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Biomedical subjects

H Watari

Publications and source records attributed to H Watari.

At least 55 records · Page 3Linked to original sources

1H-NMR studies on water structures in the rat brain tissues with cerebral tumour or ischaemia.

Spin-lattice relaxation times (T1) of water protons and cross-relaxation times (T(IS)) between irradiated protein protons and observed water protons were measured to study water structure in various rat brain tissue. A tumour-bearing rat model and an incomplete forebrain ischaemia model of stroke prone spontaneously hypertensive rats (SHRSP) were used for the experiments. T(IS) and T1 were measured by the inversion recovery FT method with and without f2-irradiation at 7.13 p.p.m. and gamma H2/2 pi approximately 69 Hz, respectively, using a 360 MHz FT-NMR spectrometer. All of experiments on T1 and T(IS) yielded a single kind of component. In spite of small changes in T1 and water content, T(IS) values in peritumoural oedematous and ischaemic brain tissue were significantly shorter than those of normal brain and far shorter than those of brain tumour, indicating a large amount of hydration of marco-molecules in the oedematous and/or ischaemic rat brain tissue. T(IS) might be sensitive parameter for studying the water structure in various brain tissue.

Animals

Leakage of cytoplasmic enzymes from rat heart by the stress of cardiac beating after increase in cell membrane fragility by anoxia.

The effects of spontaneous beating after anoxia and the pumping stress induced by a left ventricular balloon on the leakage of myocardial enzymes from the isolated perfused rat heart were investigated. Beating of the heart was arrested by perfusion with high-K+ medium. When the beating was arrested during reoxygenation after anoxia, the leakage of lactate dehydrogenase (LDH) was significantly lower than during reoxygenation with spontaneous cardiac beating. After changing from K+ arrest to spontaneous beating by perfusion with low-K+ medium during reoxygenation, the leakage of LDH increased markedly. Imposition of left ventricular wall stress on the K(+)-arrested heart by repetitive passive distension during aerobic perfusion and after 20 min and 60 min of anoxia caused LDH leakages of 1.0, 4.6 and 21.0 units/g in 30 min, respectively. Under this mechanical stress, the release of LDH as a percentage of its total myocardial activity coincided well with that of cytoplasmic aspartate aminotransferase (AST), while the percentage release of mitochondrial AST was much less. These results appeared to indicate that the leakage of cytoplasmic enzymes during reoxygenation is accelerated by cardiac beating because of fragility of the cell membranes developing during the preceding anoxia.

Adenosine Triphosphate

Energetic recovery from hypothermic preservation in the rat liver.

Changes in energy metabolism induced by norepinephrine were observed in liver perfused immediately after dissection and in liver stored in Euro-Collins solution at 0 degrees C for 24 hr. Oxygen consumption, glucose output, ATP content, and tissue pH were measured in isolated perfused rat liver at 37 degrees C. In fresh liver, a two-fold increase in glucose output and oxygen consumption was induced by norepinephrine (1 microM), while changes in ATP content and tissue pH were minimal. In stored liver, the cumulative increments in glucose output and oxygen consumption induced by norepinephrine were 30 and 27% those of fresh liver, respectively. ATP content of the unstimulated liver was 76% that of the fresh liver, then decreased to 50% during stimulation. A transient decrease in tissue pH (0.14 pH unit) was significant compared with that of fresh liver. These results suggest that norepinephrine stimulation is useful for assessing energetic and functional recovery of reperfused liver following hypothermic storage.

Adenosine Triphosphate

Adenosine triphosphate compartmentation in the rat heart: a 31P spin-lattice relaxation study.

Longitudinal relaxation times (T1) of phosphorus compounds in the perfused rat heart and erythrocytes were measured using the 31P Driven-Equilibrium Single-Pulse Observation of T1 relaxation (DESPOT) method at 33 degrees C. Both creatine phosphate in the heart and the three phosphate groups of adenosine triphosphate (ATP) in erythrocytes showed single-exponential relaxation. The three phosphate groups of ATP in the heart, however, had two T1 components. The T1 values of the short and the long T1 components of the beta-phosphate of ATP were ca. 0.4 and 14 s, respectively. The fraction with the long T1 represented ca. 30% of the total ATP content. These results suggested that there were two major pools of intracellular ATP in the rat heart which could be determined by 31P NMR spectroscopy.

Adenosine Triphosphate

Structural transition of bovine plasma albumin in the alkaline region--the N-B transition.

Bovine plasma albumin (BPA) has approximately one SH group (Cys-34) which catalyzes the intramolecular SH, S-S exchange reaction in the alkaline region at low ionic strength, resulting in the formation of the aged form. So, the N-B transition at ionic strength above 0.20 and below 0.10 was studied using BPA and iodoacetamide-blocked BPA (IA-BPA), respectively. (1) pH profiles of [theta]262 and [theta]268 of BPA in 0.20 M KCl showed the characteristic changes in the pH region 7.0-9.0, corresponding to the N-B transition. On going from pH 7.0 to 9.0 in 0.10 M KCl or NaCl, IA-BPA did not show significant changes in rotational relaxation times of tryptophyl fluorophors, CD-resolved secondary structures, spin-echo 1H-n.m.r. spectra and cross-relaxation times (TIS) between irradiated and observed protein protons, which might reflect the rigidity of the domains and/or subdomains. On the other hand, rotational relaxation times of 1-anilino-8-naphthalenesulfonate-IA-BPA complex (IA-BPA-ANS0.9, molar ratio of ANS to IA-BPA = 0.9/1) showed significant decreases from 131 to 114 ns on going from the N- to the B-forms in 0.10 M KCl. The above results and reported experimental evidence might indicate that on going from the N- to the B-forms in 0.10 M KCl or NaCl, the mutual movement of subdomains, connected with a flexible hinge region (Brown & Shockley (1982)) might increase without loss in the helicity and the rigidity of subdomains. (2) The N-B transition of IA-BPA in the absence of salt was quite different from those in 0.10 M KCl or NaCl. Decreases in the helicity and the intramolecular rigidity, as monitored by TIS-measurements, were observed on going from the N- to the B-forms.

Animals

Potassium-39 nuclear magnetic resonance observation of intracellular potassium without chemical shift reagents during metabolic inhibition in the isolated perfused rat heart.

The intracellular potassium content of perfused rat heart was measured by potassium-39 nuclear magnetic resonance (NMR) spectroscopy at 33 degrees C with an inversion recovery technique based on the fact that the spin-lattice relaxation time (T1) of the intracellular potassium (8.3 msec at 8.45 T) is much faster than that of the extracellular potassium (68 msec). Intracellular potassium decreased to 60.2 +/- 4.3% of the control level (mean +/- SEM, n = 6) at 40 minutes from the start of metabolic inhibition (2 mM cyanide, 0 mM glucose). Removal of cyanide restored intracellular potassium to 94.2 +/- 3.9% at 30 minutes from the restart of oxidative metabolism. The cumulative potassium loss was determined from the flow rate and potassium concentration of the coronary effluent, which reached 139 +/- 12 mumol/g dry wt during 40 minutes of metabolic inhibition. This value was calculated as 41.8% of intracellular potassium in the control heart and agreed with the decrement of intracellular potassium measured by NMR. During the metabolic inhibition and recovery period, a linear correlation was observed between the changes in 39K NMR-observed intracellular potassium and the cumulative potassium loss. The present results evaluate the inversion recovery technique as a method to successfully monitor the myocardial intracellular potassium.

Animals

[Partial volume effect in MRI--a phantom study].

According to the direction and the thickness of the imaging slice in tomography, the border between the tissues becomes unclear (partial volume effect). In the present MRI experiment, we examined border area between fat and water components using phantom in order to investigate the partial volume effect in MRI. In spin echo sequences, the intensity of the border area showed a linear relationship with composition of fat and water. Whereas, in inversion recovery and field echo sequences, we found the parameters to produce an extremely low intensity area at the border region between fat and water. This low intensity area was explained by cancellation of NMR signals from fat and water due to the difference in the direction of magnetic vectors. Clinically, partial volume effect can cause of mis-evaluation of walls, small nodules, tumor capsules and the tumor invasion in the use of inversion recovery and field echo sequences.

Magnetic Resonance Imaging

Vector electron paramagnetic resonance spectroscopy with first and second harmonic displays of ferrihemoglobin.

Superimposed plots of electron paramagnetic resonance spectra with the first and second harmonic displays of ferrihemoglobin at pH 9.1 and 90 K were measured at 20 degree intervals of phase angle using a phase-sensitive detector. The high spin signal in the g = 6 region was observed in both displays, and a small splitting of the signal was found in the calculated amplitude spectrum of the second harmonic display, with g values of 5.95 and 6.05. Low spin signals were observed at g = 2.55, 2.25 and 1.82 in both harmonic displays. A signal in the g = 2.05 region was observed only in the second harmonic display. The signal is probably associated with the low spin spectrum; however, its origin is obscure.

Electron Spin Resonance Spectroscopy

[Significance of brain renin for pathogenesis of hypertension: effect of antihypertensive drugs on active and inactive renins in the brain of spontaneously hypertensive rat].

Relationship between blood pressure and brain renin was studied in four groups of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY); as controls (n = 5), administered captopril (n = 5), trichlormethiazide (n = 5) and atenolol (n = 5). 1) Inactive renin in the hypothalamus of captopril-administered SHR was significantly lower than that of control SHR and captopril-administered WKY. On the other hand, active renin in the hypothalamus, thalamus and striatum of captopril-administered SHR was significantly lower than that of control SHR and captopril-administered WKY. 2) Inactive renin in the hypothalamus of trichlormethiazide administered SHR was significantly lower than that of control SHR and trichlormethiazide-administered WKY. On the other hand, active renin in the hypothalamus, thalamus and midbrain of trichlormethiazide-administered SHR was significantly lower than that of control SHR and trichlormethiazide-administered WKY. 3) Inactive renin in the hypothalamus of atenolol-administered SHR was significantly lower than that of control SHR and atenolol-administered WKY. On the other hand, active renin in the hypothalamus, thalamus and midbrain of atenolol-administered SHR was significantly lower than that of control SHR and atenolol-administered WKY. These results suggest that the production and/or activation of renin in the hypothalamus, thalamus, midbrain and striatum play an important role in the initiation and/or development of hypertension of SHR by the local generation of angiotensin II.

Animals

Direct measurement of K movement by 39K NMR in perfused rat mandibular salivary gland stimulated with acetylcholine.

The intracellular K content (Kin) of the isolated perfused rat mandibular gland was measured by 39K NMR spectroscopy at 25 degrees C, using an inversion recovery technique based on the fact that the spin-lattice (T1) relaxation of Kin is much faster than that of the extracellular K. Kin decreased by 30-34% of the resting level and reached a plateau level during secretion evoked by a sustained infusion of 1 mumol/l acetylcholine. Addition of 1 mmol/l ouabain decreased Kin by an additional 41% of the resting level. The K net flux to the blood and saliva was calculated from the K concentrations and flow rates of the effluent and the saliva. At an initial stage of secretion the gland lost K to the vascular side at a rate of 12.6 +/- 1.8 mumol/g-min (mean +/- SEM, n = 7). During sustained secretion, the gland took K up from the vascular side at a rate of 3.3 +/- 0.7 mumol/g-min (n = 7), and the same amount of K was secreted into the saliva (4.7 +/- 1.1 at 5-10 min, 2.8 +/- 0.8 mumol/g-min at 20-30 min), resulting in no net K movement from the gland. Addition of 1 mmol/l ouabain stopped salivary secretion and caused a transient K release to the vascular side at a maximum rate of 12.8 +/- 1.1 mumol/g-min. Withdrawal of acetylcholine and ouabain induced K uptake from the vascular side (6.5 +/- 0.7 mumol/g-min) and the amount of K released was completely restored when Kin recovered completely.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine

An experimental approach to 31P spin-lattice relaxation time measurement in biological systems.

The accuracy of measurement of the spin-lattice relaxation time (T1) in biological systems using the variable nutation angle method is discussed using computer simulations. This method could reduce the time required for measurement to 10% of that using the inversion recovery method. In the perfused salivary gland of the rat (310 K), the T1 values of sugar phosphates, creatine phosphate, gamma-phosphate, and beta-phosphate of nucleotide triphosphates determined by this method were 5.2, 4.8, 0.9, and 0.8 s, respectively. During secretion of saliva, the T1 value of sugar phosphates decreased to 3.7 s, which could be due to an increased chemical exchange of phosphate in glycolysis.

Animals

Internal motion of a tryptophan residue in Streptomyces subtilisin inhibitor: deuterium nuclear magnetic resonance in solution.

Deuterium NMR spectroscopy was used to study internal motions of a deuterium-labeled single tryptophan (Trp) residue (per subunit) of Streptomyces subtilisin inhibitor (SSI) in solution. The free inhibitor with the five ring protons of the Trp replaced with deuterons showed a narrow resonance component (56 Hz) of about one-quarter of the total intensity, in addition to the broad resonance component (about 600 Hz) at 25 degrees C, showing that it exits in an equilibrium mixture of two conformers, in one of which the tryptophan side chain is highly mobile. In analogy to the two structures of SSI found in the crystal, these two conformers were attributed to the one in which the contact between the alpha-lobe and the beta-lobe of the subunit is tight and the other in which the same contact is loose. When SSI forms a complex with subtilisin BPN', the broad component becomes invisibly broad, but the narrow component increases with even further narrowing, suggesting that the binding to the enzyme favors the "loose" conformer over the "tight" conformer.

Bacterial Proteins

NMR-invisible ATP in rat heart and its change in ischemia.

The subcellular compartmentalization of adenosine 5'-triphosphate (ATP) in isolated perfused rat heart and its relation to energy depletion in ischemia were examined by 31P nuclear magnetic resonance (31P-NMR) spectroscopy and chemical analyses. The signal intensities of the beta-phosphate of ATP and creatine phosphate in the 31P-NMR were standardized by the intracellular volume ratio measured with 23Na-NMR to determine the actual content of each. During aerobic perfusion the ATP content determined by NMR (13.7 +/- 2.2 mumol/g dry weight) was significantly lower than that found by chemical analysis (22.4 +/- 0.7 mumol/g dry weight), while the creatine phosphate contents determined by the two methods were the same. During ischemia at 33 degrees C, the signal of the beta-phosphate of ATP in the 31P-NMR spectrum decreased progressively, disappearing completely after 16 min. But at this time 5.7 +/- 1.7 mumol/g dry weight of myocardial ATP was still detected by chemical analysis. These results indicated that there were two different compartments of intracellular ATP in the heart, only one of which is detectable by 31P-NMR spectroscopy, and that during ischemia the ATP that is detectable, which seems to be the free ATP in the cytosol, decreased more rapidly than the ATP in the other compartment.

Adenosine Triphosphate

Phosphorous compounds studied by 31P nuclear magnetic resonance spectroscopy in the taenia of guinea-pig caecum.

1. In the isolated taenia (0.4-0.6 g) of guinea-pig caecum, the intracellular phosphorous compounds and pH were investigated using 31P nuclear magnetic resonance (NMR) under various metabolic conditions. 2. The ratios of the intracellular concentration of phosphocreatine ([ PCr]) and inorganic phosphate [( Pi]) to nucleotide triphosphate ([ NTP]) were 1.71 +/- 0.14 and 0.58 +/- 0.11 (n = 25), respectively, in normal solution (32 degrees C). The intracellular pH estimated from the chemical shift of Pi was 7.05 +/- 0.06 (n = 25), agreeing well with those previously obtained. 3. In the absence of glucose, the [PCr] and [NTP] were decreased to almost a half after 150 min exposure to 40 mM-K+ solution, while [Pi] was increased 3-fold. These changes were much faster than the rate of decline in tension. When glucose was readmitted, the contractile response to K+ fully recovered in 50 min. However, this was accompanied with only a partial recovery of [PCr] and [Pi], but no recovery of [NTP]. The intracellular pH was lowered by about 0.2 of a unit, suggesting an increase in glycolysis. 4. In Ca2+-free solution, respiratory inhibition with hypoxia or CN (1 mM) only decreased [PCr], leaving [NTP] nearly unchanged. On the other hand, respiratory inhibition in excess-K+ solution containing Ca2+ (2.4 mM) severely depleted PCr and decreased [NTP] to 40%. Increasing glucose to 50 mM did not prevent these changes, although it increased tension development. 5. The simultaneous decrease of [NTP] and [PCr] during K+ contracture suggests that the activity of creatine phosphokinase is low. The recovery from respiratory inhibition was much better for [PCr] than for [NTP]. Slow, but perfect, recovery of all NTP peaks was produced by adding 1 mM-adenosine to normal solution. 6. It was suggested that tension development is closely related to the turnover rate of ATP, and not to its concentration, and that deamination of adenosine is a limiting factor in the recovery of ATP after excessive consumption.

Adenosine

Phosphorus nuclear magnetic resonance in isolated perfused rat pancreas.

Phosphorus nuclear magnetic resonance spectroscopy was applied to measure phosphorus energy metabolites in isolated perfused rat pancreas. The gland was perfused with a modified Krebs-Henseleit solution at room temperature (25 degrees C). 31P resonances of creatine phosphate (PCr), ATP, ADP, inorganic phosphate (Pi), and phosphomonoesters (PMEs) were observed in all the preparations of pancreas. In different individual preparations, the resonance of PCr varied, but those of ATP were almost the same. The initial levels of PCr and ATP in individual preparations, however, remained almost unchanged during perfusion with the standard solution for 2 h. When the perfusion was stopped, the levels of ATP and PCr decreased, while the levels of PME and Pi increased. At that time, the Pi resonance shifted to a higher magnetic field, indicating that the tissue pH decreased. On reperfusion, the tissue levels of phosphorus compounds and the tissue pH were restored to their initial resting levels. Continuous infusion of 0.1 microM acetylcholine caused marked and sustained increases in the flow of pancreatic juice and protein output. During the stimulation the tissue levels of phosphorus compounds remained unchanged, while the tissue pH was decreased slightly (0.05 pH unit).

Adenine Nucleotides

Dissociation of fluid secretion and energy supply in rat mandibular gland by high dose of ACh.

The effects of a high dose of acetylcholine (ACh) on oxygen consumption (VO2) and changes in phosphorus energy metabolites during secretion were studied in isolated perfused mandibular gland of rats at 24 degrees C. Sugar phosphates (SP), Pi, phosphocreatine (PCr), and ATP were identified by phosphorus-31 nuclear magnetic resonance spectroscopy. One micromole ACh induced a tachyphylactic secretory response, a persistently elevated VO2, and decreased PCr and ATP; 1 mM ACh caused an initial burst of secretion that was followed by suppression of secretion and a rapid increase in the VO2 to the same level as that with 1 microM ACh. These findings indicate a dissociation between secretion and VO2. During stimulation with 1 mM ACh, the level of PCr first decreased and then partially recovered, but the level of ATP continued to decrease and the levels of Pi and SP increased markedly. These findings suggest compartmentalization of creatine phosphokinase (CPK) systems and the possibility that a high concentration of ACh interferes with the transport of PCr between one CPK system near adenosinetriphosphatase and another system near mitochondria in acinar cells.

Acetylcholine

In vivo 31P MRS in new antineoplastic agents evaluation on experimental tumor models.

Two new antineoplastic agents, a nitrosourea and a DNA-bis-intercalator have been studied in vivo by 31P magnetic resonance spectroscopy on a rat glioma and Walker carcinoma. On rat glioma, spectra are modified when the tumor is treated by the nitrosourea, showing the depletion of high-energy phosphates. On Walker carcinoma both drugs delay the tumor evolution to necrosis, showing important levels of high-energy phosphates on NMR spectra. There appears to be a great dependence upon energy metabolism during chemotherapy, depending on the nature and physiology of the observed tumor.

Animals