The alpha (1D)-adrenoceptor antagonist BMY 7378 is also an alpha (2C)-adrenoceptor antagonist

1 We have investigated the actions of the alpha(1D)-adrenoceptor selective antagonist BMY 7378 in comparison with yohimbine at alpha(1)- and alpha(2)-adrenoceptors. 2 In rat aorta (alpha(1D)-adrenoceptor), BMY 7378 (pA(2) of 8.67) was about 100 times more potent than yohimbine (pA(2) of 6.62) at antagonizing the contractile response to noradrenaline. 3 In human saphenous vein (alpha(2C)-adrenoceptor), BMY 7378 (pA(2) of 6.48) was approximately 10 times less potent than yohimbine (pA(2) of 7.56) at antagonizing the contractile response to noradrenaline. 4 In prostatic portions of rat vas deferens, BMY 7378 (10 mum) did not significantly affect the concentration-dependent inhibition of single pulse nerve stimulation-evoked contractions by xylazine (an action at prejunctional alpha(2D)-adrenoceptors). 5 In ligand-binding studies, BMY 7378 showed 10-fold selectivity for alpha(2C)-adrenoceptors (pK(i) of 6.54) over other alpha(2)-adrenoceptors. 6 It is concluded that BMY 7378, in addition to alpha(1D)-adrenoceptor selectivity in terms of alpha(1)-adrenoceptors, shows selectivity for alpha(2C)-adrenoceptors in terms of alpha(2)-adrenoceptors.     

α1‐ and α2‐Adrenoceptor hyporesponsiveness in isolated bisected vas deferens of bile duct‐ligated rats

1 It has been suggested that cholestasis accompanied with changes in autonomic balance and hyporesponsiveness in muscarinic and adrenergic receptors of some organs, e.g. cardiovascular system. Increased plasma levels of epinephrine and norepinephrine has been shown during cholestasis suggesting augmented activity of sympathetic nervous system. In this study we evaluate both α₁ and α₂ responsiveness in isolated rat vas deferens, as a tissue with rich adrenergic innervations. 2 Epididymal and prostatic halves of vas deferens responsiveness have been studied to phenylephrine and clonidine respectively in three groups of un‐operated, sham‐operated (sham), and bile duct‐ligated (BDL) rats. 3 Our results indicate that in vas deferens of BDL animals, the concentration‐response curve of both phenylephrine and clonidine shifted to rightward compared to control group, while the position of concentration‐response curve of sham group did not change significantly (P > 0.05). EC₅₀ of phenylephrine and IC₅₀ of clonidine were increased showing a decreased responsiveness of tissue to phenylephrine (P < 0.05) and clonidine (P < 0.001) in BDL rats. 4 In this study, both subtype of α‐adrenoceptors (α₁ and α₂) has been studied in cholestatic rat vas deference. Our results showed that cholestasis induce hyporesponsiveness to phenylephrine and clonidine. These results are consistent with previous reports, suggesting the hyporesponsiveness of α₁‐adrenoceptors in pulmonary artery and papillary muscle and mesenteric beds. Our conclusion is that the cholestasis induces hyporesponsiveness to phenylephrine and clonidine in epididymal (α₁‐adrenoceptors) and prostatic (α₂‐adrenoceptors) halves of rat vas deferens respectively. Although the logical explanation to this hyporesponsiveness is the down regulation but it has been suggested that it is not because of down regulation.     

Selective agonists reveal alpha(1A)- and alpha(1B)-adrenoceptor subtypes in caudal artery of the young rat

Multiple alpha(1)-adrenoceptors were evaluated in caudal artery of the young Wistar rat using selective agonists and antagonists. Arteries were exposed to the selective alpha(1A)-adrenoceptor agonist, A-61603 (N-[5-(4,5-dihydro-1H-imidazol-2-yl)-2-hydroxy-5,6,7,8-tetrahydronaphthalen-1-yl] methanesulfonamide) or to phenylephrine and to prazosin (alpha(1)-adrenoceptor antagonist), or the selective alpha(1A)-adrenoceptor antagonists 5-methylurapidil, RS 100329 (5-methyl-3-[3-[4-[2-(2,2,2,-trifluoroethoxy)phenyl]-1-piperazinyl]propyl]-2,4-(1H)-pyrimidinedione), RS 17053 (N-[2(2-cyclopropylmethoxy) ethyl]-5-chloro-alpha, alpha-dimethyl-1H-indole-3-ethanamide), and the selective alpha(1D)-adrenoceptor antagonist BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4.5] decane-7,9-dione). Results showed a 100-fold higher potency of A-61603 for the alpha(1)-adrenoceptor present in the artery, compared with phenylephrine. Prazosin displaced both agonists with high affinity, whereas 5-methylurapidil, RS 100329 and RS 17053 displaced A-61603 with high affinity, indicating the presence of alpha(1A)-adrenoceptors. The selective alpha(1A)-adrenoceptor antagonists blocked phenylephrine responses with low affinity, suggesting that phenylephrine activated a second receptor population in caudal artery. BMY 7378 antagonized with low affinity both A-61603 and phenylephrine-induced contractions, indicating absence of alpha(1D)-adrenoceptors in the vessel. The results suggest that functional alpha(1B)-adrenoceptors are present in caudal arteries of the young Wistar rat.     

Selective ligands for opioid receptors: β-Cyclopropylalanyl containing analogs of enkephalin

The synthesis and resolution of the amino acid β-cyclopropylalanine (Cpr) and its incorporation into four enkephalin analogs is reported. The analogs prepared were: Tyr - l - Cpr - Gly - Phe - Pen (des - COOH - Nle = n - pentylamide = Pen) (l -Cpr²-Pen⁵-ENK), Tyr-d -Cpr-Gly-Phe-Pen (d -Cpr²-Pen⁵-ENK), l -Cpr-Tyr-d -Ala-Gly-Phe-Pen (l -Cpr⁰-d -Ala²-Pen⁵-ENK) and d -Cpr-Tyr-d -Ala-Gly-Phe-Pen (d -Cpr⁰-d -Ala²-Pen⁵-ENK). Each was tested for its ability to inhibit the field stimulated guinea pig ileum (GPI) and rat vas deferens (RVD) and the results compared to the effect d -Ala²-d -Leu⁵-enkephalin (DADLE) has on the same preparations. The results show that at concentrations up to 10^-5 m all four analogs, as well as DADLE, are full agonists on the GPI preparation. The concentrations necessary to produce a 50% inhibition of the twitch response were, DADLE, 3.5 °× 10^-8 m ; l - Cpr⁰-d -Ala²-Pen⁵-ENK, 6.0 × 10^-8 m ; d -Cpr²-Pen⁵-ENK, 1.1 × 10^-7 m ; l -Cpr²-Pen⁵-ENK, 1.2 × 10^-6 m and d -Cpr⁰-d -Ala²-Pen⁵-ENK, > 10^-5 m . On RVD a different result was observed with only DADLE (1.3 × 10^-6 m ) and l -Cpr⁰-Pen⁵-enkephalin (1.8 × 10^-6 m ) showing full agonist activity. d -Cpr²-Pen⁵-ENK was a partial agonist (29 · 5% inhibition of the twitch at 10^-5 m ) while d -Cpr⁰-d -Ala²-Pen⁵-ENK and l -Cpr²-Pen⁵-ENK did not inhibit the twitch at concentrations up to 10^-5 m . These compounds which were inactive or of low potency on each preparation were also tested as antagonists. Only d -Cpr²-Pen⁵-ENK was an antagonist (pA₂ = 6.09) versus DADLE on RVD while d -Cpr⁰-d -Ala²-Pen⁵-ENK was inactive as an antagonist on both GPI and RVD. d -Cpr²-Pen⁵-ENK, therefore, represents the first enkephalin analog to be categorized as a mixed agonist-antagonist.     

Chloroethylclonidine reveals that alpha (1 A)-adrenoceptors mediate contraction in aorta of alpha (1 D)-adrenoceptor knockout mice

1 We have characterized the alpha(1)-adrenoceptor subtypes present in isolated aorta of the alpha(1D)-adrenoceptor knockout (KO) mice, by chloroethylclonidine (CEC)-induced alkylation and their protection by selective alpha(1)-adrenoceptor antagonists. 2 The alpha(1D)-adrenoceptor is involved in the contractile response to noradrenaline in wild type (WT) mouse aorta. 3 In WT mice 5-methylurapidil (5-MU, an alpha(1A)-adrenoceptor antagonist) or BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl] ethyl]-8-azaspiro[4.5] decane-7,9 dione, a selective alpha(1D)-adrenoceptor antagonist), protected the receptors from CEC-induced (alpha(1B/D)-adrenoceptor) alkylation, the combination of both antagonists resulted in complete protection, while AH11110A (1-[biphenyl-2-yloxy]-4-imino-4-piperidin-1-yl-butan-2-ol, an alpha(1B)-adrenoceptor antagonist) did not protect. 4 In aorta of KO mice there was a 19-fold rightward shift in noradrenaline effective concentration (EC(50)) compared with WT; while 5-MU alone or in combination with AH11110A protected alpha(1)-adrenoceptors to the same extent. 5 The data indicate that alpha(1A)-adrenoceptors mediate contraction and suggest their role in maintaining homeostasis in the alpha(1D)-adrenoceptors KO mice.     

The in vitro and in vivo pharmacological activity of Boiga dendrophila (mangrove catsnake) venom

The great taxonomic and prey base diversity of colubrids (non-front-fanged snakes) suggests that their venoms may represent a 'literal gold mine' for scientists eager to find novel pharmacological probes. While pharmacological characterization is lacking for most of these venoms, this is even more so with regard to activity of colubrid venoms on the mammalian autonomic nervous system. This study characterizes the activity of venom from the colubrid, Boiga dendrophila using in vitro smooth muscle preparations and the anaesthetized rat. In the prostatic segment of the rat vas deferens, cumulative additions of venom (1-150 microg ml(-1)) induced concentration-dependent inhibition of electrically evoked (0.2 Hz, 0.3 ms, 70-100 V) twitches. The inhibitory effect of venom (100 microg ml(-1)) was attenuated by 8-phenyltheophylline (8-PT) (20 microM) and 8-cyclopentyl-1, 3-dipropylxanthine (20 microM) but not idazoxan (1 microM), or a combination of ranitidine (0.2 microM) and thioperamide (10 microM). The inhibitory effect of venom (100 microg ml(-1)) was augmented by dipyridamole (10 microM) but abolished by pretreatment with adenosine deaminase (7.5 units/100 microl) suggesting that it contains components with adenosine A(1) receptor activity, most likely adenosine. In isolated segments of guinea-pig ileum, venom (10-100 microg ml(-1)) caused concentration-dependent contractions which were inhibited by the muscarinic receptor antagonist atropine (0.1 microM) but not by the histamine receptor antagonist mepyramine (0.5 microM). In the anaesthetized rat, venom (5-7.5 mg kg(-1), i.v.) caused a hypotensive effect. Our data suggest that the venom contains components with purinergic and muscarinic receptor activity.     

β-Endorphin: peripheral opioid activity of homologues from six species

The peripheral opioid activity of six homologous β-endorphins (β-EPs) were assayed on the guinea pig ileum and the vas deferens of the mouse, the rat and the rabbit. In the guinea pig ileum assay, human β-EP (βh-EP) was less potent than camel, turkey, and ostrich β-EPs, of the same potency as equine β-EP and more active than des-acetyl salmon β-EP. In the rat vas deferens, mammalian β-EPs showed higher activity than those from the bird and the fish, whereas in the mouse vas deferens assay, βh-EP is more active than those from other species. In the rabbit vas deferens, however, all homologous β-EPs show very weak activity. The relative potency of β-EP homologues obtained from rat vas deferens assay is in good correlation with the analgesic potency, while the receptor binding activity does not correlate with any of the four bioassays, but appears to be related to the charge properties of the peptides.     

Responsiveness, affinity constants and beta 2-adrenoceptor reserves for isoprenaline on portal veins from normo- and pre- and hypertensive rats

1. This study used contractility methods with the portal veins of 5- and 14-week-old Wistar-Kyoto normotensive rats (WKY) and spontaneously hypertensive rats (SHRs). The SHRs are prehypertensive at 5 weeks. 2. The first part of our study was to determine whether the responsiveness to isoprenaline and forskolin was altered in the maturation of portal veins from normo- and prehypertensive rats. The responses to forskolin were similar on the portal veins of 5- and 14-week-old WKY and SHRs. 3. The sensitivity and maximum responses to isoprenaline were similar on portal veins of 5- and 14-week-old WKY. The sensitivity and maximum responses to isoprenaline were lower on the portal veins of 5-week-old SHRs (pD2 = 8.25, maximum = 85%) than age-matched WKY (pD2 = 8.79, maximum = 96%); these differences are not caused by hypertension. At 14 weeks, the sensitivity was similar (WKY pD2 = 8.74, SHR pD2 = 8.65) but the maximum responses to isoprenaline were lower on the portal veins SHRs (77%) than WKY (97%). Thus, the sensitivity to isoprenaline increases with the development of hypertension in the SHR portal vein. 4. The second part of the study was to determine whether the affinity for isoprenaline at beta2-adrenoceptors and the fractional beta2-adrenoceptor occupancy-response relationships on the portal vein were altered in maturation from normo- and pre-hypertensive rats. The effects of bromoacetylalprenololmenthane (BAAM), an irreversible beta-adrenoceptor blocker, on the isoprenaline responses of 5- and 14-week-old WKY and SHRs were studied. Maturation of the WKY portal vein between 5 and 14 weeks was associated with a loss of affinity for isoprenaline (from pKA of 7.13 to 7.87), and increase in beta2-adrenoceptor reserve (from 72 to 92% at the 95% response). There were similar affinity and reserve findings in the maturation of the SHR portal vein. Thus, there are major changes in beta2-adrenoceptor structure and reserve in maturation on the portal vein that are irrespective of the development of hypertension.     

NO对去甲吗啡在离体豚鼠回肠上依赖性耐受性产生的作用

目的观察 ＮＯ合酶（ ＮＯ ｓｙｎｔｈａｓｅ,ＮＯＳ）抑制剂左旋硝基精氨酸（ＮＧ－ｎｉｔｒｏ－Ｌ－ａｒｇｉｎｉｎｅ,ＮＯＡｒｇ）和 ＮＯ合成前体左旋精氨酸（Ｌ－ａｒｇｉｎｉｎｅ,Ｌ－Ａｒｇ）对去甲吗啡（ｎｏｒｍｏｒｐｈｉｎｅ,ＮＭ）在离体豚鼠回肠上依赖性耐受性产生的影响,研究ＮＯ在阿片类药物依赖性耐受性产生中的作用。方法以离休豚鼠回肠为实验对象,４℃孵育６ ｈ, ２４ ｈ后,分别测定纳洛酮（ｎａｌｏｘｏｎｅ,Ｎｘ）与氯化乙酰胆碱（ａｃｅｔｙｌｃｈｏｌｉｎｅ,Ａｃｈ）引起的收缩高度比值,作为依赖性产生的指标。在耐受性产生实验中,分别测定回肠空白组和药物组４℃孵育３ｈ后对ＮＭ抑制电刺激收缩的剂量反应曲线。结果回肠分别经ＮＭ,ＮＭ加ＮＯＡｒｇ,ＮＭ加Ｌ－Ａｒｇ中孵育６ｈ后,纳洛酮均可使其产生戒断收缩,且两组与Ａｃｈ的收缩高度比值差异无显著性（ｎ＝６）, Ｐ＞０．０５。而孵育２４ ｈ后,ＮＭ,ＮＭ加 Ｌ－Ａｒｇ组收缩高度比值分别为 ０． ８９±０． ０２, ０． ９０±０． ０２, Ｐ＞ ０． ０５,而ＮＭ加ＮＯＡｒｇ药物组对纳洛酮的戒断收缩反应明显减弱,收缩高度比值为 ０．４７± ０．０５（ｎ＝ ６, Ｐ＜ ０．０１）。可见ＮＭ加ＮＯＡ?     

Cyclic enkephalin analogs containing various para‐substituted phenylalanine derivatives in place of Tyr1 are potent opioid agonists*

Abstract: The cyclic enkephalin analog H‐Tyr‐c[d ‐Cys‐Gly‐Phe(pNO₂)‐d ‐Cys]NH₂ is a highly potent opioid agonist with IC₅₀s of 35 pm and 19 pm in the guinea‐pig ileum (GPI) and mouse vas deferens (MVD) assays, respectively. The Phe¹‐analog of this peptide showed 370‐fold and 6790‐fold lower agonist potency in the GPI and MVD assays, respectively, indicating the importance of the Tyr¹ hydroxyl‐group in the interaction with μ and δ opioid receptors. In the present study, the effect of various substituents (‐NH₂, ‐NO₂, ‐CN, ‐CH₃, ‐COOH, ‐COCH₃, ‐CONH₂) introduced in the para‐position of the Phe¹‐residue of H‐Phe‐c[d ‐Cys‐Gly‐Phe(pNO₂)‐d ‐Cys]NH₂ on the in vitro opioid activity profile was examined. Most analogs showed enhanced μ and δ agonist potencies in the two bioassays, except for the Phe(pCOOH)¹‐analog, which was weakly active, probably as a consequence of the negative charge. The most potent compounds were the Phe(pCOH₃)¹‐ and the Phe(pCONH₂)¹‐analogs. The latter compound showed subnanomolar μ and δ agonist potencies and represents the most potent enkephalin analog lacking the Tyr¹ hydroxyl‐group reported to date. Taken together, these results indicate that various substituents introduced in the para‐position of Phe¹ enhance opioid activity via hydrogen bonding or hydrophobic interactions with the receptor. Comparison with existing structure‐activity relationship on phenolic hydroxyl replacements in morphinans indicates that these nonpeptide opiates and some of the cyclic enkephalin analogs described here may have different modes of binding to the receptor.     

In vitro pharmacology of the opioid peptides, enkephalins and endorphins.

In the guinea-pig ileum methionine-enkephalin, normorphine and morphine are equipotent in depressing electrically evoked contractions; leucine-enkephalin has about 25% of the activity. The mouse vas deferens is more sensitive to the enkephalins which are 30 to 60 times more potent than morphine. Fragments of beta-lipotropin61-91 (beta-endorphin) having sequences up to LPH76 are more potent in the mouse vas deferens than in the guinea-pig ileum but beta-endorphin is about equipotent in the two preparations. None of the peptides has antagonist activity. Methionine-enkephalin and normorphine are equipotent in inhibiting [3H]-naloxone binding by homogenate of guinea-pig brain in the absence of Na+ while leucine-enkephalin has only 25% of this activity. In the guinea-pig ileum, naloxone antagonises normorhine and the enkephalins equally well whereas in the mouse vas deferens about ten times more naloxone is required for the enkophalins that for normorphine. Methionine-enkephalin depresses output of acetylcholine in the guinea-pig ileum and of noradrenaline in the mouse vas deferens.     

眼镜蛇神经毒素对大鼠脑内脑啡肽含量及豚鼠回肠和小鼠输精管收缩的影响

连续注射1wk眼镜蛇神经毒素后,用放射免疫测定法测定对照组及神经毒素组大鼠5个脑区亮啡肽样免疫反应物质及甲硫啡肽样免疫反应物质的含量。神经毒素组两种脑啡肽样物质在下丘脑、中脑+丘脑两个脑区的含量变化均出现明显升高(p<0.05),海马及后脑的含量均无显著变化。吗啡在很低浓度对低频电刺激豚鼠回肠及小鼠输精管收缩有明显抑制作用。神经毒素的存在与否并不影响吗啡的效应。结果提示,神经毒素可能并不直接与吗啡受体产生相互作用,但其镇痛机理可能涉及中枢内源性阿片肽能系统。     

Pharmacological profiles of β-funaltrexamine (β-FNA) and β-chlornaltrexamine (β-CNA) on the mouse vas deferens preparation

The profiles of action of β-funaltrexamine (β-FNA) and β-chlornaltrexamine (β-CNA) have been assessed in the mouse vas deferens preparation. β-FNA, but not β-CNA, demonstrated a reversible agonist action that appeared to be mediated via κ-receptor interaction. β-CNA produced an irreversible antagonism of μ-, κ- and δ-mediated agonist actions, whereas β-FNA irreversibly antagonized μ-mediated agonist effects only. This selective action of β-FNA could also be seen following administration in vivo. β-CNA and particularly β-FNA should prove valuable in the elucidation of multiple opioid receptors.     

Evidence of alpha1-adrenoceptor functional changes in omental arteries of patients with end-stage renal disease

1 Alpha1-Adrenoceptor (alpha1-AR) subtypes were characterized in isolated omental arteries obtained after abdominal surgery in patients with end-stage renal disease (ESRD) or with Diabetes Mellitus type 2 plus ESRD (ESRD-DM). 2 Omental arteries from patients with ESRD and ESRD-DM elicited a significant increase in sensitivity to phenylephrine with a pD(2) (-log EC50) of 6.7 and 6.6, respectively, vs. the control (5.8, P < 0.001). 3 Stimulation with phenylephrine was conducted in the presence or absence of selective alpha1-AR competitive antagonists: 5-methylurapidil (alpha1A-), AH11110A (1-[biphenyl-2-yloxy]-4-imino-4-piperidin-1-yl-butan-2-ol; alpha1B-) and BMY7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro [4.5] decane-7,9-dione; alpha(1D)-). The relative abundance of mRNA for all three alpha(1)-ARs was determined. 4 The maximal contractile responses to phenylephrine were: E(max) 1.59 +/- 0.17, 1.48 +/- 0.08 and 1.55 +/- 0.14 g for the ESRD, ESRD-DM and control groups, respectively. 5 Functionally, there was an increment in the affinity for the alpha(1A)-AR antagonist (pA2: control 7.45, ESRD 8.36, ESRD-DM 8.0; P < 0.01), and a reduction in the alpha1B-AR antagonist affinity (8.3 for controls, 7.6 for ESRD and 7.3 for ESRD-DM; P < 0.01) associated with renal disease. The affinities for the alpha1D-AR antagonist were similar among the studied groups (8.5 for the controls, 8.7 for the ESRD and 8.1 for the ESRD-DM groups). 6 Renal disease increased mRNA expression of alpha(1B)-ARs and reduced both alpha1A- and alpha(1D)-ARs subtypes in ESRD and ESRD-DM patients. 7 The results suggest that human omental arteries exposed to chronic uraemia show vascular hypersensitivity to phenylephrine, because of functional alpha1-AR changes.     

Roles of c-Src in alpha1B-adrenoceptor phosphorylation and desensitization

1 The role of the protein tyrosine kinase, c-Src, on the function and phosphorylation of alpha1B-adrenoceptors (alpha1B-AR) and their association with G-protein-coupled receptor kinase (GRK) isozymes was studied. 2 Inhibitors of this kinase (PP2 and Src Inhibitor II) decreased ( approximately 50-75%) noradrenaline- (NA) and phorbol myristate acetate-mediated receptor phosphorylation. Expression of a dominant-negative mutant of c-Src similarly reduced receptor phosphorylation induced by the natural agonists, active phorbol esters and endothelin-1 (ET-1). 3 c-Src, GRK2, GRK3 and GRK5 coimmunoprecipitate with alpha1B-ARs in the basal state. In cells treated with NA or phorbol myristate acetate the amount of coimmunoprecipitated GRK2 and GRK3 increased ( approximately 2- to 3-fold), while treatment with ET-1 only augmented the amount of coimmunoprecipitated GRK2 ( approximately 2-fold). The Src inhibitor, PP2, markedly attenuated all these increases. 4 Cell pretreatment with PP2 amplified the increase in intracellular-free calcium observed with NA, in the basal state and after the stimulation (desensitization) induced by ET-1. 5 The data suggest a role of c-Src in alpha1B-AR desensitization/phosphorylation and in the interaction of these ARs with GRKs.     

Opposite effects of endogenous nitric oxide and prostaglandin F2 alpha in the rat mesenteric bed

1. The relationship between the effects of endogenous nitric oxide (NO) and prostanoids on the noradrenaline (NA)-induced contractions and the mechanisms involved were investigated in the rat perfused mesenteric bed, using NG-nitro-L-arginine methyl ester (l-NAME), a NO synthase inhibitor and sodium nitroprusside (SNP), a NO donor. 2. The constrictor responses to NA were reduced to 50% by the cyclooxygenase inhibitor 10 microm indomethacin as well as by 1 microM SNP. When indomethacin and SNP were perfused simultaneously the contractions were further reduced. 3. The NA-induced contractions were increased by the addition of 400 microM L-NAME and the addition of either indomethacin or SNP abolished such increases. The simultaneous perfusion of both agents further reduced the contractions. 4. Removal of the endothelium increased NA-induced contractions to a similar extent as L-NAME and this increase was abolished by indomethacin as well as by SNP. 5. The perfusion of 10 microM NA augmented the release of prostaglandin (PG) F2 alpha by the mesenteric bed without modifications in any other prostanoid. In the presence of L-NAME, this effect was further increased. However, SNP abolished the NA-induced stimulation of PGF2 alpha release. 6. In de-endothelialized preparations NA also stimulated PGF2 alpha production as observed in intact preparations. This effect was more marked in the presence of L-NAME; in contrast, SNP abolished the stimulation. 7. In conclusion, the present results suggest an opposite action between NO and PGF2 alpha on the NA-induced contractions in the rat mesenteric bed.     

Acute withdrawal induced by adenosine A1‐receptor activation in isolated guinea‐pig ileum: role of opioid receptors and effect of cholecystokinin

Objectives In isolated guinea‐pig ileum, the μ‐opioid acute withdrawal response is under control of several neuronal systems, including the κ‐opioid and the A₁‐adenosine systems, which are involved in the μ‐withdrawal response inhibitory control. After μ‐opioid system stimulation, indirect activation of both κ‐opioid and A₁‐adenosine systems is prevented by the peptide cholecystokinin‐8 (CCk‐8). Guinea‐pig ileum exposed to A₁‐adenosine agonist (CPA), shows a withdrawal contracture precipitated by the A₁‐adenosine antagonist (CPT). We investigated this response. Methods We investigated the involvement of the opioid system in the A₁‐adenosine acute withdrawal response in guinea‐pig ileum, the potential induced cross‐dependence between the A₁ and the opioid system and also the interaction between the CCk‐8 and A₁ systems. Key findings We found that in the guinea‐pig ileum preparation exposed to CPA, μ‐ and κ‐opioid antagonists increased the withdrawal response to CPT. Tissues exposed to CPA showed a contractile response to the opioid receptor antagonist naloxone only after complete removal of the A₁‐agonist. In the presence of CPA, the response to CCk‐8 was inhibited while a significant increase in CPT response intensity was observed. Conclusions In guinea‐pig ileum, stimulation of the A₁ system indirectly activates both μ‐ and κ‐opioid systems; this indirect activation is significantly, albeit not completely, antagonised by CCk‐8. Cross dependence between A₁ and opioid systems was also observed.     

A comparison of the effects of the 5HT1A antagonists MM-77 and WAY-100635 on the mouse isolated vasa deferentia

1. Experiments were carried out to characterize the possible adrenergic properties of the 5-HT(1A) antagonists WAY 100635 and MM-77 using the mouse isolated vasa deferentia preparation. 2. When vasa deferentia were preincubated for 10 min in the presence of MM-77 (10(-8)-10(-6) m) or WAY100635 (10(-8)-7 x 10(-7) m), a concentration-dependent inhibition of the contractile response to submaximal electrical field stimulation (10 Hz, 50 V, 50 ms) was observed with pIC(50) values of 7.05 +/- 0.01 and 6.85 +/- 0.1 respectively. 3. MM-77 (10(-8)-10(-6) m) antagonized the contractile responses of the vasa deferentia to phenylephrine (PE) (10(-6)-10(-3) m) in a concentration-dependent manner. Schild plots of these data were linear and yielded a mean rhoA(2) value of 6.81 +/- 0.084. The mean slope was 1.42 +/- 0.22. 4. WAY100635 (10(-8)-10(-6) m) antagonized the contractile responses of the vasa deferentia to PE (10(-6)-10(-3) m) in a concentration-dependent manner. Schild plots of these data were linear and yielded a mean rhoA(2) value of 7.05 +/- 0.08. The mean slope was 0.97 +/- 0.1. 5. The results suggest that while WAY100635 acts as a competitive antagonist at alpha(1)-adrenoceptors, MM-77 displays non-competitive antagonist characteristics at this receptor subtype. 6. These results may have important implications for the use of these compounds as 5-HT(1A) receptor antagonists in in vivo studies.     

Alpha1A-adrenoceptors predominate in the control of blood pressure in mouse mesenteric vascular bed

1 The pressor action of the alpha1A-adrenoceptor agonist, A61603 (N-[5-(4,5-dihydro-1H-imidazol-2-yl)-2-hydroxy-5,6,7,8-tetrahydronaphthalen-1-yl] methanesulfonamide) or the alpha1-adrenoceptor agonist phenylephrine, and their blockade by selective alpha1-adrenoceptor antagonists in the mouse isolated mesenteric vascular bed were evaluated. 2 A61603 showed a approximately 235-fold higher potency in elevating perfusion pressure in mesenteric bed compared to phenylephrine. 3 The alpha1A-adrenoceptor selective antagonist RS 100329 (5-methyl-3-[3-[4-[2-(2,2,2,-trifluoroethoxy) phenyl]-1-piperazinyl] propyl]-2,4-(1H)-pyrimidinedione), displaced with high affinity agonist concentration-response curves to the right in a concentration-dependent manner. 4 The alpha1D-adrenoceptor selective antagonist BMY 7378 (8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4.5] decane-7,9-dione), did not displace A61603 nor did it block the phenylephrine-induced pressor response. 5 The alpha1B/D-adrenoceptor alkylating antagonist chloroethylclonidine (CEC), caused a rightward shift of the phenylephrine concentration-response curve and reduced its maximum response; however, CEC only slightly modified A61603 evoked contraction. 6 The results indicate that the isolated mouse mesenteric vascular bed expresses alpha1A-adrenoceptors and suggest a very discrete role for 1B-adrenoceptors.     

Alpha2-adrenoceptor subtypes involved in the regulation of catecholamine release from the adrenal medulla of mice

BACKGROUND AND PURPOSE: This study was carried out to elucidate which alpha(2)-adrenoceptor subtypes mediated the inhibition of noradrenaline and adrenaline release from the adrenal medulla of mice. EXPERIMENTAL APPROACH: Isolated adrenal medullae from wild-type and alpha(2A), alpha(2B) and alpha(2C)-adrenoceptor knockout (KO) mice were placed in superfusion chambers. Catecholamine overflow was evoked by 1,1-dimethyl-4-phenylpiperazinium (500 microM) in absence or in presence of the alpha(2)-adrenoceptor agonist medetomidine. The effect of medetomidine was tested in presence of the alpha-adrenoceptor antagonists rauwolscine, WB 4101, spiroxatrine, phentolamine and prazosin. KEY RESULTS: In wild-type mice, medetomidine reduced noradrenaline and adrenaline overflow in a concentration-dependent manner (EC(50) in nM: 1.54 and 1.92; E(max) in % of inhibition: 91 and 94, for noradrenaline and adrenaline, respectively). The pK (D) values of the antagonists for noradrenaline overflow did not correlate with pK(D) values at alpha(2A), alpha(2B), or alpha(2C) binding sites. The pK (D) values of the antagonists for adrenaline overflow correlated positively with pK(D) values at alpha(2C) binding sites (opossum kidney cells). The effect of medetomidine (100 nM) on noradrenaline overflow was significantly reduced in all three alpha(2)KO mice (57, 54, 44 % inhibition, for alpha(2A), alpha(2B), and alpha(2C), respectively), whereas the effect of medetomidine on adrenaline overflow was greatly reduced in alpha(2C)KO mice (14 % inhibition). CONCLUSIONS AND IMPLICATIONS: In the adrenal medulla of mice, all three alpha(2)-adrenoceptor subtypes (alpha(2A), alpha(2B), and alpha(2C)) play an equal role in the inhibition of noradrenaline overflow, whereas the alpha(2C)-adrenoceptor is the predominant alpha(2)-adrenoceptor subtype involved in the inhibitory mechanism controlling adrenaline overflow.