Yasufumi Kataoka

Time course of nitric oxide synthase activity in neuronal, glial, and endothelial cells of rat striatum following focal cerebral ischemia.

Summary1. The time course of nitric oxide synthase (NOS) activity in neuronal, endothelial, and glial cells in the rat striatum after middle cerebral artery (MCA) occlusion and reperfusion was examined using a histochemical NADPH-diaphorase staining method.2. In sham-operated rats, neuronal cells of the striatum exhibited strong NADPH-diaphorase activities. When rats were subjected to MCA occlusion for 1 hr, neuronal damage, including neurons with positive NADPH-diaphorase activities, appeared in the striatum at 3 hr after and extended to all areas of the striatum 3–4 days after reperfusion.3. NADPH-diaphorase activities in the endothelial cells increased in the damaged part of striatum from 3 hr after, peaked at 1–2 days after MCA occlusion/reperfusion, then gradually decreased.4. In parallel with the development of neuronal damage, some astrocytes and a high proportion of microglia/macrophages located in the perisite and in the center of the damaged striatum, respectively, exhibited a moderate to high level of NADPH-diaphorase activities. Most of these activities disappeared at 4 days after MCA occlusion.5. These findings provided evidence that an inappropriate activation of NOS in endothelial cells and microglia/macrophages, in response to MCA occlusion/reperfusion, is closely associated with initiation and progression of ischemic neuronal injury in the striatum.

Involvement of the dorsal hippocampus in mediation of the antianxiety action of tandospirone, a 5-hydroxytryptamine1A agonistic anxiolytic

The effect of tandospirone, a 5-hydroxytryptamine (5-HT)1A agonist/anxiolytic, injected directly into dorsal hippocampus, on Vogel-type conflict behavior in rats was investigated and the findings were compared with the effects of diazepam and zopiclone. Tandospirone (30 micrograms/2 microliters and 60 micrograms/2 microliters) and diazepam (40 micrograms/2 microliters) but not zopiclone (20 micrograms/2 microliters), produced a potent anticonflict action in rats. The anticonflict action of tandospirone (30 micrograms/2 microliters), injected into the dorsal hippocampus, was significantly blocked by (-)-propranolol (5 mg/kg i.p.). The present findings provide evidence that suggests that tandospirone has an antianxiety action, presumably by stimulating 5-HT1A receptors in the dorsal hippocampus.

Increased production of endothelins in the hippocampus of stroke-prone spontaneously hypertensive rats following transient forebrain ischemia: Histochemical evidence

Summary1.The effect of transient forebrain ischemia on endothelin-1 (ET-1) and endothelin-3 (ET-3) production in the hippocampus of stroke-prone spontaneously hypertensive rats (SHRSPs) was investigated using immunohistochemical techniques.2.In SHRSPs subjected to 10-min bilateral carotid occlusion, neuronal degeneration in the CA1 pyramidal cell layer of the hippocampus was detectable at 4 days and remarkable at 7 days after reperfusion.3.Coinciding with neuronal degeneration, ET-1- and ET-3-like immunoreactivities were intense in the CA1 pyramidal-cell layer, the stratum lacunosum moleculare, and the CA4 subfield of the hippocampus. Almost all of the immunostained cells had morphological characteristics of astrocytes.4.The possibility that ET has a role in the development of neuronal cell death following transient forebrain ischemia warrants further attention.

Conflict situation increases serotonin release in rat dorsal hippocampus : in vivo study with microdialysis and Vogel test

The release of serotonin (5-hydroxytryptamine, 5-HT) in the dorsal hippocampus was measured using an in vivo microdialysis method in rats subjected to the Vogel type conflict test. The conflict situation significantly increased 5-HT release in the dorsal hippocampus. Midazolam (0.75 and 1.5 mg/kg i.p.) suppressed the dosage-dependently this increased 5-HT release, an inhibition closely associated with the attenuation of conflict behavior. These findings suggest that the activation of serotonergic neuronal activity in the dorsal hippocampus is linked to mediation of anxiety-related behavior.

Endothelin-3 stimulates the release of catecholamine from cortical and striatal slices of the rat

Endothelin-3 (ET-3) evoked the release of dopamine/noradrenaline from cortical slices and dopamine from striatal slices in a concentration-dependent manner. This action peaked slowly and was long-lasting in real-time monitoring, being different from the high K(+)-evoked response. The striatal response to 10 microM of ET-3 was reduced by extracellular Ca2+ depletion to 40% of control and by Ca2+ antagonists, especially nifedipine and flunarizine, to 40% of control. Our findings suggest that ET has a physiological significance in the brain as a neuromodulator for catecholaminergic transmission.

Activation of Protein Kinase C Suppresses the γ‐Aminobutyric AcidB Receptor‐Mediated Inhibition of the Vesicular Release of Noradrenaline and Acetylcholine

Abstract: Modulation of the γ‐aminobutyric acidB (GABAB) receptor‐mediated response by protein kinase C (PKC) was examined with regard to inhibition by stimulation of the GABAB receptor of stimulation‐evoked release of noradrenaline (NA) from slices of cerebellar cortex and of acetylcholine (ACh) from strips of ileum. 12‐O‐Tetradecanoylphorbol 13‐acetate (TPA) potentiated the high K+‐evoked Ca2+‐dependent release of NA and ACh, but not the ouabain‐evoked release, even in the presence of external Ca2+. The potentiating effect was antagonized by sphingosine, thereby suggesting that PKC participates in the exocytotic‐vesicular release of neurotransmitters, but does not do so in case of a nonvesicular release. GABA inhibited the high K+‐evoked release of NA and ACh, but not the ouabain‐evoked Ca2+‐independent release. The effect of GABA was mimicked by baclofen and was antagonized by phaclofen, thereby suggesting that stimulation of the GABAB receptor inhibits the vesicular but not the nonvesicular release of neurotransmitters. TPA suppressed the GABAB receptor‐mediated inhibition of high K+‐evoked release of NA and ACh. The effect of TPA was antagonized by sphingosine. These results indicate that stimulation of the GABAB receptor inhibits the stimulation‐evoked Ca2+‐dependent release of neurotransmitters and that activation of PKC suppresses the GABAB receptor‐mediated response.

Involvement of glial endothelin/nitric oxide in delayed neuronal death of rat hippocampus after transient forebrain ischemia.

Abstract1. We examined time- and cell-type-dependent changes in endothelin (ET)-1-like immunoreactivity, ET receptors binding and nitric oxide (NO) synthase (NOS) activity in CA1 subfields of the hippocampus of stroke-prone spontaneously hypertensive rats subjected to a 10-min bilateral carotid occlusion and reperfusion.2. Microglia aggregated in accord with neuronal death and expressed a high density of ETB receptors and an intense NOS activity in the damaged CA1 pyramidal cell layer, 7 days after the induced transient forebrain ischemia. The increased NOS activity and ETB receptor in microglia disappeared 28 days after this transient ischemia.3. In contrast to microglia, astrocytes presented a moderate level of ET-1-like immunoreactivity, ETB receptors, and NOS activity in all areas of the damaged CA1 subfields, 7 days after the ischemia. These events were further enhanced 28 days after the ischemia.4. In light of these findings, the possibility that the microglial and the astrocytic ETB/NO system largely contributes to development of the neuronal death and to reconstitution of the damaged neuronal tissue, respectively, in the hippocampus subjected to a transient forebrain ischemia would have to be considered.

Specific binding sites for 125I-endothelin-1 in the porcine and human spinal cord.

Specific binding sites for 125I-endothelin-1 (125I-ET-1) in the spinal cord were investigated using quantitative receptor autoradiographic and chemical cross-linking methods. The binding sites were highly concentrated in porcine and human spinal cord areas corresponding anatomically to the dorsal horn (Rexeds laminae I-III), an area around the central canal (lamina X) and the principal part of the intermediolateral nucleus (IMLp). The localization of the binding sites differed from those of 125I-omega-conotoxin GVIA (125I-CgTx) and 125I-Bolton-Hunter substance P (125I-BH-SP), with the exception that the IMLp shared 125I-ET-1 with 125I-CgTx and 125I-BH-SP binding sites. Specific 125I-ET-1 binding sites in the areas examined were characteristically single and of high affinity. There were no differences between the potencies of unlabeled ET family peptides, ET-1, ET-2, ET-3 and sarafotoxin S6b at inhibiting 125I-ET-1 binding to the areas. Chemical cross-linking studies showed that 125I-ET-1 and 125I-ET-3 mainly bound to a protein with molecular mass of 43 kDa in the porcine and human thoracic spinal cord membranes. The present finding shows the neuronal significance of this newly discovered peptide in the spinal cord.

Localization and Characterization of Endothelin Receptors in Human Gliomas: A Growth Factor?

Localization and characterization of endothelin receptors in surgical specimens of human gliomas (6 benign astrocytomas and 7 glioblastomas multiforme) and in normal human cortices were studied using quantitative receptor autoradiographic methods. Low numbers of [125I]endothelin-1 [( 125I]ET-1) binding sites were detected in the gray matter of the human frontal cortex, with little binding in the white matter. Conversely, relatively high numbers of [125I]ET-1 binding sites were homogeneously present in tissue sections derived from astrocytomas, whereas higher numbers of [125I]ET-1 binding sites were heterogeneously located on groups of cells with a pseudopalisading appearance and pleomorphic astrocytes in glioblastoma multiforme. Necrotic areas within the tissue sections derived from glioblastoma were devoid of binding. Binding of [125I]ET-1 to gliomas and normal gray matter was specific. Unlabeled ET-1 and its natural analogs (ET-2 and ET-3) inhibited the binding of [125I]ET-1 to these lesions in a concentration-dependent manner and with similar high potencies. Possibly related substances, such as ion channel regulators (omega-conotoxin, apamin, and tetrodotoxin), a Ca2+ channel blocker (nicardipine), and growth factors (epidermal growth factor and insulin-like growth factor I), did not affect the binding to tissue sections derived from gliomas or from normal frontal cortices. Scatchard analysis revealed the presence of a single class and high-affinity binding sites for endothelin in normal cortex and in gliomas. There was no significant difference in the binding affinities: dissociation constants (Kd) were 2.1 +/- 0.5 nM in 6 astrocytomas, 2.5 +/- 0.4 nM in 7 glioblastomas, and 1.4 and 1.5 nM in two normal cortices.(ABSTRACT TRUNCATED AT 250 WORDS)

Endothelin-triggered brain damage under hypoglycemia evidenced by real-time monitoring of dopamine release from rat striatal slices

The role of endothelin in the pathogenesis of hypoglycemic brain damage in rats was evaluated using an in vitro model with which we could directly monitor the release of dopamine from striatal slices. There was no evidence of impairment in case of non-exposure of the slices to endothelin during 20-40 min of hypoglycemia. The response all but disappeared in striatal slices stimulated with endothelin 10(-5) M twice during 20 min of hypoglycemia. Endothelin-triggered hypoglycemic damage was not observed in the absence of extracellular Ca2+ or in the presence of nifedipine 10(-6) M. Our findings provide strong evidence that endothelin is one etiological factor in the development of hypoglycemic/ischemic brain injury, as a result of interaction with specific receptors which activate the voltage-sensitive Ca2+ channel.