This study is to investigate the mechanism of inhibitory effect of imipramine on the calcium utilization in single cells isolated from canine detrusor. 2 mm thick smooth muscle chops were incubated in 0.12% collagenase solution at 36degreeC, and aerated with 95% O2/5% CO2, and then cell suspension was examined Acetylcholine (ACh) evoked a concentration-dependent contraction of the isolated detrusor cells in normal physiologic salt solution (PSS), and the ACh-induced contraction was significantly inhibited by imipramine. In Ca2+-free PSS, ACh-induced contraction was less than those in normal PSS and it was not affected by the pretreatment with imipramine. Ca2+-induced contraction in Ca2+-free PSS was supressed by imipramine, but addition of A 23187, a calcium ionophore, overcomed the inhibitory effect of imipramine. High potassium-depolarization (40 mM KCl) evoked cell contraction, which was inhibited by imipramine. Caffeine, a releasing agent of the stored Ca2+ from sarcoplasmic reticulum, evoked a contraction of the cells that was not blocked by the pretreatment with imipramine. These results suggest that imipramine inhibits the influx of calcium in the detrusor cells through both the receptor-operated- and voltage-gated-calcium channels, but does not affect the release of calcium from intracellular storage site.
Acetylcholine ; Caffeine ; Calcimycin ; Calcium* ; Collagenases ; Imipramine* ; Muscle, Smooth ; Sarcoplasmic Reticulum

OBJECTIVE: The oligosaccharide moieties of glycoproteins and proteoglycans have a vital function in blastocyst differentiation. Concanavalin (ConA), a lectin, is known to bind on the preimplantation embryos, especially on blastocyst. In this study, we investigated whether ConA can modulate the trophoblast development and about the regulating mediator. Also, we investigated whether expansion is enough for hatching procession of the mouse blastocyst. METHOD: Embryos were collected at 72 h post hCG injection and chemicals were treated after 24 h (96 hr post hCG injection). ConA or calcium ionophore A23187 were exposed to blastocyst and than analysis the developmental process for 48 hr. Intracellular free-Ca2+ concentration in trophectoderm was measured with confocal laser microscope after exposing to ConA or calcium ionophore A23187. ConA-pretreated blastocyst exposed to the calcium ionophore A23187 and then analyzed the developmental process. Otherwise ouabain was treated to the blastocyst to block the Na+/K+-ATPase activity. RESULTS: In contrast to the control blastocyst, the ConA-exposed blastocysts developed beyond the expansion stage with significantly high rate (90.4%) at 12 h post administration. ConA induced an increase the intracellular Ca2+ concentration in trophectoderm. Calcium ionophore A23187 also stimulated expansion of blastocyst. Most of the control blastocysts developed to the hatching stage at 144 h post hCG injection. However, strongly 65% of the ConA-exposed embryos were arrested at expanded stage at same time point. The developmental progression rates to hatching stage of both ConA- and calcium ionophore A23187-expose blastocysts were significantly lower than that of the control. However ConA-pretreated embryos developed to the hatching stage like control embryos. Ouabain showed a tendency to delayed the progress to expansion stage but did not inhibit the development to the hatching stage. CONCLUSION: ConA-mediated expansion is the result of the increase of intracellular free-calcium in blastocyst stage embryo. It is suspected that expansion of the blasocyst is a essential indirect factor in hatching and the calcium may triggering the cellular mechanisms for the both expansion and hatching progression.
Animals ; Blastocyst* ; Calcimycin ; Calcium* ; Concanavalin A* ; Embryonic Structures ; Glycoproteins ; Mice* ; Ouabain ; Proteoglycans ; Trophoblasts

Tri- and tetra-cyclic antidepressants are known to cause dry mouth among other several major complications. The present study was designed to compare the degree of reduced salivation due to antidepressants and to explore whether intracellular calcium-increasing agents ameliorate the salivation. Effects of antidepressants and agents increasing intracellular calcium on the cholinergic submandibular secretion and blood flow induced by the chorda stimulation or intra-arterial acetylcholine were observed in anesthetized cats. Effects of antidepressants and calcium-mobilizing agents on K+ efflux were also observed in excised gland slices. The results obtained were as follows: 1. Salivary secretion in response to the chorda stimulation (3 V, 20 Hz, 1 msec) was significantly attenuated by antidepressants in a dose-dependent manner, whereas the blood flow was not affected. 2. Salivary secretion and increased blood flow evoked by intra-arterial acetylcholine (20 microgram/kg) were markedly diminished by antidepressants, the magnitude of which was amitryptyline>imipramine >mianserin in order. 3. Cholinergic salivation was significantly decrease by cyclopiazonic acid, a calcium pump inhibitor of the endoplasmic reticulum, or by BAPTA/AM, a specific intracellular calcium chelator. 4. Caffeine and ryanodine potentiated the cholinergic salivation and ameliorated the depressed salivary secreation due to antidepressants. 5. Calcium ionophore A 23187 ameliorated the depressed salivation due to antidepressants. 6. Antidepressants inhibited the K+ efflux, which were restored by caffeine or A 23187. These results suggest that the depressed salivary secreation due to antidepressants is ameliorated by increasing intracellular calcium levels.
Acetylcholine ; Animals ; Antidepressive Agents ; Caffeine ; Calcimycin ; Calcium* ; Cats ; Endoplasmic Reticulum ; Mouth ; Ryanodine ; Salivation*

Cultured cortical primary astroglia treated with zinc died while rapidly detached from culture plates, a distinct part of zinc-treated astroglia. In the present study, we investigated the mechanism underlying the rapid change in the morphologic integrity of zinc-treated astroglia. Among the early cellular events occurring in zinc-treated astroglia, strong activation of p38 MAPK and JNK was evident. Although inhibitors of p38 (SB203580 and SB202190) or JNK (SP600125) did not protect zinc-insulted astroglia from cell death, the p38 inhibitors, but not the JNK inhibitor, suppressed actin filament and cell morphology disruption. The Ca2+ ionophore, A23187, also suppressed actin filament and cell morphology disruption, but not cell death, of zinc-insulted astroglia. However, A23187 did not inhibit p38 MAPK activation in zinc-treated astroglia. Together these results suggest that zinc influx in astroglia results in rapid loss of the morphologic integrity via mechanisms regulated by p38 kinase and/or Ca2+ signaling.
Actin Cytoskeleton ; Astrocytes ; Calcimycin ; Cell Death ; p38 Mitogen-Activated Protein Kinases ; Phosphotransferases ; Zinc

OBJECTIVE: To evaluate the basophil histamine releasability in response to IgE- and non- IgE-mediated stimuli in children with atopic asthma. Met: Basophil histamine releasability was measured in Dermatophagoides farinae (D. farinae)-sensitive atopic asthmatics, D.farinae-sensitive healthy atopics, non-atopic asthmatics, and healthy non-atopics. Basophils were stimulated with D.farinae, goat antihuman IgE antibody, formyl-Met-Leu-Phe(fMLP), and Calcium ionophore A23187. Histamine was measured by automated fluorometric technique. RESULTS: Sponianeous histamine release was higher in atopic asthmatics compared to healthy non-atopics. Histamine release by D.farinae and by anti-IgE antibody was higher in atopic asthmatics compared to the other groups. There was no difference in histamine release by fMLP among all groups. Histamine release by Calcium ionophore was higher in healthy atopics and non-atopic asthmatics compared to healthy non-atopics. The atopics showed correlation between histamine release by D.farinae, by anti-IgE antibody and total serum IgE levels. CONCLUSIONS: Spontaneous and IgE-mediated histamine release were related to the presence of both atopy and asthma, whereas non-IgE mediated histamine release was different depending on the stimuli.
Asthma* ; Basophils* ; Calcimycin ; Calcium ; Child* ; Dermatophagoides farinae ; Goats ; Histamine Release ; Histamine* ; Humans ; Immunoglobulin E

It has been implicated that leukotrienes play roles in the pathogenesis of gastritis and gastric ulceration associated with Helicobacter pylori (H. pylori). Rebamipide is being used as an antiulcer drug but it's mechanism of action has not been understood well. One possible mechanism of action of this drug is to inhibit the cellular release of leukotrienes by various stimuli, particularly H. pylori. In the present study, attempts were made to test this possibility and the results are as follows. When Kato III cells (gastric adenoma cells) were stimulated by H. pylori, leukotriene D4 (LTD4) was released and rebamipide inhibited this release dose-dependently. Similar experiment was performed on neutrophils because the infilteration of neutrophils is a common phenomenon in H. pylori-infected gasrtric tissues. Neutrophils released LTD4 when these cells were stimulated by H. pylori and rebamipide also inhibited this release. Furthermore, rebamipide inhibited the release of LTD from neutrophils induced by calcium ionophore A23187 and arachidonic acid. The results suggest that rebamipide has the action to inhibit the release of LTD4 from various cells and this action may contribute in part to prevent the ulcerogenesis induced by H. pylori.
Adenoma ; Arachidonic Acid ; Calcimycin ; Calcium ; Gastritis ; Helicobacter pylori* ; Helicobacter* ; Leukotriene D4* ; Leukotrienes ; Neutrophils ; Stomach Ulcer

PURPOSE: The release of histamine from human basophils is controlled by an intrinsic, as yet unidentified, cellular property termed "releasability." We carried out this study to ascertain whether there was any difference in the releasability of basophils from asthmatic children compared to those from normal children. We intended also to clarify the correlation between the releasability and the atopic status of asthma. METHODS: We selected nineteen atopic asthmatic, eighteen nonatopic asthmatic and fourteen normal children for this study. Suspensions of leukocytes were isolated and stimulated with calcium ionophore A23187, anti-IgE and D. pteronyssinus antigen. After incubation, the supernatant was assayed for histamine with an automated fluorometric technique. RESULTS: Basophil histamine release with anti-IgE was different in three groups. Anti-IgE caused significantly more basophil histamine release in asthmatic children than in nomal children. Atopic asthmatic group showed greater basophil histamine release with anti-IgE than nonatopic asthmatic group. D. pteronyssinus antigen caused the significant amount of histamine release only in atopic asthmatic group. CONCLUSIONS: Our data suggests that basophils from asthmatic children are characterized by a specific increase in IgE mediated histamine releasability. The difference of histamine releasability with anti-IgE between atopic and nonatopic asthmatic children may be due to the heterogeneity of IgE bound to cell surface, or may be due to the degree of the basophil activation by cytokines such as IL3. The specific release of histamine with D. pteronyssinus antigen in atopic asthmatic group suggests that the basophil histamine release test can be used to diagnose the causing antigen.
Asthma ; Basophils* ; Calcimycin ; Calcium ; Child* ; Cytokines ; Histamine Release ; Histamine* ; Humans ; Immunoglobulin E ; Leukocytes ; Population Characteristics ; Suspensions

Leukotrienes (LTs) are known to act as a mediator provoking tissue response in inflammation. This finding implicates that LTs also play important roles in the pathogenesis of H. pylori-induced gastritis and gastric ulceration. Rebamipide is being currently used as a therapeutics for gastritis and peptic ulcer, but their mechanisms of action have not been known clearly yet. One possibility is that their therapeutic effects are ascribed to interfering with the H. pylori-induced release of LTs from neutrophils and gastric mucosal cells. In the present study, this possibility was tested using LTB4 as the test material in human neutrophils and Kato III cells(gastric adenoma cells as a substitute for gastric mucosal cells). The release of LTB4 from both neutrophils and Kato III cells was time and H. pylori-dose dependent. The maximum release of LTB4 was induced by neutrophils and Kato III cells when these cells incubated with H. pylori 4.8 X 108 cells/ml for 30 min. But in the presence of rebamipide the release of LTB4 from these cells was suppressed in dose dependent manners. The release was completely suppressed at 1.0 mM of rebamipide in neutrophils and 2.0 mM of this drug in Kato III cells, respectively. We also obtained the results that the release of LTB4 was induced by A23187 (Ca2+ ionophore) and the A23187-induced release was also inhibited by rebamipide. It seems that the mechanism of action of rebamipide is through its interaction with the level of intracellular Ca2+. In view of the roles of LTB4 in inflammatory reaction and the roles of H. pylori in gastritis and peptic ulcer, the effects of this drug observed in this study may contribute to their therapeutic action in these gastric disorders.
Adenoma ; Calcimycin ; Gastritis ; Helicobacter pylori* ; Helicobacter* ; Humans ; Inflammation ; Leukotriene B4* ; Leukotrienes ; Neutrophils ; Peptic Ulcer ; Stomach Ulcer

BACKGROUND: The potent bronchodilatory effects of ketamine on airway smooth muscle tone are important in the management of patients with asthma, but its mode of action is unclear. In the present study we evaluated that effects on isolated guinea pig tracheal smooth muscle. METHODS: Changes of isometric contraction of strip were measured. (1) Serial stimulation with acetylcholine(ACh) in Krebs solution or with A23187, nifedipine, ketamine were evaluated. After that, ACh stimulation was induced in Ca2+ free solution. (2) In Ca2+ free solution, ACh contraction was obtained(L1) and emptied by repetitive ACh stimulation. Internal stores were refilled by Ca2+ with ACh stimulation. During the incubation period, A23187, nifedipine, ketamine, cyclopiazonic acid + ketamine was added and tested for their ability to inhibit refilling. Refilling was evaluated by ACh produced contraction (L2) with ratio (L2/L1). (3) Effects of ketamine on the contraction induced by caffeine were also checked. RESULTS: Ketamine inhibited amplitude dose-dependently by successive application of ACh in modified Krebs solution and Ca2+ free solution. Ca2+ influx through voltage gated channels were inhibited with nifedipine but not with A23187. ACh sensitive internal store were different when A23187, nifedipine and ketamine were applied in Ca2+ free solution. Refilling of internal store were potentiated by A23187, but decreased by nifedipine and ketamine. Caffeine produced contractions in the presence of ketamine were not significantly different from control. CONCLUSION: We concluded that the inhibitory effects of ketamine in guinea pig trachea were by acting through voltage and receptor gated channels in dose-depedent manner and these effects may be interferences of intracellular second messengers system.
Animals ; Asthma ; Caffeine ; Calcimycin ; Guinea Pigs* ; Guinea* ; Humans ; Isometric Contraction ; Ketamine* ; Muscle, Smooth ; Nifedipine ; Second Messenger Systems ; Trachea*

This study aimed to evaluate the effect of seminal vesicle fluid (SVF) on the acrosome reaction (AR) occurred spontaneously or induced by Ca2+ ionophore A23187, follicular fluid, and progesterone in mouse epididymal sperm. SVF was divided into high (MW>10 kM)) and low (MW<10 kD) fractions by ultrafiltration. The low MW fraction of SVF decreased the rate of spontaneous AR, however the high MW fraction did not. It suggested that the low MW fraction of SVF might have contained decapacitation factor(s) responsible for prolonging of time need for capacitation. When sperm preincubated for 60 min in the presence of SVF, the rate of AR induced by A23187 was decreased, but prolongation of preincubation time for 120 min significantly potentiated the AR by A23187. It suggested that addition of SVF into sperm preincubation medium imposed the epididymal sperm a condition similar to ejaculation. AR induced by human follicular fluid or progesterone was also inhibited by SVF. It suggested that substance in SVF might have affected AR of mouse sperm by inhibiting the interaction between AR inducing ligands and sperm surface receptors involved in acrosomal exocytosis.
Acrosome Reaction* ; Acrosome* ; Animals ; Calcimycin ; Ejaculation ; Exocytosis ; Female ; Follicular Fluid ; Humans ; Ligands ; Male ; Mice* ; Progesterone ; Seminal Vesicles* ; Spermatozoa* ; Ultrafiltration