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T Hukuhara

Publications and source records attributed to T Hukuhara.

At least 19 recordsLinked to original sources
PubMed2001

Enhanced fusion of a nucleopolyhedrovirus with cultured cells by a virus enhancing factor from an entomopoxvirus.

Fusion of Pseudaletia unipuncta nucleopolyhedrovirus with an armyworm cell line (SIE-MSH-805-F) was studied by means of three fluorescence assays that are based on the relief of fluorescence self-quenching of octadecylrhodamine B chloride (R18). A gradual increase in fluorescence intensity indicative of virus-cell fusion was observed by spectrofluorometry when R18-labeled polyhedron-derived virus was incubated with cultured cells. The fusion was enhanced by the virus enhancing factor (EF) from Pseudaletia separata entomopoxvirus. Lysosomotropic agents had little effect on the virus-cell fusion. The percentage of positively fluorescent cells, as determined by flow cytometry, gradually increased after the addition of labeled virus and was higher in the presence of the EF than in its absence. Confocal microscopy of cultured cells that had been combined with labeled virus showed that the fluorescence appeared first on their surface. The plasma membrane of cultured cells had specific affinity to the EF, as revealed by indirect immunofluorescence microscopy.

Animals↗
PubMed2001

A bacterially produced virus enhancing factor from an entomopoxvirus enhances nucleopolyhedrovirus infection in armyworm larvae.

Using an Escherichia coli expression system, pGEX-2T, that expresses foreign sequences as fusion proteins with a glutathione S-transferase (GST) carrier, we have expressed a virus enhancing factor (EF) from Pseudaletia separata entomopoxvirus, which enhances P. unipuncta multi nucleopolyhedrovirus (PsunMNPV) infection in larvae of the armyworm, P. separata. The lysates of transformed E. coli cells, which were not active in enhancing PsunMNPV infection, became active when treated with either trypsin or thrombin. The GST-EF fusion protein in a lysate was purified with a bulk GST purification module and cleaved into the EF and GST moieties with thrombin. Removal of the GST moiety with glutathione-Sepharose 4B resulted in a highly purified EF preparation, which enhanced PsunMNPV infection in armyworm larvae and PsunMNPV fusion with an armyworm cell line, SIE-MSH-805-F.

Animals↗
PubMed1999

Increased baculovirus susceptibility of armyworm larvae feeding on transgenic rice plants expressing an entomopoxvirus gene.

We have introduced an entomopoxvirus gene encoding a virus enhancing factor (EF) into rice, which resulted in high-level accumulation of the EF in the transgenic plants. The introduced gene was stably inherited in the progeny of the primary transformants, as shown by analysis of their genomic DNA. Bioassays for insect susceptibility to baculovirus infection showed that armyworm larvae feeding on the transgenic rice had increased susceptibility to a Nucleopolyhedrovirus. Thus, introduction of the EF gene into plants can be used as a strategy to increase the effectiveness of baculoviruses in insect pest management.

Animals↗
PubMed1998

Detection of a virus enhancing factor in the spheroid, spindle, and virion of an entomopoxvirus.

Spheroids, spindles, and virions of an entomopoxvirus (EPV) enhanced the infectivity of a nuclear polyhedrosis virus (NPV) when they were perorally administered to larvae of the armyworm, Pseudaletia separata. Spheroids and spindles at the same dose exhibited nearly the same enhancing activity. When the dose of spheroids or spindles was reduced 10 times, the median infectious dose of the NPV was increased approximately 100 times. An antiserum against an enhancing factor detected the homologous antigen in spheroids, spindles, and tissue-derived EPV virions but not in spheroid-derived virions.

Animals↗
PubMed1998

[Entomopoxviruses].

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Acid Anhydride Hydrolases↗
PubMed1996

Cloning and sequencing of the gene for an enhancing factor from Pseudaletia separata entomopoxvirus.

An enhancing factor (EF) has been shown to increase the susceptibility of the armyworm, Pseudaletia separata, to this nuclear polyhedrosis virus. Using amino acid microsequencing data obtained from the purified EF protein, we cloned and sequenced the gene for EF. The deduced amino acid sequence exhibited a significant similarity to major spindle proteins, fusolins, whose function is unknown, from Heliothis armigera, Choristoneura biennis and Melolontha melolontha entomopoxviruses.

Amino Acid Sequence↗
PubMed1990

Selective actions of anesthetic agents on membrane potential trajectory in bulbar respiratory neurons of cats.

The effects of two anesthetic agents, halothane and thiopental, on the membrane potential trajectory of respiratory-related neurons in the ventral respiratory group were investigated in decerebrate cats, of which the carotid sinus and vagal afferents were denervated. Infusion of halothane (2% for 90 s) depolarized the membrane in nearly half of the inspiratory (12/21), post-inspiratory (10/26) and expiratory (4/6) neurons and caused hyperpolarization in the rest of the population. Thiopental (2.5 mg/kg i.v.) produced depolarization in 11 inspiratory and 10 post-inspiratory neurons and hyperpolarization in 1 expiratory, 4 inspiratory and 7 post-inspiratory neurons. In both hyperpolarized and depolarized neurons, reduction of the respiratory membrane potential fluctuations and an increase of input resistance were commonly observed. Both drugs depressed spontaneous firing in most of the neurons studied. An increase of firing was observed in 9 out of 47 depolarized cells. These two contrasting effects on the membrane potential trajectory occurred similarly in the known groups of respiratory neurons, but the response of a given cell was consistent for the two anesthetic agents. The present results demonstrate that the anesthetic drugs exert various influences on the ventral respiratory group neuron population in maintaining the membrane potential trajectory and discharge activity. This may reflect a functional heterogeneity in the bulbar respiratory network of neurons.

Action Potentials↗
PubMed1990

Replication of an entomopoxvirus in two lepidopteran cell lines.

Pseudaletia separata entomopoxvirus replicated in two lepidopteran cell lines, SIE-MSH-805-F and BM-N. Microscopic examination, and the virus passage tests, of infected cultures indicated that the virus replicated more readily in the former cell line. Virus release by exocytosis occurred in both cell lines. A sequence of virus morphogenesis in the cultured cells was described, based on electron microscopic observations of thin sections. The nucleus of infected cells contained spherical inclusions, and the cytoplasm contained virions, immature virus forms, spheroids, and spindles. A portion of the virions in the cytoplasm was occluded within spheroids, which were often associated with crystallogenic matrix. Virions acquired a coat prior to their occlusion.

Animals↗
PubMed1990

[Physiological and neuropharmacological aspects of the central respiratory mechanisms].

The present physiological and neuropharmacological views and the essentials of the experimental results on the anatomical localization, functional and neuronal organization of the central respiratory mechanisms, classically expressed as the respiratory centers, in the brain stem were reviewed and discussed. The brain stem neural mechanism for central regulation of breathing is regarded as a complex neuronal mechanism consisting of several functional subsystems subserving different functions. One of its functions is the generation of respiratory rhythm. The subsystem for respiratory rhythm-generating mechanisms is located in the medullary reticular formation outside the DRG and VRG regions, which are thought to be premotor neuron pools. Rhythmic activity originating in the medulla is dominant in terms of the spontaneity over other rhythmic activity in the pontine and spinal cord mechanisms. Evidences for heterogeneity of the functional properties of brain stem respiratory neurons have been demonstrated. Neuronal mechanisms involving respiratory neurons identified as members of the primary respiratory neuron population or neuronal networks consisting of different types of respiratory neurons located in the lateral region of the bulbar reticular formation may play important roles in the generation of respiratory rhythms. These aspects contribute to the understanding of the neurophysiological basis, providing important prerequisites for further neuropharmacological studies on neurotransmission within the neuronal network of the central respiratory mechanisms.

Animals↗
PubMed1989

Location of a synergistic factor in the capsule of a granulosis virus of the armyworm, Pseudaletia unipuncta.

A synergistic factor (SyF), which enhanced the infection of nuclear polyhedrosis viruses, was purified from capsules of a Pseudaletia unipuncta granulosis virus (Hawaiian strain) by immune affinity chromatography. The isolated SyF consisted primarily of a protein with molecular mass 98 kDa. The recovery rate depended on the alkali used to dissolve the capsules: the highest rate occurred with 0.05 M Na2CO3-0.05 M NaCl, followed in turn with 0.02-0.05 M NaOH and 0.04 M NaOH-0.05 M glycine. The solubilized components from untreated capsules contained 98- and 100-kDa proteins in addition to the matrix protein (29 kDa) and its decomposed products, while those from heat-treated capsules contained only the 100-kDa protein. Virons liberated from the capsules with the glycine buffer contained three proteins (33, 98, and 100 kDa) serologically related to the SyF. Immunoelectron microscopy of infected tissue and purified virions revealed the localization of the SyF antigens on the viral envelope.

Animals↗
PubMed1989

Diazepam potentiates postsynaptic inhibition in bulbar respiratory neurons of cats.

The purpose of this study was to assess the effect of benzodiazepine on inhibitory postsynaptic potentials (IPSPs) of medullary respiratory neurons in decerebrate, paralyzed cats. Diazepam (0.05 and 0.1 mg/kg i.v.) reversibly increased the IPSP waves occurring during the inactive phase of the respiratory cycle in all inspiratory and postinspiratory neurons examined. Input resistances of these neurons were reduced at that phase. The reversal potential for the IPSP wave was unaltered. Intracellular injection of chloride ions reversed the IPSP to depolarization, and diazepam produced a purely depolarizing effect. The drug effects observed during the active phase of each neuron include a decrease in the firing rate and a shortening of the burst activity. The firing threshold and shape of these spikes, however, remained unaltered. These results suggest that diazepam depresses the bulbar respiratory neuronal activities specifically by potentiating the periodic postsynaptic inhibition.

Animals↗
PubMed1988

Medullary inspiratory neurons with stable respiratory rhythm and little correlation to phrenic high-frequency oscillation.

Respiratory neurons with a highly stable rhythm have been proposed to be involved in the central neural mechanism responsible for the respiratory rhythmogenesis. In the present study it was examined whether the stably discharging inspiratory neurons are related to the high-frequency oscillation (HFO) in the phrenic nerve activity. Experiments were performed on 27 vagotomized rabbits anesthetized with diethyl ether, immobilized with gallamine triethiodide, and ventilated artificially. Spike-triggered averaging was used to evaluate the degree of the correlation between inspiratory unit spikes and the phrenic HFO. Twenty out of the 35 inspiratory units examined were related to HFO and were located in the medullary reticular formation (n = 19; n, number of units) and in the vicinity of the nucleus tractus solitarius (n = 1). The correlation to HFO decreased after the intravenous injection of thiamylal. The inspiratory units with little correlation to HFO were located in the reticular formation of the pons (n = 3) and medulla (n = 12). The inspiratory neurons with a stable respiratory rhythm (n = 4) had little correlation to HFO and located in the lateral region of the medullary reticular formation. These results suggest that the central neural mechanisms responsible for phrenic HFO are not directly related to the respiratory rhythmogenesis.

Action Potentials↗
PubMed1988

Organization of the brain stem neural mechanisms for generation of respiratory rhythm--current problems.

The brain stem neural mechanism for central regulation of breathing is regarded as a complex neuronal mechanism consisting of several functional subsystems subserving different functions. One of its functions is the generation of the respiratory rhythm. Evidence indicates with certainty that the subsystem for respiratory rhythm-generating mechanisms is located in the medullary structure outside the DRG and VRG regions which have been postulated for many years as the hypothetical site generating respiratory rhythm. DRG and VRG are thought to be premotor neuron pools. Rhythmic activity originating in the medulla is dominant in terms of the spontaneity over other rhythmic activity in the pontine mechanisms as well as those in the spinal cord. Evidences for heterogeneity of functional properties of respiratory neurons in the brain stem are demonstrated. Possible functional differentiation among respiratory neurons is suggested. Neuronal mechanisms involving respiratory neurons identified as members of primary respiratory neuron populations or neuronal networks consisting of various types of respiratory neurons located in the lateral region of the bulbar reticular formation may play important roles in generation of respiratory rhythms. Precise neural processes within the neuronal mechanisms for respiratory rhythm generation are rather equivocal and remain to be determined by further investigation.

Animals↗
PubMed1987

Dopaminergic control of the cough reflex as demonstrated by the effects of apomorphine.

The effect of apomorphine on the cough reflex induced by electrical stimulation of the superior laryngeal nerve was studied in cats anesthetized with pentobarbital. Intravenous administration of apomorphine in doses which ranged from 0.1 to 1.0 mg/kg decreased the number of coughs in a dose-dependent manner. Haloperidol (0.3 mg/kg i.v.) did not significantly change the number of coughs. However, haloperidol administered 15 min prior to administration of apomorphine abolished the apomorphine-induced decrease in the number of coughs. These results suggest that dopaminergic mechanisms could have an important role in the regulation of the cough reflex.

Animals↗
PubMed1986

[Effects of naloxone on the cough depressant activities of antitussive drugs].

Effects of naloxone on the cough depressant action of antitussive drugs were investigated in pentobarbital anesthetized male and female cats. Respiration and cough reflex were measured using a pneumotachograph via a cannula inserted into the trachea. The cough reflex was elicited by electrical stimuli to the superior laryngeal nerve. An i.v. administration of morphine (1 mg/kg) significantly inhibited the cough reflex for at least 60 min. The antitussive effect of morphine was antagonized by naloxone (400 micrograms/kg). The inhibitory effect of morphine on the respiratory frequency was also antagonized by naloxone. The cough reflex was significantly inhibited by fominoben (5 mg/kg, i.v.) and dextromethorphan (3 mg/kg, i.v.). However, the respiratory frequency was increased by these two drugs. An i.v. administration of naloxone prevented the antitussive effects and the excitatory effects in respiratory frequency of fominoben and dextromethorphan. The present study suggests that the cough depressant actions of antitussive drugs may be mediated by endogenous opiates and/or by neurotransmitters which are modified by endogenous opiates.

Animals↗
PubMed1986

Effects of methysergide on the cough reflex.

The present study had two basic purposes: to observe the effect of methysergide on the cough reflex and to investigate the effect of methysergide on the antitussive effect of dextromethorphan. Male and female cats were anesthetized with pentobarbital-Na. Respiration and cough reflex were measured using a pneumotachograph via a cannula inserted into the trachea. The cough reflex was elicited by electrical stimuli to the superior laryngeal nerve. Methysergide (3 mg) injected into the vertebral artery increased the number of coughs and respiratory frequency. Dextromethorphan in a dose of 3 mg inhibited the cough reflex. Methysergide (1 and 3 mg) reduced the antitussive effect of dextromethorphan in a dose-dependent manner, but did not inhibit the excitatory effect on respiratory frequency. These findings might indicate that the central serotonergic system has an inhibitory role on the cough reflex and may be related to the antitussive mechanisms of dextromethorphan.

Animals↗