The effects of potassium channel opener P1075 on the human saphenous vein and human internal mammary artery.

Because adrenergic contractions can contribute to the development of life-threatening spasm of coronary artery bypass graft, this study was performed to investigate the effect of adenosine 3-phosphate (ATP)-sensitive K channel (KATP) opener P1075 on contractions of isolated human saphenous vein (HSV) and human internal mammary artery (HIMA). Phasic contractions were evoked by electric field stimulation (20 Hz) and noradrenaline. The sustained contractions were evoked by phenylephrine. The presence of pore-forming Kir6.1 and Kir6.2 subunits of the KATP channels in the HIMA and only Kir6.2 in the HSV was confirmed immunomorphologically. P1075 inhibited in the HSV only, the electrical field stimulation contractions more strongly than noradrenaline contractions. In addition, the phenylephrine contractions of HSV were more sensitive to P1075 in comparison to those of HIMA. Glibenclamide, a KATP channel blocker antagonized the vasodilatation produced by P1075 in both grafts differently, because its effect was more prominent on the P1075-induced inhibition of contractions of HSV than of HIMA. We conclude that P1075 has a vasorelaxant effect and inhibited adrenergic contractions of the tested grafts. This effect is graft and vasoconstrictor selective and seems to be mediated by Kir6.1- and/or Kir6.2-containing KATP channels. Thus, P1075 can be considered as a potential drug in the prevention of graft spasm.

Determining conditions for nitric oxide synthesis in Caco-2 cells using Taguchi and factorial experimental designs.

Intestinal inflammation correlates well with the increased synthesis of nitric oxide (NO), which is attributed mainly to the up-regulation of inducible nitric oxide synthase (iNOS). We optimized the use of interferon gamma (IFN-gamma), tumour necrosis factor alpha (TNF-alpha), interleukin 1beta (IL-1beta), lipopolysaccharide (LPS), and phorbol myristate acetate (PMA) as inducers to stimulate NO synthesis in Caco-2 cells using a Taguchi design. The results indicated that IFN-gamma was the most important inducer of iNOS in Caco-2 cells. Treating Caco-2 cells with both IFN-gamma and PMA using an optimal mixture of 8000 U/ml IFN-gamma and 0.1 microg/ml of PMA resulted in a synergistic induction of NO synthesis. Further experiments using a 5-factor/2-level factorial design including Caco-2 growth conditions such as cell passage, culture medium composition, cell seeding time and density, and stimulation time were also performed. Cell seeding and stimulation times significantly (P<0.05) affected NO synthesis, whereas culture medium and seeding density did not appreciably affect NO synthesis in Caco-2 cells. Western blotting and RT-PCR findings confirmed that the optimal mixture of IFN-gamma and PMA effectively up-regulated iNOS mRNA and protein. The induced NO, iNOS mRNA, and protein were all inhibited by the iNOS selective inhibitor, aminoguanidine (AG).

Selective inhibition of ion transport mechanisms regulating intracellular pH reduces proliferation and induces apoptosis in cholangiocarcinoma cells.

BACKGROUND: Cells within the acidic extracellular environment of solid tumours maintain their intracellular pH through the activity of the Na(+)/H(+) exchanger and the Na(+) dependent Cl(-)/HCO(3)(-) exchanger. The inhibition of these mechanisms could therefore inhibit cancer cell growth. AIM: We evaluated the effect of two selective inhibitors of these transporters (cariporide and S3705) on proliferation and apoptosis of human cholangiocarcinoma cells (HUH-28 and Mz-ChA-1 cells) as a function of external pH (7.4 and 6.8). METHODS/RESULTS: HUH-28 cells incubated for 24h at external pH 7.4 or 6.8 without inhibitors maintained intracellular pH at physiological level, whereas incubation with cariporide and/or S3705 caused the intracellular pH of cells to drop. Incubation of HUH-28 cells with cariporide and/or S3705 was able to reduce proliferation, evaluated by a colorimetric ELISA method, and to induce apoptosis, evaluated by measuring caspase-3 activity and Annexin-V staining, and these effects were more evident at external pH 6.8. S3705 but not cariporide was able to inhibit serum-induced phosphorylation of ERK1/2, AKT and BAD, intracellular molecules involved in cancer cell proliferation and survival. Similar results were obtained in Mz-ChA-1 cells. CONCLUSIONS: (1) Inhibition of intracellular pH regulatory mechanisms by cariporide and S3705 reduces proliferation and induces apoptosis in cholangiocarcinoma cells; and (2) these drugs might have potential therapeutic value against cholangiocarcinoma.

Effects of corticosteroid and neuraminidase inhibitors on survival in patients with respiratory distress induced by influenza virus.

BACKGROUND/PURPOSE: Neuraminidase inhibitors (NAIs) including oseltamivir and peramivir are used for influenza treatment. A systemic corticosteroid is usually administrated for acute respiratory distress syndrome. The aim of this study was to investigate the effect of a systemic corticosteroid and its interaction with NAIs in patients with influenza infection and respiratory distress. METHODS: A retrospective survey of hospitalized patients infected with influenza from January 2012 to May 2014 was conducted in a medical center in Taiwan. RESULTS: Eighty-six patients were hospitalized during the study period. Forty-eight patients had respiratory distress and 39 of them (81.3%, 39/48) were supported by a mechanical ventilator. All patients with respiratory distress received oseltamivir; 60.4% (29/48) and 31.3% (15/48) of them received a corticosteroid and salvage intravenous peramivir, respectively. All-cause mortality was 29.1% (14/48), 20% (3/15), and 31% (9/29) in patients with respiratory distress, patients who received salvage peramivir, and patients who received a systemic corticosteroid, respectively. Salvage peramivir seemed to improve prognosis in patients with H1pdm09 or type B virus infection and respiratory distress (p = 0.05). Early initiating corticosteroid had a worse prognosis than initiation after 72 hours of NAI treatment (p = 0.024). In particular, a systemic corticosteroid seemed to lead to a shorter survival time in patients with chronic lung disease (p = 0.05). CONCLUSION: Salvage peramivir provided a better prognosis than monotherapy with oseltamivir in patients who were infected with H1pdm09 or type B virus and who developed respiratory distress. A systemic corticosteroid should be administered after initiating NAI therapy, especially in patients with chronic lung disease.

Combination of thiol antioxidant Silibinin with Brostallicin is associated with increase in the anti-apoptotic protein Bcl-2 and decrease in caspase 3 activity.

The cytotoxic activity of Brostallicin was previously shown to be enhanced in the presence of high glutathione and glutathione transferase levels. We hypothesized that thiol antioxidants, N-acetylcysteine and Silibinin, could potentiate Brostallicin's cytotoxicity in a similar way. HepG2 and CNE-2 cells were treated with N-acetylcysteine, Silibinin and Brostallicin, either alone or in combination. Surprisingly, we found that NAC and Silibinin had adverse effects on Brostallicin's cytotoxicity. The mechanism underlying the interaction involved the apoptotic pathway as we demonstrated an increase in Bcl-2 protein levels and decrease in caspase 3 activity with the Silibinin-Brostallicin combination.

Identification of a high-affinity binding site for dinotefuran in the nerve cord of the American cockroach.

The binding of the neonicotinoid insecticide dinotefuran to insect nicotinic acetylcholine receptors (nAChRs) was examined by a centrifugation method using the nerve cord membranes of American cockroaches and [3H]dinotefuran (78 Ci mmol-1). The Kd and Bmax values of [3H]dinotefuran binding were estimated to be 13.7 nM and 14.8 fmol 40 microg-1 protein respectively by Scatchard analysis. Epibatidine, an nAChR agonist, showed a rather lower affinity to the dinotefuran binding site (IC50=991 nM) than dinotefuran (IC50=5.02 nM). Imidacloprid and nereistoxin displayed lower potencies than dinotefuran but higher potencies than epibatidine. The potencies of five dinotefuran analogues in inhibiting the specific binding of [3H]dinotefuran to nerve cord membranes were determined. A good correlation (r2=0.970) was observed between the -log IC50 values of the tested compounds and their piperonyl butoxide-synergised insecticidal activities (-log LD50 values) against German cockroaches. The results indicate that a high-affinity binding site for dinotefuran is present in the nerve cord of the American cockroach and that the binding of ligands to the site leads to the manifestation of insecticidal activity.

DTG-induced circling behaviour in rats may involve the interaction between sigma sites and nigro-striatal dopaminergic pathways.

The distribution of sigma sites in brain areas enriched in dopamine, and the finding that circling behaviour can be elicited by specific sigma ligands such as DTG (di-o-tolylguanidine) suggest a modulatory role of these sites in the dopaminergic system. The present study was carried out to investigate further this hypothesis. Circling behaviour induced in rats by unilateral intranigral injection of DTG (10 nmol/rat) was decreased by haloperidol (0.1 mg/kg, i.p.), clebopride (0.25 mg/kg, i.p.) and SCH 23390 (0.03 mg/kg, s.c.) indicating that an interaction between sigma sites and the midbrain dopaminergic system may be involved in this rotational behaviour. Microdialysis experiments in freely moving rats showed that unilateral intranigral injection of DTG (5, 10, 20 nmol/rat) produced increases in extracellular levels of dopamine metabolites (DOPAC, HVA) in the ipsilateral striatum which correlated with the number of rotations. In addition intranigral injection of DTG (10 nmol/rat) produced increases in tissular dopamine metabolite levels in the ipsilateral striatum without affecting dopamine metabolite levels in limbic structures. These results indicate that sigma sites may be involved in the modulation of the dopaminergic motor system.

Vasoactive systems in L-NAME hypertension: the role of inducible nitric oxide synthase.

OBJECTIVES: The contribution of the renin-angiotensin system (RAS) and the sympathetic nervous system (SNS) to blood pressure (BP) maintenance was evaluated in rats with N(omega)-nitro-L-arginine methyl ester (L-NAME) hypertension. Furthermore, we studied the extent of nitric oxide (NO) synthesis inhibition and the participation of remaining NO in the counterbalance of pressor systems, with a special reference to inducible nitric oxide synthase (iNOS). METHODS: Wistar rats subjected to chronic L-NAME treatment (40 mg/kg per day for 4 weeks) were used. A consecutive blockade of RAS (captopril) and SNS (pentolinium) was followed by acute L-NAME injection. Dimethylguanidine or aminoguanidine were used to affect NO synthesis by iNOS. RESULTS: L-NAME hypertensive rats had borderline augmentation of depressor response to captopril injection, but their BP fall after pentolinium was considerably enhanced compared with controls. Residual BP (recorded after simultaneous blockade of the RAS and the SNS) was elevated by 20-40% in hypertensive rats. Pronounced inhibition of NO synthase activity (50% reduction in the aorta and myocardium) was detected in L-NAME hypertensive rats in which the BP rise elicited by acute L-NAME injection was considerably attenuated (by 60-80%). In contrast, acute administration of dimethylguanidine [mixed endothelial NO synthase (eNOS)/iNOS inhibitor] to hypertensive rats induced a major BP rise similar to that caused by L-NAME injection in controls. Aminoguanidine (a selective iNOS inhibitor) caused a substantial BP rise in L-NAME hypertensive rats only. CONCLUSION: The contribution of SNS to BP maintenance in L-NAME hypertension is more important than that of RAS. In L-NAME hypertensive rats the iNOS becomes a major source of hemodynamically important NO production, which is still insufficient to compensate prevailing vasoconstriction.

In vivo effects of the immunosuppressant 15-deoxyspergualin on hematopoiesis in murine allogeneic bone marrow chimeras. Its thrombopoietic activity and reversal of adverse effects with granulocyte colony-stimulating factor and/or erythropoietin.

When 15-deoxyspergualin (DSG), a potent immunosuppressant, was administered into [BALB/c-->C3H/He] bone marrow chimeras from day 14 to day 25, increased thrombopoiesis was induced on day 20 to day 33, accompanied by marked leukocytopenia and anemia. The mean platelet counts in DSG-treated and control [BALB/c-->C3H/He] bone marrow chimeras on day 25 were (114.1 +/- 0.5) x 10(4)/microliter versus (58.6 +/- 2.6) x 10(4)/microliter (1.9-fold increase). Colony-forming units-megakaryocyte (CFU-Meg) were not significantly increased in DSG-treated bone marrow chimeras. Colony-forming units-granulocyte/macrophage (CFU-GM) and burst-forming units-erythroid (BFU-E) were decreased during DSG-treatment whereas CFU-Mix colony formations were rather increased, and more primitive hematopoietic progenitor cells (highly proliferative potential colony-forming units [CFU-HPP]) were not decreased in the same time period. Since CFU-GM and BFU-E colony formations were increased immediately after the cessation of DSG treatment, followed by the rebound of leukocyte counts and the recovery of hemoglobin (Hb) levels, the leukocytopenia and anemia appeared to be induced by a cytostatic effect of DSG. The adverse effect of DSG was partly reversed by the simultaneous administration of granulocyte colony-stimulating factor (G-CSF) and/or erythropoietin (EPO), suggesting the need for the administration of these cytokines in the case of bone marrow transplants treated with DSG. Furthermore, it was of note that DSG modulated hematopoiesis and stimulated the production of thrombopoietin (TPO)-like cytokine(s) as well as interleukin-3 (IL-3).

Pinacidil inhibits the ryanodine-sensitive outward current and glibenclamide antagonizes its action in cells from the rabbit portal vein.

Pinacidil, a potassium-channel opener, inhibited the ryanodine-sensitive oscillatory outward potassium current induced by Ca released from an intracellular store. Glibenclamide, a blocker of the ATP-sensitive K-channel, prevented the action of pinacidil, suggesting the presence of an additional site (to K channels) for the vasodilator actions of pinacidil at which glibenclamide can act as an antagonist.

The antagonism of adrenergic neurone blockade by amphetamine and dexamphetamine in the rat and guinea-pig.

1. In isolated rat mesentery preparations, intra-arterial injection of the following drugs rapidly suppressed vasoconstrictor responses to sympathetic nerve stimulation: bretylium (75-100 mug), guanethidine (10-20 mug) and bethanidine (20-30 mug); with phenoxypropylguanidine (15-30 mug) the onset of blockade was slower. The blockade caused by these or higher concentrations was rapidly abolished by intra-arterial injection of amphetamine (100 mug) as also was the blockade caused by infusing bretylium or guanethidine for 10-20 min. Partial blockade was produced by 20 mug of reserpine and this was only slightly and briefly antagonized by amphetamine.2. In mesentery preparations taken from rats 24 h after subcutaneous injection of bretylium 50 mg/kg, guanethidine 10 mg/kg, phenoxypropylguanidine 10 mg/kg or reserpine 0.1 mg/kg, responses to sympathetic nerve stimulation were greatly impaired. Only in the preparations from the bretylium-treated rats did amphetamine antagonize the blockade. The adrenergic neurone blocking effect of bethanidine 10 mg/kg was evident at 12 h but not at 24 h after injection.3. In rat mesentery amphetamine did not cause vasoconstriction but briefly potentiated the vasoconstrictor effect of sympathetic nerve stimulation. Responses to noradrenaline were not importantly affected.4. The contractile responses of the rat inferior eyelid caused by stimulation of the cervical sympathetic nerve was greatly reduced 17-27 h after subcutaneous injection of bretylium 300 mg/kg, bethanidine 30 mg/kg, guanethidine 10 mg/kg or reserpine 0.3 mg/kg. Intravenous dexamphetamine (0.5 mg/kg) powerfully antagonized the effect of bretylium, weakly antagonized the blockade by bethanidine and guanethidine and caused no change in the response of reserpine-treated animals.5. The vas deferens taken from guinea-pigs 24 h after subcutaneous injection of either bretylium or guanethidine showed greatly impaired responses to hypogastric nerve stimulation. Amphetamine largely restored the contractile response in bretylium-treated rats but caused only weak antagonism in the guanethidine-treated animals.

The effects of sigma ligands and of neuropeptide Y on N-methyl-D-aspartate-induced neuronal activation of CA3 dorsal hippocampus neurones are differentially affected by pertussin toxin.

1. The in vivo effects of the high affinity sigma ligands 1,3-di(2-tolyl)guanidine (DTG), (+)-N-cyclopropylmethyl-N-methyl-1,4-diphenyl-1- ethyl-but-3-en-1-ylamine hydrochloride (JO-1784), (+)-pentazocine and haloperidol, as well as of those of neuropeptide Y (NPY), on N-methyl-D-aspartate (NMDA)- and quisqualate (Quis)-induced neuronal activations of CA3 pyramidal neurones were assessed, using extracellular unitary recording, in control rats and in rats pretreated with a local injection of pertussis toxin (PTX), to evaluate the possible involvement of Gi/o proteins in mediating the potentiation of the neuronal response to NMDA by the activation of sigma receptors in the dorsal hippocampus. 2. Microiontophoretic applications as well as intravenous injections of (+)-pentazocine potentiated selectively the NMDA response in control rats as well as in PTX-pretreated animals. In contrast, the PTX pretreatment abolished the potentiation of the NMDA response by DTG, JO-1784 and NPY. Moreover, microiontophoretic applications of DTG induced a reduction of NMDA-induced neuronal activation. Neither in control nor in PTX-treated rats, did the sigma ligands and NPY have any effect on Quis-induced neuronal response. 3. In PTX-treated rats, the potentiation of the NMDA response induced by (+)-pentazocine was suppressed by haloperidol, whereas the reduction of the NMDA response by DTG was not affected by haloperidol. 4. This study provides the first in vivo functional evidence that sigma ligands and NPY modulate the NMDA response by acting on distinct receptors, differentiated by their PTX sensitivity.

The relation between the adrenergic neurone-blocking and noradrenaline-depleting actions of some guanidine derivatives.

1 The effects of some guanidine derivatives, (-)-N-(1-phenylethyl) guanidine (PEG), guanethidine and debrisoquine have been investigated on the content and subcellular distribution of noradrenaline in cat spleen and on the overflow of noradrenaline from this organ during stimulation of the splenic nerve.2 PEG and guanethidine, at a dose of 5 mg/kg, produced adrenergic neurone blockade within 15 min and the same dose of debrisoquine produced blockade within 30 minutes.3 All three compounds produced a decrease of similar magnitude in the noradrenaline content of the high-speed particulate (P(2)) and supernatant (S) fractions of splenic homogenates; these actions were temporally correlated with the adrenergic neurone-blocking action of the compounds.4 PEG did not produce any further decrease in the noradrenaline content of the subcellular fractions at times up to 18 h after its administration; adrenergic neurone blockade was maintained throughout this period but had disappeared after 7 days when the noradrenaline content of the subcellular fractions was restored to control levels.5 Guanethidine, in contrast, caused a marked progressive loss of the transmitter from all subcellular fractions-an effect which was maximal 18 h after its administration but continued, as did the adrenergic neurone-blocking action, for at least 72 hours. This additional loss of noradrenaline, over and above that seen after 15 min, is unlikely to be connected with the adrenergic neurone-blocking action of the drug.6 Dexamphetamine both prevented and antagonized the neurone blockade and the subcellular noradrenaline-depleting action of PEG and guanethidine. The restoration of nerve function after administration of dexamphetamine to animals pretreated with 5 mg/kg of either of these compounds was temporally correlated with a selective repletion of the noradrenaline content of the supernatant fraction of the spleen.7 Larger doses (15 mg/kg) of PEG or guanethidine produced a selective depletion of noradrenaline in only the supernatant fraction of the spleen. This depletion was temporally correlated with the adrenergic neurone-blocking action of these compounds. The lack of effect of the compounds at this dose level on the noradrenaline contained in the P(2) fraction may be due to ;stabilization' of the store of noradrenaline in vivo which gives rise to this fraction on homogenization.8 It is suggested that the guanidine-type adrenergic neurone-blocking agents displace the noradrenaline which is readily available for release by nerve impulses, and that it is this action that may account for their sympathomimetic properties.9 The ability of these guanidines to impair the release of noradrenaline by nerve impulses could occur because whilst they are present within the neurone the ;nerve-releasable store', which may in these experiments appear in the supernatant fraction after homogenization, may be unable to refill with transmitter.

Characterization of responses to cromakalim and pinacidil in smooth and cardiac muscle by use of selective antagonists.

1. In dog isolated coronary artery (precontracted with endothelin, 10 nM) cromakalim (0.1-30 microM) and pinacidil (1-30 microM) produced concentration-dependent vasorelaxant responses. The effects of these compounds could be blocked by glibenclamide (3 microM), phentolamine (30 microM) or alinidine (30 microM) to a similar extent, indicating that both agents alter vascular tone through the same mechanism in this preparation. 2. The ability of the antagonists glibenclamide, phentolamine and alinidine to block the response to cromakalim in a number of smooth muscle types from the guniea-pig was determined. Cromakalim (0.1-30 microM) produced concentration-dependent relaxant responses in thoracic aorta (precontracted with endothelin, 30 nM), ileum (precontracted with K+, 25 mM) and trachea (spontaneously contracted). Responses to cromakalim in all tissues could be blocked by the three antagonists. However, significantly higher concentrations of the antagonists were required to block responses in the thoracic aorta than in the ileum or trachea. Given that the rank order of potency of the antagonists was similar in all tissues (i.e. glibenclamide greater than phentolamine = alinidine), this result may suggest vascular K+ channels opened by cromakalim are quantitatively but not qualitatively different in vascular compared with non-vascular smooth muscle. Glibenclamide was approximately 10 times more potent than phentolamine or alinidine. 3. Cromakalim had minimal functional effects on the rat spontaneously beating right atrial (rate) or electrically driven left ventricular strip (force) preparations. Similarly the three antagonists studied failed to alter force generation in the right ventricular strip. However alinidine and phentolamine did produce a dose-related bradycardia in the spontaneously beating right atria. This effect appears to be unrelated to blockade of the K+ channel opened by cromakalim since glibenclamide, the most potent K+ channel antagonist studied, failed to produce the same response. 4. It would appear that the K+ channel opened by cromakalim is present in a number of vascular and non-vascular smooth muscle. Based on the potency of the three antagonists studied, there appears to be little heterogeneity in the process activated by cromakalim in vascular and non-vascular smooth muscle.

Low dose of 1,3-di(2-tolyl)guanidine (DTG) attenuates MK-801-induced spatial working memory impairment in mice.

MK-801 (30-100 micrograms/kg, SC) impaired spontaneous alternation behavior of mice, a behavior related to the spatial working memory. 1,3-Di-(2-tolyl)guanidine (DTG), (+)-pentazocine and (+)-SKF 10,047 (100 micrograms/kg, SC), putative sigma agonists, administered 10 min before MK-801, partially but significantly reversed the impairment, without affecting the concomitant hyperlocomotion. The antagonizing effects by DTG were prevented by BMY-14802 (5 mg/kg, IP), a purported sigma antagonist. These findings suggest that, at low doses, sigma ligands may modulate the N-methyl-D-aspartate dependent memory processes.

Regulation of biosynthesis of guanidinosuccinic acid in isolated rat hepatocytes and in vivo.

To clarify the metabolic pathway of guanidinosuccinic acid (GSA), we investigated the relationship between GSA synthesis and the urea cycle. In isolated rat hepatocytes, GSA formation increased as urea concentration was raised; the effect of urea was not modified by the addition of ammonium chloride. Other urea cycle intermediates, including arginine and cyanate, a degradation product of urea, failed to stimulate GSA synthesis. Ornithine and arginine, which stimulate urea synthesis, strongly inhibited urea-stimulated GSA synthesis in the presence of 10 mM ammonium chloride, but the inhibitory effect of ornithine was not observed when ammonium chloride was not present. Citrulline (5 mM) strongly inhibited urea-stimulated GSA synthesis with or without ammonium chloride. D,L-norvaline, which inhibits urea cycle enzymes, strongly inhibited GSA synthesis. Following urea injection, hepatic GSA levels also increased in vivo, but there was little change in hepatic arginine. However, the addition of ornithine or D,L-norvaline inhibited the production of hepatic GSA, although arginine was increased substantially. These results indicate that GSA synthesis occurs in rat hepatocytes and is stimulated by urea. The data also suggest that the urea cycle enzymes catalyze some of the biochemical reactions in the GSA synthetic pathway.

Sigma receptor-mediated emetic response in pigeons: agonists, antagonists and modifiers.

In order to more fully characterize sigma ligand-induced emesis in the pigeon, the effects of a number of compounds were tested alone or in combination with ditolyguanidine (DTG). The drugs tested could be categorized into three types: agonists, which produced the emetic response (DTG > amitriptyline > BD 737 > thioridazine), antagonists, which effectively antagonized the effects of DTG (haloperidol > BMY 14802 > BD 1139 > chlorpromazine), and agents which did not produce the emetic response on their own, but potently enhanced the emetic effect of DTG (BD-1008 > or = phencyclidine > (+)-n-allylnormetazocine > or = propranolol). Chronic haloperidol resulted in a markedly diminished emetic response to DTG, which returned to control levels by 24.5 days. Haloperidol, but not BMY 14802, was effective in antagonizing the lethal effects of DTG. These data suggest further in vivo evidence for a functional mediation by sigma sites of the emetic response to DTG in the pigeon, and may provide in vivo evidence for potential allosteric modification of sigma ligands.

Camostate- and caerulein-induced delay of gastric emptying in the rat: effect of CCK receptor antagonists.

The effect of camostate, a potent releaser of endogenous cholecystokinin (CCK), and of caerulein, an amphibian peptide mimicking the biological actions of CCK, as well as of selective CCK receptor antagonists on gastric emptying of liquids was studied in the rat. Oral administration of camostate (200 mg/kg with the liquid test meal preceded by the same dose 10 min before the meal) significantly delayed gastric emptying of saline, an effect which was completely blocked by previous administration of the CCKA receptor antagonist, devazepide, at a dose (1 mg/kg i.v.) unable to modify the emptying rate when administered alone. Caerulein (0.03-30 nmol/kg i.v.) also delayed the emptying rate in a dose-dependent manner, with an ID50 of 3.94 nmol/kg. The effect of the peptide was also inhibited by devazepide. The CCKB receptor antagonist, L365,260 (3R-(+)-N-(2,3-dihydro-1-methyl-2-oxo-5-phenyl-1H-1, 4-benzodiazepine-3-yl)-N'-(3-methylphenyl)-urea; 3 mg/kg i.v.), was completely unable to modify the CCK (both endogenous and exogenous)-induced delay in gastric emptying. Repeated (7 days) camostate administration did not modify the gastric motor response to endogenous CCK, thus, suggesting that adaptation did not take place. These results demonstrate that endogenous and exogenous CCK delays gastric emptying of liquids through stimulation of CCKA receptors and suggest that adaptation of the gastric motor response to CCK does not occur.

Cellular pharmacodynamics of the cytotoxic guanidino-containing drug CHS 828. Comparison with methylglyoxal-bis(guanylhydrazone).

N-(6-(4-chlorophenoxy)hexyl)-N'-cyano-N"-4-pyridylguanidine (CHS 828) is a new guanidino-containing compound with antitumoral activity both in vitro and in vivo. Its activity profile differs from those of standard cytotoxic drugs but the mechanism of action is not yet fully understood. CHS 828 is presently in early phase I and II clinical trials. In the present study, the pharmacodynamic effects at the cellular level of CHS 828 was compared to another compound containing two guanidino groups, methylglyoxal-bis(guanylhydrazone) (MGBG). MGBG is known to inhibit the synthesis of polyamines, which are important in, e.g., proliferation and macromolecular synthesis. The concentration-response relationship of CHS 828 closely resembled that of MGBG and the drugs were similar with respect to inhibition of DNA and protein synthesis. On the other hand, CHS 828 induced a significant increase in cellular metabolism while MGBG did not. The cytotoxic effect of MGBG was reversed by the addition of exogenous polyamines, while that of CHS 828 was unaffected. Unlike MGBG, there was also no effect of CHS 828 on the levels of decarboxylating enzymes in the polyamine biosynthesis. In conclusion, CHS 828 does not appear to share any major mechanisms of action with the polyamine synthesis inhibitor MGBG. Further studies will be required to define the exact mechanism of action of CHS 828.

Relaxation by calcitonin gene-related peptide may involve activation of K+ channels in the human uterine artery.

The vasodilatory role of calcitonin gene-related peptide in activating K+ channels was examined in isolated, suffused human uterine arteries. Calcitonin gene-related peptide produced a concentration-dependent relaxation of norepinephrine (1 microM)-induced contractions. Calcitonin gene-related peptide was antagonized by glybenclamide (1-100 microM), an inhibitor of ATP-sensitive K+ channels, but not by tetraethylammonium (1 mM), an inhibitor of calcium(2+)-activated K+ channels. Glybenclamide (10 microM) produced a 6.7 fold and an 11-fold shift to the right of calcitonin gene-related peptide (0.1 to 100 nM) in uterine arteries from pregnant patients (n = 3) and nonpregnant patients (n = 6), respectively. Calcitonin gene-related peptide (10 nM) less effectively (P < 0.05) relaxed contractions produced by KCl (50 mM) (29.4 +/- 1.6%) than by norepinephrine and glybenclamide (10 microM) did not reverse this relaxation (22.2 +/- 6.8%, n = 4 nonpregnant patients). Pinacidil (1 microM), an ATP-sensitive K+ channel opener, relaxed norepinephrine-induced contractions of uterine arteries. Glybenclamide (10 microM) also antagonized pinacidil. These results suggest that calcitonin gene-related peptide relaxes norepinephrine-contracted human uterine arteries, at least in part, by activation of a K+ channel, perhaps of the ATP-sensitive type.