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K Muramoto

Publications and source records attributed to K Muramoto.

At least 73 records · Page 4Linked to original sources
PubMed1995

Substitution of lysine for arginine in the N-terminal 217th amino acid residue of the H gamma II of Staphylococcal gamma-hemolysin lowers the activity of the toxin.

The staphylococcal toxin gamma-hemolysin consists of two protein components, LukF and H gamma II. Staphylococcus aureus P83 was found to have five components, LukF, LukF-PV, LukM, LukS, and H gamma II for leukocidin or gamma-hemolysin. H gamma II of S. aureus P83 was demonstrated to be a naturally-occurring analogous molecule of H gamma II [H gamma II(P83)], in which the 217th arginine residue was replaced by lysine. The H gamma II(P83) showed about 50% of the hemolytic activity of normal H gamma II in the presence of LukF.

Amino Acid Sequence↗
PubMed1994

A substrate of ecto-protein kinase is microtubule-associated protein 1B in cortical cell cultures undergoing synaptogenesis.

Synapse formation between cultured rat cortical neurons is inhibited by the continuous application of K-252b, an ecto-protein kinase inhibitor, which cannot permeate the cell membrane. In order to identify the phosphorylated membrane proteins which are necessary for synapse formation, endogenous substrates for ecto-protein kinase activity were investigated. To detect phosphorylation of proteins containing extracellular domains, [gamma-33P]ATP was applied to the medium for brief periods. Proteins were then separated by SDS polyacrylamide gel electrophoresis and detected by autoradiography. Some bands showed immediate phosphorylation and this phosphorylation was suppressed by the addition of K-252b to the medium. We examined partial amino acid sequences of these substrates. The band with the highest molecular weight, whose phosphorylation was strongly inhibited by K-252b, was identified as microtubule-associated protein (MAP) 1B. These results suggest the possibility that the phosphorylation of extracellular domains of MAP1B is involved in synaptogenesis between cortical neurons.

Adenosine Triphosphate↗
PubMed1994

Cloning and analysis of the gene encoding lectin from the acorn barnacle Megabalanus rosa.

In the acorn barnacle Megabalanus rosa, two types of galactose-binding C-type lectins (BRA-2 and BRA-3) have been identified. Here, we report the isolation and characterization of BRA-2 cDNA and genomic clones. In contrast to the BRA-3 gene, which consists of four exons, BRA-2 is encoded by a single exon, implying differences in the physiological roles of the two lectins.

Amino Acid Sequence↗
PubMed1994

Endoglycoceramidase treatment inhibits synchronous oscillations of intracellular Ca2+ in cultured cortical neurons.

Gangliosides are major components of nerve cell membranes and are especially rich in synaptic areas. In order to evaluate the role of endogenous gangliosides in synapse formation, endoglycoceramidase (EGCase) was used to remove oligosaccharides of gangliosides from the cell surface. We have reported previously that synapse formation between cultured rat cerebral cortical neurons can be estimated by the synchronous oscillation of synaptic activity monitored by fura-2 calcium imaging. Continuous application of endoglycoceramidase (EGCase) together with its activator protein dose-dependently decreased the frequency of synchronous oscillations without any morphological changes in neurons and their neurites. The result suggests that oligosaccharides liberated from glycosphingolipids on cultured cortical cell surface with EGCase are important for synapse formation between cortical neurons.

Animals↗
PubMed1994

Successive inactivation of the force-generating units of sodium-driven bacterial flagellar motors by a photoreactive amiloride analog.

Like amiloride, 6-iodoamiloride (6-IA) competitively and reversibly inhibits rotation of the Na(+)-driven flagellar motors of alkalophilic Bacillus cells. However, when 6-IA-treated cells are irradiated with UV light, motility is irreversibly inhibited. This treatment does not alter the membrane potential or affect Na(+)-coupled alpha-aminoisobutyrate transport. An increase in the Na+ concentration during UV irradiation substantially protects the motors from irreversible inhibition. Thus, photoactivated 6-IA seems to bind specifically and covalently at or around the Na(+)-interaction site of the force-generating units of the motors to inhibit motor rotation irreversibly. Rotation of each motor, which is monitored using tethered alkalophilic Bacillus cells, is also inhibited by photoactivated 6-IA. In this case, however, the rotation rate during UV irradiation decreases stepwise, suggesting the presence of several independently functioning force-generating units in a motor. From the data of 14 tethered cells, the number of units/motor is estimated to be 5-9.

Amiloride↗
PubMed1994

Aluminum promotes the aggregation of Alzheimer's amyloid beta-protein in vitro.

Amyloid beta-protein is the major component of senile plaques in the brains of Alzheimer's disease and has an intrinsic tendency to form insoluble aggregates. The aggregation of amyloid beta-protein has been suggested to enhance its neurotoxicity and to play a key role in the amyloid deposition. Here we show, using gel-electrophoresis and immunoblotting, that the aggregation of synthetic amyloid beta-protein (beta 1-40) is promoted by aluminum, a suspected risk factor in Alzheimer's disease. High molecular weight aggregates were observed, and the amount of precipitated protein was estimated using high performance liquid chromatography. The results suggest the possibility that aluminum directly influences the process of aggregation and the deposition of senile plaques.

Aluminum Chloride↗
PubMed1994

Ostrich (Struthio camelus) carboxypeptidase A: purification, kinetic properties and characterization of the pancreatic enzyme.

1. Carboxypeptidase A beta and carboxypeptidase A tau-type from the pancreas of the ostrich were purified by water extraction of acetone powder, aminobenzylsuccinic acid affinity and hydroxylapatite chromatography. 2. The final preparations were homogeneous when subjected to SDS-PAGE and PAGE. The M(r) values obtained from SDS-PAGE for CPA beta and CPA tau-type were 34,600 and 34,400, respectively. 3. The effects of inhibitors (1,10 phenanthroline and indole-3-acetic acid), pH and temperature on CPA activity were examined. Ki-values for CPI, PPA, D-phe, D-trp and aminobenzylsuccinic acid were determined. 4. Km, kcat and kcat/Km values were determined for hipp-phe, cbz-gly-phe, cbz-(gly)2-phe, cbz-gly-leu, cbz-(gly)2-leu and cbz-(gly)2-val. 5. N-terminal sequencing and amino acid analysis were performed for CPA beta and CPA tau-type.

Amino Acid Sequence↗
PubMed1994

Myosin II is involved in transmitter release at synapses formed between rat sympathetic neurons in culture.

The presynaptic function of myosin II was studied at cholinergic synapses formed between rat superior cervical ganglion neurons in culture. Immunofluorescent staining showed that myosin II was colocalized with synaptophysin at the presynaptic nerve terminals. Antimyosin II antibody introduced into presynaptic neurons inhibited synaptic transmission. Transmission was also inhibited in a dose-dependent manner by two inhibitors of myosin light chain kinase: a peptide, SM-1, and an organic inhibitor, wortmannin. The inhibition produced by these agents was dependent on presynaptic activity. Extracellularly applied wortmannin also blocked synaptic transmission, but its effects were slower in onset. Wortmannin also decreased postsynaptic potentials and post-tetanic potentiation in intact superior cervical ganglia. These results suggest a model in which myosin light chain kinase phosphorylates myosin, and the resultant change in actin-myosin interactions is involved in neurotransmitter release.

Acetylcholine↗
PubMed1994

Preparation of a photoactivatable fluorescent derivative of lactose and its application to photoaffinity labeling of a conger eel lectin.

A photoactivatable heterobifunctional fluorescent reagent, 1-azido-5-naphthalene sulfonyl (ANS) hydrazide, was synthesized and characterized. ANS-hydrazide reacted with lactose to form a photoactivatable hydrazone. The derivative (ANS-lactose) had the same binding affinity for a conger eel lectin as lactose judging from the hemagglutinating-inhibition assay with rabbit erythrocytes. ANS-lactose was used for photoaffinity labeling of a conger eel lectin. The photolabeled lectin was digested with chymotrypsin to isolate photolabeled peptides by reversed-phase HPLC by monitoring fluorescence. A major labeled peptide was located at positions 31-45 in the lectin by amino acid analysis and N-terminal sequencing. The identified segment was close to the highly conserved region throughout animal beta-galactoside-binding lectins.

Affinity Labels↗
PubMed1994

Gas-phase microsequencing of peptides and proteins with a fluorescent Edman-type reagent, fluorescein isothiocyanate.

A fluorescent Edman-type reagent, fluorescein isothiocyanate (FITC), was adapted to a gas-phase sequencer fitted with a miniature reaction chamber. Fluorescein thiohydantion amino acids were identified by on-line gradient HPLC with fluorescence monitoring. The optimized protocol for the coupling reaction using FITC and phenylisothiocyanate, and the degree of washing to remove the excess reagents were established. Myoglobin (5 pmol) and lysozyme (5 pmol) were analyzed by the sequencer to obtain a 35-51% initial yield (I.Y.) and 82-88% repetitive yield (R.Y.) for the former, and 51-66% I.Y. and 91-92% R.Y. for the latter. These figures permitted 20-25 amino acid residues to be identified from the N-terminus of 5 pmol samples applied to the sequencer.

Amino Acid Sequence↗
PubMed1994

The Cleveland Clinic-Nimbus total artificial heart. In vivo hemodynamic performance in calves and preclinical studies.

In vitro function of the Cleveland Clinic-Nimbus electrohydraulic total artificial heart met National Heart, Lung, and Blood Institute hemodynamic guidelines for such devices. In a series of in vivo experiments, we implanted the total artificial heart in eight calves (mean weight 87 kg), one for a short-term experiment and seven for long-term experiments. The mean blood flow during support was 7.7 +/- 1.6 L/min with left atrial pressure 13 +/- 6 mm Hg, right atrial pressure 13 +/- 4 mm Hg, and aortic pressure 97 +/- 9 mm hg. Maximum pump flow (9.6 L/min) occurred after 4 days of support as a result of the high resting cardiac output of the animals. A 10% to 15% right pump stroke-volume limit effectively balanced atrial pressures, and afterload insensitivity was confirmed by the in vivo studies. Calves tolerated treadmill exercise studies well, with an average duration of 22 minutes and an average top speed of 2.1 mph. The experiments were terminated after 1 day to 120 days of support (mean 32 days). Most experiments were terminated as a result of correctable mechanical problems. In a separate study of six adult human patients undergoing orthotopic cardiac transplantation, five showed an excellent fit for the Cleveland Clinic-Nimbus total artificial heart. Further studies using chest roentgenograms, chest measurements, and transesophageal echocardiography should help predict fit of the total artificial heart in potential candidates. Initial candidates for a "vented-electric" version of the Cleveland Clinic-Nimbus total artificial heart are patients for whom univentricular (left ventricular assist device) support is not appropriate, but who require mechanical support as a bridge to cardiac transplantation.

Animals↗
PubMed1993

Frequency of synchronous oscillations of neuronal activity increases during development and is correlated to the number of synapses in cultured cortical neuron networks.

It has been proposed that synchronous oscillations of groups of neurons corresponding to sensory information and changes in temporal pattern of oscillations are important for processing of the information in the cortex. However, it has not been determined yet how the temporal or spatial pattern of such oscillations are regulated. We observed spontaneous synchronous oscillations of Ca2+ transients, which were caused by bursts of action potentials of neurons, even in cultured cortical neurons. The frequency of synchronous Ca2+ oscillations increased with development of synapses in cultured neurons and was highly correlated to the number of synapses formed in the same culture.

Animals↗
PubMed1993

Acorn barnacle Megabalanus rosa lectin (BRA-3): cDNA cloning, gene structure and seasonal changes of mRNA and protein levels.

We have isolated cDNA clones coding for a lectin (BRA-3) from the acorn barnacle, Megabalanus rosa. Sequence comparison of the cDNA clones has revealed polymorphism in the BRA-3 mRNA, which results from single-nucleotide (nt) differences at three positions. All three differences are within the coding region and cause conservative amino acid (aa) changes. The BRA-3 gene is composed of four exons, and the three single-nt differences are located on different exons. In addition, the BRA-3 mRNA and BRA-3 protein levels increased during early summer in a similar fashion, indicating that BRA-3 production is regulated mainly at the level of transcription.

Amino Acid Sequence↗
PubMed1993

Characterization of peptides in sea anemone venom collected by a novel procedure.

Peptide neurotoxins were isolated from the venom obtained by electrical stimulation of the sea anemone Bunodosoma caissarum. This technique allows almost pure venom to be collected, and the animals to survive. Three neurotoxins (assayed on crustacean nerves) were isolated by gel filtration and reversed-phase high performance liquid chromatography. Hemolysins were also detected in the venom. The amino acid sequence of a major neurotoxin BcIII was determined. BcIII has 48 amino acid residues with six half-cystine residues. This sequence has homology with the type 1 long sea anemone neurotoxins. Two minor toxins (BcI and II) have similar amino acid composition and amino-terminal sequences to BcIII.

Action Potentials↗
PubMed1993

Formation and maturation of synapses in primary cultures of rat cerebral cortical cells: an electron microscopic study.

Cerebral cortical cells from fetal rats (18 days) were cultured for 3, 7, 14, 21, 28, 35 days in vitro (DIV) and the development of synapses was examined morphologically by electron microscopy. At 3 DIV, no synapses could be recognized. An immature type of synapse was found at 7 DIV which thereafter developed morphologically. The length of the synaptic contact zone (SCZ) increased with DIV from 271 +/- 11.4 nm (mean +/- SEM) at 7 DIV to 410 +/- 11.4 nm at 35 DIV. The number of synaptic vesicles per terminal also increased with DIV: 10.0 +/- 1.2 at 7 DIV, 35.7 +/- 3.4 at 21 DIV, and 53.3 +/- 4.5 at 35 DIV. The time course of numerical density of synapses was examined quantitatively by electron microscopy. Synaptic density was very low at 7 DIV. It was significantly increased at 14 DIV and thereafter showed variable changes. Four culture series showed decreases after 14 DIV, but one series showed a further increase at 21 DIV followed by a decrease at 28 DIV. The mean density of synapses at each DIV was as follows: 1780 +/- 86/10(6) microns3 at 7 DIV, 4244 +/- 595/10(6) microns3 at 14 DIV, 2285 +/- 674/10(6) microns3 at 21 DIV, 2552 +/- 646/10(6) microns3 at 28 DIV, and 2080 +/- 532/10(6) microns3 at 35 DIV. Neuronal cell density was counted in each culture to calculate the relative number of synapses per neuron. The cell density decreased with age from 301 +/- 51/10(6) microns3 at 7 DIV to 39 +/- 9/10(6) microns3 at 35 DIV.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Rat amygdaloid neuron responses during auditory discrimination.

To investigate amygdaloid contribution to stimulus-affect association and to emotional behavior, single neuron activity was recorded in the basolateral and corticomedial amygdala of the rat amygdala during discrimination of conditioned cue tones associated with positive (glucose and intracranial self-stimulation) and negative (weak electric footshock) unconditioned stimuli. Amygdaloid neurons that responded to two rewarding unconditioned stimuli responded in the same manner to both. Responses to conditioned cue tones and those to rewards were also significantly related, suggesting that amygdaloid neuronal responses to conditioned stimuli are closely related to the affective nature of the corresponding unconditioned stimulation. These might be neuronal functions that underlie stimulus-affect association in the amygdala. In random repeated trials, including acquisition to conditioned cue tones and extinction after acquisition, response plasticity was more evident in basolateral than in corticomedial neurons. Furthermore, more basolateral than corticomedial neurons differentiated glucose and its cue from intracranial self-stimulation and its cue by responding to one but not the other. More basolateral than corticomedial neurons were excited by intracranial self-stimulation and its cue and inhibited by footshock and its cue. The differences between the two nuclear groups suggest that basolateral neurons are more plastic and discriminative, and corticomedial neurons respond indiscriminately to positive and negative unconditioned or conditioned stimuli. The results indicate amygdaloid neuron involvement in stimulus-affect association. Basolateral neurons may be more involved than corticomedial neurons in discriminating and learning conditioned stimuli.

Acoustic Stimulation↗
PubMed1992

Functional and morphological changes in cultured neurons of rat cerebral cortex induced by long-term application of aluminum.

Aluminum is an environmental neurotoxin and a suspected risk factor for Alzheimer's disease. The neurotoxicity of aluminum on cultured neurons of rat cerebral cortex was investigated using an assay system for synapse formation and immunohistochemistry. The frequency of spontaneous oscillations of intracellular Ca2+, which is correlated to the number of synapses, was decreased after exposure to 100 microM of aluminum chloride for 22 days. Long-term application of aluminum (48 days) caused aggregation of cell bodies and fasciculation of processes. Processes and cell bodies were strongly stained by antibody to tau protein, which is one of the main components of Alzheimer's neurofibrillary tangles. It is suggested that the characteristics of the degeneration of cultured neurons induced by aluminum show some similarities to the pathology observed in brains with Alzheimer's disease.

Aluminum↗