Total Adenine Nucleotide Content Research Articles

Decrease in subunit aggregation of phosphoribosylpyrophosphate synthetase: a mechanism for decreased nucleotide concentrations in pyruvate kinase-deficient human erythrocytes

Pyruvate kinase (PK)-deficient RBCs have several unexplained metabolic abnormalities, such as decreased concentrations of total adenine nucleotides (AMP, ADP, and ATP) and total (oxidized and reduced) nicotinamide adenine dinucleotide (NAD). Because 5-phosphoribosyl-1- pyrophosphate (PRPP) is an intermediate in the synthesis of adenine nucleotides and NAD, we investigated PRPP synthetase (PRPPS), the enzyme responsible for PRPP synthesis. This enzyme is regulated, in part, by changes in its state of subunit aggregation. The proportion of aggregated PRPPS can be altered in vitro by ATP and 2,3- diphosphoglycerate (DPG). Because PK-deficient RBCs have decreased ATP and increased DPG concentrations, we examined the state of subunit aggregation of PRPPS in RBCs from normal and PK-deficient subjects, using gel permeation chromatography. Young normal RBCs have more aggregated PRPPS than do older RBCs. In contrast, due to their decreased ATP and increased DPG concentrations, PK-deficient RBCs contain less aggregated PRPPS than do RBCs of comparable age without PK deficiency. These data suggest that PRPPS should be less active in vivo in PK-deficient RBCs. This may play a key role in mediating the decreases in total adenine nucleotide and total NAD concentrations in these RBCs.

Decrease in Subunit Aggregation of Phosphoribosylpyrophosphate Synthetase: A Mechanism for Decreased Nucleotide Concentrations in Pyruvate Kinase-Deficient Human Erythrocytes

Pyruvate kinase (PK)-deficient RBCs have several unexplained metabolic abnormalities, such as decreased concentrations of total adenine nucleotides (AMP, ADP, and ATP) and total (oxidized and reduced) nicotinamide adenine dinucleotide (NAD). Because 5-phosphoribosyl-1-pyrophosphate (PRPP) is an intermediate in the synthesis of adenine nucleotides and NAD, we investigated PRPP synthetase (PRPPS), the enzyme responsible for PRPP synthesis. This enzyme is regulated, in part, by changes in its state of subunit aggregation. The proportion of aggregated PRPPS can be altered in vitro by ATP and 2,3-diphosphoglycerate (DPG). Because PK-deficient RBCs have decreased ATP and increased DPG concentrations, we examined the state of subunit aggregation of PRPPS in RBCs from normal and PK-deficient subjects, using gel permeation chromatography. Young normal RBCs have more aggregated PRPPS than do older RBCs. In contrast, due to their decreased ATP and increased DPG concentrations, PK-deficient RBCs contain less aggregated PRPPS than do RBCs of comparable age without PK deficiency. These data suggest that PRPPS should be less active in vivo in PK-deficient RBCs. This may play a key role in mediating the decreases in total adenine nucleotide and total NAD concentrations in these RBCs.

Intractable unphysiologically low adenylate energy charge values in synaptosome fractions: an explanatory hypothesis based on the fraction's heterogeneity.

Synaptosomes prepared and incubated in a variety of ways from rat cerebra exhibited intractable, unphysiologically low adenylate energy charge values (approximately 0.37-0.60), low total adenine nucleotide contents (approximately 8-10 nmol/mg protein), and much higher adenylate kinase apparent Keq values (approximately 3-8) as compared to intact brain tissue (values of approximately 0.90, 25 nmol/mg, and 0.74, respectively). Synaptosomes prepared from mouse, dog, and chicken cerebra had values essentially identical to those from rat. When incubated under oxygen in a physiological salt solution containing glucose, synaptosomes metabolized more glucose to lactic acid than to CO2, and the addition of 100 microM veratridine caused a two- to threefold stimulation of O2 uptake, lactate accumulation, and CO2 output. It is known that synaptosome fractions contain a substantial number (at least 30-45% by volume) of cytoplasm-containing particles devoid of mitochondria (henceforth termed "cytosolic particles"), and that approximately 80% of brain hexokinase is bound to the outer mitochondrial membrane. For the cytosolic particles, lacking oxidative phosphorylation, to maintain their "in vivo" ATP turnover would require about a 19-fold increase in the glycolytic rate, which is not possible due to limiting amounts of hexokinase, and thus these particles are postulated to be responsible for the high level of aerobic lactate accumulation and the intractable low energy charge values found in synaptosome fractions. The mitochondria-containing particles are postulated to have a normal energy charge, a submaximal glycolytic rate, and minimal lactate production, on the basis of the capacity of veratridine to stimulate synaptosomal O2 uptake and CO2 and lactate output. Calculations based on this "two populations of particles" hypothesis indicate that for synaptosome fractions in general, (1) the cytosolic particles contain approximately 35-64% of the total adenine nucleotides and maintain an energy charge of approximately 0.12; (2) the cytosolic particles and mitochondria-containing particles have adenylate kinase apparent Keq values of approximately 0.21-1.66 and 0.74, respectively, revealing that the higher apparent Keq values of the synaptosome fractions probably are not real departures from equilibrium: and (3) approximately 31-45% of synaptosome fraction protein is contained in debris, which, when taken into account, yields total adenine nucleotide contents in the cytosolic particles and mitochondria-containing particles of approximately 15-24 and approximately 11-19 nmol/mg of particle protein, respectively.

The role of high-energy phosphate in norepinephrine-induced acute renal failure in the dog.

Previous studies have demonstrated that pretreatment with mannitol, furosemide, or bradykinin can attenuate the severity of norepinephrine-induced renal functional impairment. The present studies were designed to evaluate the possibility that these agents are protective, in part, by preserving cellular metabolic integrity. The renal cortex was repetitively biopsied during the course of this study, and high-pressure liquid chromatography was used to analyze the tissue content of adenine nucleotides (expressed in nanomoles per gram of wet tissue). The adenine nucleotide charge ratio (CR) and total adenine nucleotide (TAN) content were calculated as indices of cellular metabolic integrity. In addition to the above-established protective agents, phenoxybenzamine was used to evaluate a direct toxic effect of norepinephrine on renal tissue. Inulin clearance at 3 hours post infusion (expressed as a percent of control) was 7% with norepinephrine alone and, in the protected groups, 36% with bradykinin, 61% with furosemide, 51% with mannitol, and 100% with phenoxybenzamine. There was no change in CR or TAN with phenoxybenzamine. In contrast, during norepinephrine administration CR fell significantly in all other groups. Three hours after stopping norepinephrine, CR had returned toward control values and the level of CR was significantly better in all protected groups when compared with norepinephrine alone. Similarly, the levels of TAN were significantly diminished in the norepinephrine-alone group when compared to all protected groups, and there was significantly more tubular necrosis as well. The maintenance of higher levels of TAN and the preserved ability to regenerate adenosine triphosphate in the protected groups, when compared to the norepinephrine-alone group, support the contention that these agents offer protection, at least in part, by preserving cellular metabolic integrity.

The Adenylate Energy Charge of Vesicular-arbuscular Mycorrhiza of Onions ( Allium cepa L.)

Summary Onion seedlings ( Allium cepa L. cv. Topas) were grown in the presence or absence of Glomus fasciculatus in a sand-loam soil of low phosphate content in which only mycorrhizal plants were able to take up phosphate. Total and free phosphate in roots and shoots and the adenine nucleotides in the roots were measured during the establishment of the mycorrhizal symbiosis and, subsequently, after fertilization of the soil with phosphate. The total concentration of adenine nucleotides was found to be consistently about three times higher in the mycorrhizal roots than in the non-mycorrhizal roots, even at an early stage of the fungal infection and also after the P-fertilization. In contrast, the adenylate energy charge (AEC) was found to be identical in both roots, and virtually constant.

Muscle ammonia metabolism during isometric contraction in humans.

The changes in ammonia (NH3) and amino acid contents in human skeletal muscle during isometric exercise (2/3 maximal voluntary contraction force) to fatigue have been investigated. Biopsies from musculus quadriceps femoris were obtained at rest, fatigue, and 1 and 4 min recovery. Muscle NH3 (n = 9) increased from 1.3 +/- 0.3 (mean +/- SE) mmol/kg dry muscle (dm) at rest to 3.6 +/- 0.6 at fatigue (P less than 0.01) and remained elevated during recovery, whereas the lactate increase after contraction decreased rapidly during recovery. Total adenine nucleotide (TAN) content decreased from 28.7 +/- 0.5 mmol/kg dm at rest to 25.1 +/- 0.6 at fatigue (P less than 0.001). Muscle glutamine did not change after contraction (P greater than 0.05), whereas glutamate decreased (P less than 0.001), and alanine increased (P less than 0.001). In vivo AMP deaminase activity (measured by the rate of TAN decrease) was positively correlated with the percentage of fast-twitch fibers (r = 0.92; P less than 0.001) and the ATP turnover rate (r = 0.75; P less than 0.001) but was not related to the muscle lactate content (r = 0.27; P greater than 0.05). Phosphocreatine decreased to 6.1 +/- 0.7 mmol/kg dm (range = 1-11) after contraction. It is concluded that during exercise activation of AMP deaminase in vivo occurs when a high ATP turnover rate is coupled with a low phosphocreatine level, muscle pH is of minor importance for direct activation of AMP deaminase in vivo, and increases in NH3 do not have an important influence on glycolysis.

Changes in adenylate energy charge and total adenine nucleotide concentration in mouse testis induced by cadmium administration

Subcutaneous administration of CdCl 2 (3 mg/kg body weight) caused a decrease in adenylate energy charge, (ATP + 0.5ADP)/(ATP + ADP + AMP), and a depletion of the adenine nucleotide pool in the testis of mice. This preceded a decrease in LDH-X activity in the testicular extracts. These results biochemically support the hypothesis that the testicular damage induced by cadmium is secondary to anoxia caused by vascular lesion.

ATP, ADP and AMP in plasma from peripheral venous blood

ATP, ADP and AMP in concentrations at least 1 μmol/1 have been found by high pressure liquid chromatography (HPLC) in plasma from peripheral venous blood. Total adenine nucleotide concentrations of about 15–20 μmol/1 can be found in some conventional clinical samples of blood using EDTA as an anticoagulant. EDTA prevented adenine nucleotide conversion to inosine in plasma. In order to estimate concentrations in vivo, the contribution derived from the cell breakage inherent even in careful venous blood sampling has been estimated by extrapolation to zero ‘haemoglobin’ concentration in plasma and minimum values in samples of small volume. Available results appear to be consistent with the release of small amounts of ATP in or near the peripheral circulation at the time of venepuncture. In CSF, ATP, ADP and AMP concentrations were < 0.05 μmol/1 suggesting that membrane activity in the central nervous system is not associated with non-specific leakage. The high Km variant of lymphocyte ecto-5′-nucleotidase was not associated with a significantly higher concentration of its substrate AMP in plasma. However, this enzyme may function on the lymphocyte in the thymus and spleen.

Evaluation of Kidney Viability during Hypothermic Perfusion or Cold Storage

The effect of up to 6 days of hypothermic perfusion and up to 72 h of cold storage on tissue viability were studied from measurements of sodium potassium pump activity and adenosine nucleotide content. For the hypothermic perfusion two different albumin-based perfusates were used, one with, the other without addition of caprylic acid and amino acids. No difference could be found in the parameters studied between perfusion with these two perfusates. For both perfusates total adenine nucleotide content (TAN) decreased by approximately 60% during perfusion while energy charge potential (ECP) and the ATP/ADP ratio remained constant. During cold storage a decrease of both TAN and ECP and the ratio ATP/ADP was seen, the main decrease occurring within the 1st h. The transmembrane potassium influx as deduced from potassium content in incubated cortical slices during steady state conditions decreased during 6 days of hypothermic storage to 24.5 +/- 2.4 mumol/kg which was a significantly higher level than after 24 h of cold storage. Thus, addition of extra substrate to the perfusate does not seem to improve kidney cortex viability during hypothermic perfusion. Parenchymal viability is better preserved with continuous hypothermic perfusion, compared to simple cold storage in a solution of the same ionic composition as that used for perfusion.

Possible mechanisms for reoxygenation-induced recovery of myocardial high-energy phosphates after hypoxia.

Changes in several factors responsible for high-energy phosphate production and metabolism in the heart perfused under hypoxic and subsequent reoxygenated conditions were studied using rabbit heart Langendorff preparation. A marked decline in myocardial ATP and creatine phosphate contents was observed with prolonged periods of hypoxia lasting from 15 to 60 min. Upon reoxygenation after 15 or 30 min hypoxia, creatine phosphate levels were fully recovered, whereas ATP contents were partially restored. Possible mechanisms responsible for reoxygenation-induced differential recovery of high-energy phosphate contents were investigated. Mitochondrial function for generating ATP was depressed upon hypoxia for longer than 15 min hypoxia, and the decreased function was found to be irreversible upon reoxygenation even after 15 min hypoxia. However, mitochondrial ability to generate ATP in the heart receiving 60 min hypoxia was still observed to some extent. Creatine phosphokinase activity of the myocardium exposed to hypoxic solution for 60 min showed only 19% depression. A release of creatine phosphokinase from the perfused heart was observed after more than 30 min of hypoxic perfusion or during reoxygenated perfusion after 60 min hypoxia. Changes in creatine phosphokinase activities of the myocardium and of the perfusate were not associated with those in myocardial high-energy phosphate contents. Hypoxia also induced significant release of adenine nucleotide metabolites from the perfused heart in a biphasic manner. Substrates responsible for the release of the metabolites were found to be mainly inosine and partly hypoxanthine. The metabolite release was also supported by our finding of a decrease in total adenine nucleotide contents of the myocardium upon hypoxia. The present results suggested a crucial role of hypoxia-induced release of adenine nucleotide metabolites in a differential recovery of ATP and creatine phosphate upon reoxygenation.

Effect of uridine supply on glycogen resynthesis after ischaemia in the isolated perfused rat heart.

Isolated rat hearts were subjected to 30 min low flow ischaemia (0.5 ml X min-1). During reperfusion, uridine (5 X 10(-5) mol X litre-1) was added to the perfusion medium for 30 min. The concentrations of creatine phosphate, adenine nucleotides, uridylic nucleotides and glycogen were determined at the end of the experiments. The purpose of this work was to study the effects of uridine supply on the concentration of energetic compounds during reperfusion recovery. Low flow ischaemia induced a breakdown of creatine phosphate, adenosine triphosphate, and total adenine nucleotides by 53%, 23% and 15% respectively. The creatine phosphate content was restored during reperfusion without uridine, but the adenine nucleotides remained unchanged. The uridylic nucleotides and the glycogen were also degraded during ischaemia by 56% and 53% for uridine triphosphate and glycogen respectively. Reperfusion without uridine induced a partial resynthetisation of uridylic nucleotides but glycogen stores were not significantly restored. When tested in oxygenated hearts, uridine supply induced a fall in creatine phosphate concentration and an enhancement of uridine triphosphate level but it had no effect on adenosine triphosphate, uridine diphosphate glucose or glycogen concentrations. If supplied during reperfusion, the nucleoside induced the complete restoration of myocardial ATP, total adenine nucleotide content, an increase in the uridylic nucleotide concentration and the resynthetisation of glycogen to supra-normal value.

The effects of cyanide and iodoacetate intoxication and ischaemia on enzyme release from the perfused rat heart

Isolated rat hearts perfused in the presence of iodoacetate show inhibition of glycolysis and release enzymes into the perfusate. Hearts perfused with cyanide, a mitochondrial inhibitor, show acceleration of glycolysis and no enzyme release. The adenine nucleotide content of the iodoacetate, but not the cyanide-perfused hearts was reduced. These results indicate that the membranes were permeable in the former treatment group. The adenylate energy charge and the ATP content of both the cyanide and iodoacetate treatment groups were similar but, as the extent of enzyme release was quite different, it appears that the energy state of the cell was not the prime factor controlling membrane integrity. Isolated perfused hearts were rendered ischaemic by placing a one-way ball valve in the aortic outflow tract. ATP concentration declined, as did ADP after an initial rise of short duration. AMP concentrations rose as the time of ischaemia increased. At the time at which enzyme release was first determined, the intracellular total adenine nucleotide content began to decline, suggesting that the membrane had become permeable to both small and large molecules. Glycolysis was stimulated by the hypoxia induced in the preparation and then this increase became inhibited. The point at which this inhibition was observed was also the point at which membrane permeability was evident. Taken together, the data from these experiments suggest that the energy derived from the activity of the glycolytic pathway may be important to the heart for maintenance of membrane function, particularly in ischaemia.

Citrulline synthesis: Regulation by alterations in the total mitochondrial adenine nucleotide content

The total adenine nucleotide content of rat liver mitochondria was varied in vitro over a wide range in order to investigate a possible relationship between net changes in the total matrix ATP + ADP + AMP content and the overall rate of citrulline synthesis. Isolated mitochondria were specifically depleted of matrix adenine nucleotides by incubating with inorganic pyrophosphate ( G. K. Asimakis and J. R. Aprille, 1980, Arch. Biochem. Biophys. 203, 307–316 ); alternatively, matrix adenine nucleotides were increased by incubating mitochondria with 1 m m ATP at 30 °C. No exogenous ATP or ADP was included in the subsequent incubations for the determination of citrulline synthesis. Rates varied from 0.1 to 1.6 μmol citrulline/mg protein/h as a linear function of total adenine nucleotide content in the range 2–15 nmol (ATP + ADP + AMP)/mg protein. Further increases in the matrix ATP + ADP + AMP content caused no further increase in citrulline synthesis rates. Changes in the total adenine nucleotide content were reflected in proportional changes in both the ATP and ADP content of the matrix. The ATP ADP ratio did not change significantly. Therefore, the variations in citrulline synthesis were most simply explained as the effect of different concentrations of ATP on the activity of carbamoyl-phosphate synthetase. It was concluded that net changes in the total adenine nucleotide content can contribute to the control of citrulline synthesis. These findings are significant in the context of recent evidence which shows that the matrix adenine nucleotide pool size is under hormonal control.

Transmural Gradient in High-Energy Phosphate Content in Patients with Coronary Artery Disease

In 16 patients undergoing elective coronary artery bypass, transmural biopsies were performed during bypass but before global ischemia. Subendocardial and subepicardial halves were separately assayed in each sampled tissue. Adenosine triphosphate (ATP) levels, total adenine nucleotide content (ΣAd), and creatine phosphate (CP) content were significantly higher ( p < 0.005) in the subepicardium than the subendocardium in regions of the heart distal to major occlusions: 35.36 ± 2.12 nmole/mg versus 28.7 ± 1.7 (ATP), 42.24 ± 2.04 versus 35.6 ± 1.6 (ΣAd), and 29.99 ± 4.32 versus 16.35 ± 3.48 (CP). The opposite was true in two hearts with normal coronary arteries, in which high-energy phosphates tended to be higher in the subendocardium than the subepicardium. A transmural metabolic gradient therefore exists in regions of the myocardium distal to significant coronary occlusive disease. The subendocardium's relative depression in metabolic reserve could determine its susceptibility to ischemic damage and influence techniques designed to preserve the heart during ischemia.

Large particle fluxes and the vertical transport of living carbon in the upper 1500 m of the northeast Pacific Ocean

The distribution, production, and flux of organic carbon were measured at a station in the eastern North Pacific Ocean, off Point Sur, California. Adenosine triphosphate (ATP) and total adenine nucleotide (A T) concentrations were measured to estimate the contribution of biomass (living) carbon to the total particulate organic pools. Two separate classes of particulate materials were considered: (1) ‘suspended’ or the relatively abundant, low density, fine-grained particles commonly collected in standard water bottles and (2) sediment trap particles, which represent the larger, relatively rare particles not usually sampled (in a statistical sense) when using standard water bottle collections and which require the use of in situ particle traps. The study indicates that living organisms associated with the sedimenting particles can contribute more than 100 times the biomass carbon normally associated with the ‘suspended’ fraction. The significance of the downward vertical flux of viable organisms associated with large particles may require that we alter our present concepts of the production and cycling of carbon in the open ocean.

Acute postnatal regulation of pyruvate carboxylase activity by compartmentation of mitochondrial adenine nucleotides

The total adenine nucleotide content of the mitochondrial matrix increases 3–4-fold within a few hours of birth in rat liver. This provides a mechanism for the acute postnatal regulation of pyruvate carboxylase, which is located in the matrix compartment and which requires ATP as a substrate. The sudden increase in pyruvate carboxylase activity is proposed to account for a rapid 4–5-fold increase in the gluconeogenic rate of the newborn rat.

Oscillations and control features in glycolysis: numerical analysis of a comprehensive model.

We present an analysis of glycolysis based on experimental findings and an interpretation based on concepts of efficiency, resonance response, and control features available in highly nonlinear reaction kinetics. We begin with a model for the glycolytic mechanism that is comprehensive, includes a large number of known activations and inhibitions of enzymes by metabolites, and couples the phosphofructokinase (PFKase) and the pyruvate kinase (PKase) reactions. The PFKase and PKase reactions and the coupling between them are modeled according to experimental information, but we do not attempt to model the glyceraldehyde-3-phosphate dehydrogenase-3-phosphoglycerate kinase reaction. We use experimental data to obtain the best estimates for the kinetic parameters and test the model by calculating the concentration variations of the intermediate metabolites. We confirm oscillatory behavior and calculate the ATP/ADP ratio and the free-energy dissipation for an extended range of the kinetic parameters as a function of the driving force for the glycolytic pathway, a measure of which is the total adenine nucleotide concentration. We find agreement of the calculated results with experimental findings except for the insufficiently represented reactions. Our model shows that the average ATP/ADP ratio is increased and the average free-energy dissipation is decreased in an oscillatory compared with a steady state mode of operation. The average values of the ATP/ADP ratio and of the free energy dissipation change abruptly past the onset of sustained oscillations.

218 ACUTE POSTNATAL REGULATION OF PYRUVATE CARBOXYLASE ACTIVITY BY COMPARTMENTATION OF ADENINE NUCLEOTIDES

In the term rat liver, crossover studies reveal that pyruvate carboxylase (PC) is rate-limiting for gluconeogenesis. Within 1 hr of birth, the rate of gluconeogenesis increases 4 fold, even though there is no net synthesis of PC. (Biochem. J. 124:265(1971) This suggests that PC is inactive at birth, and becomes active within a short time. PC is localized in the mitochondrial compartment, and requires ATP as a substrate. Since the total adenine nucleotide content (ATP+ADP+AMP) of the matrix increases right after birth (Archiv. Biochem. Biophys. 201:564(1980), we considered whether the resulting increase in matrix ATP concentration might cause an increase in PC activity. We found that PC activity measured in intact isolated mitochondria increased ∼ 3 fold within a few hours of birth, in parallel to a ∼ 3 fold increase in the total matrix adenine nucleotide content. The increase in PC activity could be brought about in vitro by incubating isolated newborn mitochondria with exogenous ATP under conditions shown to promote the net accumulation of adenine nucleotides. Conversely, adult mitochondria specifically depleted of matrix adenine nucleotides had proportionately lower PC activity. We concluded that acute regulation of PC can occur by compartmentation of adenine nucleotides. This mechanism probably provides for the rapid 4-5 fold increase in gluconeogenesis which occurs within 1-2 hrs of birth, without the need for new enzyme synthesis. [Supported by NIH HD11697 and NS 14936.]

The ATP and Total Adenine Nucleotide Content of Four Unicellular and Colonial Green Algae

The ATP and total adenine nucleotide content (TANC) of two unicellular and two colonial green algae were measured in conditions of nitrogen and phosphorus deficiency and during exponential growth. The ATP content ranged from 0.07 to 0.44% (mean = 0.25%) of the cellular organic carbon during exponential growth and the TANC varied from 0.28 to 0.79% (mean = 0.53%). ATP and TANC decreased during nitrogen and phosphorus deficiency to values between 2 and 31% of the content in exponentially growing cells. Large errors are expected when cellular organic carbon is calculated from ATP data in natural environments dominated by green algae, whereas TANC gives a better estimate of biomass.

Effects of lactic acid accumulation and ATP decrease on muscle tension and relaxation.

The present study was undertaken to evaluate the effect of lactic acid accumulation on peak tension and relaxation rate of the isometric twitch. Isolated extensor digitorum longus muscle from rat was stimulated electrically at a frequency of 2/s under anaerobic conditions. Comparison was made with muscles in which glycolysis was blocked with iodoacetic acid (IAA). Stimulation of unpoisoned muscles for 3 min decreased tension to 50% and increased relaxation time to 250% of the initial value. Lactate increased 15-fold, and muscle pH decreased from 7.10 to 6.76. Stimulation of IAA-poisoned muscles for 1 min decreased tension to 50% but did not increase the relaxation time. Stimulation of IAA-poisoned muscle resulted in a pronounced decrease (about 50%) of the ATP and total adenine nucleotide content in muscle, whereas only a small decrease (10-15%) occurred in unpoisoned muscle. The main findings in the present study were that tension decline in unpoisoned muscle is closely related to decrease in muscle pH and increase in ADP but not to ATP content per se and that slowing of relaxation rate is closely related to decrease in muscle pH but not to muscle content of ATP or creatine phosphate.