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Biomedical subjects

H Takagi

Publications and source records attributed to H Takagi.

At least 1,009 records · Page 56Linked to original sources
PubMed1987

A Met-enkephalin releaser (kyotorphin)-induced release of plasma membrane-bound Ca2+ from rat brain synaptosomes.

The neuropeptide kyotorphin (Tyr-Arg) released 45Ca2+ from synaptosomal membrane preparations of the rat lower brainstem. This dipeptide also decreased plasma membrane-bound Ca2+ ([Ca2+]m), determined using chlorotetracycline, in the slice and synaptosome of the rat lower brainstem. Taken together with other data indicating that kyotorphin induces an increase in synaptosomal intracellular Ca2+ ([Ca2+]i) in a Ca2+ channel-independent manner, it is suggested that kyotorphin may mobilize the [Ca2+]m and thus increase the [Ca2+]i in the nerve terminals of the rat lower brainstem.

Analgesics↗
PubMed1987

Kyotorphin (tyrosine-arginine) synthetase in rat brain synaptosomes.

Kyotorphin (Tyr-Arg) is a unique neuropeptide which produces analgesia by releasing Met-enkephalin from slices of the brain and spinal cord. Recent studies revealed that kyotorphin possesses the properties of neurotransmitter/neuroregulator. In the present study, we identified a kyotorphin synthetase in the soluble fraction of rat brain synaptosomes (synaptosol) and characterized it. The enzyme partially purified with Sephacryl S-300 showed an absolute requirement for ATP, MgCl2, tyrosine, and arginine. The optimal pH was 7.5-9.0 and the pI was determined to be 6.1-6.2 by isoelectric focusing. The Km was 25.6 microM for tyrosine, 926 microM for arginine, 294 microM for ATP, and 442 microM for MgCl2. The Vmax was 34.0 pmol/mg of protein/h. The apparent molecular size of this "kyotorphin synthetase" further purified by the DE52 column was 240,000-245,000 daltons, estimated using TSKgel G4000SW column chromatography. The enzyme reaction is represented by the following equation: Tyr + Arg + ATP + MgCl2 + kyotorphin synthetase----Tyr-Arg (kyotorphin) + AMP + PPi + MgCl2 + kyotorphin synthetase. The regional distribution and subcellular localization of the synthetase showed a close correlation to that of kyotorphin levels in the rat brain. The amounts of kyotorphin formed from amino acids by the synthetase in the dialyzed synaptosol was 3.0-4.0 times higher than that from precursor proteins by processing enzymes within the 30 min incubation.

Analgesics↗
PubMed1987

The specific opioid kappa-agonist U-50,488H inhibits low Km GTPase.

DAGO, a specific opioid mu-agonist stimulated the low Km GTPase activity of membranes of the guinea-pig striatum at 1-1000 nM and this effect was antagonized by naloxone 10-100 nM. On the other hand, U-50,488H, a specific opioid kappa-agonist inhibited the low Km GTPase activity of membranes of the guinea-pig cerebellum at 1-100 nM. As this effect was antagonized by MR2266 but not by naloxone, the findings suggest that opioid mu- and kappa-receptor stimulation is probably linked to the stimulation and inhibition of low Km GTPase, respectively.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗
PubMed1987

Requirement of pro-sequence for the production of active subtilisin E in Escherichia coli.

Subtilisin E, an alkaline serine protease of Bacillus subtilis 168, is first produced as a precursor, pre-pro-subtilisin, which consists of a signal peptide for protein secretion (pre-sequence) and a peptide extension of 77 amino acid residues (pro-sequence) between the signal peptide and mature subtilisin. When the entire coding region for pre-pro-subtilisin E was cloned into an Escherichia coli expression vector, active mature subtilisin E was secreted into the periplasmic space. When the pre-sequence was replaced with the E. coli OmpA signal peptide, active subtilisin E was also produced. When the OmpA signal peptide was directly fused to the mature subtilisin sequence, no protease activity was detected, although this product had the identical primary structure as subtilisin E as a result of cleavage of the OmpA signal peptide and was produced at a level of approximately 10% of total cellular protein. When the OmpA signal peptide was fused to the 15th or 44th amino acid residue from the amino terminus of the pro-sequence, active subtilisin was also not produced. These results indicate that the pro-sequence of pre-pro-subtilisin plays an important role in the formation of enzymatically active subtilisin. It is proposed that the pro-sequence is essential for guiding appropriate folding of the enzymatically active conformation of subtilisin E.

Amino Acid Sequence↗
PubMed1987

Purified opioid mu-receptor is of a different molecular size than delta- and kappa-receptors.

Opioid mu-receptors were solubilized from rat brains with 0.5% Triton X-100 in the presence of 0.32 M sucrose and then purified with 6-succinyl morphine-Affi-Gel 102 column. Purified materials showed a major band with mol.wt. 58,000 Da and a minor band with mol.wt. 41,000 Da. Affinity cross linking with [3H]DAGO, a selective mu-opioid agonist, to the neural membranes and purified materials revealed that opioid mu-binding protein has a mol.wt. of 58,000 Da, a value totally different from that of delta- (mol.wt. 18,500 Da) and kappa- (mol.wt. 36,000 Da) binding protein.

Animals↗
PubMed1987

Subconvulsive doses of intracisternal bicuculline methiodide, a GABAA receptor antagonist, produce potent analgesia as measured in the tail pinch test in mice.

Bicuculline methiodide, a GABAA antagonist produced potent analgesia in the tail pinch test when it was given intracisternally (i.c.) but not intrathecally (i.t.). The ED50 was 5 ng/mouse. This analgesia was antagonized by i.c. muscimol, a GABAA agonist. On the contrary, muscimol (i.t.) produced bicuculline-reversible analgesia. These findings suggest that brain GABA may transmit the nociceptive information while GABA in the spinal cord may inhibit it.

Analgesics↗
PubMed1987

Release of substance P from the spinal dorsal horn is enhanced in polyarthritic rats.

Using in situ spinal dorsal horn perfusing method in decerebrated rats, we measured the release of immunoreactive substance P (iSP) in polyarthritic rats. The spontaneous release of iSP from the dorsal horn in the polyarthritic rats was significantly accelerated over that in control non-inflamed rats. Passive movements of the inflamed ankle joint evoked a significant increase in the release of iSP, while a similar movement of the non-inflamed joint led to no such increase. These results suggest that the facilitated release of iSP from the primary afferent terminals in the spinal dorsal horn in polyarthritic rats possibly plays a role in transmission of chronic pain of inflamed joints.

Animals↗
PubMed1987

Calcitonin gene-related peptide promotes mechanical nociception by potentiating release of substance P from the spinal dorsal horn in rats.

In vitro superfusion with capsaicin (5 X 10(-7) M) of slices of the dorsal half of the rat spinal cord produced a significant increase in a release of immunoreactive substance P (iSP). Calcitonin gene-related peptide (CGRP: 10(-6) M) significantly potentiated the capsaicin-induced release of iSP. On the other hand, when CGRP (5 nmol/rat) was intrathecally injected, the peptide produced a significant hyperalgesia to mechanical noxious stimuli (pinching the hind paw), but aversive responses and potentiation of substance P-induced aversive responses were never observed. These findings suggest that in the rat spinal dorsal horn, CGRP potentiates the release of substance P from the primary afferent terminal and promotes the transmission of nociceptive information induced by mechanical noxious stimuli.

Animals↗
PubMed1987

A novel proenkephalin processing carboxypeptidase and its activation by cyclic AMP dependent protein kinase.

Carboxypeptidase B-like enzymes cleaving Met-enkephalin-Arg from synaptosomes of the rat striatum purified using a DEAE-cellulose column and Met-Arg-CH-Sepharose 4B affinity column proved to be different from enkephalin-convertase, lysosomal carboxypeptidase B-like enzyme, pancreas carboxypeptidase B and carboxypeptidase Y, in effects of inhibitors and activators, pH optimum (7.5-8.5) and molecular size (50,000). This enzyme, named "Processin CP-E" was activated by cAMP dependent protein kinase, and the Vmax was increased from 4.3 to 13.3 microM/min/mg protein, while the Km (28.2 microM) was unchanged.

Animals↗
PubMed1987

Nigrostriatal dopamine neurons receive substance P-ergic inputs in the substantia nigra: application of the immunoelectron microscopic mirror technique to fluorescent double-staining for transmitter-specific projections.

A direct synaptic contact between nigrostriatal dopamine neurons and substance P axons in the substantia nigra was demonstrated using the immunoelectron microscopic mirror technique combined with the fluorescent double-staining method for transmitter-specific projections. Substance P-immunoreactive terminals were found to make synaptic contact with nigral cells exhibiting tyrosine hydroxylase immunoreactivity and retrograde fluorescent labeling following injection of biotinylated lectin into the neostriatum. It appears that substance P afferents directly affect nigrostriatal dopamine neurons in the substantia nigra via the synaptic contacts.

Animals↗
PubMed1987

Intra-arterial long term infusion chemotherapy via transverse cervical artery for intra-abdominal malignancies.

An original method of intra-arterial catheter placement for long term infusion chemotherapy, using a transverse cervical artery, is described. Twelve patients with far advanced intra-abdominal malignancies, which precluded abdominal explorations, were catheterized and treated for prolonged periods. All patients were ambulated during the infusion chemotherapy and there were few catheter associated vascular complications. This method is a simple and safe procedure and seems superior to the conventional method using axillary or femoral arteries.

Abdominal Neoplasms↗
PubMed1987

A stomach oncofetal antigen recognized by monoclonal antibody GC302.

A monoclonal antibody, GC302, was established by fusing murine myeloma NS/1 cells with the splenocytes of a BALB/c mouse immunized with a human gastric cancer cell line, NU-GC-3. The serological specificity of GC302 was analyzed by an anti-mouse Ig mixed-hemadsorption (MHA) test on a panel of human cell lines, and an immunoperoxidase method using the frozen sections of tumors and normal tissues of adult and fetus. GC302 reacted with cancers of the stomach and colorectum but did not react with hepatocellular carcinomas, melanomas, or astrocytomas in the MHA tests. By the immunoperoxidase method, GC302 was found not to react with normal adult gastric mucosa, but to react with the mucosa in the fetal stomach, intestinal metaplasia, and almost all of the cancer of the stomach. GC302 also reacted with the normal mucosa of the intestine, colon, and rectum as well as with cancers of these origins. In normal liver sections, the antibody reacted with the bile ducts, but not with the hepatic cells. These results indicate that the antigen detected by GC302 is characterized as an oncofetal antigen in the stomach, and also as a differentiation antigen whose localization discriminates between the gastrointestinal tracts of the forgut origin and those of the midgut and hindgut origin. The molecular weight of the GC302 antigen was estimated to be ca. 40,000 by the Western blot analysis. Periodic acid treatment on the antigen suggested that the antigenic determinant is a carbohydrate.

Animals↗
PubMed1987

A novel method for the synthesis of kyotorphin, Tyr-Arg, and 3H-Tyr-Arg, catalyzed by tyrosyl-tRNA synthetase from Bacillus stearothermophilus.

A novel method of dipeptide synthesis is described that can be carried out in aqueous solution and does not require complicated protecting and deprotecting procedures. An analgesic neuropeptide named kyotorphin, H-Tyr-Arg-OH, was synthesized from unprotected tyrosine and arginine in a new enzymatic reaction catalyzed by immobilized tyrosyl-tRNA synthetase from Bacillus stearothermophilus. The reaction could be a useful tool in the syntheses of radioisotope-labeled oligopeptides to be used in receptor binding assays. 3H-Kyotorphin was prepared by this method at a yield of 72% and could be used in receptor binding assays after a single chromatographic separation.

Amino Acyl-tRNA Synthetases↗
PubMed1987

Hormonal status of gastric cancer. II. Abnormal constitution of urinary steroids in gastric cancer patients.

We undertook a case-control study regarding the excretions of 14 urinary steroids in gastric cancer (GC) patients. The results are as follows: the levels of androgens, progestins, and two corticosteroids were, relative to tetrahydrocortisol, significantly depressed in GC patients of both sexes compared with the corresponding normal controls. The deviation profile of urinary steroids was not affected by radical gastrectomy. Evidence indicated that observed changes of GC urines were the steroidal expression of a decrease of endogenous testosterone combined with an increase of endogenous hydrocortisone; there was also evidence that the hormonal environment of our GC patients was endocrinologically homologous to that of rice-fed or salty rice-fed mice. Epidemiological inquiry revealed that GC patients having more access to rice-rich or salt-rich diets were taller and less obese than were rural healthy controls. In agreement with the anthropometric data of those cancer patients is the finding that the specific death rate of GC (as calculated for each of 15 prefectures of northern Japan) was positively correlated for each sex with the mean heights, but not with the mean weights, of 14-year-old youths of those areas. This paper discusses the possible relevancy of the hormonal and epidemiological aspects of GC patients to gastrocarcinogenesis in light of steroid physiology.

Adult↗
PubMed1987

Possible mechanism involved in the inhibitory action of U-50, 488H, an opioid kappa agonist, on guinea pig hippocampal CA3 pyramidal neurons in vitro.

Intracellular recordings of the CA3 pyramidal neurons in the guinea pig hippocampus were made in vitro. U-50 488H (100 microM), a selective opioid kappa agonist, decreased the synaptic response produced by stimulation of the mossy fibers but did not affect the membrane potential, the input resistance and the generation of the Na+ spikes or the Ca2+ spikes. Iontophoretically applied U-50 488H depressed the depolarization produced by L-glutamate. U-50 488H (100 microM) also depressed the consistent depolarization produced by veratrine (3 X 10(-5) g/ml) and this effect was partially antagonized by naloxone. Moreover, application of U-50 488H led to a disappearance of the anomalous rectification. These results suggest that U-50 488H depresses the synaptic activities of the CA3 pyramidal neurons by inhibiting a subtype of the Na+ channel, "Na+ channel type II" which slowly closes, of the soma and/or the dendrites of the neurons.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗