Purification and properties of hypoxically induced lactate dehydrogenase from barley roots

Using Affigel Blue and oxamate-agarose affinity chromatography, lactate dehydrogenase (LDH) was purified 2000-fold from hypoxically induced barley roots. Molecular weights of the native and sodium dodecyl sulfate-denatured LDH protein were 157 and 40 kilodaltons, respectively, indicating a tetramer. Purified barley LDH was very similar in size and kinetic properties to potato LDH. However, their amino acid compositions differed substantially and antibodies raised against barley LDH did not cross-react with potato LDH on immunoblots, implying that the barley and potato LDHs are not closely related proteins. In vivo [³⁵S] methionine labeling and immunoprecipitation experiments indicated that hypoxia increased the rate of LDH protein synthesis, and immunoblot analysis showed that LDH protein levels rose during hypoxia. We conclude that increased enzyme synthesis plays a major part in the induction of LDH enzyme activity by low O₂ levels in barley roots.     

Identification and characterization of a hypoxically induced maize lactate dehydrogenase gene

In cereal root tissue, hypoxia induces the enzyme lactate dehydrogenase (LDH); (S)-lactate:NADH oxidoreductase, EC 1.1.1.27). In barley, both biochemical and genetic data indicate that five isozymes are induced under hypoxia. These isozymes are tetramers and arise from the random association of the products of two Ldh genes. The induction of LDH activity in root tissue has been shown to be correlated to an increase in LDH protein and Ldh mRNA.In order to more fully characterize the hypoxic induction of LDH, we have isolated a maize Ldh genemic clone which has strong homology at both the amino acid and nucleotide level to the barley LDH cDNA clones. The Ldh1 gene consists of two exons separated by a 296 bp intron, has the expected eukaryotic regulatory signals and a sequence that has strong homology to the maize anaerobic regulatory element.     

beta-Amylase from Mustard (Sinapis alba L.) Cotyledons : Immunochemical Evidence for Synthesis de Novo during Photoregulated Seedling Development

In barley (Hordeum vulgare L.), alcohol dehydrogenase (ADH) and lactate dehydrogenase (LDH) are induced by anaerobiosis in both aleurone layers and roots. Under aerobic conditions, developing seeds of cv Himalaya accumulate ADH activity, which survives seed drying and rehydration. This activity consists almost entirely of the ADH1 homodimer. Activity of LDH also increases during seed development, but the level of activity in dry or rehydrated seeds is very low, indicating that this enzyme may not be involved in anaerobic glycolysis during the initial stages of germination. In contrast to ADH, the LDH isozymes present in developing seeds are similar to those found in uninduced and induced roots. Developmental expression of ADH and LDH was monitored from 0 to 24 days postgermination. Neither activity was induced to any extent in the germinating seeds; however, both enzymes were highly induced by anoxia in root tissue during development. Based on gel electrophoresis, this increase in activity results from the differential expression of different Adh and Ldh genes in root tissue. The changes in ADH and LDH activity levels were matched by changes in the amount of these particular proteins, indicating that the increase in activity results from de novo synthesis of these two proteins. The level of inducible LDH activity in an ADH1⁻ mutant was not found to differ from cv Himalaya. We suggest that although the ADH⁻ plants are more susceptible to flooding, they are not capable of responding to the lack of ADH1 activity by increasing the amount of LDH activity in root tissue.     

Hypoxic induction of alcohol and lactate dehydrogenases in lupine seedlings

Seedlings of lupine (Lupinus luteus L. cv. Juno) were exposed for up to 96 hours to 1 to 2 kPa partial pressure oxygen (hypoxic treatment) and activities of alcohol dehydrogenase (ADH), lactate dehydrogenase (LDH) and their isoform profiles were determined. Roots of lupine seedlings were grown in a nitrogen flushed nutrient solution while their shoots were in air. Prolonged hypoxia led to a reduction of root elongation. This was accompanied by reduced increase in dry weight suggesting that insufficient carbohydrate supply was the cause of retarded growth of lupine roots. Hypoxically treated roots showed induction of ADH and LDH acivities. The maximum increase in LDH activity was low (2-fold) in contrast to ADH activity, which increased up to 7-fold. Hypoxic treatment of roots did not affect the activities of ADH and LDH in hypocotyls and cotyledons. Analysis of ADH and LDH activity gels indicated in roots 1 and 2 isoforms, respectively. The level of isozymes of both enzymes increased in roots upon exposure to hypoxic stress. Differences in isoenzymatic spectrum of ADH and LDH between roots, hypocotyls and cotyledons indicate organ specificity of isozymes of both enzymes. The importance of alcohol and lactate fermentation in roots to cope with hypoxic stress is discussed.     

Effect of root applied 24-epibrassinolide on carbohydrate status and fermentative enzyme activities in cucumber (<Emphasis Type="Italic">Cucumis sativus</Emphasis> L.) seedlings under hypoxia

The effects of 24-epibrassinolide (EBR) added to nutrient solution on growth of cucumber (Cucumis sativus L.) under root-zone hypoxia were investigated. Cucumber seedlings were hydroponically grown for 8 days in normoxic and hypoxic nutrient solutions with and without addition of EBR at 1 μg l⁻¹. EBR exerted little influence on plant performance in the normoxic nutrient solution, while the chemical alleviated root-zone hypoxia-induced inhibition of root and shoot growth and net photosynthetic rate (Pn). EBR added to hypoxic nutrient solution caused an increase in the concentration of fructose, sucrose, and total soluble sugars in the roots but not in the leaves. Root-zone hypoxia enhanced the activities of lactate dehydrogenase (LDH), alcohol dehydrogenase (ADH), and pyruvate decarboxylase in the roots. Interestingly, EBR further enhanced ADH activity but lowered LDH activity in hypoxic roots. These results suggest that EBR added to hypoxic nutrient solution may stimulate the photosynthate allocation down to roots and the shift from lactate fermentation to alcohol fermentation in hypoxic roots, resulting in the increase in ATP production through glycolysis and the avoidance of cytosolic acidosis and eventually enhanced tolerance of cucumber plants to root-zone hypoxia.     

Anaerobic induction of alanine aminotransferase in barley root tissue

Alanine aminotransferase, otherwise called glutamate-pyruvate aminotransferase (GPT), activity increases up to fourfold during several days of anaerobic induction in barley (Hordeum vulgare L.) roots, reaching a maximum activity of 13 international units per gram fresh weight. This increase in activity paralleled the increase in alcohol dehydrogenase activity in the same root tissue. Upon return to aerobic conditions, the induced GPT activity declined with an apparent half-life of 2 days. The isozyme profile of GPT in barley root tissue comprised one band of activity; in maize there were three bands of activity, the bands with greater mobility had much lower activity. Native polyacrylamide gel electrophoresis indicated that the induction of GPT activity results from an increase in the level of activity of these bands; no other activities were detected. When root tissue was induced under different levels of hypoxia (0%, 2%, 5%, and 21% O₂), changes in GPT activity were found to increase with lower levels of oxygen. Comparisons of GPT induction in barley, maize (Zea mays), rye, (Secale cereale) and wheat (Triticum aestivum) indicate that this enzyme is induced in the root tissue of all of these cereals; however, anaerobic root conditions do not result in the induction of GPT activity in leaf tissue. The dependence of GPT induction on high levels of nitrate in the media was tested by comparing activity levels in Hoagland solution and a nitrate-free nutrient solution. GPT activity was induced to similar levels under both conditions. These results indicate that alanine aminotransferase shows a very similar pattern of induction to alcohol dehydrogenase in barley root tissue and may be important in anaerobic glycolysis.     

Physical and Functional Association of Lactate Dehydrogenase (LDH) with Skeletal Muscle Mitochondria

The intracellular lactate shuttle hypothesis posits that lactate generated in the cytosol is oxidized by mitochondrial lactate dehydrogenase (LDH) of the same cell. To examine whether skeletal muscle mitochondria oxidize lactate, mitochondrial respiratory oxygen flux (JO₂) was measured during the sequential addition of various substrates and cofactors onto permeabilized rat gastrocnemius muscle fibers, as well as isolated mitochondrial subpopulations. Addition of lactate did not alter JO₂. However, subsequent addition of NAD⁺ significantly increased JO₂, and was abolished by the inhibitor of mitochondrial pyruvate transport, α-cyano-4-hydroxycinnamate. In experiments with isolated subsarcolemmal and intermyofibrillar mitochondrial subpopulations, only subsarcolemmal exhibited NAD⁺-dependent lactate oxidation. To further investigate the details of the physical association of LDH with mitochondria in muscle, immunofluorescence/confocal microscopy and immunoblotting approaches were used. LDH clearly colocalized with mitochondria in intact, as well as permeabilized fibers. LDH is likely localized inside the outer mitochondrial membrane, but not in the mitochondrial matrix. Collectively, these results suggest that extra-matrix LDH is strategically positioned within skeletal muscle fibers to functionally interact with mitochondria.     

Metabolic and molecular responses in Nile tilapia,Oreochromis niloticus during short and prolonged hypoxia

The strictly aquatic breathing Nile tilapia, Oreochromis niloticus is an extremely hypoxia-tolerant fish. To augment our understanding of the effects of hypoxia on anaerobic glycolysis in the Nile tilapia, we studied the effect of short-term for 1 day (trial 1) and long-term for 30 days (trial 2) hypoxia on a selected glycolytic enzymes activity and mRNA expression in liver and white muscle. The hypoxic oxygen concentrations used in the two trials were 2, 1, and 0.5 mg O₂ L^?1 for comparison with a control normoxic group 8 mg O₂ L^?1. The activity of phosphofructokinase (PFK), pyruvate kinase (PK), and lactate dehydrogenase (LDH) in liver and white muscle except liver LDH decreased in trial 1 and increased in trial 2. Assessments of mRNA levels in trial 1 revealed that PFK was downregulated and LDH was upregulated in liver and white muscle, while PK fluctuated between upregulation in liver and downregulation in white muscle. Meanwhile, PK and LDH were upregulated while PFK was similar to control values in both tissues in trial 2. Comet assay results demonstrated an increase in DNA damage that was directly proportional to increasing hypoxic concentrations. This damage was more pronounced in trial 1. This suggests that the Nile tilapia cope better with long-term hypoxic conditions, possibly as an adaptive response.     

G proteins as regulators in ethylene-mediated hypoxia signaling

Waterlogging or flooding are frequently or constitutively encountered by many plant species. The resulting reduction in endogenous O₂ concentration poses a severe threat. Numerous adaptations at the anatomical, morphological and metabolic level help plants to either escape low oxygen conditions or to endure them. Formation of aerenchyma or rapid shoot elongation are escape responses, as is the formation of adventitious roots. The metabolic shift from aerobic respiration to anaerobic fermentation contributes to a basal energy supply at low oxygen conditions. Ethylene plays a central role in hypoxic stress signaling, and G proteins have been recognized as crucial signal transducers in various hypoxic signaling pathways. The programmed death of parenchyma cells that results in hypoxia-induced aerenchyma formation is an ethylene response. In maize, aerenchyma are induced in the absence of ethylene when G proteins are constitutively activated. Similarly, ethylene induced death of epidermal cells that cover adventitious roots at the stem node of rice is strictly dependent on heterotrimeric G protein activity. Knock down of the unique Gα gene RGA1 in rice prevents epidermal cell death. Finally, in Arabidopsis, induction of alcohol dehydrogenase with resulting increased plant survival relies on the balanced activities of a small Rop G protein and its deactivating protein RopGAP4. Identifying the general mechanisms of G protein signaling in hypoxia adaptation of plants is one of the tasks ahead.Key words: submergence, hypoxia, ethylene, G protein, reactive oxygen species, H₂O₂     

Does tolerance of wheat to salinity and hypoxia correlate with root dehydrogenase activities or aerenchyma formation?

The effects of NaCl salinity (100 or 150 mol m^-3) and hypoxia on seedlings of several wheat varieties (Lyallpur-90, SARC-1, Pato, Tchere, Pb-85, Siete Cerros, Chinese Spring and a Chinese Spring × Thinopyrum elongatum amphidiploid) were studied in solution culture. In vivo studies of activities of different enzymes (alcohol dehydrogenase (ADH), lactate dehydrogenase (LDH) and cytochrome oxidase (COase)) extracted from Pato and Pb-85 included the effect of salinity with and without hypoxia, while during in vitro studies, NaCl, glycinebetaine and proline were added to the assay mixture. The extent of aerenchyma formation was also determined in Pato, Chinese Spring and a Chinese Spring × Thinopyrum elongatum amphidiploid. Imposition of hypoxia greatly induced ADH and LDH activity in roots of wheat seedlings after a week-long exposure. However, exposure of roots to salinity also slightly increased LDH and ADH activity compared with the non-saline control. On a relative basis, Pato had higher ADH activity under hypoxic (21×) or saline-hypoxic stress (20×) than in aerated conditions. Hypoxia alone or in the presence of salts decreased COase activity in both Pato and Pb-85. The in vitro studies revealed that NaCl (on an equimolar basis at up to 500 mol m^-3) is more disruptive than glycinebetaine or proline. LDH was more sensitive to NaCl than ADH. Aerenchyma development was higher near the root-shoot interface compared to near the root tip. Salinity under hypoxic conditions significantly reduced aerenchyma development near the root tip and root-shoot interface compared to hypoxia alone. Neither enzyme activity nor aerenchyma formation could account for varietal differences in tolerance to hypoxia alone or in combination with salinity.     

Respiratory and metabolic responses of the Australian Yabby Cherax destructor to progressive and sustained environmental hypoxia

The Australian Yabby, Cherax destructor, inhabits occasionally hypoxic water. The respiratory gas, acid-base, metabolite and energetic status of this crayfish was assessed during progressive hypoxia and during 3 h at a water PO₂ of 1.33 kPa. The O₂ affinity of haemocyanin from C. destructor was increased by lactate (Δlog P ₅₀/Δlog[lactate] = −0.111) and by Ca (Δlog P ₅₀/Δlog[Ca] = −0.62) but not by urate. While the non-bicarbonate buffering capacity was low (Δ[HCO₃ ⁻]/ ΔpH=−4.89) the haemocyanin had a low sensitivity to pH changes (ϕ = −0.33). The crayfish showed a compensatory hyperventilation, which induced a respiratory alkalosis, until the water O₂ partial pressure declined below 2.67 kPa, after which the O₂ uptake rate was approximately 10% of normoxic rates. The high haemocyanin-O₂ affinity maintained haemolymph O₂ content during progressive hypoxia despite the normally low arterial O₂ partial pressure of C. destructor. During severe hypoxia, pH decreased but increased lactate aided in maintaining haemocyanin-O₂ saturation. The importance of regulated haemocyanin-O₂ affinity in hypoxic C. destructor was reduced by lowered metabolism, including reduced cardiac output, and the consequent reduction in O₂ requirement. Anaerobiosis became important only at very low PO₂ but thereafter proceeded rapidly, supported by a marked hyperglycaemia. There was no depletion of adenylates, even after 3 h of severe hypoxia. The tail muscle of C. destructor held small amounts of glycogen which would sustain anaerobiosis for a only a few hours. Hypometabolism seems an important hypoxic response but severe hypoxia may encourage the crayfish to breathe air. Accepted: 26 February 1998     

Quantitative phosphoproteomic analysis of prion-infected neuronal cells

Following cultivation of distinct mesenchymal stem cell (MSC) populations derived from human umbilical cord under hypoxic conditions (between 1.5% to 5% oxygen (O₂)) revealed a 2- to 3-fold reduced oxygen consumption rate as compared to the same cultures at normoxic oxygen levels (21% O₂). A simultaneous measurement of dissolved oxygen within the culture media from 4 different MSC donors ranged from 15 μmol/L at 1.5% O₂ to 196 μmol/L at normoxic 21% O₂. The proliferative capacity of the different hypoxic MSC populations was elevated as compared to the normoxic culture. This effect was paralleled by a significantly reduced cell damage or cell death under hypoxic conditions as evaluated by the cellular release of LDH whereby the measurement of caspase3/7 activity revealed little if any differences in apoptotic cell death between the various cultures. The MSC culture under hypoxic conditions was associated with the induction of hypoxia-inducing factor-alpha (HIF-1α) and an elevated expression of energy metabolism-associated genes including GLUT-1, LDH and PDK1. Concomitantly, a significantly enhanced glucose consumption and a corresponding lactate production could be observed in the hypoxic MSC cultures suggesting an altered metabolism of these human stem cells within the hypoxic environment.     

Oxygen dynamics in a salt-marsh soil and in Suaeda maritima during tidal submergence

Habitats occupied by many halophytes are not only saline, but are also prone to flooding and yet surprisingly few studies have evaluated submergence tolerance in halophytes. Sediment, floodwater, and intra-plant O₂ dynamics were evaluated during tidal submergence for the leaf-succulent halophyte Suaeda maritima (L.) Dum. For S. maritima growing in soil just above the mud flat in a UK salt marsh, the soil was only moderately hypoxic just prior to tidal inundation, presumably owing to drainage and O₂ entry facilitated by frequent, large cracks. O₂ declined to very low levels following high tide. By contrast, mud flat sediment remained waterlogged, lacked cracks, and was anoxic. Plant O₂ dynamics were investigated using field-collected plants in sediment blocks transported to a controlled-submergence system in a glasshouse. Submergence during night-time resulted in anoxia within leaves, whereas during day-time O₂ was produced by underwater photosynthesis. The thin lateral roots of S. maritima presumably access some O₂ from hypoxic sediments, but could also experience transient episodes of severe hypoxia/anoxia, especially as any internal O₂ movement from shoots would be small owing to the low gas-filled porosity in roots. Fermentative metabolism to lactate, producing some ATP in the absence of O₂, might contribute to tolerance of transient O₂ deficits. Lactate was high in root tissues, whereas ethanol production (tissue and incubation medium contents) was low, both in comparison with values reported for other species. Our findings demonstrate the importance of tolerance to transient waterlogging and submergence for the halophyte S. maritima growing in a tidal salt marsh.     

Oxygen-Sensitive K+ Channels Modulate Human Chorionic Gonadotropin Secretion from Human Placental Trophoblast

Human chorionic gonadotropin (hCG) is a key autocrine/paracrine regulator of placental syncytiotrophoblast, the transport epithelium of the human placenta. Syncytiotrophoblast hCG secretion is modulated by the partial pressure of oxygen (pO₂), reactive oxygen species (ROS) and potassium (K⁺) channels. Here we test the hypothesis that K⁺ channels mediate the effects of pO₂ and ROS on hCG secretion. Placental villous explants from normal term pregnancies were cultured for 6 days at 6% (normoxia), 21% (hyperoxia) or 1% (hypoxia) pO₂. On days 3–5, explants were treated with 5mM 4-aminopyridine (4-AP) or tetraethylammonium (TEA), blockers of pO₂-sensitive voltage-gated K⁺ (K_V) channels, or ROS (10–1000μM H₂O₂). hCG secretion and lactate dehydrogenase (LDH) release, a marker of necrosis, were determined daily. At day 6, hCG and LDH were measured in tissue lysate and ⁸⁶Rb (K⁺) efflux assessed to estimate syncytiotrophoblast K⁺ permeability. hCG secretion and ⁸⁶Rb efflux were significantly greater in explants maintained in 21% pO₂ than normoxia. 4-AP/TEA inhibited hCG secretion to a greater extent at 21% than 6% and 1% pO₂, and reduced ⁸⁶Rb efflux at 21% but not 6% pO₂. LDH release and tissue LDH/hCG were similar in 6%, 21% and 1% pO₂ and unaffected by 4-AP/TEA. H₂O₂ stimulated ⁸⁶Rb efflux and hCG secretion at normoxia but decreased ⁸⁶Rb efflux, without affecting hCG secretion, at 21% pO₂. 4-AP/TEA-sensitive K⁺ channels participate in pO₂-sensitive hCG secretion from syncytiotrophoblast. ROS effects on both hCG secretion and ⁸⁶Rb efflux are pO₂-dependent but causal links between the two remain to be established.     

Effects of intermittent hypoxia different regimes on mitochondrial lipid peroxidation and glutathione-redox balance in stressed rats

The purpose of this study was to compare the influence of two regimes of intermittent hypoxia (IH) [repetitive 5 cycles of 5 min hypoxia (7% O₂ or 12% O₂ in N₂) followed by 15 min normoxia, daily for three weeks] on oxidative stress protective systems in liver mitochondria. To estimate the effectiveness of hypoxia adaptation at the early and late preconditioning period, we exposed rats to acute 6-h immobilization at the 1^st and 45^th days after cessation of IH. We showed that severity of hypoxic episodes during IH might initiate different adaptive programs. Moderate hypoxia during IH prevents mitochondrial glutathione pool depletion induced by immobilization stress, maintains GSH-redox cycle via activation of glutathione peroxidase, glutathione-S-transferase, glutathione reductase, NADP⁺-dependent isocitrate dehydrogenase, and increases Mn-SOD activity. Such regimen of hypoxic preconditioning caused the decrease of mitochondrial superoxide anion generation as well as of basal and stimulated in vitro lipid peroxidation and this protective effect remained for 45 days under renormoxic conditions. Hypoxic adaptation in a more severe regimen exerted beneficial effects on the mitochondrial antioxidant defense system only at its later phase.     

1H-NMR study of the metabolome of a moderately hypoxia-tolerant fish, the common carp (Cyprinus carpio)

All 20.000 different fish species vary greatly in their ability to tolerate and survive fluctuating oxygen concentrations in the water. Especially fish of the genus Carassius, e.g. the crucian carp and the goldfish, exhibit a remarkable tolerance to limited/absent oxygen concentrations. The metabolic changes of anoxia-tolerant crucian carp were recently studied and published. Contrary to crucian carp, the hypoxia-tolerant common carp cannot survive a complete lack of oxygen (anoxia). Therefore, we studied the ¹H-NMR-based metabolomics of brain, heart, liver and white muscle extracts of common carp, subjected to anoxia (0 mg O₂ l^?1) and hypoxia (0.9 mg O₂ l^?1) at 5 °C. Specifically, fish were exposed to normoxia (i.e. 9 mg O₂ l^?1; controls 24 h, 1 week and 2 weeks), acute hypoxia (24 h), chronic hypoxia (1 week) and chronic hypoxia (1 week) with normoxic reoxygenation (1 week). Additionally, we also investigated the metabolic responses of fish to anoxia for 2 h. Both anoxia and hypoxia significantly changed the tissue levels of standard energy metabolites as lactate, glycogen, ATP/ADP and phosphocreatine. Remarkably, anoxia induced increased lactate levels in all tissues except for the heart whereas hypoxia resulted in decreased lactate concentrations in all tissues except for brains. Furthermore, hypoxia and anoxia influenced amino acids (alanine, valine/(iso)leucine) and neurotransmitters levels (GABA, glutamate). Lastly, we also detected ‘other’ i.e. previously not reported compounds to play a role in the present context. Scyllo-inositol levels changed significantly in heart, liver and muscle, providing novel insights into the anoxia/hypoxic responses of the common carp.     

Metabolic Control of Anaerobic Glycolysis (Overexpression of Lactate Dehydrogenase in Transgenic Tomato Roots Supports the Davies-Roberts Hypothesis and Points to a Critical Role for Lactate Secretion

Roots of all plants examined so far have the potential for both ethanol and lactate fermentation. A short burst of lactate fermentation usually occurs when plant tissues are transferred from normoxic to anoxic conditions. According to the Davies-Roberts hypothesis, the consequent pH drop both initiates ethanol fermentation and blocks further production of lactate by inhibiting lactate dehydrogenase (LDH). However, the role of LDH in this pH control mechanism is still a matter of debate. To perturb the control system in a defined way, a barley LDH cDNA under the control of the cauliflower mosaic virus 35S promoter was introduced into tomato (Lycopersicon esculentum Mill. cv VFMT) using Agrobacterium rhizogenes. The transgenic root clones expressed up to 50 times the LDH activity of controls. The fermentative metabolism of these clones was compared using roots grown previously in normoxic conditions or roots given a 3-d hypoxic pretreatment. During the transition from normoxia to anoxia, lactate accumulation was no faster and no more extensive in transgenic roots than in controls. Similarly, during prolonged anoxia the flux of 14C from [U-14C] glucose to lactate and ethanol was not modified by the expression of the transgene. However, in both transgenic and control roots, hypoxic pretreatment increased the flux to lactate and promoted lactate export to the medium. These results show that LDH has a very low flux control coefficient for lactate fermentation, consistent with the Davies-Roberts hypothesis. Moreover, they suggest that lactate secretion exerts major control over long-term lactate glycolysis in vivo.     

Ultrastructure and metabolism of skeletal muscle fibres in the tench: Effects of long-term acclimation to hypoxia

Summary Tench (Tinca tinca) were acclimated to either aerated (P _{O 2} 17.6 KPa) or hypoxic (P _{O 2} 1.5 KPa) water for 6 weeks.Acclimation to hypoxia resulted in a decrease in mitochondrial volume fraction in both slow (22.9 to 15.0 %) and fast glycolytic (4.5 to 1.8 %) myotomal muscles fibres (P<0.01).Intermyofibrillar mitochondrial populations (4.4 to 1.2% slow; 0.6 to 0.04% fast fibres) were affected to a greater extent than those in the subsarcolemmal zone (18.5 to 13.8% slow; 3.9 to 1.8% fast fibres). After acclimation to hypoxia, cytochrome-oxidase activities decreased by 31 and 33 % in slow and fast fibres, respectively, but were maintained in the liver.Fibre size remained unchanged and actively differentiating fibres were observed in muscles from both groups of fish. Hypoxia resulted in a significant increase in myofibrillar volume fraction in both slow (43.1 to 56.1 %) and fast glycolytic fibres (73.1 to 82.7%) (P<0.05).Glycogen concentrations (mg/100g tissue) for liver (6616) slow muscle (1892) and fast muscle (334) were similar for fish acclimated to aerated or hypoxic water. Acclimation to hypoxia increased carnitine palmitoyl transferase activity (moles substrate utilised g·dry wt_-1 min^-1) in slow (0.42 to 1.1), fast glycolytic muscle (<0.01 to 0.15) and liver (1.1 to 3.7) indicating an enhanced capacity for fatty acid oxidation.Phosphofructokinase activities of fast glycolytic fibres were similar in fish acclimated to either aerated or hypoxic water, consistent with an unaltered capacity for anaerobic glycogenolysis. Hexokinase activities (moles substate utilised, g·dry wt^-1 min^-1) decreased in fast fibres (1.2 to 0.4) but were maintained in the slow muslce (2.1 to 2.5) and liver (4.5 to 4.8) of hypoxic fish. The activities of phosphofructokinase in slow muscle and phosphofructokinase, pyruvate kinase and lactate dehydrogenase in liver were two times higher in fish acclimated to hypoxia. An enhanced capacity for glycolysis in these tissues may reflect a reduced threshold for anaerobic metabolism during activity and/or an adaptation for acute exposure to anoxia in fish acclimated to hypoxia.Abbreviations/Glossary CO cytochrome oxidase activity - CPT carnitine palmitoyltransferase activity - HK hexokinase activity - LDH lactate dehydrogenase activity - PFK phosphofructokinase activity - PK pyruvate kinase activity - Vv volume fractions of cell components - normoxic fish acclimated to aerated water - hypoxic fish acclimated to reduced oxygen tensions - P _{O 2} partial pressure of oxygen tension A preliminary account of part of this work was presented at theXth European Meeting on Muscle and Cell Motility held at Galway, Ireland, in September 1981