European Journal of Pharmacology (v.388, #2)

LY354740 {(1S,2S,5R,6S)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylate monohydrate}, a selective group II metabotropic glutamate (mGlu) receptor agonist, was recently reported to attenuate the behavioral effects of phencyclidine (PCP) in rats. In the present study, LY354740 failed to attenuate the discriminative stimulus properties of PCP and its disruption of prepulse inhibition of the acoustic startle response, at a dose range which decreased startle responding. The suggestion that mGlu group II receptor activation induces antipsychotic effects may be premature.
Keywords: Drug discrimination; Glutamate receptor; Schizophrenia;

The conserved aspartic acid that is required for ligand binding to the dopamine D2 receptor is followed by three tandem sulfur-containing amino acids. While previous point mutation studies did not reveal any single one of these residues as being critical for ligand binding, we now show that simultaneously substituting all three with isovolumetric, non sulfur-containing amino acids results in large decreases in the binding affinity for dopamine, (−)-raclopride and 7-(-4(4-(2,3-dichlorophenyl)-1-piperazinyl)butyloxy)-3,4-dihydro-2(1H)-quinolinone (aripiprazole), but not for methylspiperone or allosteric modulators.
Keywords: Mutagenesis; Allosteric modulator; Catecholamine;

The dopamine D1 receptor agonist SKF-82958 serves as a discriminative stimulus in the rat by Colin N Haile; Galen Carey; Geoffrey B Varty; Vicki L Coffin (125-131).
We examined the discriminative stimulus effects of the high-efficacy dopamine D1 receptor agonist (±)6-chloro-7,8-dihydroxy-3-ally1-phenyl-2,3,4,5-tetrahydro-1H-3benzazepine hydrobromide (SKF-82958) in rats trained to discriminate SKF-82958 (0.03 mg/kg) from vehicle in a two-lever food-reinforced drug discrimination task. SKF-82958 produced dose-related increases in responding to the SKF-82958 appropriate lever with full substitution occurring at the training dose. Pretreatment with the dopamine D1/D5 receptor antagonist (−)-trans-6,7,7a,8,9,13b-hexahydro-3-chloro-2hydroxy-N-methyl-5H-benzo-[d]naphtho-{2,1-b}azepine (SCH-39166) (0.01 mg/kg) attenuated the discriminative stimulus effects of SKF-82958. Pretreatment with the dopamine D2 receptor antagonist raclopride (0.03 mg/kg) had no effect. The high-efficacy dopamine D1 receptor agonist R(+)6chloro-7,8-dihydroxy-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrobromide (SKF-81297) fully substituted for SKF-82958, whereas the low-efficacy dopamine D1 receptor agonist (±)1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazepine-7,8-diol hydrochloride (SKF-38393) produced only partial substitution. The dopamine D2 receptor agonist trans-(±)-4,4a,5,6,7,8,8a,9-octahydro-5-propyl-1H-propyl-1H-pyrazolo[3,4-g]quinoline dihydrochloride (quinpirole) and the indirect dopamine agonist cocaine did not substitute fully for the SKF-82958 discriminative stimulus cue. These results demonstrate that the high-efficacy dopamine D1 receptor agonist SKF-82958 can serve as an effective discriminative stimulus in the rat, and that these effects are mediated by a dopamine D1-like receptor mechanism.
Keywords: Dopamine; Dopamine D1 receptor agonist; Drug discrimination; SKF-82958; SCH-39166; Cocaine; (Rat);

Adrenomedullin decreases extracellular signal-regulated kinase activity through an increase in protein phosphatase-2A activity in mesangial cells by Narayanan Parameswaran; Ponnal Nambi; Carolyn S Hall; David P Brooks; William S Spielman (133-138).
Adrenomedullin is a recently identified peptide hormone that has receptors in a number of different systems including renal mesangial cells. We reported recently that adrenomedullin can cause a decrease in extracellular signal-regulated kinase (ERK) activity and increase jun amino-terminal kinase (JNK) and P38 mitogen-activated protein kinase (P38 MAPK) acitivities in rat mesangial cells. Associated with these responses we also reported that adrenomedullin can decrease proliferation and increase apoptosis in mesangial cells. The major aim of the present study was to examine the mechanism of decrease in ERK activity by adrenomedullin and to identify the role of protein phosphatase 2A (PP2A) in the decrease in ERK activity, using okadaic acid [9,10-Deepithio-9,10-didehydroacanthifolicin], a selective inhibitor of PP2A at low nanomolar concentrations. The adrenomedullin-induced decrease in [3H]-thymidine incorporation and increase in apoptosis were reversed by okadaic acid at the concentration that selectively inhibits PP2A. Okadaic acid completely reversed the ERK inhibition caused by adrenomedullin, suggesting that PP2A may be involved in the adrenomedullin-mediated changes in proliferation, apoptosis and ERK activity. PP2A activity in mesangial cells was increased over time following exposure to adrenomedullin. The tyrosine phosphorylation of ERK did not change significantly following adrenomedullin treatment although the ERK activity was decreased significantly. This suggests that the decrease in ERK activity is not mediated through a decrease in MEK (a dual phosphorylating kinase upstream of ERK) or by an increase in MKP-1/2 (a dual specificity phosphatase) activities. Thus we conclude that the mechanism of adrenomedullin-induced decrease in ERK activity in rat mesangial cells is at least in part mediated by an increase in PP2A activity.
Keywords: Okadaic acid; Adrenomedullin; ERK (extracellular signal-related kinase); JNK (jun amino-terminal kinase); P38 MAPK (P38 mitogen-activated protein kinase);

σ Receptor ligands attenuate N-methyl-d-aspartate cytotoxicity in dopaminergic neurons of mesencephalic slice cultures by Seiichiro Shimazu; Hiroshi Katsuki; Chikako Takenaka; Michiko Tomita; Toshiaki Kume; Shuji Kaneko; Akinori Akaike (139-146).
We investigated the potential neuroprotective effects of several σ receptor ligands in organotypic midbrain slice cultures as an excitotoxicity model system. When challenged with 100-μM N-methyl-d-aspartate (NMDA) for 24 h, dopaminergic neurons in midbrain slice cultures degenerated, and this was prevented by (5R, 10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,b]-cyclohepten-5,10-imine (MK-801; 1–10 μM). Concomitant application of ifenprodil (1–10 μM) or haloperidol (1–10 μM), both of which are high-affinity σ receptor ligands, significantly attenuated the neurotoxicity of 100 μM NMDA. The σ1 receptor-selective ligand (+)-N-allylnormetazocine ((+)-SKF 10047; 1–10 μM) was also effective in attenuating the toxicity of NMDA. The effect of R(−)-N-(3-phenyl-1-propyl)-1-phenyl-2-aminopropane hydrochloride ((−)-PPAP), a σ receptor ligand with negligible affinity for the phencyclidine site of NMDA receptors, was also examined. (−)-PPAP (3–100 μM) caused a concentration-dependent reduction of NMDA cytotoxicity, with significant protection at concentrations of 30 and 100 μM. In contrast, (+)-SKF 10047 (10 μM) and (−)-PPAP (100 μM) showed no protective effects against cell death induced by the Ca2+ ionophore ionomycin (1–3 μM). These results indicate that σ receptor ligands attenuate the cytotoxic effects of NMDA on midbrain dopaminergic neurons, possibly via inhibition of NMDA receptor functions.
Keywords: σ Receptor; Organotypic slice culture; Mesencephalon; N-methyl-d-aspartate (NMDA); Dopaminergic neuron;

Effect of glipizide on dopamine synthesis, release and metabolism in PC12 cells by Itschak Lamensdorf; Le-Ping He; Amotz Nechushtan; Judith Harvey-White; Graeme Eisenhofer; Rusnak Milan; Eduardo Rojas; Irwin J Kopin (147-154).
Sulfonylureas block ATP-dependent K+ channels (K/ATP channels) in pancreatic β cells and brain γ-aminobutyric acid (GABA) containing neurons causing depolarization-evoked insulin or GABA release. In high concentrations, sulfonylureas also inhibit catecholamine release from bovine adrenal chromaffin cells and isolated guinea pig aorta. In this study, we examined the effect of glipizide, a sulfonylurea, on dopamine release from PC12 cells and found that neither basal nor K+-stimulated dopamine release was affected. Although PC12 cells expressed mRNA for the K/ATP channel, functional K/ATP channels could not be demonstrated electrophysiologically, consistent with the lack of effect of glipizide on dopamine release. Glipizide did, however, increase cytoplasmic retention of the acidic dopamine metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), indicating blockade of their outward transport. The cellular accumulation of DOPAC was accompanied by reduced tyrosine hydroxylase activity and reduced formation of dopamine and its metabolites presumably by a negative feedback effect of the increased cytoplasmic concentrations of DOPAC.
Keywords: Dopamine; Tyrosine hydroxylase; Sulfonylurea; Glipizide; DOPAC (3,4-dihydroxyphenylacetic acid); HVA (homovalinic acid);

Characterization and modulation of [125I]iberiotoxin-D19Y/Y36F binding in the guinea-pig urinary bladder by Eduardo J Molinari; James P Sullivan; Yieh-Ping Wan; Jorge D Brioni; Murali Gopalakrishnan (155-161).
The radioligand binding characteristics of the Ca2+-activated K+ channel ligand [125I]iberiotoxin-D19Y/Y36F were examined in guinea-pig urinary bladder membranes. Saturation analysis revealed a single class of high affinity binding sites in the bladder with a K D value of 45.6 pM and a B max value of 112 fmol/mg protein. Specific binding was displaced by unlabeled iberiotoxin and penitrem A, but not by blockers of other classes of K+ channels including α-dendrotoxin, margatoxin and apamin. The indole alkaloids, paxilline and verruculogen, significantly increased binding by 4.5- and 4.3-fold, respectively. Tetraacetic acid derivatives such as ethylenediamine tetraacetic acid and ethyleneglycoltetraacetic acid enhanced specific [125I]iberiotoxin-D19Y/Y36F binding about 2.5-fold, which was not attributable to calcium chelation. This increase was due to a significant change in ligand binding affinity (K D=6.3 pM), but not due to a change in the B max, indicating that these compounds may enhance toxin binding via allosteric interactions. Collectively, these results demonstrate that the binding sites for [125I]iberiotoxin-D19Y/Y36F present in the urinary bladder shows a pharmacological profile typical of maxi-K+ channels and can be modulated, not only by previously known indole alkaloids, but also by tetraacetic acid analogs.
Keywords: Maxi-K+ channel; Radioligand binding; Iberiotoxin; Urinary bladder;

Topiramate potentiates the antiseizure activity of some anticonvulsants in DBA/2 mice by Giovambattista De Sarro; Santo Gratteri; Federico Bonacci; Sebastiano A Musumeci; Maurizio Elia; Angela De Sarro (163-170).
Topiramate (1–50 mg/kg, intraperitoneally (i.p.)) was able to antagonize audiogenic seizures in DBA/2 mice in a dose-dependent manner. Topiramate at dose of 2.5 mg/kg i.p., which per se did not significantly affect the occurrence of audiogenic seizures in DBA/2 mice, potentiated the anticonvulsant activity of carbamazepine, diazepam, felbamate, lamotrigine, phenytoin, phenobarbital and valproate against sound-induced seizures in DBA/2 mice. The degree of potentiation induced by topiramate was greatest for diazepam, phenobarbital and valproate, less for lamotrigine and phenytoin and not significant for carbamazepine and felbamate. The increase in anticonvulsant activity was associated with a comparable increase in motor impairment. However, the therapeutic index of the combination of all drugs+topiramate was more favourable than that of antiepileptics+ saline, with the exception of carbamazepine or felbamate+topiramate. Since topiramate did not significantly influence the total and free plasma levels of the anticonvulsant drugs studied, we suggest that pharmacokinetic interactions, in terms of total or free plasma levels, are not probable. However, the possibility that topiramate can modify the clearance from the brain of the anticonvulsant drugs studied cannot be excluded. In addition, topiramate did not significantly affect the hypothermic effects of the anticonvulsants tested. In conclusion, topiramate showed an additive effect when administered in combination with some classical anticonvulsants, most notably diazepam, phenobarbital, lamotrigine, phenytoin and valproate.
Keywords: Epilepsy; Topiramate; Carbamazepine; Phenytoin; Valproate; Felbamate; Anticonvulsant; Audiogenic seizure; DBA/2 mouse;

Effects of zatebradine and propranolol on canine ischemia and reperfusion-induced arrhythmias by Hisaya Naito; Yasuyuki Furukawa; Daisuke Chino; Chikaomi Yamada; Keitaro Hashimoto (171-176).
1,3,4,5-Tetrahydro-7,8-dimethoxy-3[3-[[2-(3,4-dimethoxyphenyl)-ethyl]methylamino]propyl]-2H-3-benzazepin-2-one-hydrochloride (Zatebradine) is a specific bradycardiac agent, blocking the hyperpolarization-activated pacemaker current (I f), and thus has no negative inotropic effect. The purpose of this study was to examine whether zatebradine is effective against ischemia and reperfusion-induced arrhythmias in dogs compared to propranolol. Arrhythmia was induced by ligation of the left anterior descending coronary artery followed by reperfusion. Ischemia-induced biphasic arrhythmias were suppressed in both zatebradine and propranolol groups. During ischemia, fatal ventricular fibrillation occurred in four dogs in the control group, 0 in the zatebradine group, and two dogs in the propranolol group. Of the 31 dogs subjected to reperfusion, mortality rates in the zatebradine, propranolol, and control groups were 56%, 75%, and 86%, respectively, and there were no significant differences. In the heart beating 10 beats/min faster than the predrug heart rate by atrial pacing, both zatebradine and propranolol attenuated ischemia-induced arrhythmias but did not affect reperfusion arrhythmias. Our results suggest that I f and/or β-adrenoceptors rather than the bradycardiac action might be related to the antiarrhythmic effects during ischemia, but that they do not play a role in the generation of the reperfusion-induced ventricular arrhythmias.
Keywords: Zatebradine; Ischemia-induced arrhythmia; Reperfusion arrhythmia; Propranolol; (Dog);

Characterization of bradykinin B2 receptor antagonists in human and rat urinary bladder by Stefania Meini; Riccardo Patacchini; Sandro Giuliani; Massimo Lazzeri; Damiano Turini; Carlo Alberto Maggi; Alessandro Lecci (177-182).
The effect of three selective bradykinin B2 receptor antagonists, MEN11270 (H-dArg-Arg-Pro-Hyp-Gly-Thi-c(Dab-dTic-Oic-Arg)c(7γ–10α)), Icatibant (H-dArg-Arg-Pro-Hyp-Gly-Thi-Ser-dTic-Oic-Arg-OH), and FR173567 ((E)-3-(6-acetamido-3-pyridyl)-N-[N-[2,4-dichloro-3-[(2-methyl-8-quinolinyl) oxymethyl] phenyl]-N-methylaminocarbonylmethyl]acrylamide) was evaluated in the human and rat urinary bladder in vitro and in vivo in anaesthetized rats. Bradykinin evoked a concentration-dependent contraction of human (pD2=7.2) and rat (pD2=7.7) detrusor muscle strips. In human preparations, all the antagonists tested produced a rightward-shift in the concentration–response curve for bradykinin. Schild plot analysis yielded pKB values of 8.4, 8.4 and 8.6 for MEN11270, Icatibant, and FR173567, respectively. In the rat preparations the three antagonists (at 100 nM concentration), produced a shift to the right which gave apparent pA2 values of 8.2, 8.0 and 8.1 for MEN11270, Icatibant, and FR173567, respectively. In anaesthetized rats, both MEN11270 and Icatibant (1–10 nmol/kg i.v.) dose dependently reduced the bradykinin (100 nmol/kg i.v.)-induced urinary bladder contraction, their effect being prompt and long-lasting. In contrast, FR173567 (100 nmol/kg i.v.) produced a partial and short-lasting inhibition of bradykinin-induced bladder contractions. The present findings indicate that all the antagonists tested recognize with similar potencies the bradykinin B2 receptors expressed in the detrusor muscle of both humans and rats. MEN11270 and Icatibant possess a higher potency and longer duration of action in vivo than FR173657, suggesting that the activity of this non-peptide antagonist in vivo is hampered by factors unrelated to its affinity for bradykinin B2 receptors.
Keywords: Bradykinin B2 receptor; Bradykinin receptor antagonist; Cystitis; MEN 11270; Urinary bladder; Urinary bladder;

Nociceptin/orphanin FQ, the endogenous ligand of the opioid receptor-like (ORL1) receptor, caused contractions in the isolated colon of the mouse. Tetrodotoxin and the nitric oxide (NO) synthase inhibitor Nω-nitro-l-arginine also produced contractions which were quantitatively similar to those seen in response to nociceptin. In the presence of either tetrodotoxin or Nω-nitro-l-arginine, nociceptin was unable to cause a further contraction, whereas the muscarinic receptor agonist carbachol elicited a contractile response. Nociceptin had no contractile activity in colonic preparations contracted by Nω-nitro-l-arginine then relaxed by addition of the NO donor sodium nitroprusside. These data suggest that nociceptin causes contractions of the mouse proximal colon by inhibiting the tonic, neuronal release of NO.
Keywords: Nociceptin; ORL1 receptor; Colon, mouse, isolated; Nitric oxide (NO);

The effect of endothelin-1 on lipopolysaccharide-induced cyclooxygenase 2 expression in association with prostaglandin E2 by Keiji Shimada; Taizo Kita; Yukio Yonetani; Akio Suzumura; Toshikatsu Nakashima (187-194).
We demonstrated previously that endothelin-1 (10−14 to 10−8 M) promotes lipopolysaccharide-induced cyclooxygenase 2 expression and prostaglandin E2 production through endothelin ETB receptors effects which are up-regulated by lipopolysaccharide. In the present study, we confirmed these findings and showed that prostaglandin E2 (10−6 to 10−5 M) inhibited the lipopolysaccharide plus endothelin-1-induced cyclooxygenase 2 expression more profoundly as compared to its inhibition of the lipopolysaccharide-induced cyclooxygenase 2 expression. The endothelin ETB receptor selective antagonist, N-cis-2,6-dimethylpiperidino-carbonyl-l-γ-methyl-leucyl-d-l-methoxycarbonyl-tryptophanyl-d-norleucine (BQ788), partly inhibited this suppression. Interestingly, the expression of endothelin ETB receptors in macrophages was increased by lipopolysaccharide plus prostaglandin E2 (10−8 to 10−5 M) about 1.6-fold compared with that evoked by lipopolysaccharide stimulation alone. We also showed that treatment with endothelin-1 at 10−14 M (15 min) elevated an intracellular cyclic AMP concentration in macrophages stimulated by lipopolysaccharide or lipopolysaccharide plus prostaglandin E2 (10−6 M) for 6 h, and the elevation in the latter cells was more pronounced. These results suggested that endothelin-1 shows an opposite modulation of lipopolysaccharide-induced cyclooxygenase 2 expression in macrophages through endothelin ETB receptors, depending on the level of extracellular prostaglandin E2, and the changes of intracellular cyclic AMP by endothelin-1 may be involved in this mechanism.
Keywords: Endothelin-1; Cyclooxygenase 2; Prostaglandin E2; Lipopolysaccharide; Macrophage;