to understand since the opposite, namely an increase in mesolimbic activity, has been implicated in drug abuse as well as in the rewarding effect of direct mesolimbic stimulation. In order to evaluate how the activity of the mesolimbic system depends on mesolimbic GABAergic influence, a GABAA receptor antagonist, bicuculline methiodide, was unilaterally injected into the ventral tegmental area and its effect on self-stimulation thresholds derived from stimulations applied to the same area was evaluated. Microinjection of 15, 20 and 30 ng increased the stimulation threshold. This decrease in stimulation efficiency lasted no more than 15 min after which baseline levels were obtained. Such a decrease is paradoxical considering that the manipulation should have released the ventral tegmentum from a tonic inhibitory influence. The metabolic consequences of repeated injections of 30 ng bicuculline were furthermore evaluated by cytochrome oxidase histochemistry. The staining was found to be weak around the injection site and dense in the ipsilateral nucleus accumbens. Release of a tonic GABAergic inhibition added to some cytotoxic damage probably resulted in an increased metabolic activity of this system. The presently reported paradoxical response of the ventral tegmentum and mesolimbic system to a GABAergic challenge may account for the paradoxical relationship between some behavioral properties of the mesolimbic system and GABAergic drugs.

Animals↗