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Biomedical subjects

H Hatanaka

Publications and source records attributed to H Hatanaka.

At least 199 records · Page 11Linked to original sources
PubMed1992

Interleukin-6 and leukemia inhibitory factor promote the survival of acetylcholinesterase-positive neurons in culture from embryonic rat spinal cord.

Interleukin-6 (IL-6) and leukemia inhibitory factor (LIF) promoted the survival of acetylcholinesterase (AChE)-positive neurons in culture from embryonic E15 rat spinal cord. Half of the AChE-positive neurons died during 3-7 days in culture in the absence of IL-6 and LIF. However, IL-6 at a concentration of 5 ng/ml completely prevented the death of AChE-positive neurons. LIF at a concentration of 5 U/ml also stimulated the survival of neurons, although to a lesser extent than IL-6. IL-6 and LIF also increased the numbers of process-bearing neuron-like cells in culture. The dose-dependencies of IL-6 and LIF with regard to the survival of total neuron-like cells were different from those for AChE-positive neurons.

Acetylcholinesterase↗
PubMed1992

Interleukin-6 as a neurotrophic factor for promoting the survival of cultured catecholaminergic neurons in a chemically defined medium from fetal and postnatal rat midbrains.

Interleukin-6 (IL-6, human recombinant) promoted the survival of catecholaminergic neurons from fetal and postnatal rat midbrains as assessed by an immunohistochemical staining for tyrosine hydroxylase (TH) in culture using a chemically defined medium. The maximal dose of IL-6 for the cell survival of postnatal P15 rat mesencephalic TH-positive neurons in culture for 7 days was 50 ng/ml. The survival-promoting effects on P15 cultures were observed both in high- and low-density cultures. The survival effect of IL-6 on the cultured P15 TH-positive neurons was significant for only 4-15 days in vitro. However, the viable number of TH-positive neurons with IL-6 was less than that of the control at early points in the culture process (1-2 days in vitro). Continuous presentation of IL-6 was required for promoting survival. The optimal dose of IL-6 for the survival of fetal E16 midbrain TH-positive neurons was 5 ng/ml, and the survival promoting effect was less than that for the P15 cultures. The maximal dose of IL-6 for the survival of P2 TH-positive neurons was 5 ng/ml and that of P8 was 50 ng/ml, indicating that the response of rat mesencephalic TH-positive neurons to IL-6 changes during the first postnatal week.

Animals↗
PubMed1992

Neurotrophic effect of brain-derived neurotrophic factor on basal forebrain cholinergic neurons in culture from postnatal rats.

We examined the effect of brain-derived neurotrophic factor (BDNF) on cholinergic neurons in culture from postnatal rat basal forebrain by assay of choline acetyltransferase (ChAT) activity and cytochemical staining for acetylcholinesterase (AChE). BDNF was found to increase the ChAT activities but failed to promote the survival of AChE-positive neurons in cultures from neonatal (P3) rats, suggesting that its main role is cholinergic differentiation. In contrast, an enhancement of the survival of AChE-positive neurons and of ChAT activity was observed in cultures from P15-16 rats, suggesting that BDNF's main action is the maintenance of cholinergic neurons. Our results indicate a similarity between BDNF and nerve growth factor effects on the responses of cholinergic neurons of postnatal rat basal forebrain in culture.

Animals↗
PubMed1992

Nerve growth factor (NGF)-mediated up-regulation of low-affinity NGF receptor gene expression in cultured basal forebrain cholinergic neurons from postnatal 3-day-old rats.

We examined the effect of nerve growth factor (NGF) on the expression of low-affinity NGF receptor (LNGFR) in cultured P3 basal forebrain cholinergic neurons, on which NGF acts to promote differentiation. Based on the results of the RT-PCR (reverse transcriptase-polymerase chain reaction) method, over a 2-fold increase in the LNGFR mRNA level was found in NGF-treated cultures compared with control cultures at one day after the addition of 100 ng/ml NGF. This increase was maintained for up to 3 days after the addition of NGF. The increase in LNGFR mRNA was found even at 1 ng/ml NGF (= 40 pM), indicating that the up-regulation of the LNGFR mRNA level in cultured cholinergic neurons occurred at an NGF concentration near the Kd value for the high-affinity NGF receptor. In addition, immunohistochemical staining showed stronger staining with a monoclonal anti-LNGFR antibody of the cholinergic neurons in NGF-treated cultures than those in control cultures. Our results suggest that NGF can up-regulate LNGFR expression at the mRNA and protein levels in cultured P3 basal forebrain cholinergic neurons and that this mechanism may play an important role in potentiating the effect of NGF on these neurons.

Acetylcholine↗
PubMed1992

Culture of neuronal cells from postnatal rat brain: application to the study of neurotrophic factors.

1. The authors developed a primary culture technique for neuronal cells from postnatal rat brains and studied the effects of neurotrophic factors on the naturally developed neurons. 2. We demonstrated changes in the neurotrophic role of nerve growth factor (NGF) during the developmental stages of the rat: NGF was shown to act as a differentiation factor in the early stages and as a survival factor later. 3. It appeared that interleukin-6 (IL-6) supported the survival of septal cholinergic neurons obtained from 10-day-old rats. IL-6, however, did not induce the differentiation of embryonic rat septal cholinergic neurons. IL-6 improved the survival of mesencephalic catecholaminergic neurons from postnatal and embryonic rat brains, which have known not to be response to NGF.

Animals↗
PubMed1991

Expression and localization of smg p25A (= rab3A) in cultured rat hippocampal cells.

We have studied the expression of smg p25A and synaptophysin in cultured hippocampal neurons isolated from 5-day-old rat brain by an immunocytochemical technique. In a dispersed cell culture seeded on astrocyte monolayers, well-branching neurite proliferation was observed along with age in culture. The synaptophysin immunoreactivity was present in the neuronal cell bodies and neurites at 1 and 5 days in vitro (DIV) and was eventually localized to discrete areas along neurites at 15 DIV while the immunoreactivity in cell bodies became less prominent. On the other hand, the smg p25A immunoreactivity was observed in the neuronal cell bodies and neurites at 1 through 15 DIV. The immunoreactivity for smg p25A or synaptophysin was not observed in astrocytes and this finding was confirmed by an immunoblot analysis. These results indicate that smg p25A as well as synaptophysin is present exclusively in neurons and suggest that these two synapse-associated proteins have different sites of function and different kinetics of synthesis, transport, and/or turnover in cultured hippocampal neurons.

Animals↗
PubMed1991

Expression of a hybrid Cu/Zn-type superoxide dismutase which has high affinity for heparin-like proteoglycans on vascular endothelial cells.

Since plasma levels of enzymes, such as superoxide dismutase (SOD), that scavenge reactive oxygen species are low, surface membranes of endothelial and parenchymal cells of various tissues are often exposed to oxidative stress. To dismutase superoxide radicals efficiently in and around vascular endothelial cells, we constructed a fusion gene encoding a hybrid SOD (HB-SOD) consisting of human Cu/Zn-SOD and a C-terminal basic peptide that binds to heparin-like proteoglycans. The fusion gene was expressed in yeast, and the resulting HB-SOD was highly purified. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis, HB-SOD revealed a protein band with an apparent molecular weight of 20,000. HB-SOD bound to endothelial cells of aortic segments by a mechanism which was inhibited by heparin but not by antithrombin III. When injected intravenously to rats, 125I-labeled HB-SOD rapidly disappeared from the circulation; the rate of disappearance was decreased by heparin. Less than 1% of the injected HB-SOD was found in the urine 20 min after administration at which time more than 70% of SOD was excreted in its intact form. Immunohistochemical studies revealed that HB-SOD predominantly bound to heparin-like proteoglycans on endothelial cells of the artery and other tissues. HB-SOD might permit studies on pathophysiological roles of superoxide radicals in and around vascular endothelial cells in vivo.

Amino Acid Sequence↗
PubMed1991

Interleukin-6 improves the survival of mesencephalic catecholaminergic and septal cholinergic neurons from postnatal, two-week-old rats in cultures.

Interleukin-6 (human recombinant) supported the survival of cultured mesencephalic, catecholaminergic and septal cholinergic neurons from postnatal, two-week-old (P13-P15) rats. Significantly, more catecholaminergic neurons, stained by monoclonal anti-tyrosine hydroxylase antibody, were found in cultures supplemented with interleukin-6 at a concentration of 5 ng/ml than in cultures not treated with interleukin-6. The optimal dose used was 50 ng/ml. The survival effect of interleukin-6 on postnatal rat, tyrosine hydroxylase-positive neurons was observed both in cultures using serum-containing and serum-free medium. Contents of dopamine and noradrenaline in cultures with interleukin-6 were also larger than in control cultures. Interleukin-6 also increased the survival of cultured embryonic (E17) rat midbrain tyrosine hydroxylase-positive neurons. The effect on these neurons was, however, smaller, and the optimal dose of interleukin-6 was nearly 5 ng/ml. Interleukin-6 also supported the survival of cultured postnatal (P13) rat septal cholinergic neurons, visualized by acetylcholinesterase staining. The concomitant addition of mouse nerve growth factor (100 ng/ml) and interleukin-6 (50 ng/ml) had a synergetic effect on the survival of acetylcholinesterase-positive neurons in culture. Our data suggest that the survival of cultured tyrosine hydroxylase-positive, mesencephalic, and acetylcholinesterase-positive, septal neurons from postnatal two-week-old rats was supported by interleukin-6, just as there was a different dose dependency of interleukin-6 on the cultured postnatal neurons compared with embryonic neurons.

Animals↗
PubMed1991

Possible alteration of the blood-brain barrier by boron-neutron capture therapy.

In the course of re-assessment of boron-neutron capture therapy (BNCT) for malignant brain tumors, fractionation of neutron irradiation has been proposed. The authors have used BNCT with a single fraction technique during the past 21 years and now decided to study some effects of fractionation. Twenty-two healthy mouse brains were irradiated with thermal neutrons after boron-10 injection (mercaptoundecahydrododecaborate). A second dose of boron-10 was administered and its uptake in the boron-neutron-capture-irradiated brains was determined. A tendency towards increased boron uptake in the moderately BNCT-treated brains was noticed, which may result in increased brain damage if fractionated neutron irradiation is used.

Animals↗
PubMed1990

High oxygen atmosphere for neuronal cell culture with nerve growth factor. I. Primary culture of basal forebrain cholinergic neurons from fetal and postnatal rats.

Cholinergic neurons cultured from postnatal days 11-13 (P11-P13) rat basal forebrain showed better survival in the culture condition using a 50% O2 atmosphere with and without nerve growth factor (NGF) than in a low (10 or 20%) O2 atmosphere. Except for the culture at a low cell density, the beneficial effect of the highly oxidized culture condition was found in the culture from P3 neurons, but not from embryonic day 18 neurons. The survival of microtubule-associated protein 2 (MAP2)-positive neurons in culture from P3 basal forebrain regions was more enhanced in a 50% O2 atmosphere than in 20% and also 10% O2 atmosphere. The viable number of the MAP2-positive neurons in a 10% O2 condition was about half of that in a 20% condition. These results suggest that the response of the cultured neurons to an incubator O2 concentration changes during the neuronal development in CNS from fetal to postnatal stages.

Acetylcholine↗
PubMed1990

High oxygen atmosphere for neuronal cell culture with nerve growth factor. II. Survival and growth of clonal rat pheochromocytoma PC12h cells.

When clonal rat pheochromocytoma PC12h cells were cultured in a 50% O2 atmosphere, cells gradually died during the cultivation. On the other hand, the addition of NGF at the final concentration of 50 ng/ml could rescue the cells from death. The culture in a 40% O2 atmosphere had little effect on the growth of PC12h cells, as compared with the culture in a normal 20% O2 condition. A very high O2 concentration, as 60%, caused severe damage to PC12h cell growth, and the restoration of cell growth by NGF seemed to be insufficient. PC12h cells were fully differentiated and extended dense long neurites by NGF even in a 50% O2 atmosphere. However, the neurite extension in the culture in a 60% O2 atmosphere was suppressed. The cell-saving effect of NGF on cell death in culture under a 50% O2 atmosphere was dose-dependent, and the ED50 value of NGF was 5 ng/ml. Basic fibroblast growth factor and epidermal growth factor also had a potent effect to rescue the cell death in the high O2 culture, but insulin had no effect. Since the differentiation effects of NGF on PC12h cells are thought to offer a model system to investigate the effect of NGF on neurons, the present observations suggest that a protection machinery for high O2 toxicity to neurons may exist in the neuronal differentiated PC12h cells by NGF, but not in the undifferentiated cells.

Animals↗
PubMed1990

Nerve growth factor rapidly induces expression of the 68-kDa neurofilament gene by posttranscriptional modification in PC12h-R cells.

We have isolated a subclonal cell line of PC12, PC12h-R, which is characterized by its rapid response to nerve growth factor (NGF) in extending its neurites. The mRNA levels of the 68-kDa neurofilament (NF-L) increased severalfold as early as 3 h after the addition of NGF, but this was not accompanied by an increase in the transcription rate. Analysis of the stabilities of 3.5- and 2.3-kilobase (kb) NF-L mRNAs using actinomycin D revealed that 2.3-kb mRNA was degraded much faster than the 3.5-kb species and that NGF apparently stabilized NF-L mRNAs. These results show that the NF-L gene can be regulated posttranscriptionally, a finding that may be important in responding to the acute demand for neurofilament syntheses.

Adrenal Gland Neoplasms↗
PubMed1990

Monitoring of acetylcholine released from postnatal rat basal forebrain cholinergic neurons cultured on membrane filter by cell bed perfusion system and HPLC-ECD.

A method for the determination of release of endogenous neurotransmitter from cultured neurons has been developed. Basal forebrain cholinergic neurons from postnatal rats were cultured on polyethyleneimine-coated membrane filter with nerve growth factor. The membrane filter, on which the neurons lived and extended neurites, was packed in a cell bed chamber for a closed perfusion. The perfusate was fractionated and the content of acetylcholine released from cultured neurons was measured by high-performance liquid chromatography-electrochemical detection (HPLC-ECD). Acetylcholine release evoked by glutamate and high K+ was 1.5- and 3-fold greater than the spontaneous level. This method for the determination of the neurotransmitter released from cultured neurons is very advantageous for investigating regulation of transmitter release in central neurons.

Acetylcholine↗