Pharmacological profile of TP-680, a new cholecystokininA receptor antagonist.

1. The pharmacological characteristics of a newly developed serine derivative (R)-1-[3-(3-carboxypyridine-2-yl) thio-2-(indol-2-yl)carbonylamino]propionyl-4-diphenylmethyl- piperazine (TP-680), a cholecystokinin type A (CCKA) receptor antagonist, were studied and compared with those of MK-329 and loxiglumide. 2. TP-680 showed approximately 2 and 22 times greater selectivity for peripheral CCKA receptors relative to brain CCK (CCKB) receptors than MK-329 and loxiglumide, respectively, when IC50 values for inhibition of [125I]-CCK-8 binding in isolated acini and cerebral cortex were compared. 3. TP-680 was approximately 17 times less potent than MK-329, but was 106 times more potent than loxiglumide in inhibiting 100 pM CCK-8-stimulated amylase release from rat pancreatic acini. The antagonism produced by TP-680 was specific for CCK in that the effects of other receptor secretagogues or agents bypassing receptors were not altered. 4. TP-680 caused a parallel rightward shift of the dose-response curve for CCK-8-stimulated amylase release as did MK-329 and loxiglumide. However, in contrast to MK-329 and loxiglumide, TP-680 suppressed the maximal responses of CCK-8-induced amylase release in a concentration-dependent fashion, indicating that TP-680 is an unsurmountable antagonist. 5. Repeated washing of acini after a 30 min treatment with TP-680 restored the responsiveness but not the sensitivity, causing a residual inhibition on the action of CCK-8. 6. The addition of loxiglumide prior to or together with application of TP-680 protected CCK receptors from unsurmountable and irreversible antagonism by TP-680. 7. Our results indicate that TP-680 is a potent and the most selective CCKA receptor antagonist for the pancreas reported to date.

Stimulatory effects of vanadate on amylase release from isolated rat pancreatic acini.

The effects of vanadate on exocrine pancreatic function were examined in isolated rat pancreatic acini. Vanadate caused a concentration-dependent stimulation of amylase release above a concentration of 1 mM. Co-incubation of vanadate with vasoactive intestinal polypeptide, 8-bromoadenosine 3':5'-cyclic monophosphate, and the Ca2+ ionophore A23187 produced a synergistic pattern of amylase release, whereas co-incubation with cholecystokinin octapeptide (CCK-8), carbamylcholine, and 12-O-tetradecanoylphorbol 13-acetate produced an additive effect. Vanadate alone had no influence on acinar cyclic AMP content, Ca2+ efflux, or intracellular Ca2+ concentration. However, preincubation with vanadate prevented the plateau phase of CCK-8-induced Ca2+ transient increase from returning to baseline. Moreover, depletion of the intracellular Ca2+ pool by pretreatment of acini with CCK-8 in Ca2+-free medium (plus ethyleneglycol bis[beta-aminoethylether]-N,N'-tetraacetic acid) had no effect on subsequent stimulation by vanadate, although it abolished the response to both CCK-8 and carbamylcholine stimulation. The protein kinase C (PKC) inhibitors staurosporine and calphostin C significantly inhibited vanadate-stimulated amylase release, whereas the protein tyrosine kinase inhibitor genistein had no inhibitory effect. Moreover, vanadate caused a significant translocation of PKC from cytosol to membrane fraction in pancreatic acinar cells. This translocation was inhibited significantly by staurosporine and calphostin C but not by genistein. These results suggest that vanadate acts directly on pancreatic acini and stimulates amylase release by activating PKC without an effect on Ca2+ mobilization, cyclic AMP, or protein tyrosine kinase.

Serum pepsinogen can predict response to H2-receptor antagonist in patients with functional dyspepsia.

BACKGROUND: Therapy for the relief of symptoms of functional dyspepsia is unpredictable. AIM: To identify which patients may benefit from antisecretory therapy. METHODS: Twenty-seven patients with functional dyspepsia were selected to receive H2-receptor antagonist (H2RA) treatment for 4 weeks. Serum pepsinogen A, pepsinogen C and gastrin were measured, and Helicobacter pylori status was determined. Symptoms were assessed at baseline and after H2RA treatment. RESULTS: Fourteen patients were identified as H2RA responders and the remaining patients were non-responders. No differences were found between responders and non-responders with regard to serum pepsinogen A, pepsinogen C, gastrin and H. pylori status. However, the pepsinogen A/C ratio was significantly higher in responders than in non-responders. Ten of the 13 functional dyspepsia patients (77%) with a high value of the pepsinogen A/C ratio (> or = 4.5) achieved symptom resolution by H2RA, compared with only one of the eight patients (13%) with a low value of the pepsinogen A/C ratio (< or = 3.0). CONCLUSIONS: The serum pepsinogen A/C ratio seems to identify those functional dyspepsia patients for whom acid control provides benefit. This ratio may be a practical tool for the management of functional dyspepsia patients.

Metabolic abnormalities in the genetically obese and diabetic Otsuka Long-Evans Tokushima Fatty rat can be prevented and reversed by alpha-glucosidase inhibitor.

The recently developed Otsuka Long-Evans Tokushima Fatty (OLETF) rat is known to develop insulinopenic diabetes after a prolonged period in a condition resembling non-insulin-dependent diabetes mellitus (NIDDM). We examined the effect of pharmacological intervention with a potent intestinal alpha-glucosidase inhibitor, acarbose, on the metabolic and histopathologic changes in this rat model. The first two groups of rats received an acarbose-rich diet (150 mg/100 g normal chow) from 12 weeks of age (ie, before the onset of diabetes) or from 28 weeks of age (ie, after the onset of diabetes), while a third group received the acarbose-rich diet for the initial 16 weeks only (from 12 to 28 weeks of age). A control group received standard rat chow. Acarbose-fed rats gained less weight or lost weight despite increased food intake when switched to the acarbose-rich diet. Acarbose also reduced visceral adipose depots and fasting triglyceride (TG), glucose, and insulin levels. At the end of the study at 72 weeks, the pancreatic wet weight and insulin content were significantly higher in the treated groups versus control rats. The morphological changes observed in control rats, such as atrophy of the pancreas and reduced number and size of islets, were not present in acarbose-treated rats. Rats fed acarbose from 12 to 28 weeks of age gradually gained weight after switching to standard chow, and hyperinsulinemia, hyperglycemia, and hyperlipidemia appeared (in that order). The pancreatic insulin content in these rats was significantly higher and the visceral adipose depot was significantly smaller than in control rats. Our study demonstrates that acarbose prevented and reversed the metabolic derangement and histopathological changes in genetically diabetic rats. Moreover, treatment with acarbose even for a short period produced a marked delay in the development of insulin insensitivity and frank diabetes.

Pharmacologic profile of TS-941, a new benzodiazepine derivative cholecystokinin-receptor antagonist, in in vitro isolated rat pancreatic acini.

We investigated the pharmacologic characteristics of a newly developed benzodiazepine derivative (S)-(-)-N-[2,3-dihydro-2-oxo-5-phenyl-1-[(1H-tetrazol-5-yl)methyl] -1H-1,4-benzodiazepine-3-yl]-2-indolecarboxamide (TS-941), a cholecystokinin type A (CCK-A)-receptor antagonist, in the isolated rat pancreatic acini and compared with those of well-known CCK-A-receptor antagonists, devazepide and loxiglumide. TS-941 inhibited CCK-8-stimulated amylase release concentration dependently, as did devazepide and loxiglumide, with a half-maximal inhibition (IC50) at 78.6 +/- 10.3 nM. TS-941 was approximately 23 times less potent than devazepide (IC50, 3.4 +/- 0.3 nM), but was 50 times more potent than loxiglumide (IC50, 3,966 +/- 544 nM) in inhibiting 100 pM CCK-8-stimulated amylase release from rat pancreatic acini. TS-941 had a fivefold lower selectivity than devazepide for pancreatic CCK (CCK-A) over brain CCK (CCK-B) receptors but fourfold greater than loxiglumide when IC50 values for inhibition of [125I]CCK-8 binding in isolated acini and cerebral cortex were compared. The antagonism produced by TS-941 was specific for CCK in that the effects of other receptor secretagogues or agents bypassing receptors were not altered. TS-941 caused a parallel rightward shift of the entire dose-response curve for CCK-8-stimulated amylase release without altering the maximal increase, as did devazepide and loxiglumide. TS-941, whether added at the beginning or 20 min after the CCK-8 stimulation, inhibited amylase release. TS-941 caused a concentration-dependent residual inhibition of the action of CCK-8. The acini, once incubated with a high concentration of TS-941 (10 microM; 127 times IC50) for 30 min, was 10-fold less sensitive to CCK-8 than the acini preincubated without TS-941, whereas the sensitivity and the responsiveness to CCK-8 stimulation of those incubated with a low concentration of TS-941 (1.0 microM) were similar to the control acini. These results indicate that TS-941 is a potent, competitive, and selective CCK-A receptor antagonist for the pancreas.

Role of endogenous cholecystokinin and cholecystokinin-A receptors in the development of acute pancreatitis in rats.

Recent studies provide significant evidence that cholecystokinin (CCK) is involved in the induction and development of acute pancreatitis in experimental animals. However, the results obtained with specific CCK-A (peripheral) receptor antagonists are still controversial. The present studies were undertaken to evaluate the involvement of endogenous CCK and the CCK-A receptors in the development of severe acute pancreatitis induced in Otsuka Long-Evans Tokushima Fatty (OLETF) rats that have a selective defect in the CCK-A receptor. Three models of severe acute pancreatitis were induced by retrograde intraductal infusion of 4% sodium taurocholate, by the closed duodenal loop, or by a single intraperitoneal injection of 500 mg/100 g body weight of L-arginine in OLETF rats and control Long-Evans Tokushima Otsuka (LETO) rats. Plasma CCK levels rose up to 4- to 14-fold over the preloading values after the onset of acute pancreatitis in all three models in both groups of rats. However, histologic alterations as well as the magnitudes of increase in serum amylase and lipase activity and the pancreatic wet weight were significantly less in the OLETF rats than those in the LETO rats. In addition, 72 h after the onset of arginine pancreatitis, massive destruction of pancreatic parenchyma with a significant reduction in serum amylase and lipase activities and pancreatic wet weight was observed in the LETO rats, whereas these changes were not seen in OLETF rats. These results suggest that endogenous CCK and CCK-A receptors play a role in the development of severe acute pancreatitis in rats.

Portal vein thrombosis associated with antiphospholipid syndrome.

Portal vein thrombosis is a rare occurrence, and often an underlying hypercoagulable state can be found. Recently, there has been growing interest and recognition of the antiphospholipid syndrome in association with acquired hypercoagulable state. This syndrome consists of the association of lupus anticoagulant or antiphospholipid antibodies with arterial or venous thrombosis, thrombocytopenia, and spontaneous abortion. We report a case of portal vein thrombosis associated with the antiphospholipid syndrome. In our patient, chronic liver disease, hepatobiliary infection, abdominal malignancies, myeloproliferative disorders, and inherited coagulation disorders were excluded. This case report suggests that serum antiphospholipid antibodies should be investigated in patients with portal vein thrombosis of unexplained etiology.

Pancreatic cancer complicated by disseminated intravascular coagulation associated with production of tissue factor.

A 54-year-old man was diagnosed as having pancreatic cancer and disseminated intravascular coagulation. His plasma tissue factor level on the 11th hospital day was 996 pg/ml (normal range, 120-270 pg/ml). He was treated with gabexate mesilate, antithrombin III, and low-molecular-weight heparin. However, he died of multiple organ failure on the 17th hospital day. The histological finding was poorly differentiated ductal adenocarcinoma of the pancreas, and the production of tissue factor in this lesion was revealed. Tissue factor is a factor that initiates blood coagulation; thus, its expression in pancreatic cancer is one of the causes of coagulation abnormalities in this disease. Although one report has demonstrated immunoreactivity for tissue factor in pancreatic cancer, the patient's detailed clinical course was not mentioned in that report. This is the first report to prove that pancreatic cancer produced tissue factor in a patient with disseminated intravascular coagulation.

Defects of cholecystokinin (CCK)-A receptor gene expression and CCK-A receptor-mediated biological functions in Otsuka Long-Evans Tokushima Fatty (OLETF) rats.

Recent studies in genetically obese and diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats suggest defects of cholecystokinin (CCK)-A receptor gene expression and CCK-A receptor-mediated biological functions such as pancreatic juice, protein, and gastric acid secretion. The present studies were undertaken to further examine CCK-A receptor gene expression and CCK-A receptor-mediated biological functions in the pancreas, stomach, and brain of OLETF rats. Expression of the CCK-A receptor gene could not be detected in the stomach, pancreas and brain by the reverse-transcription polymerase chain reaction (RT-PCR) method and Southern blotting of the PCR products. Southern blot analysis of genomic DNA from OLETF and control Long-Evans Tokushima Otsuka (LETO) rats with CCK-A receptor fragment as a probe revealed different restriction bands. Expression of the CCK-B receptor gene was observed in the stomach, pancreas, and brain in both OLETF and LETO rats by the RT-PCR method, with expression of the CCK-B receptor gene markedly enhanced in OLETF rats compared with that in LETO rats. Consistent with the defect of CCK-A receptor gene expression, CCK-A receptor-mediated biological functions were not observed in these organs. Perfused exocrine and endocrine pancreas of OLETF rats were insensitive to CCK stimulation but not to carbamylcholine stimulation. Basal gastric acid and pepsinogen secretions in OLETF rats were higher than in LETO rats. OLETF rats showed a significantly higher average daily food intake, gained body weight faster, and were heavier than LETO rats. The present study confirmed that OLETF rats have CCK-A receptor gene anomalies and demonstrated deficient CCK-A receptor-mediated biological function in the pancreas, stomach, and brain.

Testicular dysfunction induced by cadmium and its improvement caused by selenium in the mouse.

Histological observation revealed widespread and severe necrosis at 3 days after the final injection of Cd (0.012 mmol/Kg. daily for 2 consecutive days). Two months later, the changes were further aggravated. In addition, severe atrophy was found, and there was an increase in materials which could be stained by the Kossa method. These changes were prevented by the simultaneous injection of Se (0.024 mmol/Kg, daily for 2 consecutive days). Two months later, the Cd group showed no longer its reproduction. As a result of the Cd injection, at 3 days lipoperoxide concentration, expressed as substances reacting with thiobarbituric acid, was increased significantly, and accompanied by a decrease of glutathione. The levels of lipoperoxide and glutathione values were restored to the control level by the Se injection. At 3 days, testicular Zn and Mg decreased significantly, but Ca and Fe increased markedly in the Cd group. Ca increased mainly in the slow-speed centrifugation fraction, while Mg decreased evenly. In the Cd + Se group, there was no detectable change of metal concentration. However, Se stimulated the Cd uptake into the testis and vice versa. The testicular Cd and Se contents were maintained at a constant level for 2 months. The proportions of Cd existing in the cytosol fraction were 80% and 45% in the Cd group and the Cd + Se group, respectively. Furthermore, the recovery rates for cytosolic Cd in the F2 fractions corresponding to metallothionein were 60% and 75% in the Cd and Cd + Se groups, respectively. Our results suggest that Se has multiple functions against testicular Cd toxicity and that the toxicity once blocked by Se does not occur for a relatively long time.

Force-velocity relation of sliding of skeletal muscle myosin, arranged on a paramyosin filament, on actin cables.

To investigate in vitro ATP-dependent sliding of regularly arranged myosin molecules on actin filaments, we prepared thick hybrid filaments in which myosin molecules isolated from rabbit skeletal muscle were arranged around the paramyosin core (length, 10-20 micron; diameter,

Experimental studies of moderate temperature burns.

The pathology and characteristics of moderate temperature burns have been investigated in rats with a heating apparatus capable of applying thermal doses varying in temperature and exposure time, to which the responses have been observed both macroscopically and microscopically. The macroscopic examination showed only erythema and greyish necrosis. The microscopic changes were divided into six grades of severity from 0 to 5. From the time-temperature threshold curves made by microscopic observations, the critical temperatures for superficial and deep dermal burns, and full skin thickness burns were 37.8 degrees C, 41.9 degrees C and 47.9 degrees C, respectively. According to the data, almost all commercially available topical heaters are considered hazardous. Susceptibility to thermal injury was markedly increased by compression and ischaemia, the effects of which are thought to be due mainly to rapid heat accumulation (heat damage) and hypoxaemia (hypoxic damage).

Numerous huge iron particles appeared around micro blood vessels under intestinal epithelial cells in infant mice fed excess iron with protein.

In infant mice (2 weeks old) fed a diet with excess iron for one week, numerous huge iron particles around micro blood vessels under the basement membrane of intestinal epithelial cells were observed by electron microscopy. The frequency of occurrence of these particles (0.24 +/- 0.04 micron diameter) was markedly higher in mice fed a casein-based diet than in mice fed an amino acid-based diet. The quantity of these particles in both groups decreased in proportion to the term of 2 or 3 weeks. Changes in morphological features, such as opening of the intercellular junctions between intestinal epithelial cells, were also observed in these experimental groups. On the other hand, in mice fed the casein-based diet with excess iron, fat globules appeared in the intestinal epithelial tissue (intestinal epithelial cells, interstitial tissue, lympha) and the occurrence of these increased gradually in proportion to the term of feeding. These fat globules were not observed in mice fed the amino acid-based diet with excess iron. These phenomena might be elicited temporally in infant mice fed excess iron together with protein. The mechanisms of fat globule formation remain unclear.

Investigation of morphological changes in absorptive cells in young adult and infant mice fed different amounts of iron for a long-term.

The changes in fine structure of the intestinal tract in young adult (4 week-old) and infant (2 week-old) mice fed a diet containing different amounts of iron salt (Fe-0, Fe-2.5, Fe-25: 0, 2.5 and 25 mg Fe/100 g diet, respectively) for a long-term (1 or 2 weeks) were investigated. The hepatic iron levels in infant mice fed Fe-25 for 2 weeks were significantly higher than those observed after 1 week of feeding, but there was no such increase in young adult mice during the feeding period. Observations of fine structure indicated typical signs of impairment of enterocytes due to excess iron such as the opening of intercellular junctions between adjacent epithelial cells and the marked appearance of eosinophilic leukocytes outside the basement membrane in young adult and infant mice fed Fe-25. The frequency of the opening in intercellular junctions increased in young adult mice fed Fe-25 for 2 weeks, but decreased in infant mice. On the contrary, under iron-deficient conditions, the frequency in infant mice was higher than that in young adult mice. The appearance of eosinophilic leukocytes indicated that some immunological reaction was elicited in both groups of mice fed Fe-25 for 2 weeks.

[Multiple intracranial aneurysms following radiation therapy for pituitary adenoma; a case report].

A report is made here on a rare case in which occurrence of multiple cerebral aneurysm was observed after radiotherapy. The case was that of a female aged 51 who was hospitalized with the chief complaint of consciousness disorder. The patient was discharged from the hospital 1 year before, after undergoing subtotal extirpation of a tumor through the transsphenoidal sinus in a case of pituitary adenoma, and post-operative radiotherapy (topical 50 Gy). Mild hyperlipidemia associated with hypothyroidism was observed by blood biochemical test during the patient's hospitalization, and multiple cerebral infarction was also observed by CT scanning and MR imaging. Therefore, conservative treatments including intensified endocrine-supplementing treatment, centering on thyroid hormone, were attempted. The patient's thyroid gland function and hyperlipidemia improved but the consciousness disorder persisted. Exacerbation of the consciousness disorder occurred suddenly 6 weeks after the patient's hospitalization. Subarachnoid hemorrhage and hydrocephalus were observed by CT scanning. Cerebral angiography detected a saccular aneurysm in the trifurcation of the right middle cerebral artery, 3 fusiform aneurysms in the periphery of the right middle cerebral artery, 2 fusiform aneurysms in the posterior cerebral artery and irregularity of wall width in the central artery including the terminal region of the internal carotid artery. These findings were not observed at the initial hospitalization and were considered to have been formed newly afterwards. Further, these findings were observed unexceptionally in all the radiated fields. The patient died 8 weeks after hospitalization, and no autopsy finding was obtained. From the above, we presumed that radiation vasculopathy caused by the radiation therapy made one year previously, had led to the formation of multiple cerebral aneurysms.(ABSTRACT TRUNCATED AT 250 WORDS)

Surface ultrastructural features of isolated perfused rat hearts during calcium paradox.

Cellular damage caused by calcium (Ca++) paradox is of significant importance under both experimental and clinical conditions. Electron microscopy was used to study the surface changes of the isolated rat hearts perfused with normal Krebs-Henseleit (KH) medium, Ca++-free KH medium, Ca++-free KH medium followed by normal KH medium. The hearts perfused with Ca++-free medium and normal KH medium showed elongated parallel myofibrils with transverse ridges spaced at regular intervals. The major change in Ca++- free hearts consisted of wide cellular separation. For the Ca++- paradox hearts, wht myofibrils were twisted and narrowed at several locations. The myocardial fiber surfaces were bulged out, frequently with sarcolemmal ruptures and holes. Supercontraction of cells caused breaks in the sarcolemma and aggregation of mitochondria at the cell periphery. These results suggest that Ca++-depletion primarily affects the myocardial cell junctions which, following reintroduction of Ca++, accelerate the entry of Ca++ into the cells, supercontraction of cells and stretching of sarcolemma.

Supramaximal CCK and CCh concentrations abolish VIP potentiation by inhibiting adenylyl cyclase activity.

Exocrine pancreatic secretion stimulated by vasoactive intestinal polypeptide (VIP), which acts through the adenylyl cyclase-cAMP pathway, is potentiated by stimulation with other secretagogues such as CCK and carbachol (CCh). However, the potentiating effect is abolished by the same secretagogues at supramaximal concentrations. In the present study, we examined the mechanisms by which supramaximal concentrations of CCK octapeptide (CCK-8) or CCh reduce the VIP-induced potentiation of amylase secretion from isolated rat pancreatic acini. VIP-stimulated amylase secretion was potentiated by submaximal stimulatory concentrations of CCK-8 and CCh but was reduced by the same reagents at higher concentrations. Supramaximal concentrations of CCK-8 or CCh also reduced forskolin-induced potentiation of amylase release but did not reduce that induced by 8-bromo-cAMP. Moreover, supramaximal concentrations of CCK-8 or CCh inhibited VIP-stimulated intracellular cAMP production as well as adenylyl cyclase activity. 12-O-tetradecanoylphorbol 13-acetate (TPA) also reduced the magnitude of the potentiation of amylase release caused by VIP plus CCK-8 or CCh, although TPA itself decreased neither VIP-stimulated adenylyl cyclase activity nor intracellular cAMP accumulation. These results indicate that supramaximal concentrations of CCK-8 and CCh reduce the potentiating effect of VIP and forskolin on amylase secretion by inhibiting the adenylyl cyclase activity. In addition, protein kinase C is suggested to be partly implicated in this inhibitory mechanism. The mechanisms that lead to such inhibition may be interlinked but distinct from each other.

Effect of ONO-3144, a free radical scavenger, on reoxygenation injury in anoxic myocardium.

Thus, ONO-3144 treatment affords great protection against reoxygenation injury in the anoxic myocardium through the scavenging mechanism that includes .OH or closely related species of free radicals and also may inhibit thromboxane synthetase in the anoxic myocardium. We would hypothesize that free radical formation generated through the conversion of PGG2 to PGH2 should play an important role in the reoxygenation injury of the anoxic myocardium.