European Journal of Pharmacology (v.436, #1-2)

Possible involvement of oxidative stress in hypoxia-induced adrenomedullin secretion in cultured rat cardiomyocytes by Fumiki Yoshihara; Takeshi Horio; Toshio Nishikimi; Hisayuki Matsuo; Kenji Kangawa (1-6).
Although hypoxia induces adrenomedullin gene expression in cultured rat cardiac myocytes, it is still unknown whether oxidative stress is involved in the hypoxia-induced adrenomedullin production. We investigated whether oxidative stress might participate in hypoxia-induced adrenomedullin secretion and whether adrenomedullin might have a protective effect on damaged myocytes. Hypoxia increased adrenomedullin secretion and its gene expression in cardiac myocytes, but not in nonmyocytes. Furthermore, oxidative stress (hydrogen peroxide) also increased adrenomedullin secretion from myocytes. N-acetyl-l-cysteine, a free radical scavenger, completely inhibited the stimulation of adrenomedullin secretion by hydrogen peroxide, and this agent reduced the stimulation of adrenomedullin secretion by hypoxia. Lactate dehydrogenase leakage, a marker of cell injury, was significantly increased with the exposure to hydrogen peroxide and adrenomedullin significantly reduced this leakage. These findings suggest that an oxidative stress may be involved, in part, in the increased adrenomedullin secretion from cardiac myocytes under hypoxic condition. Adrenomedullin secreted from myocytes may play a cell protective role in an autocrine manner.
Keywords: Hypoxia; Oxidative stress; Adrenomedullin; Cardiomyocyte;

Troglitazone suppresses cell growth of KU812 cells independently of PPARγ by Akihito Abe; Yoshimitsu Kiriyama; Masako Hirano; Toshiaki Miura; Hiroyuki Kamiya; Hideyoshi Harashima; Yukiko Tokumitsu (7-13).
We examined the effects of troglitazone, one of thiazolidinedione derivatives on human basophilic leukemia cell line KU812. Troglitazone caused the suppression of cell growth, which was suggested to result from the decrease in cyclin E and the hyperphosphorylated form of retinoblastoma tumor suppressor gene product (pRb). In addition, troglitazone caused a decrease in histamine secretion due to the reduced expression of histidine decarboxylase mRNA. Peroxisome proliferator-activated receptor (PPAR)-γ mRNA was undetectable by reverse transcription-polymerase chain reaction (RT-PCR) in KU812 cells. These findings suggested that troglitazone suppressed cell growth and histamine synthesis independently of PPARγ.
Keywords: Thiazolidinedione; PPAR (Peroxisome proliferator-activated receptor); KU812 cell; Cell growth; Histamine secretion;

We investigated the role of angiotensin II type 1 (AT1) receptors in angiotensin II-induced actin reorganization and the signaling pathways of the response in pleural mesothelial cells. The effects of angiotensin II on actin reorganization in pleural mesothelial cells were evaluated by dual fluorescence labeling of filamentous (F) and monomeric (G) actin with fluorescein isothiocyanate (FITC)-labeled phalloidin and Texas Red-labeled DNase I, respectively. Angiotensin II (10 μM) induced actin reorganization in the presence and the absence of extracellular Ca2+. An angiotensin AT1 receptor antagonist ([Sar1,Ile8]angiotensin II) inhibited angiotensin II-induced actin reorganization. Pretreatment with C3 exoenzyme or tyrosine kinase inhibitors significantly reduced angiotensin II-induced actin reorganization. However, pertussis toxin, phosphatidylinositol-3-kinase and protein kinase C inhibitors had no effect on these responses. These results suggest that angiotensin II-induced actin reorganization in pleural mesothelial cells is extremely dependent on the angiotensin AT1 receptor coupled with pertussis toxin-insensitive heterotrimeric G proteins, Rho GTPases and tyrosine phosphorylation pathways.
Keywords: Actin; Angiotensin II; Angiotensin AT1 receptor; Mesothelial cell; Pleura; Pertussis toxin; Protein kinase C; Tyrosine kinase;

Molecular cloning and expression of the porcine trigeminal ganglion cDNA encoding a 5-ht1F receptor by Pankaj Bhalla; Hari S Sharma; Thierry Wurch; Petrus J Pauwels; Pramod R Saxena (23-33).
Using a combination of reverse transcription polymerase chain reaction (RT-PCR) and inverse-PCR techniques, we amplified, cloned and sequenced a full-length porcine 5-hydroxytryptamine 1F (5-ht1F) receptor complementary DNA (cDNA) derived from porcine trigeminal ganglion. Sequence analysis revealed 1101 base pairs (bp) encoding an open reading frame of 366 amino acids showing a high similarity (>90%) with the 5-ht1F receptor sequences from other species, including human. The recombinant porcine 5-ht1F receptor was expressed in African green monkey kidney cell lines (COS-7 cells) and its ligand binding profile was determined using [3H]5-HT. The affinities of several agonists (LY334370 (5-(4-fluorobenzoyl)amino-3-(1-methylpiperidin-4-yl)-1H-indole fumarate)>CP122638 (N-methyl-3 [pyrrolidin 2(R)-yl methyl]-1H-indol-5-ylmethyl sulphonamide)=naratriptan=5-HT>eletriptan>sumatriptan>frovatriptan=avitriptan>dihydroergotamine>zolmitriptan>5-carboxamidotryptamine>rizatriptan>alniditan=donitriptan>L694247 (2-[5-[3-(4-methylsulphonylamino)benzyl-1,2,4-oxadiazol-5-yl]-1H-indole-3-yl] ethylamine) and putative antagonists (methiothepin>GR127935 (N-[4-methoxy-3-(4-methyl-1-piperazinyl) phenyl]-2′-methyl 4′-(5-methyl-1,2,4-oxadiazol-3-yl) [1,1-biphenyl]-4-carboxamide hydrochloride)>ritanserin>SB224289 (2,3,6,7-tetrahydro-1′-methyl-5-[2′-methyl-4′(5-methyl-1,2,4-oxadiazol-3-yl) biphenyl-4-carbonyl] furo [2,3-f] indole-3-spiro-4′-piperidine hydrochloride)>BRL155572 ([1-(3-chlorophenyl)-4-[3,3-diphenyl (2-(S,R) hydroxypropanyl)piperazine] hydrochloride)>ketanserin=pindolol) correlated highly with those described for the recombinant human 5-ht1F receptor (Spearman correlation coefficient; r s=0.942). Nevertheless, as compared to the human homologue, some triptans (i.e. sumatriptan, zolmitriptan and rizatriptan) displayed a 10- to 15-fold lower affinity for the porcine 5-ht1F receptor. Using RT-PCR technique, the expression of porcine 5-ht1F receptor mRNA was observed in cerebral cortex, trigeminal ganglion and several blood vessels, but not in skeletal muscles. In conclusion, we have cloned and established the amino acid sequence and ligand binding profile of the porcine 5-ht1F receptor as well as the distribution of its mRNA. This information may be helpful in exploring the role of 5-ht1F receptor in physiological processes and diseases, such as migraine.
Keywords: 5-ht1F receptor; Cloning; 5-HT (5-hydroxytryptamine, serotonin); Ligand binding; Migraine; Sequencing; Trigeminal ganglion;

In human neuroblastoma SH-SY5Y cell preparations, σ1 receptor agonists (+)-pentazocine and 1S,2R-(−)-cis-N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1-pyrrolidinyl)cyclohexylamine (BD737) competed for [3H]haloperidol binding with K i values of 67±10 and 14±10 nM, respectively. (+)-Pentazocine or BD737 up to 10 μM did not affect basal levels of intracellular Ca2+ concentration ([Ca2+]i) in these cells, but they significantly reduced muscarine-induced [Ca2+]i changes in a dose-related manner. However, the reduction by (+)-pentazocine was not reversed by the σ1 receptor antagonist haloperidol. Further studies showed (+)-pentazocine, BD737 and haloperidol could compete for [3H]quinuclidinyl benzilate binding in SH-SY5Y cells with K i values of 0.51±0.06, 0.32±0.07 and 4.4±2.3 μM, respectively. Thus, the inhibitory effects on muscarine-induced [Ca2+]i changes by (+)-pentazocine and BD737 in SH-SY5Y cells were likely due to blockade of muscarinic receptors, not due to σ1 receptor activation by these ligands.
Keywords: SH-SY5Y neuroblastoma; σ Receptor; Ca2+; Muscarine; (+)-Pentazocine; Haloperidol;

Ambroxol inhibits platelet-derived growth factor production in human monocytic cells by Mitsuyoshi Utsugi; Kunio Dobashi; Yasuhiko Koga; Ken Masubuchi; Yasuo Shimizu; Katsuaki Endou; Tsugio Nakazawa; Masatomo Mori (47-51).
Several growth factors, including platelet-derived growth factor (PDGF), have been implicated in the mechanism of lung and airway remodeling. We investigated the effect of ambroxol, trans-4-[(2-amino-3,5-dibromobenzyl) amino] cyclohexanol hydrochloride, on the lipopolysaccharide-induced PDGF production in human monocytic cells, THP-1. Ambroxol inhibited the lipopolysaccharide-induced PDGF-AB production via PDGF-A mRNA expression. Lipopolysaccharide activated p44/42 extracellular signal-regulated kinase (ERK), and ambroxol attenuated the lipopolysaccharide-induced p44/42 ERK activation. Furthermore, mitogen-activated protein kinase kinase (MEK)-1-specific inhibitor, 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one (PD 98059), blocked the lipopolysaccharide-induced p44/42 ERK activation and PDGF production. These findings indicate that ambroxol inhibits the lipopolysaccharide-induced PDGF production due to the suppression of p44/42 ERK activity.
Keywords: Ambroxol; PDGF (Platelet-derived growth factor); ERK (p44/42 extracellular signal-regulated kinase); Monocytic cell;

Nitric oxide (NO) and serotonin (5-hydroxytryptamine; 5-HT) are important neuromodulators that are involved in a myriad of biochemical reactions. In this work, we describe a novel model co-culture system to study the interactions between NO and 5-HT. NO derived from cytokine stimulated Bv2 microglial cells depleted 5-HT from RBL-2H3 cells. Reduction of 5-HT content by NO derived from the NO donor S-nitroso-N-acetylpenicillamine (SNAP) was concentration-dependent, independent of intracellular Ca2+ and inhibited by reduced glutathione (GSH). Collectively, these data indicate that this cell co-culture system is a viable model to study the mechanisms of interaction between nitrergic and serotonergic pathways.
Keywords: Nitric oxide (NO) synthase; 5-HT (5-hydroxytryptamine, serotonin); Neuromodulation;

Effects of chronically administered venlafaxine on 5-HT receptor activity in rat hippocampus and hypothalamus by Eitan Gur; Eliyahu Dremencov; Louis D Van De Kar; Bernard Lerer; Michael E Newman (57-65).
The effects of chronic administration of the mixed serotonin [5-hydroxytryptamine (5-HT)]/norepinephrine re-uptake inhibitor venlafaxine (5 mg/kg daily by osmotic minipump for 28 days) on the sensitivity of somatodendritic 5-HT1A autoreceptors on serotonergic neurons innervating the hypothalamus, and on 5-HT1B autoreceptors in both hypothalamus and hippocampus, were determined using in vivo microdialysis in freely moving rats. Venlafaxine induced a reduction in sensitivity of 5-HT1B autoreceptors in hypothalamus, but did not affect the sensitivity of 5-HT1A autoreceptors, or of 5-HT1B autoreceptors in hippocampus. The corticosterone and oxytocin responses to the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT, 0.05 or 0.2 mg/kg), a measure of postsynaptic 5-HT1A receptor activity in the hypothalamus, were reduced in animals administered 5 or 10 mg/kg venlafaxine daily by intraperitoneal injection for 21 days. This desensitization of post-synaptic 5- HT1A receptors in the hypothalamus may be a consequence of increased 5-HT levels induced by desensitization of the presynaptic 5-HT1B receptors. These results taken together with those of previous studies suggest that the hypothalamus might be an important site of drug action, and that venlafaxine has an overall mechanism similar to that of selective serotonin re-uptake inhibitors.
Keywords: 5-HT (5-hydroxytryptamine, serotonin); Venlafaxine; Antidepressant; Hippocampus; Hypothalamus; 5-HT1A receptor; 5-HT1B receptor; Microdialysis;

Methiothepin, a nonselective 5-HT receptor antagonist was utilized to explore the 5-HT modulation of dorsal vagal complex–TRH (thyrotropin releasing hormone) analogue stimulated gastric functional parameters. Intracisternal methiothepin pretreatment (200, 0.1 nmol) produced significant inhibition (70%, 44%, respectively) of the TRH analogue [p-Glu-His-(3,3′-dimethyl)-Pro NH2; RX 77368 (12 pmol)]-induced gastric acid output compared to vehicle pretreatment. Intracisternal pretreatment with methysergide (nonspecific 5-HT receptor antagonist) or combined cyanopindolol (5-HT1A and 1B receptor antagonist)+ritanserin (receptor antagonist of the 5-HT2 family) did not alter the dorsal vagal complex–RX 77368 response. Unilateral dorsal vagal complex pretreatment with methiothepin (50 nmol/50 nl) attenuated ipsilateral dorsal vagal complex–TRH analog (12 pmol) induced gastric secretory response by 57%. The gastric secretagogue response to stimulation of the raphe obscurus (mediated by TRH release into the dorsal vagal complex) was inhibited 50% by pretreatment with intracisternal dorsal medullary methiothepin (0.1 nmol/10 μl). Intracisternal methiothepin (200 nmol/20 μl) also attenuated (a) dorsal vagal complex–glutamate (60 nmol/30 nl) stimulated gastric acid secretion and (b) gastric motility stimulated by dorsal vagal complex–RX 77368 (12 pmol/30 nl). The data suggest that other properties of methiothepin, alone or in addition to its 5-HT receptor antagonist effect, mediate its inhibitory actions at the dorsal vagal complex.
Keywords: Vagus; Raphe nucleus; Brain stem; Autonomic nervous system; 5-HT (5-hydroxytryptamine, serotonin);

Differential involvement of 5-HT2A receptors in the discriminative-stimulus effects of cocaine and methamphetamine by Patrik Munzar; Zuzana Justinova; Scott W Kutkat; Steven R Goldberg (75-82).
Involvement of 5-HT2A receptors in the discriminative-stimulus effects of cocaine versus methamphetamine was studied in Sprague Dawley rats (n=10) trained to discriminate 10 mg/kg cocaine, i.p., from saline under a fixed-ratio 10 (FR10) schedule of food presentation. The ability of (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), a 5-HT2A receptor agonist, and ketanserin, a 5-HT2A receptor antagonist, to either substitute for or block the discriminative-stimulus effects of cocaine, or to shift the cocaine dose–response curve, was evaluated. DOI (0.18–1.0 mg/kg) partially substituted for the training dose of 10 mg/kg cocaine, but only at doses that decreased rates of responding. At the highest dose of DOI tested (1.0 mg/kg), there was about 65% cocaine-appropriate responding. Substitution of DOI for cocaine and DOI-induced decreases in rates of responding were completely reversed by ketanserin (3.0 mg/kg). Ketanserin (3.0 mg/kg) also produced a significant shift to the right of the cocaine dose–response curve and antagonized increases in rates of responding produced by lower doses of cocaine. Ketanserin (1.0–10.0 mg/kg), however, did not block the discriminative-stimulus effects of the training dose of cocaine. When DOI (0.3 mg/kg) was co-administered with different doses of cocaine, there was a slight leftward shift in the cocaine dose–response curve, which was not significant and appeared to reflect simple additive effects of DOI and cocaine. In contrast, the same dose of DOI (0.3 mg/kg) produced a marked and highly significant shift to the left of the methamphetamine (0.18–1.0 mg/kg) dose–response curve in the same subjects and the effects of DOI and methamphetamine were clearly more than additive. The present findings provide new evidence that there is some serotonergic modulation of cocaine's discriminative-stimulus actions, which appears to involve stimulation of 5-HT2A receptors. However, involvement of 5-HT2A receptor activity in the discriminative-stimulus actions of cocaine appears to be less pronounced than in similar actions of methamphetamine.
Keywords: 5-HT2A receptor; Cocaine; DOI; Ketanserin; Methamphetamine; Drug discrimination;

cGMP, but not cAMP, in rat hippocampus is involved in early stages of object memory consolidation by Jos Prickaerts; Jan de Vente; Wiel Honig; Harry W.M. Steinbusch; Arjan Blokland (83-87).
The present study investigates the role of cGMP and cAMP on the memory performance in the object recognition task in rats. The analogue 8-Br-GMP or 8-Br-cAMP was administered bilaterally into the hippocampus (0, 1, 3 and 10 μg in 0.5 μl saline/site) immediately after the exposure to two identical objects. After 24 h, saline-treated animals spent equal times exploring a new and the familiar object demonstrating that they did not recognize the familiar one. However, a dose-dependent improvement in object recognition was found after injection of 8-Br-cGMP. In contrast, 8-Br-cAMP did not improve the memory performance at the doses tested. These results indicate that hippocampal cGMP but not cAMP is involved in early stages of consolidation of object memory.
Keywords: Cyclic nucleotide; Phosphodiesterase; Cognition; Hippocampus;

Recent study has shown that monophosphoryl lipid A-induced delayed preconditioning enhanced preservation with cardioplegia and that the protective effects of monophosphoryl lipid A were related to stimulation of calcitonin gene-related peptide (CGRP) release. The purpose of the present study was to explore whether the elevated release of CGRP induced by monophosphoryl lipid A is secondary to stimulation of CGRP synthesis via the nitric oxide (NO) pathway and to characterize the isoform of CGRP. Sprague–Dawley rats were pretreated with monophosphoryl lipid A 24 h before the experiment, and then the left main coronary artery of rat hearts was subjected to 1 h occlusion followed by 3 h reperfusion. Infarct size, plasma creatine kinase activity, the plasma level of CGRP, and the expression of CGRP isoforms (α- and β-CGRP) mRNA in lumbar dorsal root ganglia were measured. Pretreatment with monophosphoryl lipid A (500 μg/kg, i.p.) significantly reduced infarct size and creatine kinase release. Monophosphoryl lipid A caused a significant increase in the expression of α-CGRP mRNA, but not of β-CGRP mRNA, concomitantly with an increase in plasma concentrations of CGRP, and the increased level of CGRP expression happened before stimulation of CGRP release. The effect of monophosphoryl lipid A was completely abolished by pretreatment with l-nitroarginine methyl ester (l-NAME, 10 mg/kg, i.p.), an inhibitor of NO synthase or capsaicin (50 mg/kg, s.c.), which selectively depletes transmitters in capsaicin-sensitive sensory nerves. The results suggest that the delayed cardioprotection afforded by monophosphoryl lipid A involves the synthesis and release of CGRP via the NO pathway, and that the protection is mainly mediated by the α-CGRP isoform.
Keywords: CGRP (calcitonin gene-related peptide); Monophosphoryl lipid A; Preconditioning; Ischemia–reperfusion; Capsaicin; Nitric oxide (NO);

Fluvastatin reduces modification of low-density lipoprotein in hyperlipidemic rabbit loaded with oxidative stress by Yu Yamaguchi; Sachiko Matsuno; Satomi Kagota; Jun Haginaka; Masaru Kunitomo (97-105).
The in vivo antioxidant effect of fluvastain, a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, was investigated using Watanabe heritable hyperlipidemic (WHHL) rabbits subjected to nicotine-free cigarette smoke extracts as oxidative stress. Fluvastatin was given orally at doses of 10 and 30 mg/kg per day for 5 months. The cigarette smoke extracts were prepared by bubbling the gas phase of smoke into phosphate-buffered saline and was injected daily into the rabbit ear vein. The rabbits chronically treated with the cigarette smoke extracts showed an increase in plasma lipid peroxide levels, estimated as thiobarbituric acid-reactive substances. Oxidative modification of plasma low-density lipoprotein (LDL) was assessed by anion-exchange high-performance liquid chromatographic analysis, LDL susceptibility to oxidation, LDL incorporation into macrophages and thiobarbituric acid-reactive substances levels in LDL. Treatment with fluvastatin significantly reduced these effects induced by the cigarette smoke extracts in a dose-related manner and exerted a cholesterol-lowering effect. At the end of the experiment, the cigarette smoke extracts caused accumulation of cholesteryl ester in the thoracic aorta, while fluvastatin significantly prevented this accumulation. These results indicate that fluvastatin can exert an antioxidant effect in vivo, with a strong effect on oxidative stress such as smoking, a major risk factor of atherosclerosis.
Keywords: Fluvastatin; Antioxidant effect; Cigarette smoke extract; LDL (low-density lipoprotein); Oxidatively modified; WHHL rabbit;

Venodilator action of an organotransition-metal nitrosyl complex by Kenneth S Poon; Catherine C.Y Pang (107-110).
Nitrovasodilators, such as nitroglycerin, cause endothelium-independent dilatation of arterial and capacitance vessels via the release of nitric oxide (NO). This study examined the venodilator effect of CpCr(NO)2Cl (organotransition-metal nitrosyl complex) relative to that of nitroglycerin in conscious, unrestrained rats. Organotransition-metal nitrosyl complexes have releasable NO directly attached to metal centres. The dose–response effects of CpCr(NO)2Cl and nitroglycerin on the mean arterial pressure and the mean circulatory filling pressure (index of the body venous tone) were obtained in rats continuously infused with either normal saline or noradrenaline. The results show that both CpCr(NO)2Cl and nitroglycerin reduced the mean arterial pressure in rats with normal or elevated vasomotor tone. However, maximum depressor response of CpCr(NO)2Cl was greater than that of nitroglycerin. In vehicle-treated rats, both compounds increased the mean circulatory filling pressure. In rats with elevated vasomotor tone through the infusion of noradrenaline, both agents reduced the mean circulatory filling pressure. These results show that CpCr(NO)2Cl is an efficacious depressor and venodilator agent.
Keywords: Nitroglycerin; Nitric oxide (NO); Metal nitrosyl complex; Capacitance vessel; Vasodilator; MCFP;

Effects of the superoxide dismutase-mimetic compound tempol on endothelial dysfunction in streptozotocin-induced diabetic rats by Taher Nassar; Bashir Kadery; Chaim Lotan; Nael Da'as; Yosef Kleinman; Abdullah Haj-Yehia (111-118).
Evidence exists to support the beneficial effects of superoxide dismutase on endothelial dysfunction induced by hyperglycemia in vitro. In vivo, however, studies of the effects of native superoxide dismutase preparations on the vascular complications accompanying diabetes are limited, and their therapeutic application potential has so far been disappointing. The objective of this study was to evaluate, for the first time in vivo, the effects of long-term administration of tempol, a stable superoxide dismutase-mimic compound, on diabetes-induced endothelial dysfunction in rats. Diabetes was induced by streptozotocin and rats were monitored for 8 weeks with or without treatment with tempol (100 mg/kg, s.c., b.i.d). Diabetic rats showed increased vascular levels of superoxide, which was accompanied by increased levels of the oxidative stress markers malondialdehyde and 8-epi-prostaglandin F. In addition, the vasorelaxant as well as the cGMP-producing effects of acetylcholine and glyceryl trinitrate were reduced in diabetic rats. Treatment with tempol abolished not only the differences in the vascular content of superoxide, malondialdehyde and 8-epi-prostaglandin F,  but also the differences in the relaxation and cGMP responses of aortic rings to both acetylcholine and glyceryl trinitrate between control and diabetic rats. These results support the involvement of reactive oxygen species in mediation of hyperglycemia-induced endothelial dysfunction in vivo, and provide the rationale for potential utilization of stable superoxide dismutase-mimic nitroxides for the prevention of the vascular complications accompanying diabetes.
Keywords: Diabetes; Endothelial dysfunction; Nitric oxide (NO); Superoxide; Superoxide dismutase; Tempol;

Two cysteinyl-leukotriene receptors, CysLT1 and CysLT2 receptors, have been cloned, but the contractions to cysteinyl-leukotrienes in the guinea pig lung parenchyma have been reported to be resistant to CysLT2 receptor antagonism and to be only partially inhibited by CysLT1 receptor antagonism. The receptor preferences of the individual cysteinyl-leukotrienes (leukotriene C4, D4 and E4) in the guinea pig lung parenchyma were studied in organ baths. CysLT1 receptor antagonists competitively inhibited the contraction to leukotriene E4, but exhibited only weak antagonism of contractions to leukotriene C4 and D4. In the presence of the cyclooxygenese inhibitor indomethacin and the nitric oxide synthase inhibitor N ω-nitro-l-arginine (l-NOARG), the CysLT1 receptor antagonists did not further inhibit the leukotriene D4-induced contraction. These results suggest that leukotriene E4 solely activates a CysLT1 receptor, and that the CysLT1 receptor antagonist-resistant contraction to leukotriene D4 and C4 is mediated via another CysLT receptor.
Keywords: Cysteinyl-leukotriene receptor; Lung parenchyma; Cyclooxygenase; Nitric oxide (NO);

We investigated the mechanisms of wood smoke-induced increases in nasal airway resistance (Rna) and airway reactivity in anesthetized rats. Delivery of wood smoke into a functionally isolated nasal airway produced an increase in Rna, which was attenuated by CP-96,345 [a tachykinin NK1 receptor antagonist; (2S,3S)-cis-2-(diphenylmethyl)-N-((2-methoxyphenyl)-methyl)-1-azabicyclo(2.2.2.)-octan-3-amine] or atropine. Additionally, smoke pre-exposure animals displayed a greater amplitude and a longer duration of Rna responses to capsaicin or histamine provocation, as compared to air controls. This enhanced airway reactivity to capsaicin or histamine was largely alleviated by CP-96,345 or atropine. The nasal secretory responses to capsaicin or histamine in smoke pre-exposure animals were similar to those in air controls. We concluded that (1) reflex cholinergic and tachykininergic mechanisms play important roles in wood smoke-induced increases in nasal airway resistance and airway reactivity, and (2) this nasal airway hyperreactivity might not be due to an exaggerated secretory response, but is presumably due to augmented nasal swelling.
Keywords: Smoke; Hyperreactivity; Nasal irritation; Tachykinin; Cholinergic reflex;

Nucleotide-evoked relaxation of rat vas deferens: possible mechanisms by R. Bültmann; W. Klebroff; K. Starke (135-143).
ATP causes relaxation of the K+-contracted rat vas deferens. Possible sites of action were investigated. ATP and adenosine relaxed the vas deferens precontracted with 80 mM K+; EC50 values and maximal relaxations averaged, respectively, 760 μM and 56% for ATP and 74 μM and 30% for adenosine. The adenosine P1 receptor antagonist 8-(para-sulfophenyl)theophylline (8-SPT) reduced relaxations caused by adenosine and low concentrations of ATP, as did the Rp-diastereomer of adenosine 3′,5′-cyclic phosphorothioate (Rp-cAMPS), an inhibitor of protein kinase A. The phosphodiesterase inhibitor 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (Ro 20-1724) augmented responses to adenosine and low concentrations of ATP. α,β-Methylene ADP, an inhibitor of 5′-nucleotidase, reduced relaxations caused by ATP to a similar extent as did 8-SPT. In the presence of an almost saturating concentration of adenosine, ATP caused further relaxation. Conversely, in the presence of ATP, adenosine had little effect. Like ATP, UTP and other nucleoside triphosphates relaxed the vas deferens. The P2 receptor antagonists reactive blue 2, acid blue 25 and 4,4′-diisothiocyanotostilbene-2,2′-disulphonate (DIDS) attenuated the relaxation caused by ATP; suramin, pyridoxalphosphate-6-azophenyl-2′,4′-disulphonate (PPADS), Evans blue, trypan blue, reactive red 2 and brilliant blue G had no effect. Three non-selective inhibitors of protein kinases, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7), staurosporine and (8R*,9S*,11S*)-(−)-9-hydroxy-9-carboxy-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo[a,g]cycloocta[cde]trinden-1-one (K-252b), markedly reduced the relaxation caused by ATP. The results indicate that adenosine, derived from enzymatic dephosphorylation, contributes to the relaxant effect of ATP, presumably by activation of a smooth muscle adenosine receptor linked to the accumulation of cAMP and activation of protein kinase A. Yet, the main part of the response to ATP is mediated by a site distinct from the adenosine receptor. The pharmacological properties of this site differ from known P2 receptor subtypes. Possibly, the nucleotide-evoked relaxation is due to a phosphoryl transfer catalyzed by an ecto-protein kinase.
Keywords: Vas deferens; ATP; Adenosine; Smooth muscle relaxation; P2 receptor; Adenosine A2 receptor; Ecto-protein kinase;

Inhibition of the renin–angiotensin system ameliorates genetically determined hyperinsulinemia by J.R Ortlepp; J Breuer; F Eitner; K Kluge; R Kluge; J Floege; G Hollweg; P Hanrath; H.-G Joost (145-150).
This study was performed in order to assess the potentially different effects of the angiotensin-converting enzyme inhibitor captopril and of the angiotensin II receptor antagonist irbesartan on the metabolic syndrome in an animal model. Male NZO/BL6 F1 mice were treated with captopril, irbesartan, or placebo for 10 months: Control animals treated with placebo developed a metabolic syndrome with obesity (55.5+6.3 g), hypertension (146+10 mm Hg), hyperinsulinemia (7.2+5.7 ng/ml), hypercholesterolemia (5.1+0.7 mmol/l), cardiac hypertrophy (269+44 mg) and atherosclerotic plaques in the ascending aorta (3.6+1.5 μm2). Treatment with angiotensin-converting enzyme inhibitor or angiotensin II receptor antagonist significantly (p<0.001) reduces hypertension (73+5 and 78+11 mm Hg), cardiac hypertrophy (203+26 and 202+18 mg) and atherosclerosis (2.2+0.9 and 1.8+0.8 μm2). In addition, they prevented the development of obesity (42.2+3.5 and 38.3+2.8 g) and hyperinsulinemia (3.6+1.5 and 1.8+0.4 ng/ml). In conclusion, long-term treatment with an angiotensin-converting enzyme inhibitor or an angiotensin II receptor antagonist can ameliorate obesity and hyperinsulinemia in a genetically determined mouse model.
Keywords: Angiotensin AT1 receptor; Angiotensin-converting enzyme inhibitor; Obesity; Hyperinsulinemia; Metabolic syndrome; (Mouse);