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Biomedical subjects

H Miyata

Publications and source records attributed to H Miyata.

At least 289 records · Page 16Linked to original sources
PubMed1992

Intracellular Ca2+ concentration and pHi during metabolic inhibition.

To study the changes in intracellular Ca2+ concentration ([Ca2+]i) and pH (pHi) during metabolic inhibition, rat ventricular myocytes were dual loaded with the acetoxymethyl esters of fura-2 (fura-2/AM) and 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF/AM) and perfused with 2 mM sodium cyanide (NaCN). The percent of rod-shaped cells was 30% of the control after 30 min NaCN in the absence of glucose. [Ca2+]i increased from 82 +/- 8 to 151 +/- 25 (SE) nM (P less than 0.05) when cells were shortened, and to 421 +/- 106 nM (P less than 0.05) when cells were rounded. There was a positive relationship between pCai and pHi (r = 0.425, P less than 0.01). When 50 mM glucose was added during NaCN, there were no significant changes in [Ca2+]i and the percent of rod-shaped cells after 30 min. The pHi of rod-shaped cells, however, decreased to 6.95 +/- 0.03 (P less than 0.01). Although the acidic solution (70% O2-30% CO2) decreased pHi to 6.90 +/- 0.05 (P less than 0.01), there were no changes in the cell shape or [Ca2+]i. The addition of NaCN for 30 min decreased the rod-shaped cells to 18% of the control. Mild acidosis did not cause the changes in cell shape or [Ca2+]i. There was also no protection of the NaCN-induced changes in cell shape by intracellular acidosis. It is likely that the changes in cell shape during metabolic inhibition were related to the depletion of metabolic energy and the increase in [Ca2+]i.

Analysis of Variance↗
PubMed1992

Cytoplasmic filaments in the endothelial cells of the sheathed capillary: an ultrastructural and immunocytochemical study in the pig spleen.

Cytoplasmic filaments of the endothelial cells of sheathed capillaries in the pig spleen were identified and their ultrastructure was studied. Two types of cytoplasmic filaments were found: intermediate filaments (diameter: 10 nm) which filled most of the interior of the cells, and thin filaments (diameter: 5 nm) which were located just beneath the cell membrane and filled the lateral cytoplasmic processes. In immunocytochemical preparations, the intermediate filaments were positive for vimentin and desmin, and were negative for keratin. Staining of the thin filaments with heavy meromyosin resulted in arrowhead formations. These observations suggest that the intermediate filaments maintain the cytoarchitecture, possibly protecting the cell from structural alterations induced by blood pressure changes. Concurrently, thin filaments may facilitate the passage of red blood cells and blood platelets through the interendothelial fenestrae of the sheathed endothelial cell to the reticular meshwork in the capillary sheath.

Animals↗
PubMed1992

Dependence of hypoxic cellular calcium loading on Na(+)-Ca2+ exchange.

Na(+)-Ca2+ exchange has been shown to contribute to reperfusion- and reoxygenation-induced cellular Ca2+ loading and damage in the heart. Despite the fact that both [Na+]i and [Ca2+]i have been documented to rise during ischemia and hypoxia, it remains unclear whether the rise in [Ca2+]i occurring during hypoxia is linked to the rise in [Na+]i via Na(+)-Ca2+ exchange before reoxygenation and how this relates to cellular injury. Single electrically stimulated (0.2 Hz) adult rat cardiac myocytes loaded with Na(+)-sensitive benzofuran isophthalate (SBFI), the new fluorescent probe, were exposed to glucose-free hypoxia (PO2 less than 0.02 mm Hg), and SBFI fluorescence was monitored to index changes in [Na+]i. Parallel experiments were performed with indo-1-loaded cells to index [Ca2+]i. The SBFI fluorescence ratio (excitation, 350/380 nm) rose significantly during hypoxia after the onset of ATP-depletion contracture, consistent with a rise in [Na+]i. At reoxygenation, the ratio fell rapidly toward baseline levels. The indo-1 fluorescence ratio (emission, 410/490 nm) also rose only after the onset of rigor contracture and then often showed a secondary rise early after reoxygenation at a time when [Na+]i fell. The increase in both [Na+]i and [Ca2+]i, seen during hypoxia, could be markedly reduced by performing experiments in Na(+)-free buffer. These experiments suggested that hypoxic Ca2+ loading is linked to a rise in Na+i via Na(+)-Ca2+ exchange. To show that Na(+)-Ca2+ exchange activity was not fully inhibited by profound intracellular ATP depletion, cells were exposed to cyanide, and then buffer Na+ was abruptly removed after contracture occurred. The sudden removal of buffer Na+ would be expected to stimulate cell Ca2+ entry via Na(+)-Ca2+ exchange. A large rapid rise in the indo-1 fluorescence ratio ensued, which was consistent with abrupt cell Ca2+ loading via the exchanger. The effect of reducing hypoxic buffer [Na+] on cell morphology after reoxygenation was examined. Ninety-five percent of cells studied in a normal Na(+)-containing buffer (144 mM NaCl, n = 38) and reoxygenated 30 minutes after the onset of hypoxic rigor underwent hypercontracture. Only 12% of cells studied in Na(+)-free buffer (144 mM choline chloride, n = 17) hypercontracted at reoxygenation (p less than 0.05). Myocytes were also exposed to hypoxia in the presence of R 56865, a compound that blocks noninactivating components of the Na+ current. R 56865 blunted the rise in [Na+]i typically seen after the onset of rigor, suggesting that Na+ entry may occur, in part, through voltage-gated Na+ channels.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Relation of mitochondrial and cytosolic free calcium to cardiac myocyte recovery after exposure to anoxia.

Mitochondrial calcium overload has been suggested as a marker for irreversible injury in the ischemic heart. A new technique is used to measure dynamic changes in mitochondrial free calcium concentration ([Ca2+]m) in electrically stimulated (0.2 Hz) adult rat cardiac myocytes during exposure to anoxia and reoxygenation. Cells were incubated with indo-1 AM, which distributes in both the cytosol and mitochondria. After Mn2+ quenching of the cytosolic signal, cells were exposed to anoxia, and the residual fluorescence was monitored. [Ca2+]m averaged 94 +/- 3 nM (n = 16) at baseline, less than the baseline diastolic cytosolic free calcium concentration ([Ca2+]c, 124 +/- 4 nM, n = 12), which was measured in cells loaded with the pentapotassium salt of indo-1. [Ca2+]m and [Ca2+]c rose steadily only after the onset of ATP-depletion rigor contracture. At reoxygenation 35 minutes later, [Ca2+]c fell rapidly to preanoxic levels and then often showed a transient further rise. In contrast, [Ca2+]m showed only a slight transient fall and a secondary rise at reoxygenation. At reoxygenation, cells immediately either recovered, demonstrating partial relengthening and retaining their rectangular shape and response to stimulation, or they hypercontracted to rounded dysfunctional forms. Recovery occurred only in cells in which [Ca2+]m or [Ca2+]c remained below 250 nM before reoxygenation. Early during reoxygenation, [Ca2+]m remained higher in cells that hypercontracted (305 +/- 36 nM) than in cells that recovered (138 +/- 9 nM, p less than 0.05), whereas [Ca2+]c did not differ between the two groups (156 +/- 10 versus 128 +/- 10 nM, respectively; p = NS).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

[Effect of tranilast, an anti-allergic drug, on carrageenin-induced granulation and capillary permeability in rats].

We studied the effect of tranilast on the growth of carrageenin-induced granulation and the increase in capillary permeability induced by inflammatory agents in rats. In the carrageenin-induced granulation model, tranilast (50 or 100-200 mg/kg, p.o.) decreased significantly and dose-dependently the weight and the hydroxyproline content of the granulation tissue. Tranilast, however, showed no effect on the healing day of locally wounded dorsal skin of rats. Triamcinolone (10 mg/kg, p.o.) also showed an inhibitory effect on the carrageenin-induced granulation model. Tranilast (50-400 mg/kg, p.o.) dose-dependently inhibited the enhancement of capillary permeability induced by the Ca ionophore A23187, bradykinin and xanthine oxidase. Moreover, tranilast (30 and 300 microM) suppressed superoxide production induced by FMLP in human neutrophils, but did not act as a superoxide scavenger. Considering that hypertrophic scar and keloid are conditions characterized by abnormal cell proliferation and excessive collagen accumulation accompanied with itch and pain, these results suggest that tranilast is useful as a therapeutic drug for hypertrophic scars and keloids.

Adult↗
PubMed1992

Effects of psychoactive drugs on short-term memory in rats and rhesus monkeys.

To examine the effects of drugs on short-term memory in animals, the delayed discrimination experiment in rats and the delayed matching to sample experiment in rhesus monkeys were conducted. Nicotine at 0.125 mg/kg, s.c. in rats and at 0.5 mg/kg, s.c. in monkeys increased the percentages of correct choices. Scopolamine at 0.06-0.12 mg/kg, s.c. in rats and at 0.015 mg/kg, s.c. in monkeys decreased the percentages of correct choices. However, supposedly memory-specific, delay-time-dependent disruptive effects by scopolamine were found only in monkeys. Diazepam at 0.5-2 mg/kg, s.c. did not change the correct choices in rats. However, diazepam at 1-4 mg/kg, i.g. decreased the correct choices in monkeys regardless of the delay time. Chlorpromazine at 0.25-1.5 mg/kg, s.c. showed inconsistent effects in rats. In monkeys, chlorpromazine at 0.25-0.5 mg/kg, s.c. had no effect. These results suggested that using both rats and monkeys would be useful for evaluating the effects of drugs on memory.

Animals↗
PubMed1992

[The signal transduction of serotonin secretion involves protein tyrosine phosphorylation in rat basophilic leukemia cells].

We investigated the signal transduction of serotonin secretion by stimulation with DNP-Ascaris antigen or ionomycin in rat basophilic leukemia cells (RBL-2H3). The modes of action of antigen and ionomycin for serotonin secretion were shown to be similar. The treatment of cells with antigen resulted in increased tyrosine phosphorylation of 105 and 72 KDa proteins, in particular, the tyrosine phosphorylation of 72 KDa protein seemed to correlate with serotonin secretion. Furthermore, we observed that antigen stimulation caused a marked increase in inositol polyphosphates production, which derived from the tyrosine phosphorylation of phospholipase C-gamma in RBL-2H3 cells. On the other hand, treatment with ionomycin also resulted in an increase in tyrosine phosphorylation of 72 KDa protein, but did not induce inositol polyphosphates production. These results suggested that the activation of tyrosine kinase may be related to serotonin secretion, and that intracellular Ca2+ increase may also play an important role in this activation.

Animals↗
PubMed1992

Localization and soma diameter of rat gluteus medius motoneurons.

1. Retrograde transport of horseradish peroxidase was used to examine the localization and soma diameter of motoneurons innervating the deep (predominance of oxidative fibers) and superficial (predominance of non-oxidative fibers) portions of the gluteus medius muscle in rats. 2. The motoneurons innervating the deep portion were located caudally within the gluteus medius motoneuron pool and had smaller average soma diameter than that of motoneurons innervating the superficial portion. 3. These results suggest that the location of the muscle fibers within the muscle is related to the location of motoneurons within the motoneuron pool, and that the soma diameter of motoneuron innervating oxidative muscle fibers may be smaller than those innervating non-oxidative fibers in the rat.

Animals↗
PubMed1992

[Bacteriology of infectious disease in otorhinolaryngology (1). Bacteriological study of paranasal cyst].

The bacteriology of chronic sinusitis has been reported in several studies, but there have been few bacteriological studies on paranasal cyst. Also, details of the techniques of transportation and processing of clinical specimens have only rarely been described. Therefore, this study was undertaken with attention to the above, and to obtain information on appropriate antimicrobial therapy, we conducted a dilution antimicrobial susceptibility test of isolates. Fifteen patients with paranasal cyst participated in this study. Their lesions were frontal sinus (3), ethmoid sinus (2), sphenoid sinus (2) and maxillary sinus (8). We punctured the cyst using an 18-gauge needle attached to a syringe at the time of operation, and paranasal cyst effusions were promptly transported to the laboratory. The specimen was immediately inoculated under a set of aerobic and prereduced anaerobic plates, which were incubated in appropriate conditions. Most specimens were completely processed within one hour. Antimicrobial susceptibility testing of isolates was done with MIC panel by the Sceptor system. Positive results of bacterial cultures were found in 60% of all cases (all of the frontal sinus specimens and half of those with other lesions). A total of 38 bacteria, including 24 anaerobic and 14 aerobic species, were isolated from 9 paranasal cysts, yielding an average of 4.2 species per positive case (2.9 anaerobes and 1.3 aerobes). Monomicrobial flora were found, anaerobes only in 2 cases and aerobes only in 2 others. Mixed flora were found in 5 cases in which both anaerobes and aerobes were isolated. The bacterial concentration in the maximal case was 4.9 x 10(4) CFU/ml. The organisms most frequently isolated were Propionibacterium spp. (7), Peptostreptococcus spp. (7), Staphylococcus spp. (7), Prevotella spp. (5) and Streptococcus spp. (4). On the basis of the results of antimicrobial susceptibility testing of isolates in vitro, the use of Cephems, Clavulanic acid/Amoxicillin, Norfloxacin and Chloramphenicol proved to be clinically effective.

Adult↗
PubMed1992

Photomodulation of the nucleating activity of a photocleavable crosslinked actin dimer.

The ability to generate substrate concentration jumps through photo-deprotection of amine, carboxyl and phosphate groups has been an important development for investigations of protein activity in complex systems. To broaden the versatility and applications of photo-deprotection techniques for the photomodulation of protein activity we describe the synthesis and characterisation of a reagent for generating free thiol from thioether groups and a related photocleavable, heterobifunctional crosslinking reagent. Chemical and spectroscopic studies of a model thiol protected derivative were used to show some features of thiol group photodeprotection. To demonstrate how the photocleavable crosslinking reagent may be used to modulate the activity of proteins we investigated the effect of light on the nucleating activity of crosslinked actin dimer; thus following near-ultraviolet irradiation of the actin dimer the crosslink was cleaved, presumeably at the thioether bond, resulting in the concomitant dissociation of dimer, loss of nucleating activity and creation of a concentration jump of polymerisable G-actin monomer. On the basis of this initial study we discuss applications and limitations of these reagents for the photomodulation of protein activity in vitro and in vivo.

Actins↗
PubMed1991

Antigen-induced biphasic diacylglycerol formation in RBL-2H3 cells: the late sustained phase due to phosphatidylcholine hydrolysis is dependent on protein kinase C.

Exposure to antigen (Ag) caused a biphasic 1,2-diacylglycerol (DG) production in [3H]myristic acid-labeled RBL-2H3 cells; the early, small transient phase and the second large sustained phase. The accumulation of phosphatidic acid (PA) or phosphatidylethanol (PEt) in the presence of ethanol was paralleled by the second-phase DG generation. Ag-induced formation of phosphocholine and choline in [3H]choline-labeled cells suggested the hydrolysis of phosphatidylcholine (PC) by phospholipases C and D. Treatment with phorbol myristate (PMA) or A23187 caused increases in [3H]DG and water-soluble [3H]choline metabolites. In protein kinase C (PKC) down-regulated cells, PEt formation was markedly reduced. In these cells DG production induced by Ag and A23187 was largely suppressed, thus indicating that PKC would play an important regulatory role for PC hydrolysis. However, because the A23187 treatment showed significant accumulation of water-soluble choline metabolites in PKC down-regulated cells, an increase in intracellular Ca2+ is another factor regulating PC hydrolysis. Taken together, these results may indicate that PC hydrolysis in response to Ag is dependent on PKC and Ca2+.

Animals↗
PubMed1991

Soma diameter and oxidative enzyme activity of identified alpha-motoneurons: application of a retrograde fluorescent neuronal tracer.

Using a fluorescent compound as a retrograde neuronal tracer for the identification of a specific motoneuron pool, we examined the soma diameter and oxidative enzyme activity of alpha-motoneurons innervating the extensor digitorum longus (EDL) and the soleus (Sol) muscles in the rat. An inverse relationship between soma diameter and oxidative enzyme activity was demonstrated in the EDL, while there was no significant relationship between these variables in the Sol. These results suggest that an inverse relationship between soma diameter and oxidative enzyme activity cannot always be demonstrated in all of the motoneuron pools.

Animals↗
PubMed1991

Measurement of mitochondrial free Ca2+ concentration in living single rat cardiac myocytes.

A technique that allows the continuous measurement of mitochondrial free Ca2+ ([Ca2+]m) in a single living cardiac myocyte is described. It involves the introduction of the fluorescent chelating agent indo-1 into the cell by exposure to the acetoxymethyl ester, followed by selective quenching of the fluorescence of indo-1 in the cytosol by Mn2+. The identity of the remaining fluorescence due to intramitochondrial indo-1 is established by its resistance to treatment of the cell with digitonin at concentrations that release cytosolic but not mitochondrial enzymes and by the finding that ruthenium red and carbonyl cyanide p-trifluoromethoxyphenylhydrazone prevent its response to elevated cytosolic free Ca2+ ([Ca2+]c). [Ca2+]m is found to be low (less than 100 nM) in unstimulated cells and to rise in procedures that chronically elevate [Ca2+]c, such as Na+ replacement. The gradient [Ca2+]m/[Ca2+]c is less than unity at values of [Ca2+]c of less than 500 nM but rapidly increases at higher values of [Ca2+]c. Although there is no detectable increase in [Ca2+]m during a single electrical stimulation, [Ca2+]m increases up to 600 nM as the pacing frequency is raised to 4 Hz in the presence of norepinephrine; this increase occurs over the course of many contractions. It is concluded that these findings are consistent with an increase in [Ca2+]m acting as a signal to increase dehydrogenase activity, and hence flux through oxidative phosphorylation, in response to increased work loads.

Animals↗