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C Arús

Publications and source records attributed to C Arús.

At least 37 records · Page 2Linked to original sources
PubMed1993

A versatile perifusion system for the NMR spectroscopy of bovine retina. Assignment of resonances and effect of ischemia.

A perifusion system has been designed for metabolic nuclear magnetic resonance (NMR) spectroscopy studies of the bovine retina. This system, which allows the use of unmodified commercial probes for multinuclear studies, is shown here to maintain the perifused retina in a metabolic steady state at 25 degrees C, as deduced from its 31P and 1H NMR spectra. The 31P NMR ex vivo spectrum of the retina has been characterized, and its major resonances have been assigned in tissue extracts. We have found that excised retinal tissue subjected to 2 hr of ischemia is capable, upon perifusion with oxygenated medium, of recovering its resting spectral pattern. Thus, this system offers a versatile model for studies of physiological events related with ischemia in a neural tissue, without associated systemic effects.

Animals↗
PubMed1993

31P-MRS of quadriceps reveals quantitative differences between sprinters and long-distance runners.

Phosphorus-31 nuclear magnetic resonance spectroscopy (31P-MRS) has been used to characterize the spectral pattern of quadriceps muscle at rest for sedentaries (N = 10), sprinters (N = 12), and long-distance runners (N = 10). Intracellular pH (pHi), phosphocreatine (PCr), inorganic phosphate (Pi), phosphodiesters (PDE), adenosine triphosphate (ATP) and total phosphate (Pt), defined as the sum of the areas of PCr, PDE, Pi, and 3 times the beta-ATP resonances, were obtained from all spectra. The pHi and ATP/Pt ratio were the same in the three groups of people studied. The PCr/Pi and PCr/Pt ratios were significantly lower and the Pi/Pt ratio was significantly higher for long-distance runners than for sprinters and sedentaries, while the PDE/Pt ratio was significantly lower for sprinters than for sedentaries and long-distance runners. Furthermore, the PCr concentration for sprinters and sedentaries was significantly higher and the Pi was significantly lower than for long-distance runners. The results obtained in this study can be explained by the accepted differences in fiber type composition between sprint and endurance athletes. We suggest that 31P-MRS at rest could be used to monitor noninvasively the individual adaptive response to training in the metabolic characteristics of the athlete muscle fiber.

Adaptation, Physiological↗
PubMed1992

Effect of training on the calf muscle energy metabolism. A 31P-NMR study on four elite downhill skiers challenged with a standardized exercise protocol.

This study evaluates the effects of a 6 months' training period on the bioenergetics of the calf muscle of elite athletes by means of phosphorus-31 Nuclear Magnetic Resonance Spectroscopy (31P-NMR). Four downhill skiers, belonging to the Spanish National Team, performed a standardized exercise protocol using their right leg inside a wide-bore 2.35 Tesla magnet. The inorganic phosphate/phosphocreatine (Pi/PC) ratio and intracellular pH (pHi) were measured at steady-state during an exercise protocol composed of 5 work levels between 20% and 80% of the maximum voluntary contraction (MVC), before and after the training period. The measured values, which were markedly scattered at the beginning, regrouped after training. This was caused by a shift towards lower Pi/PC ratios and by a lower pHi acidification in three of the four subjects. This result suggests that 31P-NMR is a good tool to evaluate changes in the muscle aerobic capacity of athletes induced by training.

Adenosine Diphosphate↗
PubMed1990

Two-dimensional spectra of intact tissue: homonuclear Hartmann-Hahn spectroscopy provides increased sensitivity and information content as compared to COSY.

Hartmann-Hahn (HOHAHA) spectroscopy led to good definition of proton spin systems of metabolites such as lactate, (phospho) creatine, and carnosine from a 3-g sample of excised frog gastrocnemius muscle in a 17-min experiment. The sign of the HOHAHA cross-peak intensity appears to provide additional information about interactions between metabolites and their protein binding sites.

Animals↗
PubMed1989

Two-dimensional correlated spectroscopy (COSY) of intact frog muscle: spectral pattern characterization and lactate quantitation.

Two-dimensional scalar-correlated 1H spectra (COSY) were recorded from a model solution containing varying amounts of lactate and a fixed concentration of carnosine as the reference compound. The integrated volume of the lactate methyl to methine cross peak in the magnitude-calculated spectra showed a linear increase with respect to lactate concentration. Accordingly, the COSY cross-peak integral volume should be a useful parameter for the quantitative study of metabolism in intact tissue. We have tested this assumption in excised gastrocnemius frog muscle. Lactate, carnosine, and creatine/phosphocreatine cross peaks have been assigned in the spectrum of summer frog muscle. The change in concentration of lactate and phosphorylated compounds upon anaerobic storage of excised frog muscle has been followed by interleaving one-dimensional (1D) 1H and 31P spectra with 2D homonuclear 1H COSY spectra. The lactate concentration in the excised muscle derived from COSY agreed well with that determined from muscle extracts. Qualitative changes in the COSY pattern of the excised muscles under caffeine-induced contracture were detected. Furthermore, seasonal variations were observed in the COSY spectral pattern: winter frog muscle was found to differ from summer frog muscle by the appearance of cross peaks from phosphodiesters and mobile fatty acids.

Animals↗
PubMed1989

Chemical and computer graphics studies on the topography of the ribonuclease A active site cleft. A model of the enzyme-pentanucleotide substrate complex.

The affinity labelling of bovine pancreatic ribonuclease A with 6-chloropurine 5' ribonucleotide allowed us to postulate the existence of a new phosphate-binding subsite, P2, adjacent to the main purine-binding subsite. The study of this reaction in greater detail together with the study of a complex of the enzyme with the pentanucleotide pApUpApApG by means of model building and computer graphics indicate that at least five phosphate groups of the RNA molecule can interact with five positive regions of the enzyme. In each one a lysine residue is present: Lys-104, -66, -41, -7 and -37 appear sequentially in the 5'----3' direction. The distance between each lysine is 0.7-0.8 nm, the same distance as that found between the phosphate groups on the RNA molecule. The study also enabled many amino acid residues of the enzyme to be described as forming part of, or being near, the different binding subsites.

Binding Sites↗
PubMed1986

Observation of the terminal methyl group in fatty acids of the linolenic series by a new 1H NMR pulse sequence providing spectral editing and solvent suppression. Application to excised frog muscle and rat brain.

A new 1H NMR pulse sequence is described that combines water suppression with the selective observation of signals from coupled spin systems. The pulse sequence is easy to set up and compensates for pulse width inhomogeneity in the biological sample. Suppression of the water signal is achieved by pulses that return the water spins to their equilibrium position; spectral editing is based on the J modulation present in spin-echo spectra and its inhibition by coherent decoupling at one of the resonances of the spin system of interest. The pulse sequence, which was designed for 1H NMR spectroscopy of tissue, was tested at 470 MHz on excised frog muscle and rat brain. The lactate methyl resonance of caffeine-treated frog sartorius muscle was observed selectively by irradiation at the position of its alcoholic proton. The terminal methyl signal of linolenic acid, along with other fatty acids of the linolenic series (first double bond in the omega-3 position), was observed selectively by irradiation at the position of its omega-1 methylene group. 1H NMR spectra of rat brain were edited to reveal the terminal methyl of either linolenic series or all other fatty acids. The results suggest that the terminal methyl groups of fatty acids of the linolenic series (mostly docosahexaenoic acid, 22:6) have higher mobility than those of all other fatty acids.

Animals↗
PubMed1986

Application of high-field 1H-NMR spectroscopy for the study of perifused amphibian and excised mammalian muscles.

Frog sartorius and gastrocnemius muscles were perifused at 20 degrees C, the intracellular pH (pHi) and the concentration of phosphocreatine were determined in the resting muscle by 1H-NMR spectroscopy at 470 MHz; values of pHi = 7.31 +/- 0.05 (n = 7) and concentration of phosphocreatine = 20.4 +/- 1.1 mumol/g wet wt. (n = 6) were found. The hydrolysis of phosphocreatine and the simultaneous increase in lactate upon perifusion with 10 mM caffeine (in Ringer's solution) was followed with a time resolution of 1 min. Lactate increased at a rate of 1.0 mumol/g per min, but no pHi change was recorded during the time monitored. The lower limit for the buffering capacity of the muscle cytosol was estimated to be 16.7 mumol/g muscle per pH unit from the uncertainty in pHi determination (+/- 0.03 pH units) and from the amount of lactate produced and phosphocreatine hydrolyzed. Changes in pHi, lactate concentration and fatty acyl chain intensity were monitored by 1H-NMR spectroscopy at 361 MHz in ischemic rat skeletal muscle, excised and stored at 20 degrees C. The resonances in the 1H-NMR spectrum of a human skeletal muscle perchloric acid extract are reported and tentatively assigned.

Animals↗
PubMed1986

Two-dimensional proton magnetic resonance of human muscle extracts.

In continuation of our previous work on one-dimensional (1D) proton nuclear magnetic resonance (1H-NMR) of normal and diseased human muscle extracts we recorded the two-dimensional (2D) J-correlated proton magnetic resonance spectra of these extracts. Significant differences between normal and diseased muscle extracts, not observed in the 1D 1H-NMR spectra, were seen from their 2D connectivity contour patterns. Taurine was not present in cerebral palsy muscle extract while both normal and scoliosis muscles contained this metabolite. Only the normal muscle had carnitine. Carnosine was present in all muscles. alpha-Ketoglutarate was found only in the diseased muscle extracts. While the amino acids lysine, cysteine and glutamine were common to normal and diseased muscles, threonine was seen only in the diseased muscles. Additional small differences were detected in the 2D patterns of human muscle extracts.

Cerebral Palsy↗
PubMed1985

Effect of halothane on the natural-abundance 13C NMR spectra of excised rat brain.

Halothane increases the intensity of the 30.5- and 129-ppm resonances in 13C nuclear magnetic resonance spectra of excised rat brain, of phospholipid vesicles prepared from chloroform-methanol extract of rat brain, and of brain excised from rats anesthetized with halothane. The 13C spin-lattice relaxation times of the 30.5- and 129-ppm resonances are increased in excised brain, or phospholipid vesicles, upon addition of halothane, and they are also increased in brain excised from rats anesthetized with halothane. Excised brain and its membrane-rich subcellular fractions interact with [14C]halothane reversibly. The interaction is virtually abolished when the phospholipids are extracted from the brain. The [14C]halothane content of the brain membranes is correlated with the halothane-induced increase in the integral of the 129-ppm resonance. From this correlation and from the phospholipid content of the membranes, a halothane concentration of 3.34 mM and a partition of 0.057 mol of halothane/mol of phospholipid may be calculated in the brain of anesthetized rats.

Animals↗
PubMed1985

The separation of phosphocreatine from creatine, and pH determination in frog muscle by natural abundance 13C-NMR.

Phosphocreatine can be separated from creatine in superfused frog muscle by natural abundance 13C-NMR, based on the difference in resonance frequency of their guanidino carbons. After taking into account the longitudinal relaxation times and nuclear Overhauser enhancement factors, the integrated peak areas of the guanidino carbons could be used for determination of the phosphocreatine-to-creatine ratio in the muscle. The pH dependence of the chemical shift of the C-2 carbon in the histidine ring of carnosine was used for estimation of the intracellular pH in the intact muscle.

Animals↗
PubMed1985

Natural-abundance 13C NMR of brain.

In natural-abundance 13C NMR spectrum of excised rat brain 55 resonances were resolved. The chemical shifts of most of the resonances were well correlated with those of pure brain metabolites, determined under identical experimental conditions. These resonances were also found in the cytosol fraction and perchloric acid extract of the brain. Some brain resonances were not observed in the perchloric acid extract but only in the microsomes or cytosol.

Animals↗
PubMed1985

Proton nuclear magnetic resonance spectra of excised rat brain. Assignment of resonances.

We have recorded 1H NMR spectra of excised rat brain at 361 MHz using two different water suppression pulse sequences. The assignment of the resonances has been carried out in perchloric acid extracts and subcellular fractions. Our results show that cytosolic proteins, membrane phospholipids and 16 different metabolites contribute to the observed spectra. The new resonances assigned allow the direct observation of myo-inositol and urea. Moreover, changes in the spectral pattern upon anesthesia, ischemic exposure of the brain and age of the rat have been recorded and correlated with the compounds producing the spectra.

Aging↗
PubMed1985

Characterization of the broad resonance in 31P NMR spectra of excised rat brain.

31P NMR spectra of excised rat brain showed a broad resonance between-12 and -13 ppm. Subcellular fractions of brain, rich in membranes, exhibited the broad resonance and it was also present in isolated myelin, the major membrane component of brain. However, it was absent in brain cytosol (161,100 X g supernatant). Raising the temperature of the brain above 50 degrees C caused a gradual downfield chemical shift of the broad resonance, to about -1 ppm at 90 degrees C. An even larger downfield shift was produced by halothane or deoxycholate with concomitant narrowing of the line width of this resonance. Vesicles prepared from the phospholipids of excised brain or isolated myelin showed the broad resonance, and halothane produced the same downfield shift and peak sharpening in brain phospholipid vesicles as that in the intact brain. The chemical shift anisotropy was estimated to be 45 ppm for both myelin and the brain, as characteristic for biological membranes. The T1 and T2 relaxation times of the perpendicular 31P chemical shift tensor component of the broad resonance were 0.66 sec and 1.6 msec, respectively, in the same range as those for other biological membranes. Halothane-treatment of the brain increased both the T1 and T2 times considerably, as expected from the disruption of the phospholipid bilayer in a membrane. These data indicate that the broad resonance in the 31P NMR spectrum of excised rat brain originates exclusively from the phosphate head group of membrane bound phospholipids. Similar broad resonances were found in autopsied human brain and porcine spinal cord and to a lesser extent in excised rat liver and kidney.

Animals↗
PubMed1984

1H NMR of intact muscle at 11 T.

1H NMR spectra of intact frog, and chicken skeletal muscles, were recorded at 470 MHz with the Plateau and Guéron pulse sequence for the suppression of water [(1982) J. Am. Chem. Soc. 104, 7310]. Only a few transients were required to resolve the resonances from the protons of muscle metabolites. The previously unobserved exchangeable protons of muscles were also recorded and thereby phosphocreatine and creatine could be measured simultaneously. During aging of dissected frog muscle, changes in levels of phosphocreatine, creatine and lactic acid, and the decrease in the intracellular pH were followed by 1H NMR.

Animals↗
PubMed1984

Proton nuclear magnetic resonance of human muscle extracts.

High resolution proton nuclear magnetic resonance (NMR) spectra of normal and diseased human muscle extracts were recorded at 470 MHz. Resonances from lactic acid, creatine, glucose, ribose, purine and pyrimidine bases were identified. The longitudinal relaxation times of these resonances were determined to allow quantitation of muscle metabolites. With aid of a standardized reference capillary, inserted into the NMR tube containing the muscle extracts, the lactic acid and total creatine content of the extracts was determined. After 5 h of incubation at 37 degrees C, normal muscles contained on average 103 mumol lactic acid and 36 mumol creatine/173 mg of noncollagenous protein, equivalent to 1.0 g of fresh muscle. The lactic acid and creatine contents decreased slightly in scoliosis and idiopathic scoliosis and they decreased significantly in cerebral palsy. In an extract of a patient whose illness was diagnosed as 'scoliosis' no creatine was present, and in an extract of a patient with unknown diagnosis the creatine content was reduced to 2 mumol/173 mg of noncollagenous protein. The short time (1.7 sec to 6.5 min) and the small amount of tissue (300 mg) needed for an analysis add to the potential of proton NMR as a new technique for the characterization of muscular diseases.

Cerebral Palsy↗
PubMed1982

Evidence on the existence of a purine ligand induced conformational change in the active site of bovine pancreatic ribonuclease A studied by proton nuclear magnetic resonance spectroscopy.

The titration curves of the C-2 histidine protons of RNase A and of derivative II--a covalent derivative obtained by reaction of the enzyme with the halogenated nucleotide 9-beta-D-ribofuranosyl-6-chloropurine 5'-phosphate--in the presence of a number of purine nucleosides, nucleoside monophosphates, and nucleoside diphosphates were studied by means of proton nuclear magnetic resonance at 270 MHz. The examination of the perturbations found on the chemical shifts and pKs of the C-2 protons of His-12, -48, and -119 are consistent with the following conclusions: (1) The interaction of adenosine in the primary purine binding site of the enzyme (B2R2) induces a conformational change in the active center of the enzyme [for the nomenclature of the RNase A binding subsites, see Parés et al. [Parés, X., Llorens, R., Arús, C., & Cuchillo, C. M. (1980) Eur. J. Biochem. 105, 571-579]]. (2) The phosphate moiety of the ligands, independently of its position, probably acts as a general carrier of the nucleotide to the active center, while the substituents of the base are the generators of the specificity of the binding and control the binding equilibrium between subsites B2R2 and B1R1. (3) There is no overlapping between the binding sites occupied by the labeling nucleotide in derivative II (B3R3p2) and the primary binding site for purine mononucleotides (B2R2p1).

Adenosine↗