Inhibitory effect of a toxin okadaic acid, isolated from the black sponge on smooth muscle and platelets.

1. Effects of okadaic acid, a toxin isolated from marine sponges, on smooth muscle contraction and platelet activation were examined. 2. Contractions in rabbit aorta induced by high concentrations of K+ and noradrenaline were inhibited by 0.1-1 microM okadaic acid in a concentration-dependent manner. Spontaneous rhythmic contractions as well as high K+-induced contraction in guinea-pig taenia caeci were also inhibited by 1 microM okadaic acid. 3. High K+-induced contraction in rabbit aorta was accompanied by increased Ca2+ influx measured with 45Ca2+ and increased cytosolic Ca2+ [( Ca2+]cyt) measured with fura-2-Ca2+ fluorescence. Okadaic acid inhibited the contraction without inhibiting Ca2+ influx and produced only a small decrease in [Ca2+]cyt. 4. In a saponin-skinned taenia, Ca2+-induced contraction was not inhibited but rather potentiated by okadaic acid. 5. Okadaic acid, 1 microM, inhibited aggregation, ATP release and increased in [Ca2+]cyt induced by thrombin in washed rabbit platelets. Okadaic acid itself did not change the platelet activities. 6. Okadaic acid did not change the cyclic AMP content of rabbit aorta although the inhibitory effects of okadaic acid were similar to those of cyclic AMP. 7. Although the mechanism of the inhibitory effect of okadaic acid was not clarified in the present experiments, it is suggested that okadaic acid acts by inhibiting protein phosphatases resulting in an indirect activation of cyclic AMP-dependent protein phosphorylation.     

The inhibitory effects of 5-hydroxytryptamine on gastric acid secretion by the rat isolated stomach.

1 The effect of 5-hydroxytryptamine (5-HT) on acid secretion by a rat isolated stomach preparation has been studied. 2 5-HT at 10(-5)M in the serosal bathing fluid produced significant inhibition of the acid secretory responses to histamine, pentagastrin and isoprenaline but was without effect on basal secretion or that due to bethanechol, dibutryl cyclic adenosine 3',5'-monophosphate (db cyclic AMP) or phosphodiesterase inhibition with ICI63197. Increasing the concentration of 5-HT to 5 x 10(-5) M did not change this pattern of response whilst 5-HT at 10(-6) M did not cause consistent inhibition. 3 The inhibitory action of 5-HT could be prevented by the antagonist methysergide (2.5 x 10(-5) M). This concentration of methysergide alone did not affect responses to secretagogues or basal acid output. 4 Neither propranolol (2.5 x 10(-5) M) nor tetrodotoxin (10(-6) M) antagonized the inhibitory action of 5-HT. 5 Both indomethacin (2.8 x 10(-5) M) and ibuprofen (2.4 x 10(-4) M) antagonized the action of 5-HT. Indomethacin alone had no effect upon secretagogue responses. 6 5-HT at 10(-5) M had no inhibitory action when applied to the mucosal side of the preparation. 7 The results indicate that 5-HT can act directly on the stomach of the rat to produce inhibition of acid output. This inhibition is selective and may involve the products of cyclo-oxygenase activity.     

Effects of the dopamine receptor agonists, fenoldopam and quinpirole, in the rat stomach

The effects of the DA1-receptor agonist, fenoldopam, and the DA2-receptor agonist, quinpirole, were studied with the longitudinal muscle of rat gastric fundus and circular muscle of rat gastric corpus, as there are contrasting reports about the receptors involved in the inhibitory effect of dopamine in these tissues. Quinpirole had no effect on basal tone in the longitudinal muscle of the rat gastric fundus and did not inhibit the sustained contractions induced by electrical field stimulation or by methacholine. Fenoldopam had no effect on the tone increased by methacholine but slightly potentiated the electrically induced contraction at the highest concentrations; it concentration dependently (10(-7)-3 X 10(-5) M) increased the basal tone. The contractile effect of fenoldopam was clearly antagonized by rauwolscine 10(-6) M, yohimbine 10(-6) M and phentolamine 3 X 10(-6) M plus propranolol 10(-5) M. The 5-HT receptor antagonist, methysergide, antagonized the fenoldopam-induced contractions in a non-competitive way. Fenoldopam and quinpirole had no effect on contractions induced in the circular muscle of the rat gastric corpus by methacholine or electrical field stimulation. They induced some contraction at basal tone, at their highest concentrations. As fenoldopam and quinpirole did not mimic the inhibitory effect observed with dopamine in the same models, no evidence was found for the presence of inhibitory dopamine receptors in rat gastric muscle. The contractile effect of fenoldopam in the longitudinal muscle of the fundus is probably due to an interaction with 5-HT receptors.     

Effect of adenosine receptor agonists and other compounds on cyclic AMP accumulation in forskolin-treated hippocampal slices

The effect of adenosine analogues and some putative neurotransmitters have been studied on cyclic AMP accumulation in rat hippocampal slices treated with the adenylate cyclase activator forskolin. The effects of PGE2 and histamine were potentiated by forskolin (0.1 microM). Isoprenaline and NECA had essentially additive effects with 0.1 microM forskolin and serotonin (above 10(-4) M) inhibited forskolin-stimulated cyclic AMP accumulation. The A1-adenosine receptor selective adenosine analogue R-PIA inhibited forskolin stimulated cyclic AMP accumulation in low doses and stimulated in high. NECA, adenosine and 2-chloroadenosine uniformly stimulated cyclic AMP accumulation. 2',5'-dideoxyadenosine inhibited, but only at high concentrations. Both the stimulatory and the inhibitory effects of R-PIA were antagonized by 8-phenyltheophylline (10 microM). Enprofylline (100 microM) selectively inhibited the stimulatory effect. In the presence of enprofylline both 2-chloroadenosine showed an inhibitory effect on cyclic AMP accumulation. It is concluded that the forskolin-treated rat hippocampal slice is a useful preparation to study both stimulatory and inhibitory effects of transmitters and modulators on adenylate cyclase. The results also show that the rat hippocampus has both A1-receptors that are linked to inhibition of cyclic AMP accumulation and A2-receptors that are linked to stimulation. Furthermore, enprofylline is shown to selectively antagonize the stimulatory response, revealing inhibitory effects of compounds such as 2-chloroadenosine and adenosine.     

Caffeine and related compounds block inhibitory amino acid-gated Cl- currents in freshly dissociated rat hippocampal neurones.

1. The effects of caffeine and related compounds on responses mediated by inhibitory amino acids were investigated in freshly dissociated rat hippocampal pyramidal neurones by conventional and nystatin perforated patch-clamp techniques. 2. Glycine and gamma-aminobutyric acid (GABA) evoked Cl- currents in hippocampal neurones. The half-maximum effective concentrations (EC50) of glycine and GABA were 8.5 x 10(-5) and 5 x 10(-6) M, respectively. 3. Caffeine reversibly inhibited both 10(-4) M glycine- and 10(-5) M GABA-induced Cl-currents in a concentration-dependent manner. The half-maximum inhibitory concentrations (IC50) of caffeine were 4.5 x 10(-4) M for the glycine response and 3.6 x 10(-3) M for the GABA response. 4. Caffeine shifted the concentration-response curve of IGly to the right without affecting the maximum response. 5. The inhibitory action of caffeine did not show voltage-dependency. 6. The blocking action of caffeine was not affected by intracellular perfusion with 5 mM BAPTA or by pretreatment with the protein kinase A inhibitor, H-8. This excludes the participation of Ca2+ or cyclic AMP in the inhibitory action of caffeine. 7. Caffeine failed to inhibit the augmentations of aspartate- and N-methyl-D-aspartate (NMDA) -gated current by glycine, suggesting that caffeine has no effect on the allosteric glycine binding site on the NMDA receptor. 8. The inhibitory effects of some xanthine derivatives on IGly were compared. The inhibitory potency of those compounds on IGly was in the order of pentoxifylline > theophylline > or = caffeine > paraxanthine > IBMX > or = theobromine > dyphylline. Xanthine had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)     

Possible mechanisms of action of the antispasmodic agent tiropramide in the isolated detrusor from rats.

The effects of tiropramide hydrochloride on Ca(2+)-induced contraction, cytoplasmic free Ca2+ levels and tissue cyclic AMP concentrations were investigated to elucidate the mechanisms of its antispasmodic action in the isolated detrusor from rats. Tiropramide inhibited the Ca2+ (3 mM)-induced contractions of the isolated urinary bladder depolarized in a Ca(2+)-free medium, and the IC50 value was 3.3 x 10(-6) M. When tiropramide was added during the sustained phase of the K+ (60 mM)-contracture, IC50 values of tiropramide for the contraction and the increased fluorescence were 1.9 x 10(-5) M and 16.4 x 10(-5) M, respectively. On the other hand, the IC50 values for the K(+)-induced contraction and fluorescence after pretreatment of the isolated urinary bladder with tiropramide were 2.1 x 10(-5) M and 2.6 x 10(-5) M, respectively. Tissue cyclic AMP levels at 1 min after addition of 10(-5) M tiropramide were significantly increased. Papaverine, IBMX or forskolin potentiated the inhibitory effect of tiropramide on carbachol-induced contraction and its cyclic AMP-elevating effect. However, a good correlation between the degrees of potentiation of the inhibitory effect and the increase in cyclic AMP levels was not observed. The present results suggest that the smooth muscle relaxant activity of tiropramide in the isolated detrusor from rats may be intimately associated with predominant inhibition of Ca2+ influx and, to a lesser extent, an increase in intracellular cyclic AMP levels.     

Inhibition of cyclic AMP accumulation by alpha 2-adrenoceptors in the rat cerebral cortex

The effects of alpha 2-adrenoceptor agonist and antagonists on the accumulation of cyclic AMP were examined in rat cerebral cortex slices. Norepinephrine (10(-4) M) caused a 123 +/- 11% increase in the cyclic AMP concentration in the cortical slices, which was greater than the increase (89 +/- 7% increase) caused by isoproterenol (10(-4) M) alone. However, the cyclic AMP response to norepinephrine was completely inhibited by propranolol (10(-4) M), a beta-adrenoceptor antagonist. Yohimbine (10(-7)-10(-5) M), an alpha 2-adrenoceptor antagonist, intensified the cyclic AMP response to norepinephrine by 30%, whereas, clonidine, an alpha 2-adrenoceptor agonist, decreased the response. Treatment with reserpine (3.0 mg/kg) reduced the density of [3H]p-aminoclonidine binding sites (Bmax, 93.8 +/- 18.4 fmol/mg protein) compared to the density in non-treated rats (154.4 +/- 33.5 fmol/mg protein). The potentiating effect of yohimbine and the inhibitory effect of clonidine on the cyclic AMP response to norepinephrine were also reduced. These results suggest that alpha 2-adrenoceptors regulate the accumulation of cyclic AMP in the rat cerebral cortex in an inhibitory fashion. The results also suggest that the accumulation is mediated through beta-adrenoceptors and that this response is intensified by alpha 1-adrenoceptor stimulation.     

Interactions between prostaglandin E2 and inhibitors of platelet aggregation which act through cyclic AMP.

Prostaglandin (PG) E2 potentiates platelet aggregation at low concentrations (10(-8)-10(-6) M). It also inhibits aggregation at a higher concentration (10(-5) M), probably by acting through cyclic adenosine 3',5'-monophosphate (cyclic AMP). The mechanism of this biphasic effect of PGE2 and its implications for thrombosis are not clearly understood. Using a sensitive cyclic AMP assay, in conjunction with platelet aggregation studies, we have examined the interactions between PGE2 and other inhibitors of platelet aggregation which act through cyclic AMP. Low concentrations of PGE2 reversed the inhibition of platelet aggregation and increase in cyclic AMP levels induced by PGI2, PGD2 and adenosine (which stimulate adenylate cyclase (AC) through separate and specific platelet receptors). In contrast, low concentrations of PGE2 added to the inhibition of platelet aggregation and increase in cyclic AMP levels induced by forskolin (which stimulates AC directly) and AH-P 719 and DN-9693 (which inhibit cyclic AMP phosphodiesterase (PDE]. These results suggest that the biphasic effect of PGE2 may be mediated by interaction with two separate platelet receptors. Low concentrations appear to potentiate aggregation by acting at a receptor which is directly coupled to an inhibitory guanine nucleotide-binding protein (Gi), possibly the putative PG endoperoxide receptor. High concentrations of PGE2 appear to inhibit aggregation by acting at an additional receptor, probably the PGI2 receptor. The ease with which PGE2 reverses the effects of PGI2, PGD2 and adenosine, but adds to the effects of AH-P 719 and DN-9693, suggests that PDE inhibitors might offer greater potential than these AC stimulators as an anti-thrombotic strategy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparative inhibitory effects of niflumic acid and novel synthetic derivatives on the rat isolated stomach fundus

Novel derivatives of 2-[3-(trifluoromethyl)-analino]nicotinic acid (niflumic acid) were synthesized. The compounds were compared for their inhibitory effects on 5-hydroxytryptamine (5-HT)- and KCI-induced contraction of the rat fundus. The aim was to assess structure-activity relationships regarding the selectivity and potency of these compounds. Niflumic acid (1-100 microM) concentration-dependently inhibited 5-HT-induced tonic contractions with an IC50 value (concentration reducing the control contractile response by 50%, calculated from semi-log graphs) of 0.24 x 10(4) M (n = 9). In contrast, it was significantly less potent at inhibiting KCl-induced responses (IC50 = 1.49 x 10(4) M, n = 9). The methyl ester (NFAme) and amido (NFAm) analogues showed no selectivity between 5-HT- and KCl-induced contractions with IC50 values of 1.64 x 10(-4) M (n = 8) and 1.87 x 10(-4) M (n = 9) for 5-HT responses, and 2.61 x 10(-4) M (n = 8) and 2.55 x 10(-4) M (n = 7) for KCl-induced responses, respectively. Our results suggest that alteration of the carboxylic acid moiety of niflumic acid reduces the selectivity and potency of its inhibitory action on 5-HT-induced contractile responses of the rat fundus, possibly via a reduced interaction with calcium-activated chloride channels.     

Vasodilator effects of sodium nitroprusside, levcromakalim and their combination in isolated rat aorta

1. The endothelial modulation of the relaxant responses to the nitric oxide (NO) donor sodium nitroprusside (SNP) and the KATP channel opener levcromakalim (LEM) and the interactions between these agents were analysed in isolated rat aorta. 2. LEM-induced relaxation was unchanged by endothelium removal or by the presence of L-NAME (10-4 M) or ODQ (10-6 M). In contrast, in KCl- (25 mM), but not in noradrenaline- (NA, 10-6 M) contracted arteries, SNP-induced relaxation was augmented by endothelium removal but not by L-NAME, indomethacin, glibenclamide nor charybdotoxin plus apamin. 3. The isobolographic analysis of the interactions between exogenously activated KATP channels and cyclic GMP using mixtures of SNP and LEM revealed that there were no interactions between both drugs at the proportions at which both drugs were active. However, the points for the SNP : LEM mixtures in proportions 10:1 and 1:10,000 (i.e. at concentrations at which LEM and SNP were inactive, respectively) fell significantly above the line of additivity indicating that there were negative interactions between both drugs at these selected proportions (about 5- and 2 fold inhibition, respectively). The former interaction was sensitive to glibenclamide, whereas the latter was insensitive ODQ. The magnitude of the 10:1 SNP:LEM interaction was smaller in endothelium-intact arteries and was absent in arteries stimulated by NA. 4. In conclusion, the relaxations induced by LEM and SNP were additive. However, the presence of endothelium and low concentrations of LEM inhibited SNP-induced relaxation. Both inhibitory effects were not additive and were only observed in KCl- and not in NA-contracted aortae.     

Inhibitory effect of N-(3,4-dimethoxycinnamoyl)anthranilic acid on release of SRS from alveolar macrophages in vitro

The effects of N-(3,4-dimethoxycinnamoyl)anthranilic acid (N-5') on the release of the slow-reacting substance (SRS) by zymosan- or Ca ionophore-stimulated rat and human alveolar macrophages (AM) were examined in vitro. Disodium cromoglycate (DSCG) was used as a control. N-5' at concentrations of 10(-4)-10(-3) M significantly inhibited the release of SRS from both rat and human AM stimulated by zymosan. N-5' had almost the same inhibitory effect when added to the AM culture system at any time from 180 min before to 30 min after the addition of zymosan. N-5' (10(-4)-10(-3) M) also significantly inhibited the release of SRS by Ca ionophore-stimulated rat AM. N-5' (10(-6)-10(-3) M) had no significant effect on phagocytosis of yeast particles by rat AM. DSCG (10(-6)-10(-3) M) did not inhibit the release of SRS from the zymosan-stimulated rat AM. N-5' was concluded to have a relatively specific inhibitory effect on the non-immunological release of SRS from stimulated AM. It is postulated that N-5' inhibits the process of release of SRS from AM by acting after the initial stage.     

Implication of cyclic nucleotide phosphodiesterase inhibition in the vasorelaxant activity of the citrus-fruits flavonoid (+/-)-naringenin

The potential vasorelaxant, antioxidant and cyclic nucleotide phosphodiesterase (PDE) inhibitory effects of the citrus-fruit flavonoids naringin and (+/-)-naringenin were comparatively studied for the first time in this work. (+/-)-Naringenin (1 microM - 0.3 mM) did not affect the contractile response induced by okadaic acid (OA, 1 microM). However, (+/-)-naringenin relaxed, in a concentration-dependent manner, the contractions elicited by phenylephrine (PHE, 1 microM) or by a high extracellular KCl concentration (60 mM) in intact rat aortic rings. Mechanical removal of endothelium and/or pretreatment of aorta rings with glibenclamide (GB, 10 microM) or tetraethylammonium (TEA, 2 mM) did not significantly modify the vasorelaxant effects of this flavanone. (+/-)-Naringenin (10 microM - 0.1 mM) did not alter the basal uptake of 4) Ca2+ but decreased the influx of 45Ca2+ induced by PHE and KCl in endothelium-containing and endothelium-denuded rat aorta. (+/-)-Naringenin (10 microM - 0.1 mM) was ineffective to scavenge superoxide radicals (O*2-) generated by the hypoxanthine (HX)-xanthine oxidase (XO) system and/or to inhibit XO activity. (+/-)-Naringenin (0.1 mM) significantly increased the production of cGMP and cAMP decreased by PHE (1 microM) and high KCl (60 mM) in cultured rat aortic myocytes. (+/-)-Naringenin preferentially inhibited calmodulin (CaM)-activated PDE1, PDE4 and PDE5 isolated from bovine aorta with IC50 values of about 45 microM, 60 microM and 68 microM, respectively. In contrast, the 7-rhamnoglucoside of (+/-)-naringenin, naringin (1 microM - 0.3 mM), was totally inactive in all experiments. These results indicate that the vasorelaxant effects of (+/-)-naringenin seem to be basically related to the inhibition of PDE1, PDE4 and PDE5 activities.     

Insurmountable antagonism of AT-1015, a 5-HT2 antagonist,on serotonin-induced endothelium-dependent relaxation in porcine coronary artery

The purpose of this study was to examine the inhibitory effects of AT-1015, a newly synthesized 5-HT(2) receptor antagonist, on serotonin-induced endothelium-dependent relaxation in U 46619 (5 x 10(-9)M)-precontracted porcine coronary artery pre-incubated with ketanserin (3 x 10(-6)M), and then compare its effects with another potent 5-HT(2) antagonist, ritanserin. The investigation showed that AT-1015 (10(-8)-10(-6)M) caused rightward shift with significant inhibition of maximum relaxation response induced by serotonin in porcine coronary artery with endothelium. Ritanserin caused a rightward shift of serotonin-induced relaxation without decreasing maximum response at 10(-9) and 10(-8)M, but it inhibited the maximum relaxation response at 10(-7)M. The study showed that AT-1015 and ritanserin had no inhibitory effect on bradykinin-induced relaxation in porcine coronary artery with endothelium. Thus, these findings suggested that AT-1015 at concentrations of 10(-8)-10(-6)M caused noncompetitive blockade of serotonin-induced endothelium-dependent relaxation in porcine coronary artery. The antagonistic effects of AT-1015 on serotonin-induced relaxation were different from that of ritanserin, except at 10(-7)M ritanserin. The variation of inhibitory effects between these two 5-HT(2) antagonists may be due to the different chemical structure and/or interaction sites at the receptor.     

Effect of NAD,nicotinamide and nicotinic acid on cyclic AMP accumulation by fat cells

Summary The addition of exogenous nicotinamide adenine dinucleotide (NAD) to isolated rat fat cells at concentrations between 1 and 10 mol markedly inhibited the rise in cyclic AMP accumulation due to norepinephrine. Catabolism of NAD to adenosine did not account for the inhibitory effect since NAD was effective in the presence of adenosine deaminase. The bovine fraction V albumin in which fat cells were incubated contained a substantial amount of glycohydrolase which cleaved NAD to nicotinamide. Incubation of fat cells in albumin-free medium did not abolish the inhibitory effect of NAD, and under these conditions there was little degradation of NAD to nicotinamide during the first 10 min of incubation. Fat cells cleave NAD, in the medium to nicotinamide mononucleotide which does not inhibit cyclic AMP accumulation. NADH, NADP and nicotinic acid mononucleotide were less effective as inhibitors of cyclic AMP accumulation than NAD or deamido NAD. Ineffective as inhibitors were NADPH, nicotinamide mononucleotide, and 1-methyl nicotinamide. N-methyl nicotinamide was as potent as nicotinamide while nicotinic acid was 100- to 500fold more effective than either compound as an inhibitor of cyclic AMP accumulation. Incubation of fat cells with 100 M but not 10 M NAD for 5 min resulted in a decrease in the subsequent accumulation of cyclic AMP by rat fat cell ghosts.Supported by USPHS research grant AM-10149 from the National Institute of Arthritis, Metabolism and Digestive Diseases     

Role of central glutamate receptors, nitric oxide and soluble guanylyl cyclase in the inhibition by endotoxin of rat gastric acid secretion

1. This study examines the role of a central pathway involving glutamate receptors, nitric oxide (NO) and cyclic GMP in the acute inhibitory effects of low doses of peripheral endotoxin on pentagastrin-stimulated acid production. 2. Vagotomy or intracisternal (i.c.) microinjections of the NO-inhibitor, N(G)-nitro-L-arginine methyl esther (L-NAME; 200 microg rat(-1)) restored acid secretory responses in endotoxin (10 microg kg(-1), i.v.)-treated rats. 3. The acid-inhibitory effect of i.v. endotoxin (10 microg kg(-1), i.v.) was prevented by prior i.c. administration of the NMDA receptor antagonists, dizocilpine maleate (MK-801; 10 nmol rat(-1)) and D-2-amino-5-phosphono-valeric acid (AP-5; 20 nmol rat(-1)), or the AMPA/kainate antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX; 10 nmol rat(-1)). However, the competitive metabotropic glutamate receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine (MCPG; 20 - 1000 nmol rat(-1)) did not antagonize the effects of endotoxin. 4. I.c. administration of L-glutamate (0.1 nmol rat(-1)) inhibited pentagastrin-stimulated gastric acid secretion. Coadministration with L-NAME (200 microg rat(-1)) prevented the inhibition of gastric acid secretion by the aminoacid. 5. I.c. administration of 1H-[1,2, 4]Oxazodiolo[4,3-a]quinoxalin-1-one (ODQ; 100 nmol rat(-1)), a soluble guanylyl cyclase (sGC) blocker, reversed the hyposecretory effect of endotoxin. 6. I.c. administration of the cyclic GMP analogue 8-Bromoguanosine-3,5-cyclic monophosphate (8-Br-cGMP; 100 - 300 nmol rat(-1)) reduced gastric acid production in a dose-dependent manner. 7. We conclude that central NMDA and AMPA/kainate receptors are involved in the acid inhibitory effect of peripherally administered endotoxin. This central pathway involves synthesis of NO, which acts on the enzyme sGC.     

A comparison of the effects of the dopamine partial agonists aripiprazole and (-)-3-PPP with quinpirole on stimulated dopamine release in the rat striatum: Studies using fast cyclic voltammetry in vitro

The effects of aripiprazole, (-)-(3-hydroxyphenyl)-N-n-propylpiperidine ((-)-3-PPP) and quinpirole on single and multiple pulse stimulated dopamine release were investigated using the technique of fast cyclic voltammetry (FCV) in isolated rat striatal slices. Aripiprazole and (-)-3-PPP had no significant effect on single pulse dopamine release at concentrations from 10nM to 10μM indicating low agonist activity. The compounds failed to potentiate 5 pulse stimulated release of dopamine although inhibitory effects were seen at 10μM for aripiprazole. Both compounds were tested against the concentration-response curve for quinpirole's inhibition of stimulated single pulse dopamine release. Aripiprazole and (-)-3-PPP shifted the concentration-response curve for quinpirole to the right. In each case this was greater than a 100-fold shift for the 10μM test compound. Whilst these results indicate that both compounds show little agonist activity on dopamine release and significant antagonism of the inhibitory effect of quinpirole on dopamine release, whether they are functionally selective dopamine D(2) ligands remains controversial.     

Modulatory action of prostaglandin D2 on the release of 3H-norepinephrine from rat cerebellar slices

The modulatory action of prostaglandin D2 (PGD2) on the high-K+-induced release of 3H-norepinephrine (3H-NE) from rat cerebellar slices was investigated in relation to the presynaptic feedback mechanisms for the NE release. PGD2 (10(-7)-10(5) M) dose-dependently suppressed the release of 3H-NE. The 3H-NE release was also dose-dependently decreased by 10(-10)-5 X 10(-9) M clonidine and increased by 10(-9)-10(-6) M yohimbine, and there was an antagonism between clonidine and yohimbine, indicating the presence of an alpha 2-adrenoceptor-mediated negative feedback mechanism in the rat cerebellum. The inhibitory action of clonidine was not additive to that of PGD2, while there appeared to be an additiveness between the effects of PGD2 and yohimbine. The 3H-NE release was increased by I-isoproterenol and decreased by I-propranolol, but only at concentrations higher than 10(-6) M. PGD2 nearly abolished the actions of these beta-adrenergic agents, and the 3H-NE release remained at a level similar to that induced by 10(-5) M PGD2 alone. Based on these results, it was tentatively suggested that PGD2 inhibits the 3H-NE release by a mechanism independent of adrenoceptor-mediated feedback mechanisms.     

The effect of prolonged treatment with tricyclic antidepressants on the actions of 5-hydroxytryptamine in the hippocampal slice of the rat

The effect of long-term treatment with the tricyclic antidepressants imipramine (IMI) and desmethylimipramine (DMI) on neuronal responsiveness to 5-hydroxytryptamine (5-HT) was examined in the hippocampal slice preparation from the rat. Population spikes, evoked by electrical stimulation of the stratum radiatum, were recorded in the pyramidal cell layer of the CA1 region of the isolated hippocampus. When 5-HT (10(-7) to 2 X 10(-5) M) was applied there was an initial increase followed by a decrease in the amplitude of the population spike. On washout of 5-HT the amplitude increased transiently above control levels. Daily injection of 10 mg/kg of imipramine or desmethylimipramine, intraperitoneally, into rats for 4-5 weeks was found to produce a significant decrease in the inhibitory effect of 10(-5) M 5-HT, whereas there was no apparent change in the excitatory effects. The acute application of 10(-5) M imipramine or desmethylimipramine antagonized the inhibitory effect of 10(-5) M 5-HT without affecting the excitatory effects. Acute application of the 5-HT receptor antagonists cyproheptadine (10(-5) M) and ketanserin (7.5 X 10(-6) M) completely prevented the appearance of the inhibitory effect of 10(-5) M 5-HT without affecting the excitatory effects. It was concluded that the decreased inhibitory effect of 5-HT which was produced by chronic treatment with imipramine or desmethylimipramine was probably due to a reduction in the number of 5-HT receptors or a reduction in the post-receptor effector mechanisms for 5-HT.     

Contrasting effects of calyculin A and okadaic acid on the respiratory burst of human neutrophils

The involvement of serine/threonine protein-phosphatases in the production of superoxide (respiratory burst) by human neutrophils was investigated using calyculin A, a potent inhibitor of both protein phosphatases type 1 and 2A, and okadaic acid, which preferentially inhibits protein phosphatase type 2A. Treatment of neutrophils with calyculin A (25–75 nM) or okadaic acid (1–4 μM) had no stimulatory effect but potently enhanced total superoxide production induced by an optimal fMLP (N-formyl-methionyl-leucoyl-phenylalanine) concentration (0.1 μM). The maximal increase platacuaed with 50–75 nM calyculin A and 2–4 μM okadaic acid, reaching approximately 120 and 200% of control values, respectively. Unlike calyculin A, okadaic acid also primed the initial rate of superoxide production, suggesting that protein phosphatases may down-regulate both initiation and termination of respiratory burst. Optimal stimulation of the respiratory burst by PMA (160 nM) was inhibited by calyculin A and okadaic acid, with an IC₅₀ of 60 nM and 2 μM, respectively, although both drugs caused protein hyperphosphorylation. The inhibition was partially prevented by a nonstimulatory concentration of A23187, indicating a role of calcium in the inhibitory effects of the drugs. Unlike the optimal respiratory burst, suoptimal respiratory burst induced by PMA (1–7 nM) was enhanced by calyculin A and okadaic acid. Unprimed and primed respiratory burst were depressed by a selective antagonist of protein kinase C (GF 109203X), indicating positive regulation of these responses by protein kinase C. Thus, the use of calyculin A and okadaic acid distinguishes two regulatory processes of superoxide production. The respiratory burst induced by low PMA concentrations of fMLP was up-regulated by both calyculin A and okadaic acid, in keeping with a down-regulatory role of protein phosphatases in these responses. By contrast, intense protein kinase C activation by PMA triggered a respiratory burst which was depressed by both drugs, pointing to positive regulation of the respiratory burst by protein phosphatases.     

Release of endothelium-derived relaxing factor from pig cultured aortic endothelial cells, as assessed by changes in endothelial cell cyclic GMP content, is inhibited by a phorbol ester.

1. Cultured aortic endothelial cells of the pig respond to the endothelium-derived relaxing factor (EDRF) they release with an increase in cyclic GMP content. This response is inhibited by haemoglobin or by L-NG-monomethyl-arginine (L-NMMA), and has been used to investigate the effects of phorbol esters on EDRF release. 2. Pretreatment with phorbol-12,13-dibutyrate (PDB) but not the inactive 4 alpha-phorbol-12,13,-didecanoate (PDD), inhibited increases in cyclic GMP induced by substance P (10(-8) M) in a time and concentration-dependent manner. PDB did not affect basal cyclic GMP levels. 3. PDB (3 x 10(-7) M), but not PDD (3 x 10(-7) M), also inhibited ATP (10(-5) M)-induced increases in cyclic GMP, but did not affect those induced by bradykinin (10(-7) M). 4. Increases in cyclic GMP induced by low (10(-7) M) but not high (10(-6) M) concentrations of the calcium ionophore A23187 were inhibited by PDB (3 x 10(-7) M). This inhibitory effect was due to enhanced destruction of EDRF by superoxide anions rather than inhibition of EDRF release, as the inhibition was abolished in the presence of superoxide dismutase (SOD, 30 mu ml-1) and catalase (CAT, 100 mu ml-1). 5. SOD and CAT did not affect the inhibitory action of PDB on substance P or ATP-induced increases in cyclic GMP. 6. Increases in endothelial cell cyclic GMP content induced by sodium nitroprusside (10(-5) M) were unaffected by PDB pretreatment.(ABSTRACT TRUNCATED AT 250 WORDS)