Mouse Vas Deferens Preparations Research Articles

1-Benzyl-N-[3-[spiroisobenzofuran-1(3H),4′-piperidin-1-yl]propyl]pyrrolidine-2-carboxamide (Compound 24) antagonizes NOP receptor-mediated potassium channel activation in rat periaqueductal gray slices

Nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor, the fourth member of opioid receptor family, shares 60–70% homology with traditional opioid receptors but displays little affinity for opioids. This receptor was implicated in many neurological functions and its functional heterogeneity has been proposed. Therefore, it is imperative to develop and characterize new ligands for NOP receptors. 1-Benzyl-N-[3-[spiroisobenzofuran-1(3H),4′-piperidin-1-yl]propyl]pyrrolidine-2-carboxamide (Compound 24) is a new non-peptide ligand of NOP receptor having antagonistic actions in cloned and peripheral NOP receptors. In this study, we quantitatively characterized its effect on the native NOP receptors in the midbrain slices containing ventrolateral periaqueductal gray (vlPAG), a region with dense NOP receptors and involved in pain regulation. In vlPAG neurons, N/OFQ induced G-protein-coupled inwardly rectifying potassium (GIRK) current through NOP receptors. Compound 24, at 0.3–10 μM, attenuated N/OFQ-induced GIRK current concentration-dependently. The antagonistic potency of Compound 24 in vlPAG neurons (IC 50: 2.6 ± 0.6 μM) was, however, lower than that obtained in mouse vas deferens preparations or expressed human NOP receptors. The action kinetic of Compound 24 was slower than [Nphe 1, Arg 14, Lys 15]N/OFQ-NH 2 (UFP-101), a peptide antagonist, in the same preparation. Compound 24 had no intrinsic agonistic activity at NOP receptors at the concentration up to 10 μM. However, at concentrations higher than 3 μM, it also attenuated the GIRK current induced by [D-Ala 2, N-Me-Phe 4, Gly 5-ol]-enkephalin, a mu-opioid receptor agonist. It is concluded that Compound 24 acts as a pure antagonist at the native NOP receptors in the vlPAG with moderate potency and selectivity.

Activities of mixed NOP and μ‐opioid receptor ligands

Compounds that activate both NOP and mu-opioid receptors might be useful as analgesics and drug abuse medications. Studies were carried out to better understand the biological activity of such compounds. Binding affinities were determined on membranes from cells transfected with NOP and opioid receptors. Functional activity was determined by [(35)S]GTPgammaS binding on cell membranes and using the mouse vas deferens preparation in vitro and the tail flick antinociception assay in vivo. Compounds ranged in affinity from SR14150, 20-fold selective for NOP receptors, to buprenorphine, 50-fold selective for mu-opioid receptors. In the [(35)S]GTPgammaS assay, SR compounds ranged from full agonist to antagonist at NOP receptors and most were partial agonists at mu-opioid receptors. Buprenorphine was a low efficacy partial agonist at mu-opioid receptors, but did not stimulate [(35)S]GTPgammaS binding through NOP. In the mouse vas deferens, each compound, except for SR16430, inhibited electrically induced contractions. In each case, except for N/OFQ itself, the inhibition was due to mu-opioid receptor activation, as determined by equivalent results in NOP receptor knockout tissues. SR14150 showed antinociceptive activity in the tail flick test, which was reversed by the opioid antagonist naloxone. Compounds that bind to both mu-opioid and NOP receptors have antinociceptive activity but the relative contribution of each receptor is unclear. These experiments help characterize compounds that bind to both receptors, to better understand the mechanism behind their biological activities, and identify new pharmacological tools to characterize NOP and opioid receptors.

Opioid Peptides: Synthesis and Biological Activity of New Endomorphin Analogues

Syntheses are described of new endomorphin 1 and 2 peptoid–peptide hybrids in which Tyr1 and either one or both Phe3 and Phe4 have been replaced by N-substituted-glycine. The preparation is also described of two glycosylated Hyp2-endomorphin 2 analogues in which either 2,3,4,6-tetra-O-acetyl glucose or glucose are β-O-glycosidically linked to the hydroxyproline residue. The Hyp2-endomorphin sequences have also been elongate by adding a C-terminal β-alanine residue and several linear dimers have been prepared by coupling either the native peptides or the modified analogues. The cyclo endomorphin 2 has also been synthesized. Preliminary pharmacological experiments on isolated organ preparations showed that the agonist activities of both endomorphin 1 and 2 are not significantly affected by the Pro/Hyp substitution. Phe4/Nphe substitution in the endomorphin 1 reduced the potency on guinea pig ileum (GPI) by about 100 times and abolished the agonist activity on mouse vas deferens (MVD) preparation. The decrease of the agonist activity induced by modification of one phenylalanine residue only, either Phe3 or Phe4, is lower on endomorphin 2. Either modification of both Phe3 and Phe4 or glycosylation of the Hyp2-endomorphin 2 cancelled any agonist activity on both preparations. The linear peptide dimers [endomorphin 1]2, [endomorphin 2]2, [Hyp2-endomorphin 1]2, [Hyp2-endomorphin 2]2, [Hyp2-endomorphin 1-Hyp2-endomorphin 2]2 or [Hyp2-endomorphin 2-Hyp2-endomorphin 1]2, are 7–19 times less potent than endomorphin 1 on GPI and significantly less active than endomorphins 1 and 2 on MVD. The other afforded modifications significantly affected or abolished the agonist activity of the resulting endomorphin analogues on both GPI and MVD preparations.

Novel Nociceptin Analogues: Synthesis and Biological Activity

Syntheses are described of the nociceptin (1–13) amide [NC(1–13)-NH2] and of several analogues in which either one or both the phenylalanine residues (positions 1 and 4), the arginine residues (positions 8 and 12) and the alanine residues (positions 7 and 11) have been replaced by N-benzyl-glycine, N-(3-guanidino-propyl)-glycine and β-alanine, respectively. The preparation is also described of NC(1–13)-NH2 analogues in which either galactose or N-acetyl-galactosamine are β-O-glycosidically linked to Thr5 and/or to Ser10. Preliminary pharmacological experiments on mouse vas deferens preparations showed that Phe4, Thr5, Ala7 and Arg8 are crucial residues for OP4 receptor activation. Manipulation of Phe1 yielded peptides endowed with antagonist activity but [Nphe1] NC(1–13)-NH2 acted as an antagonist still possessing weak agonist activity. Introduction of the βAla residue either in position 7 or 11 of the [Nphe1] NC(1–13)-NH2 sequence, abolished any residual agonist activity and [Nphe1, βAla7] NC(1–13)-NH2 and [Nphe1, βAla11] NC(1–13)-NH2 acted as competitive antagonists only. Modification of both Ala7 and Ala11 abolished the antagonist activity of [Nphe1]NC(1–13)-NH2 probably by hindering receptor binding. Changes at positions 10 and 11 gave analogues still possessing agonist activity. [Ser(βGal)10] NC(1–13)-NH2 displayed an activity comparable with that of NC(1–13)-NH2, [Ser(βGalNAc)10] NC(1–13)-NH2 and [βAla11] NC(1–13)-NH2 were five and 10 times less active, respectively.

N1'-(p-[18F]Fluorobenzyl)naltrindole (p-[18F]BNTI) as a potential PET imaging agent for DOP receptors

The N1'-(p-fluorobenzyl)naltrindole 5 has been synthesized by reaction of 3-O-benzyl NTI 3 with p-fluorobenzylbromide under phase transfer catalysis. The subsequent 3-O-benzyldeprotection of 4 in HBr/CH3COOH gave the target compound 5 in three steps from naltrindole 2. p-FBNTI 5 is a novel delta opioid receptor antagonist (Ki=0.00312 nM) and antagonizes the delta opioid (DOP) agonist, DPDPE, with a Ke=1.55 nM in the mouse vas deferens preparation. Using the same synthetic strategy the synthesis of p-[18F]BNTI 10 was undertaken. The final yield was 4% and the specific activity varied in a range of 250–400 mCi/µmol. Copyright © 2006 John Wiley & Sons, Ltd.

P.6.003 Comorbid psychiatric diagnosis and substance abuse in pathological gamblers - a preliminary gender comparison study

[Phe1ψ(CH2-NH)Gly2]nociceptin-(1-13)-NH2, a pseudopeptide analog of nociceptin, has been shown to be a selective `antagonist' of the nociceptin receptor in the isolated guinea pig ileum and mouse vas deferens preparations (Guerrini et al., 1998. Br. J. Pharmacol. 123, 163–165). However, in recombinant chinese hamster ovary cells expressing the human nociceptin receptor, we find that the pseudopeptide is a potent (IC50=7.5 nM) and fully efficacious inhibitor of forskolin-induced accumulation of cAMP, thus behaving as a pure `agonist' rather than an antagonist of the receptor. The contrary behaviour of the pseudopeptide in smooth muscle and transformed cells may suggest that different nociceptin receptor types are being addressed in the two systems.

In vitro opioid activity profiles of 6-amino acid substituted derivatives of 14- O-methyloxymorphone

A series of 6-amino acid conjugates (glycine, alanine and phenylalanine) of the highly potent opioid analgesic 14- O-methyloxymorphone was developed in an effort to obtain agonists that would have potentially limited ability to cross the blood–brain barrier. Binding studies revealed that all derivatives displayed high affinities (0.77–2.58 nM) at the μ-opioid receptor in rat brain membranes. They were potent agonists in mouse vas deferens preparation (IC 50=5.52–26.8 nM). While the α-amino acid epimers are favoured by μ-opioid receptors, the β-epimers proved to have increased interaction with δ-sites. Only the β-phenylalanine conjugate showed some preference for δ- over μ-opioid receptors and δ-opioid receptor agonist activity. The relatively high δ-opioid receptor affinity of this analogue was also predicted by molecular modelling studies. The newly developed ionizable derivatives could find clinical applications as potent analgesics without the adverse actions of centrally acting opioids.

Effect of opioid receptor ligands on the μ-S196A knock-in and μ knockout mouse vas deferens

We have determined the effect of naltrexone, naloxone, [ d-Ala 2, d-Leu 5]enkephalin (DADLE), and morphine on the μ-S196A opioid receptor knock-in and μ-opioid receptor knockout mouse vas deferens preparations. The antagonists, naltrexone and naloxone, exhibited agonist activity and possessed IC 50 values that were 14- and 37-fold greater than morphine on the S196A preparation. Morphine was found to be threefold more potent at S196A relative to wild-type μ-opioid receptor. The mouse vas deferens data suggest that S196 in transmembrane helix 4 of the μ-opioid receptor modulates efficacy. It is proposed that this may be due to decreased dimerization of the receptor. Identical IC 50 values of DADLE obtained on the wild-type, S196A knock-in, and μ-opioid receptor knockout preparations support the absence of μ–δ heterodimers in the mouse vas deferens.

Opioid peptides: synthesis and biological properties of [(N gamma-glucosyl,N gamma-methoxy)-alpha, gamma-diamino-(S)-butanoyl]4-deltorphin-1-neoglycopeptide and related analogues.

The [D-Ala2]deltorphin 1 sequence in which the aspartic acid residue is replaced by the N gamma-OCH3-alpha, gamma-diamino (S) butanoyl residue was synthesized using the Fmoc-chemistry-based solid phase procedure. The resulting deltorphin analogue was chemoselectively glucosylated by reaction with unprotected D-glucose (Glc). The Asn4-, (2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-beta-D-galactopyranosyl)-Asn4- and the (2-acetamido-2-deoxy-D-galactopyranosyl)-Asn4-deltorphin I were also prepared for comparison. The affinity of the new compounds for the delta-opioid receptor was expressed by the inhibition constant (Ki) of the binding of the delta-receptor selective ligand [3H]naltrindole (NTI) to rat brain membrane preparations. The in vitro biological activity of the synthetic peptides was compared with that of the mu-opioid receptor agonist dermorphin in guinea pig ileum (GPI) preparations and with that of the delta-opioid receptor agonist deltorphin I in mouse vas deferens (MVD) preparations. The substitution of Asp4 with Asn failed to affect drastically the Ki and IC50 values for delta-sites, suggesting that an electrostatic interaction does not play an essential role in the binding to delta-opioid sites. The steric hindrance of the side chain of the residue in position 4 affects binding to delta-sites. The increase of the Ki value is smaller when the sugar-peptide linkage involves the gamma-nitrogen of the Dab residue in comparison with the Asn amide side chain.

The effect of guanethidine and local anesthetics on the electrically stimulated mouse vas deferens.

Complex regional pain syndrome is often treated with the sympatholytic guanethidine and a local anesthetic in a Bier's block. The efficacy of this treatment has been questioned. Because local anesthetics inhibit the norepinephrine uptake transporter, we hypothesized that this variable efficacy results from the local inhibiting the uptake of guanethidine. In this study, we tested this hypothesis by using a sympathetically innervated mouse vas deferens preparation. Organ bath-mounted mouse vasa deferentia were electrically stimulated in the absence and presence of guanethidine, prilocaine, procaine, and cocaine in various combinations. Prilocaine (1 mM) induced an immediate inhibition of twitch response (maximum 100% after 2 min) that fully reversed after washing. Guanethidine (3 microM) also inhibited twitching by 95% +/- 3% in 15 min, but this effect was only partially reversed after 1 h of washing (33% +/- 12% of control). When prilocaine and guanethidine were added in combination, a reversal of 80% +/- 13% (at 1 h) was observed. Procaine (300 micro M) produced a transient increase (152% +/- 14%) in response. When co-incubated with guanethidine (3 microM), the twitch was reduced to 24% +/- 4% of control and was reversed to 77% +/- 7% after 1 h. Cocaine (30 microM) inhibited the twitch response to 53% +/- 8%, which was fully reversed by 1 h of washing. When co-incubated with guanethidine, the response was reduced to 39% +/- 6% of control and was reversed to 86% +/- 10% after 1 h. In all cases, the reversal produced by the combination was significantly more intense (P < 0.05) than that produced by guanethidine alone. Local anesthetics reduce the sympatholytic actions of guanethidine, and this may explain the variable efficacy of guanethidine in the treatment of complex regional pain syndrome. In this study, with a sympathetically innervated vas deferens preparation, local anesthetics reduced the efficacy of the sympatholytic guanethidine, questioning its co-administration in the pain clinic.

Neuropeptide Y release by pumiliotoxin-B in the electrically-stimulated mouse vas deferens: an immunohistochemical study☆

Morphologic and immunohistochemical studies were conducted to ascertain whether pumiliotoxin-B (PTX-B), an indolizine alkaloid from the skin of the Neotropical dendrobatid frog, Dendrobates pumilio, affects the anatomic and immunohistochemical features of the electrically stimulated mouse vas deferens preparations. PTX-B, at a concentration of 1 μM, consistently decreased the density pattern of neuropeptide Y (NPY)-immunoreactive nerve fibers contained within the circular muscular layer. The alkaloid also induced striking morphologic changes. It enlarged the lumen of the vasa and relaxed the muscular wall. Pretreatment with prazosin or haloperidol affected neither the release of NPY nor the morphologic changes; pretreatment with tetrodotoxin and guanethidine abolished NPY release and prevented the PTX-B-induced morphologic changes. PTX-B had no appreciable effect on the density and distribution pattern of nerve fibers immunostained for vasoactive intestinal polypeptide, substance P, calcitonin gene-related peptide, enkephalin, pancreatic polypeptide, 5-hydroxy-tryptamine and tyrosine hydroxylase.

Transmitter release and uptake evoked by the amphibian skin alkaloid, pumiliotoxin-B (PTX-B), in the electrically stimulated mouse vas deferens preparation (MVD).

Upon electrical stimulation three transmitters are known to be released from the adrenergic nerve terminals of the isolated MVD preparation: two motor transmitters (noradrenaline (NA) and ATP) acting synergistically to provoke twitch contraction, and an inhibitory transmitter, the peptide NPY. The frog alkaloid pumiliotoxin-B (PTX-B) displayed two opposite effects on the electrically stimulated MVD: at low concentrations (0.1-0.3 microM) it caused twitch depression, at higher concentrations (0.5-2 microM) there was a potent twitch stimulation. Transmitters and/or receptors involved in the depressive effect could not be clearly identified, although interference with NPY is possible. On the other hand, the potent twitch stimulation caused by PTX-B may be due to exaggerated release of the same transmitters (NA and ATP) involved in twitch stimulation produced by electrical stimulation. Opening by PTX-B of the Na+ channels on the membrane of the adrenergic nerve terminals causes activation of the amine pump facilitating re-uptake of not only endogenous NA but also of exogenous catecholamines.

Phe 1ψ(CH 2-NH)Gly 2]nociceptin-(1-13)-NH 2 is an agonist of the nociceptin (ORL1) receptor

[Phe 1 ψ(CH 2-NH)Gly 2]nociceptin-(1-13)-NH 2, a pseudopeptide analog of nociceptin, has been shown to be a selective `antagonist' of the nociceptin receptor in the isolated guinea pig ileum and mouse vas deferens preparations (Guerrini et al., 1998. Br. J. Pharmacol. 123, 163–165). However, in recombinant chinese hamster ovary cells expressing the human nociceptin receptor, we find that the pseudopeptide is a potent (IC 50=7.5 nM) and fully efficacious inhibitor of forskolin-induced accumulation of cAMP, thus behaving as a pure `agonist' rather than an antagonist of the receptor. The contrary behaviour of the pseudopeptide in smooth muscle and transformed cells may suggest that different nociceptin receptor types are being addressed in the two systems.

Synthesis and Biological Evaluation of 14‐Alkoxymorphinans. Part 15. Novel δ opiod receptor antagonists with high affinity and selectivity in the 14‐alkoxy‐substituted indolomorphinan series

AbstractThe indolomorphinans 4–7 were prepared from their corresponding morphinan‐6‐one derivatives 8–11 via Fischer indole synthesis. Compounds 4 and 5 exhibited higher antagonist potency at δ opioid receptors in the mouse vas deferens preparation than the reference drug HS 378 (2), while compounds 6 and 7 were less potent.

Discovery of a structurally novel opioid kappa-agonist derived from 4,5-epoxymorphinan.

A new type of kappa-agonist, 17-cyclopropylmethyl-3, 14 beta-dihydroxy-4,5 alpha-epoxy-6 beta-[N-methyl-trans-3-(3-furyl) acrylamido]morphinan hydrochloride (1, TRK-820), was discovered by a new working hypothesis. The "message-address concept" for opioid antagonists and the "accessory site" for general antagonists were applied to design TRK-820. A unique structural feature of TRK-820, which is different from other prototypical kappa-opioid receptor agonists, is the existence of the 4,5-epoxymorphinan structure with a tyrosine-glysine moiety for endogenous opioid peptides such as dynorphins. TRK-820 exhibited high potency and high kappa-selectivity in guinea pig ileum (GPI) and mouse vas deferens (MVD) preparations. In the mouse acetic-acid-induced writhing model and mouse tail flick model of antinociception, TRK-820 was 85-140 times more potent than morphine and 85-350 times more potent than U-50488H. This structurally novel kappa-agonist showed neither aversion nor preference in the Conditioned Place Preference test, in spite of the fact that prototypes of kappa-agonists (U-50488H derivatives) demonstrated aversion.

Probes for narcotic receptor-mediated phenomena. 25. Synthesis and evaluation of N-alkyl-substituted (alpha-piperazinylbenzyl)benzamides as novel, highly selective delta opioid receptor agonists.

A series of N-alkyl- and N,N-dialkyl-4-[alpha-[(2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl] benzyl]-benzamides were synthesized and evaluated for binding affinities at mu, delta, and kappa opioid receptor subtypes. Several compounds (2e,f,h,i,m) strongly bound to the delta receptor with IC50 values in the nanomolar range. On the other hand, the binding affinities of these compounds for the mu and kappa receptors were in the micromolar or greater range indicating excellent delta opioid receptor subtype selectivities. In this series, two important structure-activity relationships were found for the delta receptor binding affinity. First, the spatial orientation of the alpha-benzylic position influenced the affinities with the alpha R derivatives 2a-n generally showing more than 10-fold greater affinity than the alpha S derivatives 3a-n. Second, the binding affinities were strongly influenced by the number of alkyl substituents on the amide nitrogen. N-Monoalkylbenzamide derivatives 2b-d showed lower affinity than N,N-dialkylbenzamide derivatives 2e-n, and the N-unsubstituted benzamide derivative 2a had the lowest affinity for the delta receptor in the series. The dramatic effect of the amide group substitution pattern on the binding affinity for the delta receptor strongly suggests that the amide function is an important structural element in the interaction of this series of compounds at the delta receptor. Selective compounds in this series were examined for binding affinity in cloned human mu and delta receptors. The results obtained generally paralleled those from the rat brain binding assay. Compounds 2e,f with potent delta binding affinities and high delta selectivities were shown to be delta agonists with high selectivity by studies in the guinea pig ileum (GPI) and mouse vas deferens (MVD) preparations. Compound 2f was the most selective compound in the rat brain and GPI/MVD assays with 1755- and 958-fold delta vs mu selectivity, respectively.

Synthesis and biological evaluation of 14-alkoxymorphinans. 14. 1 14-ethoxy-5-methyl substituted indolomorphinans with δ opioid receptor selectivity

The 5-methyl and 14-ethoxy substituted analogues (compounds 2 – 4) of the δ opioid receptor antagonist naltrindole showed similar selectivity when compared with the reference drug. Compound 2 was a δ receptor antagonist in the mouse vas deferens preparation (MVD) exhibiting considerably higher selectivity ratios than naltrindole, while compound 4 was found to be a full and potent δ receptor agonist in the MVD.

Effects of Some 7-Arylidene and 7-Heteroarylidene Morphinan-6-ones on the Antinociceptive Activity of [d-Pen2, d-Pen5];enkephalin and [d-Ala2, Glu4]deltorphin II and on Multiple Opioid Receptors

Bhargava, H. N., G.-M. Zhao, J.-T. Bian, Y. Nan, S. P. Upadhyaya, X. Wei, W. J. Dunn, III and L. Bauer. Effects of some 7-arylidene and 7-heteroarylidene morphinan-6-ones on the antinociceptive activity of [d-Pen2, d-Pen5] enkephalin and [d-Ala2, Glu4]deltorphin II and on multiple opioid receptors. peptides 18(5) 695–701, 1997.—The in vivo and functional effects of several 7-arylidene and 7-heteroarylidene morphinan-6-ones were determined at the μ-, δ-, and κ-opioid receptors using the guinea pig brain membranes, guinea pig ileum (GPI), and mouse vas deferens (MVD). In vivo effects included the antagonism by these compounds given subcutaneously on the antinociceptive actions of intracerebroventricularly injected [d-Pen2, d-Pen5]enkephalin (DPDPE) and [d-Ala2, Glu4]deltorphin II (deltorphin II), the highly selective putative δ1- and δ2- opioid receptor agonists. Finally, the partition coefficients of these compounds were estimated (CLOGP) and determined experimentally at pH 7.4 in the 1-octanol/water system. Compared with E-7-benzylidenenaltrexone (BNTX), most compounds except for E-7-(4-chlorobenzylidene)naltrexone, were more potent at δ-opioid receptors than at the μ-opioid receptor, whereas, in comparison to the κ-opioid receptor, the activities of the E-7-arylidene or E-7-heteroarylidene naltrexone derivatives at the δ-receptor were in the following order, where the 7-substituents were: 4-fluorobenzylidene- > benzylidene > 3-pyridylmethylene- > 4-pyridylmethylene- > 1-methyl-2-imidazolylmethylene > 4-chlorobenzylidene. In the MVD preparation, the potencies at the δ-opioid receptor, in comparison to BNTX, were in the following order, where the 7-substituents were: benzylidene = 1-methyl-2-imidazolylmethylene- > 4-fluorobenzylidene- = 3-pyridylmethylene- = 4-pyridylmethylene-. All compounds antagonized δ1 and δ2-opioid receptor agonist-induced analgesia. The antagonist potencies at the δ1-opioid receptor were in the following order, where the 7-substituents were: benzylidene- > 4-chlorobenzylidene- > 4-fluorobenzylidene- > 3-pyridylmethylene- > 1-methyl-2-imidazolylmethylene- ≈ 4-pyridylmethylene-, whereas at the δ2-opioid receptor, the order was benzylidene- > 4-chlorobenzylidene- > 4-fluorobenzylidene- > 3-pyridylmethylene- > 1-methyl-2-imidazolylmethylene- > 4-pyridylmethylene. In general, all compounds exhibited greater potency at the δ2- than δ1-opioid receptor. The computed partition coefficients were, as expected, greater than the apparent log P values, which were determined experimentally. Generally, the lipophilicity values in decreasing order were: 4-chlorobenzylidene- > 4-fluorobenzylidene- > benzylidene > 3-pyridylmethylene- = 4-pyridylmethylene- > 1-methyl-2-imidazolylmethylene-. In general, the benzylidene and 4-pyridylmethylene derivatives, which have medium lipophilicities, were equally effective at the δ1- and δ2- receptors; the 3-pyridylmethylene and 1-methyl-2-imidazolylmethylene derivatives had lower lipophilicities and were more selective for the δ2- than δ1- receptor; the 4-chlorobenzylidene and 4-fluorobenzylidene derivatives were more lipophilic and had intermediate activity. The plot of pED50 values for the in vivo tests for the δ1- and δ2- receptors showed that the two receptors are not independent with respect to this series of compounds.

Synthesis and biological evaluation of 14-alkoxymorphinans.12. 1 A phenethyl analogue of the μ-selective opioid receptor antagonist cyprodime

The N-phenethyl analogue (compound 4) of the μ-selective opioid antagonist cyprodime ( 1) has been prepared and evaluated in radioligand binding, bioassays (guinea-pig ileum myenteric plexus and mouse vas deferens preparations) and the rat tail flick test. It was found to possess remarkable affinity and high preference for μ opioid receptors and high antinociceptive potency which is comparable to its N-methyl analogue 3.

Effect of moderate cooling on contractile responses in mouse vas deferens and its relation to calcium.

Isolated mouse vas deferens preparations were used to study the effect of temperature on noradrenaline-induced contractions. Preparations were suspended in the organ bath containing Krebs-Henseleit solution for isometric tension recording. Contractile responses to noradrenaline were investigated in the mouse vas deferens after moderate cooling from 37 to 26 or 22 degrees C. A significant increase of the phasic contractions to noradrenaline was observed at 26 or 22 degrees C compared with responses obtained at 37 degrees C (about 12.3 and 35.6% increase at 26 and 22 degrees C, respectively). The secondary noradrenaline-induced sustained contraction was also significantly enhanced after moderate cooling to 26 degrees C. The potentiation of noradrenaline-induced contraction at 26 degrees C remained in a Ca(2+)-free EGTA (1 mM)-containing solution. However, sustained contraction was suppressed after removal of the calcium from the medium at 37 and 26 degrees C. Contraction to caffeine was significantly enhanced at 22 degrees C compared with 37 degrees C. By contrast, barium chloride-induced contraction of the vas deferens was markedly decreased after moderate cooling to 22 degrees C. In the presence of ouabain (0.1 mM), the noradrenaline-induced peak contraction was significantly increased at 37 degrees C. However, potentiation of the noradrenaline response at 22 degrees C was unaffected by the Na+/K+ pump inhibitor. Noradrenaline-induced peak contractions were depressed in the presence of vanadate (1 mM) and cyclopiazonic acid (10 microM), two Ca(2+)-ATPase inhibitors, at 37 degrees C and also at 22 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)