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N Kudo

Publications and source records attributed to N Kudo.

At least 55 records · Page 3Linked to original sources
PubMed1998

Determination of perfluorocarboxylic acids by gas-liquid chromatography in rat tissues.

A method for the determination of tissue perfluorocarboxylic acids (PFCA) was developed using gas-liquid chromatography with an electron capture detector (ECD). Perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), and perfluorodecanoic acid (PFDA) were efficiently extracted from rat liver, methylated with diazomethane, and separated on GLC. Internal standards that were added to liver homogenates were used in the quantitative analysis of PFCAs to correct the loss during the extraction and derivatization. The concentration of PFDA in rat liver 24 h after intraperitoneal administration at a dose of 20 mg/kg body weight was 113.9 +/- 11.4 microg/g liver. The value corresponds to the previously reported results that were obtained using [14C]PFDA.

Animals↗
PubMed1998

Leptomycin B inhibition of signal-mediated nuclear export by direct binding to CRM1.

Leptomycin B (LMB) is a Streptomyces metabolite that inhibits nuclear export of the human immunodeficiency virus type 1 regulatory protein Rev at low nanomolar concentrations. Recently, LMB was shown to inhibit the function of CRM1, a receptor for the nuclear export signal (NES). Here we show evidence that LMB binds directly to CRM1 and that CRM1 is essential for NES-dependent nuclear export of proteins in both yeast and mammalian cells. Binding experiments with a biotinylated derivative of LMB and a HeLa cell extract led to identifying CRM1 as a major protein that bound to the LMB derivative. Microinjection of a purified anti-human CRM1 antibody into the mammalian nucleus specifically inhibited nuclear export of NES-containing proteins, as did LMB. Consistent with this, CRM1 was found to interact with NES, when assayed with immobilized NES and HeLa cell extracts. This association was disrupted by adding LMB or purified anti-human CRM1 antibody. The inhibition of CRM1 by LMB was also observed in fission yeast. The fission yeast crm1 mutant was defective in the nuclear export of NES-fused proteins, but not in the import of nuclear localization signal (NLS)-fused proteins. Interestingly, a protein containing both NES and NLS, which is expected to shuttle between nucleus and cytoplasm, was highly accumulated in the nucleus of the crm1 mutant cells or of cells treated with LMB. These results strongly suggest that CRM1 is the target of LMB and is an essential factor for nuclear export of proteins in eukaryotes.

Biological Transport, Active↗
PubMed1998

Accumulation of various N-acylethanolamines including N-arachidonoylethanolamine (anandamide) in cadmium chloride-administered rat testis.

Changes in the levels of various molecular species of N-acylethanolamine in CdCl2-administered rat testis were examined. We found that the levels of various N-acylethanolamines including anandamide (N-arachidonoylethanolamine), an endogenous cannabinoid receptor ligand, were dramatically increased in CdCl2-admin-istered rat testis. Such changes were particularlyprominent for saturated and monoenoic species such as N-palmitoyl species (39-fold at 9 h) and N-stearoyl species (21-fold at 9 h), compared with unsaturated fatty acid-containing species such as anandamide (5-fold at 9 h). Noticeably, increased levels were observed of not only N-acylethanolamines but also several species of N-acylphosphatidylethanolamine, potential precursors for N-acylethanolamines. We confirmed that the rat testis microsomal fraction contains phosphodiesterase activity catalyzing the release of N-acylethanolamine from N-acylphosphatidylethanolamine and transacylase activity catalyzing the formation of N-acylphosphatidylethanolamine from phosphatidylethanolamine and phosphatidylcholine. These enzyme activities were not dramatically different in the microsomal fraction obtained from CdCl2-administered rat testis compared with that in the case of control rat testis, at least when estimated in cell-free assay systems, suggesting that the accessibility of the substrates to the enzymes may be increased in CdCl2-administered rat testis to generate a large amount of N-acylethanolamine. Possible pathophysiological implications of the augmented generation of N-acylethanolamine including anandamide in CdCl2-administered rat testis were discussed.

Animals↗
PubMed1998

Two testicular cDNA clones suppressed by gonadotropin stimulation exhibit ZP2- and ZP3-like structures in Japanese eel.

A single injection of human chorionic gonadotropin (HCG) can induce complete spermatogenesis in immature eel testes consisting of premitotic spermatogonia. To understand the regulatory mechanisms of spermatogenesis, we have applied a subtractive hybridization method to identify genes in which changes in expression occur after HCG treatment in vivo. The subtraction was carried out 24 hours after HCG injection. Two up-regulated and six down-regulated cDNA clones by HCG stimulation were isolated, and named eel spermatogenesis-related substance (eSRS) 1 to 8. In this paper, down-regulated cDNA clones of eSRS3 and eSRS4 were sequenced. A homology search showed that eSRS3 and eSRS4 have amino acid sequences similar to those of the ZP-domains of zona pellucida sperm-binding protein (ZP)-2 and 3, respectively. Transcripts of eSRS3 and eSRS4 have been detected only in immature testes and ovaries. Both transcripts disappeared immediately after HCG injection and were not detected in testes throughout the experimental period. To determine whether HCG action on down-regulation of eSRS3 and eSRS4 transcription is direct or mediated through 11-ketotestosterone (11-KT), a spermatogenesis-inducing steroid in eel, we investigated the effect of HCG and 11-KT on testicular eSRS3 and eSRS4 mRNA transcription in vitro. Northern blot analysis using poly(A)+ RNA extracted from cultured testis showed that both HCG and 11-KT suppressed the mRNA transcription of both eSRS3 and eSRS4. We speculate that eSRS3 and eSRS4 may play important roles in the prevention of spermatogenesis in the eel.

Amino Acid Sequence↗
PubMed1998

Development of the spatial pattern of 5-HT-induced locomotor rhythm in the lumbar spinal cord of rat fetuses in vitro.

Developmental changes in the 5-hydroxytryptamine (5-HT)-induced locomotor rhythm were examined in isolated spinal cord preparations taken from fetal rats at embryonic day (E) 16.5, E18.5 and E20.5. Motor activity was recorded from L2/L3 and L5 ventral roots. Bath application of 5-HT evoked rhythmic bursts that were synchronized in all ventral roots studied at E16.5. At E18.5, 5-HT evoked rhythmic bursts that alternated between the two sides, while the bursts in the L2/L3 ventral root were synchronous with those in the ipsilateral L5 ventral root. At E20.5, 5-HT evoked alternate rhythmic bursts in L2/L3 and L5 ventral roots, representing activity in flexors and extensors, respectively. In the presence of strychnine, 5-HT induced rhythmic bursts that were synchronized in all ventral roots studied at E18.5 and E20.5, suggesting that the change in the pattern of rhythmic motor activity that occurs with age is due to the development of glycine-mediated inhibition.

Animals↗
PubMed1998

Crystallization and preliminary X-ray diffraction studies of a rice cysteine proteinase inhibitor, Oryzacystatin-I.

Oryzacystatin-I from rice seeds was overexpressed in Escherichia coli, purified, and crystallized by the sitting-drop vapor diffusion method. Crystals obtained with 2-methyl-2,4-pentanediol as a precipitant exhibited space group I4122, with unit cell parameters of a = b = 100.0 A, c = 54.2 A, and diffracted up to 2.8 A resolution at 100 K. The crystals have one molecule per asymmetric unit.

Base Sequence↗
PubMed1997

Molecular cloning and cell cycle-dependent expression of mammalian CRM1, a protein involved in nuclear export of proteins.

Crm1 of Schizosaccharomyces pombe, a nuclear protein essential for proliferation and chromosome region maintenance, is a possible target of leptomycin B, an antifungal and antitumor antibiotic with cell cycle-arresting activity. cDNA encoding a human homolog of Crm1 was cloned. Human CRM1 (hCRM1) consisted of 1071 amino acids, of which the sequence showed 52% homology with S. pombe Crm1. hCRM1 weakly complemented the cold-sensitive mutation of S. pombe crm1-809, as did S. pombe crm1+. Overproduction of hCRM1 under the control of a series of nmt1 promoters suppressed cell proliferation in wild-type S. pombe in an expression level-dependent manner. A similar inhibitory effect was also observed for crm1+. Cells overproducing either hCRM1 or S. pombe Crm1 were distinctly larger than uninduced cells and contained compacted and fragmented nuclei. Furthermore, calcofluor staining demonstrated that most of these cells formed two septa per cell and accumulated a large amount of chitin or its related polysaccharides around the septa. Closely similar phenotypes between hCRM1- and S. pombe Crm1-induced cells indicate that the cloned cDNA encodes a functional homolog of S. pombe crm1+. Northern blot analyses with RNAs isolated from synchronized mammalian cells showed that the expression of mammalian CRM1 was initiated in late G1 and reached a peak at G2/M, although its protein level unchanged during the cell cycle. Transient expression of hCRM1 fused to the green fluorescent protein (GFP) in NIH3T3 cells showed that hCRM1 was localized preferentially in the nuclear envelope and was also detectable in the nucleoplasm and the cytoplasm. A crm1 mutation of S. pombe caused nuclear import of a GFP fusion protein containing a nuclear export signal but no change in the distribution of a GFP fusion protein containing a nuclear localization signal. All of these data suggest that CRM1 is a novel cell-cycle regulated gene that is essential for the nuclear export signal-dependent nuclear export of proteins.

3T3 Cells↗
PubMed1997

Effects of pyramidal tract stimulation on forelimb flexor motoneurons during fictive locomotion in cats.

To investigate spinal mechanisms of control of locomotion by the pyramidal system, we made intracellular recording from forelimb flexor motoneurons, and analyzed excitatory postsynaptic potentials (EPSPs) evoked by stimulation of the medullary pyramid during forelimb fictive locomotion in immobilized, decerebrate cats. We observed that (1) pyramidal stimulation evoked disynaptic EPSPs, (2) which were much bigger in the locomotor state than in the resting state, and that (3) the pyramidal EPSPs were rhythmically modulated, so that the facilitation occurred in the flexor-active phase. Last-order neurons mediating pyramidal effects presumably receive rhythmic input from the spinal locomotor network.

Animals↗
PubMed1997

Validity of electromyograms and tension as a means of motor unit number estimation.

The purpose of this study was to validate three different techniques for obtaining motor unit number estimates of the rat medial gastrocnemius muscle. These consisted of two electromyographic techniques using unprocessed and digitally averaged unitary muscle action potentials, and one mechanical technique. We also injected subunit B of cholera toxin into this muscle and counted the number of spinal motor neurons labeled by the toxin. Our results revealed that a motor unit number estimate obtained by using the unprocessed unitary muscle action potential was statistically different from the actual number of motor neurons. The other two motor unit number estimates, however, were not statistically different from the actual motor neuron number. These two methods thus seem more appropriate than the first electromyographic method for obtaining an accurate motor unit number estimate.

Animals↗
PubMed1997

Fish oil-feeding prevents perfluorooctanoic acid-induced fatty liver in mice.

The effects of perfluorooctanoic acid (PFOA) on the levels of lipids in liver and serum were compared between mice fed a diet supplemented with soy bean oil (SO), perilla oil (PO), or fish oil (FO) for 4 weeks. Hepatic content of triglyceride (TG) was significantly lower in the mice fed the FO diet than that in the mice fed either the SO or the PO diet. The treatment with PFOA caused a marked accumulation of TG in the livers of SO-fed and PO-fed mice (seven- and twofold over their respective controls), whereas a level of TG remained low in the mice fed the FO diet. Incorporation in vivo of [3H]glycerol revealed that FO-feeding reduced synthesis of TG in the liver. The administration of PFOA increased the incorporation of [3H]glycerol into hepatic phospholipid (PL) regardless of the dietary oil, while synthesis of hepatic TG from [3H]glycerol was not altered by the treatment with PFOA. Serum level of TG was reduced by the administration of PFOA to the mice fed either the SO diet or the PO diet, while no change in the level was observed in the mice fed the FO diet. These results suggest that the accumulation of TG in the livers of PFOA-treated mice is due to the inhibition of the secretion of TG into circulation. PFOA-induced hepatic accumulation of TG is prevented by the feeding of the FO diet which inhibits TG formation. Among three dietary oils, FO-feeding alone prevented the PFOA-caused accumulation of TG in the liver. The importance of docosahexaenoic acid (22:6(n - 3)) is discussed in relation to the prevention of fatty liver induced by chemicals.

Animals↗
PubMed1997

Development in neonatal rats of the sensory resetting of the locomotor rhythm induced by NMDA and 5-HT.

Developmental changes in the effects of quadriceps (Q) nerve stimulation on the locomotor rhythm induced by a mixture of N-methyl-D-aspartic acid and 5-hydroxytryptamine were examined using in vitro preparations from neonatal rats at postnatal days (P) 1-6. The effects of such stimulation on the rhythm were dependent both on stimulus strength and on the age of the animal. Low-intensity stimulation (< or =3.0 x T, where T=threshold for the monosynaptic reflex) during the flexor phase reset the rhythm via a prolongation of the flexor burst in most rats at P1-3, but via flexor burst truncation at P4-6. At any age, low-intensity stimulation during the extensor phase had no consistent effect on the ongoing rhythm. Activation of muscle afferents evoked via isometric contraction of the Q muscle caused effects similar to those obtained on low-intensity electrical stimulation in all age groups. In all age groups, high-intensity stimulation (> or =5.0 x T) caused resetting when delivered during the flexor phase via a prolongation of the flexor burst and during the extensor phase via a truncation of the extensor burst. These results suggest that the type of resetting evoked from low-threshold muscle afferents changes drastically during postnatal weekl, while effects evoked from high-threshold afferents remain unchanged.

Aging↗
PubMed1997

Urinary excretion of mefenamic acid and its metabolites including their esterglucuronides in preterm infants undergoing mefenamic acid therapy.

Urinary excretion of mefenamic acid (MA) and its two oxidative metabolites, M-I (3'-hydroxymethyl derivative) and M-II (3'-carboxyl derivative), and their glucuronides was investigated in preterm infants undergoing MA therapy. MA was given orally at a dose of 2 mg/kg and the dose was repeated every 24 h a maximum of three times. Urine was collected for up to 5 d after the last dose, and MA and the metabolites were determined by a newly developed HPLC. The cumulative amounts of MA and the metabolites excreted in the urine varied from 7 to 46% of the total dose administered, and were less than those reported in adults and children. Significant correlation was observed between the plasma half-life of MA and the cumulative amount of MA and the metabolites excreted in the urine. These results suggest that long plasma half-lives of MA observed in preterm infants are due mainly to low activity of drug metabolizing enzyme(s). In an infant who received the two regimens of MA therapy about 2 weeks apart, the plasma half-life of MA was shortened and the urinary excretion of the MA metabolites including their glucuronides was greatly increased during this period. It is suggested that the activities of both cytochrome P-450(s) and glucuronyltransferase(s) related to MA metabolism rapidly increased during the first month of the infant's life.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1997

Susceptibility of several species of Cyprinidae and Salmonidae freshwater fish to larval Gnathostoma nipponicum infection.

Susceptibility of five species of Cyprinidae and Salmonidae freshwater fish to the early third-stage larvae (EaL3) and advanced third-stage larvae (AdL3) of Gnathostoma nipponicum infection were examined. Two fish species inoculated orally with EaL3 were infected, and AdL3 were recovered from them with rate of 21.0% in Tribolodon hakonensis and 0.5% in Cyprinus carpio at 30 days postinoculation (PI). Attempts to infect five fish species with AdL3 were all successful. The recovery rate of AdL3 was 69.0% in T. hakonensis, 47.5% in Carassius auratus subsp., 35.0% in C. carpio, 53.0% in Oncorhynchus masou, and 32.0% in O. mykiss at 10 days PI. These results confirmed that the Cyprinidae and Salmonidae fish species reported here were susceptible to larval G. nipponicum infection and AdL3 had higher infectivity to them than the EaL3.

Animals↗
PubMed1997

Survey for Gnathostoma nipponicum larvae in gobiid freshwater fish and infectivity of the larvae to a gobiid fish (Chaenogobius urotaenia).

From June 1993 to November 1996, a total of 977 gobiid fish consisting of three species (Tridentiger brevispinis, Chaenogobius urotaenia, and Rhinogobiu brunneus) collected from eastern Aomori Prefecture, were examined for Gnathostoma nipponicum larvae infection. Only one species, C. urotaenia was infected with advanced third-stage larvae (AdL3), and a total of 22 larvae were recovered from 17 (3.4%) of 500 fish. The infected fish were larger than 12 cm in body length and collected in the May-June and September-November seasons. Experimental studies confirmed that C. urotaenia was susceptible to both the early third-stage larvae (EaL3) obtained from Eucyclops serrulatus and AdL3 from Misgurnus anguillicaudatus. Eight of 10 C. urotaenia inoculated orally with 10 EaL3 were positive, and 36 AdL3 were recovered from them (recovery rate: 36.0%) at 30 days postinoculation (PI). All 10 C. urotaenia inoculated with 10 AdL3 were also positive, and a total of 63 AdL3 were recovered (recovery rate: 63.0%) at 10 days PI. The main location of the larvae was the body muscles of the fish. No morphological alterations or death of the larvae were observed in this study. From these results, it seems that the C. urotaenia has characteristics suitable to be the host to the larvae and they may serve as the second intermediate and paratenic host in the natural life cycle of this nematode.

Animals↗
PubMed1996

Cadmium suppresses delta 9 desaturase activity in rat hepatocytes.

The effects of cadmium (Cd) on the fatty acid composition and delta 9 desaturation activity were studied in hepatocytes which had been cultured in serum-free medium. The aim of this study was to determine whether the inhibition of delta 9 desaturase seen in Cd-treated rat liver (Kudo et al. (1991) Toxicology, 68, 133-142) is the direct effect of Cd or not. When hepatocytes were cultured in serum-free medium, increases in the proportions of monounsaturated fatty acids such as oleic acid (18:1) were observed in the phospholipid (PL) and triacylglycerol (TG) fractions of hepatocytes, suggesting that delta 9 desaturase was induced. By contrast, essential fatty acids such as linoleic acid (18:2) and arachidonic acid (20:4) decreased. These changes were similar to those seen in the livers of essential fatty acid (EFA)-deficient rats. When hepatocytes were cultured in the presence of CdCl2, the changes in the fatty acid composition were suppressed. The effect of CdCl2 was concentration-dependent. The rate of the conversion of 18:0 to 18:1, which is catalyzed by delta 9 desaturase, was reduced by treatment with Cd, whereas the rate of conversion of 18:2 to 20:4, which is catalyzed by delta 6 desaturase, was slightly increased. Cd did not inhibit liver microsomal delta 9 desaturase activity in vitro. We concluded that Cd suppressed the induction of delta 9 desaturase in cultured hepatocytes. These phenomena can explain the selective inhibition of delta 9 desaturase in the livers of Cd-treated rats.

Animals↗
PubMed1996

Development of locomotor activity induced by NMDA receptor activation in the lumbar spinal cord of the rat fetus studied in vitro.

The development of neuronal circuits generating locomotor activity was characterized in an isolated lumbar spinal cord preparation from of fetal and neonatal rats. Locomotor activity induced by bath application of the NMDA receptor agonists, NMA and NMDA, was monitored from both sides of the corresponding lumbar ventral roots. Activation of NMDA receptors first evoked rhythmic motor activity at E15.5. NMA-induced rhythmic motor activity was not observed under synaptic blockade by TTX or cadmium ions, suggesting that this activity was evoked by synaptic drive from the interneuronal circuits in the spinal cord. At E15.5-E16.5, the rhythmic motor activity on both sides was synchronized. Phase relationship of the rhythmic motor activity between both sides was variable at E17.5-E19.5. The rhythmic motor activity was alternating on both sides at E20.5. Mid-sagittal splitting of the spinal cord did not affect the rhythm generation at all stages examined, suggesting the existence of independent rhythm-generating circuits on each side. The rhythmic motor activity in the presence of strychnine was synchronized on both sides at all stages examined. These results indicate that the changes in rhythm pattern are mediated by development of glycinergic inhibitory pathways, while the basic rhythm can be generated without the glycinergic inhibitory pathways.

Animals↗
PubMed1996

Spontaneous motoneuronal activity mediated by glycine and GABA in the spinal cord of rat fetuses in vitro.

1. Spontaneous motoneuronal activity was monitored from the lumbar ventral roots in an isolated spinal cord preparation from rat fetuses at embryonic days (E) 13.5-18.5. 2. Spontaneous bursts that were synchronized in both left and right ventral roots were observed periodically (mean interval, 1.5-2.6 min) from E14.5 to 17.5. This activity was abolished in Ca(2+)-free saline or by application of tetrodotoxin (1 microM), indicating that it was synaptically mediated. 3. The glutamate receptor blocker kynurenate (4 mM) failed to block spontaneous bursts at E14.5-15.5, though it completely abolished them at E17.5. The glycine receptor antagonist strychnine (10 microM) completely blocked spontaneous bursts at E14.5-15.5. Bicuculline, a GABAA receptor antagonist, reduced the amplitude of the spontaneous bursts. 4. At E15.5, a brief application of glycine (250 microM to 2 mM) evoked excitatory responses resembling the spontaneous bursts in both time course and amplitude. Such glycine-induced responses were not observed under Ca(2+)-free conditions, suggesting that they were synaptically evoked. These synaptic responses were not blocked by kynurenate (4 mM), but they were abolished by strychnine (10 microM). 5. It is concluded that glycine and GABA generate the earliest spontaneous motor activity of the fetus and function transiently as excitatory transmitters in the embryonic spinal cord.

Animals↗