Biomedical subjects

A Takanaka

Publications and source records attributed to A Takanaka.

At least 19 recordsLinked to original sources

PubMed1992

Culture of postimplantation rat embryos in rabbit serum for the identification of the growth factor in fractionated rat serum.

The growth factor for postimplantation rat embryos was investigated on the basis of the serum species-specificity in supporting embryonic development in culture. We used rabbit serum as a basal medium for the culture of head-fold stage rat embryos, and examined the effects of various fractions of rat serum on their development. In rabbit serum alone, rat embryos developed poorly. With the rat serum ultrafiltrate of molecular weight (MW) < 300,000, embryonic development improved, but not with the ultrafiltrate of MW < 100,000. With dialyzed rat serum or the globulin fraction of rat serum, embryonic development improved, but the albumin fraction had no effect. It was concluded from these results that some macromolecular growth factor for cultured postimplantation rat embryos was present in the globulin fraction of rat serum. The molecular weight of this growth factor was estimated to be between 65,000 and 300,000. Rabbit serum was considered to be suitable as a medium for the identification of this growth factor.

Animals↗

PubMed1992

Dopamine receptor agonists and antagonists enhance ATP-activated currents.

The effects of dopamine and related compounds on ATP-activated channels were investigated in pheochromocytoma PC12 cells. Dopamine (10 microM) enhanced an inward current activated by 100 microM ATP. A similar enhancement of the ATP-activated current was observed with apomorphine (10 microM), a non-selective dopamine receptor agonist, with (+)-SKF-38393 (10 microM), a selective dopamine D1 receptor agonist, and with (-)-quinpirole (10 microM), a selective dopamine D2 receptor agonist. Moreover, (+)-SCH-23390 (30 microM), a dopamine D1 receptor antagonist, and (-)-sulpiride (30 microM), a dopamine D2 receptor antagonist, also enhanced the ATP-activated current. The results suggest that ATP-activated channels are modulated by dopaminergic mechanisms, and that this modulation cannot be attributed to any single class of dopamine receptors.

Adenosine Triphosphate↗

PubMed1992

Extracellular adenosine 5'-triphosphate-evoked glutamate release in cultured hippocampal neurons.

Characteristics of extracellular ATP-evoked electrical responses in rat hippocampal neurons were investigated. Extracellular ATP (100 microM) induced a rapid depolarization followed by repetitive firings of spikes in these cells under whole-cell current-clamp. In whole-cell voltage-clamp experiments, ATP activated 2 types of inward currents that were inhibited by P2-purinoceptor blocker suramin (300 microM). One is a small (about -20 pA) sustained current which is insensitive to tetrodotoxin (TTX), and the other is a large (-100 to -300 pA) transient current which abolished in the presence of 3 microM TTX. The ATP-induced transient current was blocked by 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX; 30 microM), a non-N-methyl-D-aspartate (non-NMDA) receptor antagonist. ATP failed to induce the transient current in the cell which showed the desensitization to quisqualic acid (QA; 10 microM), a non-NMDA receptor agonist. These findings suggest that ATP directly activates small sustained currents, and indirectly induces the transient currents by evoking glutamate release.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed1992

Neurochemical and histological analysis of motor dysfunction observed in rats with methylnitrosourea-induced experimental cerebellar hypoplasia.

Histological and neurochemical changes related to motor dysfunction observed in rats after neonatal treatment with nitrosoureas were examined. Neonatal rats received subcutaneous injections of methylnitrosourea (MNU: 0.125 mmol/kg, s.c.) or ethylnitrosourea (ENU: 0.25 mmol/kg, s.c.) daily at 4,5,6 and 7 days post partum, a period of cerebellar granule cell, stellate cell and basket cell formation. At 14 days and 45 days after birth, MNU-treated rats displayed a lowering in motor coordination skills measured by tests of retainment ability on a rod of 26 mm diameter, chinning-climbing ability on parallel rods or retainment ability on a rotating rod. Histological examination at 14 days after birth showed a cerebellar hypoplasia with reduced cellularity of the internal granule cell layer and a disperse disposition of Purkinje cells in the granule cell layer. Cerebellar growth and cerebellar content and concentration of DNA were remarkably reduced in the MNU-treated rat. The degree of the reduction in cerebellar content of glutamic acid paralleled the degree of the cerebellar hypoplasia at 14 and 45 days after birth. In contrast, the concentrations of gamma-aminobutyric acid, acetylcholine, 5-hydroxytryptamine and norepinephrine were significantly increased by MNU treatment. ENU treatment control did not exert any significant changes in the neurotransmitters and motor coordination. These results suggest that the motor dysfunctions observed in MNU treated rats are induced by unbalanced output activities from Purkinje cells to motor neurons.

Acetylcholine↗

PubMed1992

Effect of tetrakis-mu-3,5-diisopropylsalicylatodiaquodicopper(II) on the status of reduced glutathione in freshly isolated hepatocytes.

Effects of different concentrations of tetrakis-mu-3,5-diisopropylsalicylatodiaquodicopper(II) (Cu(II)2(3,5-DIPS)4(H2O2)2) on the reduced status of glutathione (GSH), the major nonprotein thiol in tissues, were investigated using freshly isolated hepatocytes. Cu(II)2(3,5-DIPS)4 below 100 microM did not have any significant effects on either the GSH content or viability of the hepatocytes, but at 150-250 microM it decreased both parameters after 1 h of incubation. The decrease in cellular GSH was not followed by an increase in the oxidized form of GSH (GSSG) in the cell suspension. The addition of deferoxamine with Cu(II)2(3,5-DIPS)4 to the hepatocyte suspension prevented depletion in GSH content and loss of cell viability by Cu(II)2(3,5-DIPS)4. Both GSH depletion and loss of cell viability were found to be Cu(II)2(3,5-DIPS)4 dose dependent. From these results, it appears that Cu(II)2(3,5-DIPS)4 penetrated the cell membrane and acted by decreasing the GSH level by forming a copper-glutathione complex.

Animals↗

PubMed1992

N-nitrosodialkylamine dealkylation in reconstituted systems containing cytochrome P-450 purified from phenobarbital- and beta-naphthoflavone-treated rats.

Five cytochrome P-450 forms were purified from livers of rats pretreated with phenobarbital (PB) or beta-naphthoflavone (BNF), and the oxidative dealkylation of N-nitrosodialkylamines by the reconstituted cytochrome P-450 systems was measured. PB-II (P450IIB1) showed very high N-nitrosomethybutylamine (NMBA) debutylase activity, high NMBA demethylase activity and high N-nitrosomethyl-benzylamine (NMBeA) debenzylase activity, suggesting that the increase following PB treatment in hepatic microsomal NMBA debutylation and NMBeA debenzylation was due to the induction of PB-II. BNF-H (P450IA2) showed very high NMBA debutylase and high NMBeA debenzylase activities, and BNF-L (P450IA1) showed NMBA debutylase and high NMBeA debenzylase activities. These results suggested that the increase by BNF pretreatment in hepatic microsomal NMBA debutylation was due mainly to the induction of BNF-H and in some part to that of BNF-L. PB-II also showed very high dealkylation activity of lipophilic N-nitrosodialkylamines with long alkyl moieties. On the other hand, BNF-H dealkylated N-nitrosodipropylamine (NDPA), N-nitrosomethylbutylamine (NMBA) and N-nitrosoethylbutylamine (NEBA) at higher rates than N-nitrosodibutylamine (NDBA). BNF-L dealkylated NEBA at higher rates than NMBeA and NDBA. These results reveal that substrate specificity of each cytochrome P-450 form in N-nitrosodialkylamine metabolism is different from each other and several forms of cytochrome P-450 support each N-nitrosamine dealkylase activity in mammalians.

Animals↗

PubMed1992

Indium inhibits gap junctional communication between rat hepatocytes in primary culture.

The effect of indium on gap junctional communication was investigated in primary cultured rat hepatocytes. Treatment of hepatocytes with indium chloride at concentrations of 100 microM to 1 mM for 2 h resulted in dose-dependent inhibition of gap junctional communication between hepatocytes. The effect of indium on hepatocytes was also evaluated using two indices for cell viability: lactate dehydrogenase (LDH) leakage and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction. Indium did not cause any increase in LDH leakage from hepatocytes at the above concentrations, but inhibition of MTT reduction was observed at concentrations above 500 microM. These results suggest that the gap junctions between hepatocytes may be vulnerable sites to indium toxicity.

Animals↗

PubMed1992

[Studies on the teratogenic potential of p-tert-butylphenolformaldehyde resin in rats].

p-tert-Butylphenolformaldehyde resin, an adhesive, was given orally to pregnant Wistar rats by stomach intubation at the dose levels of 250, 500 and 1000 mg/kg body weight during days 7 to 17 of gestation, and the effects of the compound on dams and fetal developments were examined. No changes in general conditions, maternal body weight, food consumption, numbers of corpora lutea and implantation ratio were observed. There was no evidence of an increase in fetal death or of malformation attributable to the treatment with p-tert-butylphenolformaldehyde resin in any of dose levels examined. It was concluded that p-tert-butylphenolformaldehyde resin has no teratogenic effect in rats.

Administration, Oral↗

PubMed1991

Inhibition of Ca-channels by diazepam compared with that by nicardipine in pheochromocytoma PC12 cells.

The effects of diazepam on voltage-gated Ca channels were studied in PC12 pheochromocytoma cells using whole-cell voltage-clamp techniques. An inward current activated by a depolarizing voltage step to +10 mV from a holding potential of -60 mV in 10.8 mM Ba was larger than that activated in 10.8 mM Ca. The Ba current was completely blocked by a low concentration of Cd (30 microM) and was also sensitive to nicardipine (100 nM to 10 microM). Diazepam (1-100 microM) inhibited the Ba current in a concentration-dependent manner. Neither diazepam nor nicardipine affected the current-voltage relationship or the dependence on holding potentials of the Ba current. Both slightly accelerated the inactivation time course of the Ba current. When diazepam was applied to the cells in combination with nicardipine, the observed inhibition agreed with a value predicted assuming independent blockade by diazepam and by nicardipine. These results suggest that diazepam inhibits Ca channels in a manner similar to nicardipine, but that the binding sites for diazepam are different from those for nicardipine.

Animals↗

PubMed1991

Effects of ATP antagonists on purinoceptor-operated inward currents in rat phaeochromocytoma cells.

The effects of suramin, reactive blue 2 (RB2) and d-tubocurarine (d-TC) were investigated electrophysiologically to elucidate the mechanisms underlying their antagonism of P2 purinoceptor-mediated responses. All three compounds inhibited an adenosine triphosphate (ATP)-activated inward current in rat phaeochromocytoma PC12 cells in a concentration-dependent manner. The order of potency was RB2 greater than suramin greater than d-TC. The inhibition induced by suramin or RB2 was reversible, whereas that induced by d-TC was not reversed after a 5-min rinse. The inactivation of the ATP-activated current was accelerated by d-TC but not by suramin or RB2. RB2 administered simultaneously with ATP exerted much weaker inhibition compared to that induced by prior administration, suggesting that RB2 is a slowly acting antagonist. This was not observed for suramin or d-TC. Suramin and RB2 caused a parallel shift in the concentration/response curve for the ATP-activated current. With d-TC the maximal response of ATP was decreased but the concentration producing half-maximal response was unchanged. The voltage dependency of the ATP-activated current showed less inward rectification in the presence of d-TC. Suramin or RB2 did not affect the voltage dependency. These results suggest that suramin and RB2 reversibly block binding of ATP to receptors, whereas d-TC blocks ion permeability through the ATP-activated channel.

Adenosine Triphosphate↗

PubMed1991

Selective and competitive antagonism by suramin of ATP-stimulated catecholamine-secretion from PC12 phaeochromocytoma cells.

1. Suramin, a putative P2-antagonist, (10 to 300 microM) inhibited the adenosine 5'-triphosphate (ATP)-stimulated secretion of [3H]-noradrenaline or endogenous dopamine from phaeochromocytoma PC12 cells in a concentration-dependent manner. Suramin (300 microM) did not affect the dopamine-secretion stimulated by high K+ or nicotine. 2. Suramin shifted the concentration-response curve for ATP to the right. The antagonism was competitive with a pA2 value of 4.52. 3. ATP also stimulated an increase in intracellular Ca2+ concentration as determined by fura-2 methods. Suramin antagonized this effect over the same concentration range that antagonized the ATP-stimulated catecholamine secretion. 4. These results suggest that suramin can be used as a selective and competitive antagonist of ATP in experiments concerning mechanisms of catecholamine-secretion.

Adenosine Triphosphate↗

PubMed1991

Antagonism by reactive blue 2 but not by brilliant blue G of extracellular ATP-evoked responses in PC12 phaeochromocytoma cells.

1. The effects of reactive blue 2 and brilliant blue G, which have been shown to block extracellular ATP-evoked responses, were investigated to discover whether these compounds act as P2-purinoceptor antagonists in PC12 phaeochromocytoma cells. 2. Reactive blue 2 (10 to 100 microM) suppressed the ATP-stimulated dopamine secretion from PC12 cells in a dose-dependent manner. The concentration-response curve for ATP was shifted to the right and the maximal response was decreased by reactive blue (30 and 100 microM). Brilliant blue G (up to 100 microM) did not significantly affect the secretion. 3. Reactive blue 2 (10 to 100 microM) suppressed the ATP-activated inward current recorded from the voltage-clamped cells in a concentration-dependent manner. Brilliant blue G (up to 100 microM) did not affect the current. 4. The results suggest that reactive blue 2 but not brilliant blue G is a P2-purinoceptor antagonist in PC12 cells. The purinoceptors in these cells may be the same type as those involved in ATP-evoked smooth muscle relaxation, judging from the antagonism by reactive blue 2.

Adenosine Triphosphate↗

PubMed1991

Comparison of adenosine triphosphate- and nicotine-activated inward currents in rat phaeochromocytoma cells.

1. The adenosine triphosphate (ATP)-activated inward current was compared to the nicotine-activated inward current in nerve growth factor (NGF)-treated rat phaeochromocytoma PC12 cells. 2. Both ATP and nicotine activated an inward current at negative holding potentials. The concentration of ATP necessary to activate the inward current was about 10-fold higher than that of nicotine; the EC50 was 20.5 microM for ATP and 2.4 microM for nicotine. The maximal responses induced by ATP and nicotine were almost identical in the same cells. The current-voltage relationship for the ATP-activated current was very similar to that for the nicotine-activated current, and both currents reversed around 0 mV in a physiological saline. 3. The ATP-activated current and the nicotine-activated current were not additive; the current activated by a combined administration of ATP (100 microM) and nicotine (10 microM) was only about 20% larger than the current activated by either ATP or nicotine alone. Nicotine (100 microM) did not increase the current activated by 1 microM-ATP. 4. ATP could activate an inward current in the cells even after desensitization to nicotine had developed. 5. Hexamethonium (100 microM) selectively blocked the nicotine-activated current whereas suramin (100 microM), a purinoceptor antagonist, selectively blocked the ATP-activated current. 6. Ionic selectivity was studied by changing compositions of extracellular solutions. When external Na+ was replaced with Cs+, both ATP and nicotine activated inward currents. However, with an extracellular solution containing Tris or glucosamine as a major cation, only ATP, not nicotine, activated an inward current. 7. ATP- and nicotine-activated currents were also recorded from cells bathed in a solution containing 1.8 mM-Ca2+ as the only external cation, suggesting that both pathways are Ca2+ permeable. 8. The results suggest that the ATP-sensitive ionic pathway is not independent of the nicotine-sensitive pathway in these cells. Our working hypothesis is that ATP and nicotine activate the same channels but the binding sites and the open-states of the channels are different between these two agonists.

Adenosine Triphosphate↗

PubMed1991

Inhibition of ion channels by hirsutine in rat pheochromocytoma cells.

Effects of hirsutine, an alkaloid that produces a potent ganglion blocking effect, were investigated using rat pheochromocytoma PC12 cells. Hirsutine (1 to 10 microM) suppressed dopamine-release evoked by 100 microM nicotine. In voltage-clamped cells, hirsutine (1 to 10 microM) inhibited the inward current activated by 100 microM nicotine. Hirsutine was equipotent to hexamethonium in blocking the nicotine-activated current. The voltage-dependency of the nicotine activated current was not modified by hirsutine. Effects of hirustine on other ion channels were tested to determine its selectivity. Inward currents mediated through ATP-activated channels were scarcely affected by hirsutine (up to 100 microM). However, hirustine (10 microM) inhibited Ba currents passing through Ca channels and K currents activated by depolarizing voltage steps. The results suggest that hirsutine potently blocks nicotinic receptor-channels, but hirsutine also inhibits voltage-gated Ca and K channels. Roles of the inhibition of these channels in the pharmacological effects of hirsutine were discussed.

Adrenal Gland Neoplasms↗

PubMed1991

[Studies on the uptake of 8-anilino-1-naphthalene sulfonate by rat freshly isolated renal cells].

The addition of 8-anilino-1-naphthalene sulfonate (ANS) into rat renal cell suspension caused a rapid increase in fluorescence (Ex. 400 nm, Em. 470 nm). The increase in fluorescence seemed to be composed of two phases. When the cells were disrupted by ultrasonic wave, the fast phase (0-ca 20 sec) increased and the slow phase (after ca 40 sec) disappeared. The rate of increase in the slow phase was dependent on the concentration of ANS and on the ambient temperature. A double reciprocal plot of the rate and the concentration exhibited straight line. It was decreased by bromophenol blue and rose bengal but not by other organic anions like p-aminohippuric acid and several metabolic inhibitors. It seemed that the uptake of ANS into the renal cell is mediated by a carrier which is different from that for p-aminohippuric acid.

Anilino Naphthalenesulfonates↗

PubMed1991

[Studies on the teratogenic potential of 2,2'-isobutylidene-bis (4,6-dimethylphenol) in rats].

2,2'-Isobutylidene-bis(4,6-dimethylphenol), an antioxidant, was given orally to pregnant Wistar rats by stomach intubation at the dose levels of 5, 15 or 45 mg/kg body weight during days 7 to 17 of gestation, and the effects of the compound on dams and fetal developments were examined. In the dams at the two higher dose levels of 15 and 45 mg/kg, toxic signs (tremor, startle reflex, salivation, involuntary urination, wheezing and nostril discharge) were observed. Moreover, at the highest dose level, additional toxic signs (lacrimation and vaginal bleeding), suppression in maternal body weight gain and food consumption were observed. However, there was no evidence of an increase in malformations attributable to the treatment with 2,2'-isobutylidene-bis(4,6-dimethyl-phenol) in any of the treated groups. It was concluded that 2,2'-isobutylidene-bis(4,6-dimethylphenol) has no teratogenic effect in rats, though toxic signs were observed in treated dams of the 15 and 45 mg/kg groups.

Administration, Oral↗

PubMed1991

Inhibitory action of peripheral-type benzodiazepines on dopamine release from PC12 pheochromocytoma cells.

Characteristics of the benzodiazepine inhibition of dopamine (DA) release in PC12 cells were investigated. Diazepam inhibited DA release evoked by high concentrations of extracellular K+ in a dose-dependent manner (IC50, 10 microM). Ro 5-4864 [7-chloro-1,3-dihydro-1-methyl-5-(p-chlorophenyl)-2H-1,4-benzodiazepine- 2-one], a peripheral-type benzodiazepine, also inhibited DA release effectively. PK 11195 [1-(2-chlorophenyl)-N-methyl-N-(1-methyl-propyl)-3-isoquinoline carboxamide], a benzodiazepine generally considered a peripheral-type benzodiazepine receptor antagonist, did not antagonize the inhibition induced by diazepam, but rather inhibited DA release itself. On the other hand, the central-type benzodiazepines, clonazepam and Ro 15-1788 (ethyl-8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5a] [1,4]benzodiazepine-3-carboxylate) did not affect the DA release. Diazepam, Ro 5-4864 and PK 11195 also inhibited a Ba(++)-current carried by voltage-gated Ca++ channels, and diazepam suppressed an increase in intracellular Ca++ evoked by 80 mM extracellular K+ as measured by the fura-2 method. These results suggest that the inhibitory action of diazepam and other benzodiazepines on DA release from PC12 cells may be mediated through one type of peripheral-type benzodiazepine receptors which are coupled to voltage-gated Ca++ channels and that these receptors may not necessarily be the same as those in other tissues.

Animals↗

PubMed1990

ATP-activated single-channel currents recorded from cell-free patches of pheochromocytoma PC12 cells.

Single-channel recordings were made using cell-free membrane patches (outside-out configuration) isolated from pheochromocytoma PC12 cells. ATP (50 microM) activated single channel currents in the isolated patches and the currents inactivated with a half-decay time of about 5s. The single channel conductance was about 13 pS in external solution with 140 mM Na. The amplitudes of the single-channel currents were decreased when external Ca was increased from 1.8 to 16.2 mM, suggesting that Ca blocks ion permeation through the channel. These properties of single-channel currents may underlie those of the macroscopic current.

Adenosine Triphosphate↗