Inhibitory effect of ginger (Zingiber officinale) on rat ileal motility in vitro

Ginger (Zingiber officinale rhizome) is a widespread herbal medicine mainly used for the treatment of gastrointestinal diseases, including dyspepsia, nausea and diarrhoea. In the present study we evaluated the effect of this herbal remedy on the contractions induced by electrical stimulation (EFS) or acetylcholine in the isolated rat ileum. Ginger (0.01-1000 microg/ml) inhibited both EFS- and acetylcholine-evoked contractions, being more potent in inhibiting the contractions induced by EFS. The depressant effect of ginger on EFS-induced contractions was reduced by the vanilloid receptor antagonist capsazepine (10(-5) M), but unaffected by the alpha(2)-adrenergic antagonist yohimbine (10(-7) M), the CB(1) receptor antagonist SR141716A (10(-6) M), the opioid antagonist naloxone (10(-6) M) or by the NO synthase inhibitor L-NAME (3 x 10(-4) M). Zingerone (up to 3 x 10(-4) M), one of the active ingredients of ginger, did not possess inhibitory effects. It is concluded that ginger possesses both prejunctional and postjunctional inhibitory effects on ileal contractility; the prejunctional inhibitory effect of ginger on enteric excitatory transmission could involve a capsazepine-sensible site (possibly vanilloid receptors).     

Adenosine A<Subscript>2A</Subscript> agonist and A<Subscript>2B</Subscript> antagonist mediate an inhibition of inflammation-induced contractile disturbance of a rat gastrointestinal preparation

Adenosine can show anti-inflammatory as well as pro-inflammatory activities. The contribution of the specific adenosine receptor subtypes in various cells, tissues and organs is complex. In this study, we examined the effect of the adenosine A_2A receptor agonist CGS 21680 and the A_2BR antagonist PSB-1115 on acute inflammation induced experimentally by 2,4,6-trinitrobenzenesulfonic acid (TNBS) on rat ileum/jejunum preparations. Pre-incubation of the ileum/jejunum segments with TNBS for 30 min resulted in a concentration-dependent inhibition of acetylcholine (ACh)-induced contractions. Pharmacological activation of the A_2AR with CGS 21680 (0.1–10 μM) pre-incubated simultaneously with TNBS (10 mM) prevented concentration-dependently the TNBS-induced inhibition of the ACh contractions. Stimulation of A_2BR with the selective agonist BAY 60-6583 (10 μM) did neither result in an increase nor in a further decrease of ACh-induced contractions compared to the TNBS-induced inhibition. The simultaneous pre-incubation of the ileum/jejunum segments with TNBS (10 mM) and the selective A_2BR antagonist PSB-1115 (100 μM) inhibited the contraction-decreasing effect of TNBS. The effects of the A_2AR agonist and the A_2BR antagonist were in the same range as the effect induced by 1 μM methotrexate. The combination of the A_2AR agonist CGS 21680 and the A_2BR antagonist PSB-1115 at subthreshold concentrations of both agents found a significant amelioration of the TNBS-diminished contractility. Our results demonstrate that the activation of A_2A receptors or the blockade of the A_2B receptors can prevent the inflammation-induced disturbance of the ACh-induced contraction in TNBS pre-treated small intestinal preparations. The combination of both may be useful for the treatment of inflammatory bowel diseases.     

Internalization and recycling of human mu opioid receptors expressed in Sf9 insect cells

Internalization and recycling of G protein-coupled receptors (GPCRs), such as the mu-opioid receptor, largely depend on agonist stimulation. Agonist-promoted internalization of some GPCRs has been shown to mediate receptor desensitization, resensitization, and down-regulation. In this study, we investigated whether different mu opioid agonists displayed different effects in receptor internalization and recycling, the potential mechanisms involved in ohmefentanyl-induced internalization process. In transfected Sf9 insect cells expressing 6His-tagged wild type mu opioid receptor, exposure to 100 nM ohmefentanyl caused a maximum internalization of the receptor at 30 min and receptors seemed to reappear at the cell membrane after 60 min as determined by radioligand binding assay. Ohmefentanyl-induced human mu opioid receptor internalization was concentration-dependent, with about 40% of the receptors internalized following a 30-min exposure to 1 microM ohmefentanyl. 10 microM morphine and 1 microM DAMGO could also induce about 40% internalization. The antagonist naloxone and pretreatment with pertussis toxin both blocked ohmefentanyl-induced internalization without affecting internalization themselves. Incubation with sucrose 0.45 M significantly inhibited ohmefentanyl-induced internalization of the mu receptor. The removal of agonists ohmefentanyl and morphine resulted in the receptors gradually returning to the cell surface over a 60 min period, while the removal of agonist DAMGO only partly resulted in the receptor recycling. The results of this study suggest that ohmefentanyl-induced internalization of human mu opioid receptor in Sf9 insect cells occurs via Gi/o protein-dependent process that likely involves clathrin-coated pits. In addition, the recycling process displays the differential modes of action of different agonists.     

Kentsin: tetrapeptide from hamster embryos produces naloxone-sensitive effects without binding to opioid receptors

The opioid nature of kentsin (Thr-Pro-Arg-Lys) and its ability to alter pain perception and intestinal transit were examined. Kentsin (30,000 nM) did not inhibit electrically stimulated contractions of the guinea pig ileum (GPI) or mouse vas deferens (MVD), nor did it cause a rightward displacement of the inhibitory concentration-response curves of the mu-selective opioid agonist PL017 in the GPI or the delta-selective agonist DPDPE in the MVD. Kentsin (10,000 nM) did not displace [3H] naloxone from rat brain homogenates. These results indicate that kentsin lacks opioid agonist and mu and delta opioid antagonist properties and does not bind to opioid receptors. In vivo, kentsin produced dose-dependent analgesia in both the hotplate and abdominal stretch tests when administered intracerebroventricularly (ICV) and intrathecally but not intravenously. The central analgesic effect of kentsin was partially antagonized by the opioid antagonist naloxone. Kentsin inhibited intestinal transit in a dose-dependent manner after ICV administration only. The intestinal antitransit effect of kentsin was not blocked by pretreatment with naloxone. These results suggest that kentsin acts centrally to produce both opioid and non-opioid effects. Further, the opioid-mediated analgesic effects of kentsin involve mechanisms other than direct interaction with opioid receptors.     

Effects of interferon-alpha on cloned opioid receptors expressed in Xenopus oocytes

Interferon-α (IFNα) affects the opioid system. However, the direct action of IFNα on cloned opioid receptors remains unknown. Taking advantage of the functional coupling of cloned opioid receptors to G protein-activated inwardly rectifying K⁺ (GIRK) channels in a Xenopus oocyte expression system, we investigated the effects of recombinant IFNα on cloned μ-, δ- and κ-opioid receptors. In oocytes co-injected with mRNAs for either the δ- or κ-opioid receptor and for GIRK channel subunits, IFNα at high concentrations induced small GIRK currents that were abolished by naloxone, an opioid-receptor antagonist, compared with the control responses to each selective opioid agonist. Additionally, IFNα induced no significant current response in oocytes injected with mRNA(s) for either opioid receptor alone or GIRK channels. In oocytes expressing the μ-opioid receptor and GIRK channels, IFNα had little or no effect. Moreover, in oocytes expressing each opioid receptor and GIRK channels, GIRK current responses to each selective opioid agonist were not affected by the presence of IFNα, indicating no significant antagonism of IFNα toward the opioid receptors. Furthermore, IFNα had little or no effect on the μ/δ-, δ/κ- or μ/κ-opioid receptors expressed together with GIRK channels in oocytes. Our results suggest that IFNα weakly activates the δ and κ-opioid receptors. The direct activation of the δ- and κ-opioid receptors by IFNα may partly contribute to some of the IFNα effects under its high-dose medication.     

Morphine diminishes the constancy of spontaneous uterine contractions, antagonizes the positive inotropic effects of prostaglandin E2, but not of prostaglandin F2 alpha and inhibits prostaglandin E and F outputs from the uterus of ovariectomized rats

The effects of morphine on the constancy of spontaneous contractions (isometric developed tension = IDT and contractile frequency = CF), in uterine strips isolated from ovariectomized rats and the influence of naloxone, were explored. The inotropic responses to added prostaglandins (PGs) E2 and F2 alpha and the influences of morphine and of morphine in the presence of naloxone on PG actions, were also determined. Moreover, the synthesis and outputs of PGs E and F from uteri and the effects of morphine alone and of morphine plus naloxone, were studied. Morphine (10(-6) M) significantly depressed uterine constancy of IDT during the first hours following delivery, but its action on CF did not differ from controls. Naloxone, neither at 10(-8) M nor at 10(-6) M, altered the negative inotropic influence of morphine on IDT. Exogenous PGs E2 and F2 alpha, stimulated uterine inotropism in a concentration-dependent fashion. Morphine altered dose-response curves for exogenous PGE2, evoking a parallel surmountable shift to the right, but did not affect the inotropic action of added PGF2 alpha. This antagonistic effect of the opioid was not altered by preincubation with naloxone. Basal synthesis and outputs of PGs E and F in uteri from ovariectomized rats were significantly depressed by morphine (10(-6) M) but not altered by incubating tissues with morphine in presence of naloxone. Results are discussed in terms of a presumptive dual action of morphine on uterine motility, i.e., antagonizing PGE2 receptors and inhibiting the synthesis of some PGs by the uterus. These influences of morphine do not appear to be subserved by the activation of mu opioid receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prejunctional effects of opioids in the perfused mesentery of the rat and rabbit: interactions with alpha 2-adrenoceptors.

Prejunctional effects of opioids were examined in the perfused mesentery of two species: the rat and rabbit. Use of agonists selective for subtypes of mu, delta, and kappa opioid receptors produced no effect on contractile responses to adrenergic nerve stimulation in the rat perfused mesentery, except for small effects of the kappa agonist EKC, which may be non specific. In contrast, mu, delta and kappa receptors appear to be present in the rabbit. The mu selective agonist, DAMGO, kappa agonist, ethylketocyclazocine, and delta agonists, DPDPE and [Leu5]-enkephalin, all produced significant inhibition of contractile responses to transmural nerve stimulation. The inhibitory effect was greatest for ethylketocyclazocine. To test the possibility that prejunctional activation of alpha 2 adrenoceptors with endogenous norepinephrine might decrease the activity of prejunctional opioid receptors in the rabbit, inhibitory effects of delta and kappa selective agonists were tested in the presence of 10(-7) M yohimbine. Inhibitory responses of the kappa selective agonist ethylketocyclazocine were enhanced, while that of delta selective agonists [Leu5]-enkephalin and DPDPE remained unchanged when yohimbine was present. Thus, the effects of opioids vary and depend on the tissue and receptor subtypes they act upon. Furthermore, the enhanced inhibitory effect of opioid receptor activation in the presence of yohimbine is not found for all opioid receptors.     

The putative somatostatin antagonist cyclo-somatostatin has opioid agonist effects in gastrointestinal preparations

AimsSpecificity of receptor antagonists used is crucial for clarifying physiological/pathophysiological roles of the respective endogenous agonist. We studied the effects (somatostatin antagonist and possibly other actions) of cyclo-somatostatin (CSST), a putative somatostatin receptor antagonist on the guinea-pig small intestine, a preparation where somatostatin causes inhibition of nerve-mediated contractions.Main methodsIn isolated organ experiments, half-maximal cholinergic “twitch” contractions of the guinea-pig small intestine were evoked or tonic contractions of the rat stomach fundus strip (in the presence of physostigmine) were elicited by electrical field stimulation. The effects of somatostatin (somatostatin-14), CSST, naloxone, as well as of direct smooth muscle stimulants were examined.Key findingsSomatostatin (10 nM–1 μM) caused transient inhibition of the twitch contraction, in a naloxone-insensitive manner. Surprisingly, CSST (0.3–1 μM) also inhibited twitch contractions (more than 50% reduction at 1 μM). This effect was prevented by the opioid receptor antagonist naloxone. Responses to acetylcholine or histamine were not or only minimally inhibited by CSST (up to 3 μM). CSST (0.3 μM in the absence or 1–10 μM in the presence of naloxone) failed to inhibit the effect of somatostatin. The SST₂ receptor antagonist CYN-154806 (3 μM) attenuated the effect of somatostatin and failed to evoke naloxone-sensitive inhibition of the twitch response. The naloxone-sensitive inhibitory effect of CSST on cholinergic contractions was also confirmed in the rat stomach fundus preparation.SignificanceCyclo-somatostatin exerts opioid agonist activity in the two preparations tested, while it does not behave as a somatostatin-receptor antagonist in the guinea-pig intestine.     

Tramadol reduces the 5-HTP-induced head-twitch response in mice via the activation of mu and kappa opioid receptors

Tramadol, an atypical opioid analgesic, stimulates both opiatergic and serotonergic systems. Here we have investigated the effect of tramadol in mice on 5-hydroxyptrytophan (5-HTP)-induced head twitch response (HTR), which is an animal model for the activation of the CNS 5-HT(2A) receptors in mice. Tramadol attenuated 5-HTP-induced HTR in a dose-dependent manner as morphine. Furthermore, the nonselective opioid receptor antagonists, naloxone and diprenorphine (M5050), reversed the effect of tramadol on 5-HTP-induced HTR dose-dependently. Interestingly, in contrast to the selective delta opioid receptor antagonist NTI, beta-FNA, a selective mu receptor antagonist, and nor-BNI, a selective kappa opioid receptor antagonist, antagonized the attenuation of 5-HTP-induced HTR by tramadol. In conclusion, administration of tramadol systemically inhibits 5-HTP-induced HTR in mice by activating opiatergic system in the CNS. Our findings show that mu and kappa opioid receptors, but not delta opioid receptor, play an important role in the regulation of serotonergic function in the CNS.     

Inhibitory effect of intracerebroventricularly-administered [D-Arg(2), beta-Ala(4)]-dermorphin (1-4) on gastrointestinal transit

The inhibitory effect of intracerebroventricularly-administered [D-Arg(2), beta-Ala(4)]-dermorphin (1-4) (TAPA), a highly selective mu(1)-opioid receptor agonist, on mouse gastrointestinal transit was compared with that of morphine and [D-Ala(2), N-methyl-Phe(4), Gly(5)-ol]-enkephalin (DAMGO). When administered intracerebroventricularly 5 min before the oral injection of charcoal meal, TAPA (10-100 pmol), morphine (0.25-4 nmol), and DAMGO (20-80 pmol) dose-dependently inhibited gastrointestinal transit of charcoal. The inhibitory effect of each mu-opioid receptor agonist was completely antagonized by naloxone, a nonselective opioid receptor antagonist. The inhibitory effects of morphine and DAMGO were significantly antagonized by both beta-funaltrexamine, a selective mu-opioid receptor antagonist, and naloxonazine, a selective mu(1)-opioid receptor antagonist. In contrast, the inhibitory effect of TAPA was not affected at all by beta-funaltrexamine, naloxonazine, nor-binaltorphimine (a selective kappa-opioid receptor antagonist), or naltrindole (a selective delta-opioid receptor antagonist). These results suggest that the inhibitory effect of TAPA on gastrointestinal transit may be mediated through an opioid receptor mechanism different from that of morphine and DAMGO.     

Different effects of methionine-enkephalin on paw edema in two inbred rat strains

The effect of intraplantarly (i.pl.)-injected methionine-enkephalin (ME) on Concanavalin A (Con A)-induced paw edema in Dark Agouti (DA) and Albino Oxford (AO) rats was investigated. ME suppressed edema in DA rats, which was antagonized with naloxone (non-selective opioid receptor antagonist) and naltrindole (delta opioid receptors antagonist). Potentiating effect of ME in AO rats was blocked by naloxone, nor-binaltorphimine (kappa opioid receptors antagonist) and beta-funaltrexamine (mu opioid receptors antagonist). Dexamethasone suppressed edema in both rat strains. These findings suggest that strain-dependent differences in the effects of ME on inflammation in DA and AO rats could be related to diversity in opioid receptors expression in these strains.     

Inhibition of cholinergic neurotransmission in human airways by opioids.

Opioids reduce the cholinergic responses to electrical field stimulation (EFS) in guinea pig and canine airways by a prejunctional effect. We determined whether a similar effect operates in human airways in vitro. [D-Ala2-NMePhe4-Gly-ol5]enkephalin (DAMGO) (10(-8)-10(-6) M), a selective mu-opioid receptor agonist, inhibited the response to EFS in a dose- and frequency-dependent manner. DAMGO (10(-6) M) produced 86% inhibition at 0.5 Hz and 38% inhibition at 4 Hz, but at 32 Hz there was no significant inhibition. Another selective mu-opioid receptor agonist H-Tyr-D-Arg-Gly-Phe(4-NO2)-Pro-NH2 diacetate (BW 443C) also inhibited responses to EFS, producing 57.7% inhibition at 4 Hz at a concentration of 10(-6) M. The inhibitory effect on EFS was blocked by the opioid receptor antagonist naloxone (10(-5) M), indicating that opioid receptors are involved. DAMGO (10(-6) M) had no effect on the contractile response to exogenous acetylcholine, indicating a prejunctional effect. We conclude that mu-opioid agonists inhibit cholinergic neurotransmission in human airways in vitro, and this could have therapeutic potential in the treatment of airway disease.     

Pharmacological effects of naltriben as a ligand for opioid mu and kappa receptors in rat cerebral cortex

Naltriben (NTB) has been used to differentiate the subtypes of delta opioid receptors, delta1 and delta2. However, there is considerable evidence suggesting that NTB may act on other types of opioid receptors too. We examined the effects of NTB on the specific binding of radiolabeled ligands for opioid mu and kappa2 receptors, and the effects on the release of [3H]norepinephrine ([3H]NE) in rat cerebral cortex slices. NTB displaced the specific binding of [3H]DAMGO with Ki value of 19.79 +/- 1.12 nM in rat cortex membranes. Specific binding of [3H]diprenorphine ([3H]DIP) was inhibited by NTB with Ki value of 82.75 +/- 6.32 nM in the presence of DAMGO and DPDPE. High K+ (15 mM)-stimulated release of [3H]NE was attenuated by DAMGO in rat cerebral cortex slices. NTB (30 nM) shifted the dose-response curve of DAMGO to the right and attenuated the maximal effect. In the meantime, NTB inhibited high K+-stimulated [3H]NE release at concentrations above 100 nM. The inhibitory effect of NTB was not attenuated by CTAP (10 nM) and naloxone (3 nM) but by higher concentration of naloxone (30 nM), nor-BNI (300 nM) and bremazocine (3 nM). These results indicate that NTB, depending on the dosage, could acts not only as an antagonist at delta but also as a noncompetitive antagonist for mu receptors, and as an agonist for kappa2 receptors in rat cerebral cortex.     

Morphine reciprocally regulates IL-10 and IL-12 production by monocyte-derived human dendritic cells and enhances T cell activation

We evaluated the effect of morphine on human dendritic cells (DCs). Interestingly, immature DCs were found to express all 3 (mu, kappa, delta) opioid receptors on the cell surface. Chronic morphine treatment (10(-8) to 10(-12) M) during the development of DCs from monocytes augmented LPS-induced upregulation of HLA-DR, CD86, CD80, and CD83 and increased the T cell stimulatory capacity of DCs, which could be inhibited by naloxone, an opioid receptor antagonist. The change in surface phenotype was paralleled by a p38 MAPK-dependent decrease in IL-10 and increase in IL-12 secretion. Our data indicate that morphine exerts an immunostimulatory effect by modulating LPS-induced DC maturation.     

Role of kappa-opioid receptors in the regulation of cardiac resistance to arrhythmogenic effects of ischemia and reperfusion

It was found that pretreatment of rats with selective agonist of kappa1-opioid receptors (OR) (-)--U--50.488 decreased the incidence of ischemic (10 min) and reperfusion (10 min) ventricular arrhythmias. The selective kappa2-OR agonist GR-89696 had no effect on the incidence of ventricular arrhythmias during a 10-min coronary artery occlusion and following reperfusion in anesthetized rats. The effect of (-)--U-50.488 was abolished by the selective kappa1-OR antagonist of non-binaltorphimine and the non-selective peripheral OR antagonist naloxone methiodide. Perfusion of isolated rat heart with (-)--U-50.488 did not affect arrhythmias during ischemia and reperfusion. The authors suggest that stimulation of kappa1-opioid receptors located outside the central nervous system increases heart resistance against arrhythmogenic action of ischemia/reperfusion, antiarrhythmic action of (-)--U-50.488 being mediated through extracardiac opioid receptors.     

The cardiovascular responses to mu opioid agonist and antagonist in conscious normal and obese rats

Beta-endorphin decreases blood pressure in normal rats but increases blood pressure in obese rats. Since beta-endorphins can bind both mu opioid and kappa-opioid receptors we investigated the effect of a mu specific receptor agonist, D-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (DAMGO) and a mu specific antagonist, D-Phe-Cys-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) on cardiovascular responses in conscious control and obese rats. Rats were also implanted with telemetry transmitters and intracerebroventricular (ICV) cannulas for recording and peptide administration. The mu agonist, DAMGO, increased blood pressure (BP) in control rats. DAMGO also increased BP in obese rats but only at high concentrations. The heart rate responses paralleled the MAP responses. CTAP, the mu antagonist, paradoxically increased the MAP in both control and obese rats. The responsiveness to the mu agonist and antagonist was greater in controls. In other animals the brains were excised and the ventral medial hypothalamic area removed and mu receptor expression determined using PCR. The expression of mu opioid receptors was increased in obese rats. We conclude that the mu opioids can stimulate cardiovascular responses, but the excitatory responsiveness was not increased in conscious obese rats.     

The dynamic relationship between mu and kappa opioid receptors in body temperature regulation

Previous studies demonstrated that intracerebroventricular (icv) injection of a kappa opioid receptor agonist decreased, and a mu agonist increased, body temperature (Tb) in rats. A dose-response study with the selective kappa antagonist nor-binaltorphimine (nor-BNI) showed that a low dose (1.25 nmol, icv) alone had no effect, although a high dose (25 nmol, icv) increased Tb. It was hypothesized that the hyperthermia induced by nor-BNI was the result of the antagonist blocking the kappa opioid receptor and releasing its inhibition of mu opioid receptor activity. To determine whether the Tb increase caused by nor-BNI was a mu receptor-mediated effect, we administered the selective mu antagonist CTAP (1.25 nmol, icv) 15 min after nor-BNI (25 nmol, icv) and measured rectal Tb in unrestrained rats. CTAP significantly antagonized the Tb increase induced by icv injection of nor-BNI. Injection of 5 or 10 nmol of CTAP alone significantly decreased the Tb, and 1.25 nmol of nor-BNI blocked that effect, indicating that the CTAP-induced hypothermia was kappa-mediated. The findings strongly suggest that mu antagonists, in blocking the basal hyperthermia mediated by mu receptors, can unmask the endogenous kappa receptor-mediated hypothermia, and that there is a tonic balance between mu and kappa opioid receptors that serves as a homeostatic mechanism for maintaining Tb.     

Differential antagonism of mu agonists by U50,488H in the rat

Agonist, and antagonist effects of the proposed kappa opioid agonist, U50,488H (U50) have been studied in an experimental model of seizure activity (flurothyl-induced seizure threshold) (ST) and in the central modulation of spontaneous, volume-induced micturition contractions (bladder motility) (BM) in rats. Intracerebroventricular (i.c.v.) administration of U50 (at the doses tested) did not produce any agonist effect in either ST or in BM. In contrast, i.c.v. administration of [D-Ala2, NMPhe4, Gly-ol]enkephalin (DAGO) or etorphine, agonists with activity at mu opioid receptors, produced an elevation of ST and inhibition of BM. The elevation in ST produced by etorphine (0.004 nmol) was prevented by prior treatment with U50. In contrast, the approximately equieffective elevation in ST resulting from DAGO was not affected by U50 pretreatment. Similarly, pretreatment of rats with U50 antagonized the approximately equieffective BM effects of etorphine, but not those of DAGO. As both DAGO and etorphine are thought to exert their effects via the opiate mu receptor, the results may be consistent with the view that subpopulations of mu receptors exist within the central nervous system; these sites may be differentially associated with the kappa receptor.     

Histamine H3 receptors in the guinea pig ileum: evidence for a postjunctional location.

The effect of the selective histamine H3 receptor agonists (R)alpha-methylhistamine, (R)MHA and immepip (IMM) on intestinal smooth muscle contractility was investigated on isolated cells from the longitudinal muscle of the guinea pig ileum. (R)MHA (10(-13)-10(-8) M) and IMM (10(-13)-10(-8) M) did not significantly modify the basal length of intestinal cells; in contrast both agonists (10(-15)-10(-11) M) prevented the contraction produced by acetylcholine (10(-7) M). The (S)-isomer of alpha-methylhistamine, (S)MHA, was inactive both on basal contractility and on acetylcholine-induced contractions. The relaxant effect of (R)MHA was not modified by famotidine (10(-7) M), but totally prevented by the selective H3 receptor antagonist clobenpropit (10(-8) M), which per se did not modify either basal contractility or the contractile response to acetylcholine. These data indicate that inhibitory histamine H3 receptors are present on smooth muscle cells of the guinea pig ileum and can be activated by very low concentrations of selective agonists. It is not clear, however, whether they can have a functional importance in the regulation of intestinal contractility in an intact system.     

Agonist-Selective Protection of the Opioid Receptor-Coupled G Proteins from Inactivation by 5''-p-fluorosulphonylbenzoyl Guanosine

The guanine nucleotide analogue, 5'-p-fluorosulphonylbenzoyl guanosine (FSBG), can react covalently with GTP-binding proteins (G proteins). In rat brain membranes, FSBG causes a time-dependent loss of beta,gamma-imido[8-3H]guanosine 5'-triphosphate binding sites. Using 1 mM FSBG, the guanyl nucleotide modulation of opioid agonist binding is abolished, whereas the guanyl nucleotide sensitivity of neurotensin binding is retained. The action of FSBG can be prevented by the presence of opioid agonists, but not the antagonist naloxone. Iodoacetamide treatment of membranes in the presence of agonist, but not antagonist, can attenuate the action of FSBG in blocking guanyl nucleotide modulation of opioid agonist binding. These results suggest that FSBG covalently modifies essential thiol groups, whose exposure to the reagent is modified by agonist occupancy of the receptor, on a species of G protein linked to opioid receptors, but not on a species of G protein linked to neurotensin receptors. Thus, FSBG may have selectivity for the forms of Gi or Go, proteins associated with opioid receptors.