delta Opioid receptor subtypes activate inositol-signaling pathways in the production of antinociception

To analyze the selectivity of delta receptor subtypes to regulate different classes of G proteins, the expression of the alpha-subunits of Gi2, Gi3, Go1, Go2, Gq and G11 transducer proteins was reduced by administration of oligodeoxynucleotides (ODNs) complementary to sequences in their respective mRNAs. Mice receiving antisense ODNs to Gi2 alpha, Gi3 alpha, Go2 alpha and G11 alpha subunits showed an impaired antinociceptive response to all the delta agonists evaluated. An ODN to Go1 alpha specifically blocked the antinociceptive effect of the agonist of delta-1 receptors, [D-Pen2,5]enkephalin (DPDPE), without altering the activity of [D-Ala2]deltorphin II or [D-Ser2]-Leu-enkephalin-Thr (DSLET). In mice treated with an ODN to Gq alpha, the effects of the agonists of delta-2-opioid receptors were reduced, but not those of DPDPE. Thus, Go1 proteins are selectively linked to delta-1-mediated analgesia, and Gq proteins are related to delta-2-evoked antinociception. After impairing the synthesis of Go1 alpha subunits, DPDPE exhibited an antagonistic activity on the antinociception produced by [D-Ala2]deltorphin II. After treatment with ODNs complementary to sequences in Gq alpha or PLC-beta 1 mRNAs, the analgesic capacity of [D-Ala2]deltorphin II was diminished. However, the delta-2-agonist did not alter the antinociceptive activity of DPDPE. An ODN complementary to nucleotides 7 to 26 of the murine delta receptor reduced the analgesic potency of [D-Ala2]deltorphin II, but not that observed for DPDPE. In these mice, [D-Ala2]deltorphin II did not antagonize the effect of DPDPE. These results suggest the existence of different molecular forms of the delta opioid receptor, and the involvement of inositol-signaling pathways in the supraspinal antinociceptive effects of delta agonists.     

The delta2-opioid receptor subtype stimulates phosphoinositide metabolism in mouse periaqueductal gray matter

The delta(delta)-opioid agonists [D-Pen2,5]enkephalin (DPDPE) and [D-Ala2]deltorphin II increased the formation of inositol phosphates (IPs) in mice periaqueductal gray matter (PAG) slices pre-labeled with myo-[3H]inositol. Both delta-agonists caused an increase in IP accumulation in a dose-dependent manner (1-100 microM) and which was pertussis toxin (0.5 microg/mouse, i.c.v.) sensitive. This effect was blocked by the delta-antagonist ICI-174.864 (10 microM). The presence of subtypes of the delta-opioid receptor (delta1 and delta2) in PAG has been suggested by pharmacological studies. In this brain structure, naltrindrole 5'-isothiocyanate (5'-NTII), but not 7-benzylidenenaltrexone (BNTX), antagonized the effects of DPDPE and [D-Ala2]deltorphin II, suggesting the involvement of a population of delta receptors sensitive to the delta2-antagonist NT II on this effect. To further investigate the participation of delta-receptor subtypes in the stimulation of IPs formation, mice were injected with antisense oligodeoxynucleotides (ODNs) directed to nucleotides 7-26 or 2946 of the cloned delta-receptor mRNA, and PAG slices from these animals were used in in vitro assays. The results demonstrate that the reported increase of phosphoinositide (PI) hydrolysis depends on the agonist activation of the delta2-opioid receptor subtype in the PAG.     

Selective inhibition of [D-Ala2, Glu4]deltorphin antinociception by supraspinal, but not spinal, administration of an antisense oligodeoxynucleotide to an opioid delta receptor

Evidence in vivo has suggested the existence of subtypes of the delta opioid receptor (DOR), which have been termed delta 1 and delta 2. These proposed DOR subtypes are thought to be activated by [D-Pen2, D-Pen5]enkephalin (DPDPE, delta 1) and [D-Ala2, Glu4]deltorphin (delta 2). Recent work in which an antisense oligodeoxynucleotide (oligo) to a cloned DOR was administered by the intrathecal (i.th.) route has demonstrated a reduction in the antinociceptive actions of both i.th. DPDPE and [D-Ala2, Glu4]deltorphin, but not of [D-Ala2, NMPhe4, Gly-ol]enkephalin (DAMGO, mu agonist) in mice. The present investigation has extended these observations by administering the same DOR antisense oligo sequence by the intracerebroventricular (i.c.v.) route and evaluating the antinociceptive actions of i.c.v. agonists selective for delta, mu and kappa receptors. I.th. treatment with DOR antisense oligo, but not mismatch oligo, significantly inhibited the antinociceptive actions of both i.th. DPDPE and [D-Ala2, Glu4]deltorphin but not of i.th. DAMGO or U69,593 (kappa agonist), confirming previous data. In contrast, i.c.v. DOR antisense oligo, but not mismatch oligo, selectively inhibited the antinociceptive response to i.c.v. [D-Ala2, Glu4]deltorphin without altering the antinociceptive actions of i.c.v. DPDPE, DAMGO or U69,593. The data suggest that the cloned DOR corresponds to that pharmacologically classified as delta 2 and further, suggest that this delta receptor subtype may play a major role in eliciting spinal delta-mediated antinociception.     

Treatment with antisense oligodeoxynucleotide to the opioid delta receptor selectively inhibits delta 2-agonist antinociception

Using approaches emphasizing differential antagonism of receptor selective agonists and cross-tolerance paradigms, evidence in vivo has suggested the existence of subtypes of opioid delta receptors, which have been termed delta 1 and delta 2. Recent work has elucidated the structure of an opioid delta receptor. The present investigation attempted to continue to test the hypothesis of subtypes of delta receptors and to correlate the cloned delta receptor with the existing pharmacological classification. Synthetic oligodeoxynucleotides (oligos) complementary to the 5' end of the cloned delta receptor coding region (antisense) or its corresponding sequence (sense) were given by intracerebroventricular (i.c.v.) administration to mice, twice-daily for 3 days and antinociceptive responses to selective agonists at putative delta 1 and delta 2 receptors were subsequently determined. Treatment with antisense, but not sense, oligo significantly inhibited the response to [D-Ala2,Glu4]deltorphin (delta 2 agonist), but not to [D-Pen2,D-Pen5]enkephalin (DPDPE, delta 1 agonist). Further, subsequent administration of DPDPE elicited a full antinociceptive response in the same antisense oligo treated mice which did not show a significant response to [D-Ala2,Glu4]deltorphin while antisense oligo treated mice which responded to DPDPE did not show antinociception when tested subsequently with [D-Ala2,Glu4]deltorphin. The data suggest that the cloned delta receptor corresponds to that pharmacologically classified as delta 2 and continue to support the concept of subtypes of opioid delta receptors.     

Competitive and non-competitive NMDA antagonists block the development of antinociceptive tolerance to morphine, but not to selective mu or delta opioid agonists in mice

N-Methyl-D-aspartate (NMDA) receptor antagonists have been shown to block the development of antinociceptive tolerance to morphine. Assessment of the effects of NMDA antagonists on development of antinociceptive tolerance to selective opioid mu (mu) and delta (delta) agonists, however, has not been reported. In these experiments, selective mu and delta receptor agonists, and morphine, were repeatedly administered to mice either supraspinally (i.c.v.) or systemically (s.c.), alone or after pretreatment with systemic NMDA antagonists. Antinociception was evaluated using a warm-water tail-flick test. Repeated i.c.v. injections of mu agonists including morphine, fentanyl, [D-Ala2, NMePhe4, Gly-ol]enkephalin (DAMGO) and Tyr-Pro-NMePhe-D-Pro-NH2 (PL017) or [D-Ala2, Glu4]deltorphin, a delta agonist, or s.c. injections of morphine or fentanyl, produced antinociceptive tolerance as shown by a significant rightward displacement of the agonist dose-response curves compared to controls. Single injections or repeated administration of MK801 (a non-competitive NMDA antagonist) or LY235959 (a competitive NMDA antagonist) at the doses employed in this study did not produce behavioral toxicity, antinociception or alter the acute antinociceptive effects of the tested opioid agonists. Consistent with previous reports, pretreatment with MK801 or LY235959 (30 min prior to agonist administration throughout the tolerance regimen) prevented the development of antinociceptive tolerance to i.c.v. or s.c. morphine. Neither NMDA antagonist, however, affected the development of antinociceptive tolerance to i.c.v. fentanyl, DAMGO, or [D-Ala2, Glu4]deltorphin. Additionally, MK801 pretreatment did not affect the development of antinociceptive tolerance to i.c.v. PL017 or to s.c. fentanyl. Further, MK801 pretreatment also did not affect the development of tolerance to the antinociception resulting from a cold-water swim-stress episode, previously shown to be a delta-opioid mediated effect. These data lead to the suggestion that the mechanisms of tolerance to receptor selective mu and delta opioids may be regulated differently from those associated with morphine. Additionally, these findings emphasize that conclusions reached with studies employing morphine cannot always be extended to 'opiates' in general.     

Antinociceptive effect of dihydroetorphine in diabetic mice

The antinociceptive potency of dihydroetorphine in diabetic mice was examined. Subcutaneous administration of dihydroetorphine produced a dose-dependent antinociception in both non-diabetic and diabetic mice. The antinociceptive potency of s.c. dihydroetorphine was less in diabetic mice than in non-diabetic mice. The antinociception induced by i.c.v. dihydroetorphine (0.02 microgram) was also significantly less in diabetic mice than in non-diabetic mice. The antinociceptive effects of dihydroetorphine (10 micrograms/kg i.p.) in both diabetic and non-diabetic mice were significantly antagonized by s.c. administration of beta-funaltrexamine, a selective mu-opioid receptor antagonist. Furthermore, the antinociceptive effect of dihydroetorphine (10 micrograms/kg i.p.) in non-diabetic mice, but not in diabetic mice, was also significantly antagonized by naloxonazine, a selective mu 1-opioid receptor antagonist. The time course and the potency of the antinociceptive effect of dihydroetorphine (10 micrograms/kg i.p.) in diabetic mice were similar to those in naloxonazine-treated non-diabetic mice. Naltrindole, a selective delta-opioid receptor antagonist, or nor-binaltorphimine, a selective kappa-opioid receptor antagonist, had no significant effect on the antinociceptive effect of dihydroetorphine (10 micrograms/kg i.p.) in both diabetic and non-diabetic mice. These results suggest that dihydroetorphine produces an antinociceptive effect through the activation of both mu 1- and mu 2-opioid receptors in mice. Furthermore, the reduction in dihydroetorphine-induced antinociception in diabetic mice, as compared with non-diabetic mice, may be due to the hyporesponsive to supraspinal mu 1-opioid receptor-mediated antinociception in diabetic mice.     

Up-regulation of brain N-methyl-D-aspartate receptors following multiple intracerebroventricular injections of [D-Pen2, D-Pen5] enkephalin and [D-Ala2, Glu4]deltorphin II in mice

The effects of chronic administration of [D-Pen2, D-Pen5]enkephalin and [D-Ala2, Glu4]deltorphin II, the selective agonists of the delta 1- and delta 2-opioid receptors, on the binding of [3H]MK-801, a noncompetitive antagonist of the N-methyl-D-aspartate receptor, were determined in several brain regions of the mouse. Male Swiss-Webster mice were injected intracerebroventricularly (i.c.v.) with [D-Pen2, D-Pen5]enkephalin or [D-Ala2, Glu4]deltorphin II (20 micrograms/mouse) twice a day for 4 days. Vehicle injected mice served as controls. Previously we have shown that the above treatment results in the development of tolerance to their analgesic activity. The binding of [3H]MK-801 was determined in brain regions (cortex, midbrain, pons and medulla, hippocampus, striatum, hypothalamus and amygdala). At 5 nM-concentration, the binding of [3H]MK-801 was increased in cerebral cortex, hippocampus, and pons and medulla of [D-Pen2, D-Pen5]enkephalin treated mice. In [D-Ala2, Glu4]deltorphin II treated mice, the binding of [3H]MK-801 was increased in cerebral cortex and hippocampus. The changes in the binding were due to increases in the Bmax value of [3H]MK-801. It is concluded that tolerance to delta 1- and delta 2-opioid receptor agonists is associated with up-regulation of brain N-methyl-D-aspartate receptors, however, some brain areas affected differ with the two treatments. The results are consistent with the recent observation from this laboratory that N-methyl-D-aspartate receptors antagonists block tolerance to the analgesic action of delta 1- and delta 2-opioid receptor agonists.     

Role of intracellular calcium in modification of mu and delta opioid receptor-mediated antinociception by diabetes in mice

We examined the effects of calcium modulators on mu and delta opioid receptor agonist-induced antinociception in both diabetic and nondiabetic mice. In nondiabetic mice, intracerebroventricular (i.c. v.) pretreatment with calcium and thapsigargin, which increase intracellular calcium, reduced [D-Ala2,N-MePhe4,Gly-ol5]-enkephalin (DAMGO)-induced antinociception by shifting its dose-response curve to the right. However, in diabetic mice i.c.v. pretreatment with calcium and thapsigargin did not affect DAMGO-induced antinociception. In contrast i.c.v. administration of agents that decrease intracellular calcium, such as EGTA and ryanodine, enhanced DAMGO-induced antinociception in both diabetic and nondiabetic mice. In contrast with DAMGO i.c.v. pretreatment with calcium and thapsigargin enhanced (-)-TAN67-induced antinociception in nondiabetic mice by shifting its dose-response curve to the left. However, (-)-TAN67-induced antinociception in diabetic mice was not affected by pretreatment with calcium or thapsigargin. Moreover i.c. v. pretreatment with EGTA, but not with ryanodine, reduced (-)-TAN67-induced antinociception in nondiabetic mice. In diabetic mice i.c.v. pretreatment with both EGTA and ryanodine reduced (-)-TAN67-induced antinociception. These results suggest that cytosolic calcium has different effects on mu and delta opioid receptor agonist-induced antinociception. Further, these results suggest that the modification of mu and delta opioid receptor agonist-induced antinociception by diabetes in mice may be due to increased levels of intracellular calcium.     

Analysis of the role of delta opioid receptors in gastroprotection in the rat

The effect of delta opioid agonists - [D-Ala2, D-Leu5]-enkephalin (DADLE), [D-Pen2, D-Pen5]-enkephalin (DPDPE) and deltorphin II - on acidified ethanol induced gastric mucosal lesions was studied in the rat compared with that of morphine. It was found that DADLE, DPDPE, deltorphin II and morphine exerted a dose-dependent inhibition on the mucosal lesions injected subcutaneously, their ID50 values were 0.037, 1.8, 3.5 and 0.35 micromoles/kg, respectively. Naltrindole (10 mg/kg sc.), the selective delta opioid receptor antagonist, inhibited the gastroprotective effect of DADLE, DPDPE and deltorphin II, but it failed to antagonise the effect of morphine. Our results suggest that 1. delta receptors are involved in opioid-mediated gastroprotection, 2. ethanol-induced gastric mucosal damage in the rat may be a quick, simple in vivo model for screening opioid delta receptor agonists and antagonists in the periphery.     

Effects of mu and delta opioid agonists and antagonists on affective vocal and reflexive pain responses during social stress in rats

The present experiments evaluated the influence of intraventricular mu and delta opioid receptors on affective vocal and reflexive responses to aversive stimuli in socially inexperienced, as well as defensive and submissive responses in defeated, adult male Long-Evans rats. Defeat stress consisted of: (1) an aggressive confrontation in which the experimental intruder rat exhibited escape, defensive and submissive behaviors [i.e., upright, supine postures and ultrasonic vocalizations (USV)], and subsequently, (2) protection from the resident stimulus rat with a wire mesh screen for 10-20 min. Defeat stress was immediately followed by an experimental session with tactile startle (20 psi). The mu opioid receptor agonists morphine (0.1-0.6 microg i.c.v.) and [D-Ala2-N-Me-Phe4-Gly5-ol]-enkephalin (DAMGO; 0.01-0.3 microg i.c.v.), and the delta opioid receptor agonist [D-Pen2,5]-enkephalin (DPDPE; 10-100 microg i.c.v.) dose-dependently decreased startle-induced USV and increased tail-flick latencies in socially inexperienced and defeated rats. Of greater interest, morphine, DAMGO and DPDPE increased the occurrence of the submissive crouch posture, and defeated rats were more sensitive than socially inexperienced rats to the startle-induced USV-suppressive and antinociceptive effects of morphine and DPDPE. The antinociceptive effects of DAMGO were likewise obtained at lower doses in defeated rats. Finally, the USV-suppressive effects of morphine and DAMGO were reversed with the mu receptor antagonist naltrexone (0.1 mg/kg i.p.), but the USV-suppressive effects produced by DPDPE were not reversed with the delta receptor antagonist naltrindole (1 mg/kg i.p.). These results confirm mu, but not delta opioid receptor activation as significant in affective vocal, passive-submissive behavior, as well as reflexive antinociception. Furthermore, similar to previous studies with restraint and electric shock stress, the facilitation of mu opioid effects on vocal responses and antinociception is consistent with the proposal that defeat stress activated endogenous opioid mechanisms.     

Interaction between the mu-agonist dermorphin and the delta-agonist [D-Ala2, Glu4]deltorphin in supraspinal antinociception and delta-opioid receptor binding

1. In rats, the interaction between the mu-opioid agonist dermorphin and the delta-opioid agonist [D-Ala2, Glu4]deltorphin was studied in binding experiments to delta-opioid receptors and in the antinociceptive test to radiant heat. 2. When injected i.c.v., doses of [D-Ala2, Glu4]deltorphin higher than 20 nmol produced antinociception in the rat tail-flick test to radiant heat. Lower doses were inactive. None of the doses tested elicited the maximum achievable response. This partial antinociception was accomplished with an in vivo occupancy of more than 97% of brain delta-opioid receptors and of 17% of mu-opioid receptors. Naloxone (0.1 mg kg-1, s.c.), and naloxonazine (10 mg kg-1, i.v., 24 h before), but not the selective delta-opioid antagonist naltrindole, antagonized the antinociception. 3. In vitro competitive inhibition studies in rat brain membranes showed that [D-Ala2, Glu4]deltorphin displaced [3H]-naltrindole from two delta-binding sites of high and low affinity. The addition of 100 microM Gpp[NH]p produced a three fold increase in the [D-Ala2, Glu4]deltorphin Ki value for both binding sites. The addition of 10 nM dermorphin increased the Ki value of the delta-agonist for the high affinity site five times. When Gpp[NH]p was added to the incubation medium together with 10 nM dermorphin, the high affinity Ki of the delta-agonist increased 15 times. 4. Co-administration into the rat brain ventricles of subanalgesic doses of dermorphin and [D-Ala2, Glu4]deltorphin resulted in synergistic antinociceptive responses. 5. Pretreatment with naloxone or with the non-equilibrium mu-antagonists naloxonazine and beta-funaltrexamine completely abolished the antinociceptive response of the mu-delta agonist combinations. 6. Pretreatment with the delta-opioid antagonists naltrindole and DALCE reduced the antinociceptive response of the dermorphin-[D-Ala2, Glu4]deltorphin combinations to a value near that observed after the mu-agonist alone. At the dosage used, naltrindole occupied more than 98% of brain delta-opioid receptors without affecting mu-opioid-receptors. 7. These data suggest that in the rat tail-flick test to radiant heat, mu- and delta-opioid agonists co-operate positively in evoking an antinociceptive response. Although interactions between different opioid pathways cannot be excluded, in vitro binding results indicate that this co-operative antinociception is probably mediated by co-activation of the delta-opioid receptors at the cellular level by the mu- and delta-agonist.     

Ischemic preconditioning in the intact rat heart is mediated by delta1- but not mu- or kappa-opioid receptors

BACKGROUND: Our laboratory has previously shown that delta-opioid receptors are involved in the cardioprotective effect of ischemic preconditioning in the rat heart. However, this class of receptors consists of two subtypes, delta1, and delta2, and mu- or kappa-opioid receptors may also exist in the heart. Therefore, the purpose of the present study was to test the hypothesis that ischemic preconditioning is mediated through stimulation of one or both delta-opioid receptor subtypes. METHODS AND RESULTS: Anesthetized, open chest, male Wistar rats were assigned to 1 of 14 groups. All animals were subjected to 30 minutes of occlusion and 2 hours of reperfusion. Ischemic preconditioning was elicited by three 5-minute occlusion periods interspersed with 5 minutes of reperfusion. Two doses of 7-benzylidenenaltrexone (BNTX; 1 and 3 mg/kg i.v.), a selective delta1-opioid receptor antagonist, or naltriben (NTB; 1 and 3 mg/kg i.v.), a selective delta2-opioid receptor antagonist, were given before ischemic preconditioning. To test for a role of mu-opioid receptors, rats were pretreated with beta-funaltrexamine (beta-FNA; 15 mg/kg s.c), an irreversible mu-opioid receptor antagonist, 24 hours before ischemic preconditioning or given the mu-opioid receptor agonist D-Ala,2N-Me-Phe,4glycerol5-enkephalin (DAMGO) as three 5-minute infusions (1, 10, and 100 microg/kg per infusion i.v., respectively) interspersed with 5-minute drug-free periods before the prolonged ischemic and reperfusion periods (lowDAMGO, medDAMGO, and hiDAMGO, respectively). The involvement of kappa-opioid receptors was tested by administering one of two doses of nor-binaltorphimine (nor-BNI; 1 and 5 mg/kg i.v.) before ischemic preconditioning. Infarct size (IS) as a percent of the area at risk (AAR) was measured by triphenyltetrazolium stain. Ischemic preconditioning markedly reduced IS/AAR (14+/-4%, P<.05) compared with control (55+/-4%). NTB, beta-FNA, and nor-BNI were unable to block the cardioprotective effect of ischemic preconditioning. In addition, DAMGO had no effect on IS/AAR. However, the high dose of BNTX (3 mg/kg i.v.) significantly attenuated the cardioprotective effect of ischemic preconditioning (39+/-5%; P<.05 versus control and ischemic preconditioning). CONCLUSIONS: These results indicate that delta1-opioid receptors play an important role in the cardioprotective effect of ischemic preconditioning in the rat heart.     

Selective in vivo binding of [3H]naltriben to delta-opioid receptors in mouse brain

Naltriben (NTB) is a selective antagonist for the putative delta2-opioid receptor. We have determined the regional kinetics and pharmacological profile of [3H]naltriben in vivo in mouse brain. After i.v. administration to CD1 mice, [3H]naltriben uptake and retention were high in striatum, cortical regions and olfactory tubercles, and low in superior colliculi and cerebellum. Robust rank order correlation was found between [3H]naltriben uptake in discrete brain regions and prior delta-opioid receptor binding determinations in vitro and in vivo. [3H]Naltriben binding in vivo was saturable, and was blocked by the delta-opioid receptor antagonist naltrindole, but not by the mu-opioid receptor antagonist cyprodime or the K-opioid receptor agonist (trans)-(+/-)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]ben zeneacetamide mesylate (U50,488H). (E)-7-Benzylidenenaltrexone (BNTX), a selective antagonist for the putative delta1-opioid receptor, was 9.6- to 12.9-fold less potent than naltriben as an inhibitor of [3H]naltriben binding. Thus, the sites labeled by [3H]naltriben in vivo may correspond to the delta2-opioid receptor subtype. Such assignment is not definitive, particularly considering the 4-fold higher brain uptake of naltriben as compared to (E)-7-benzylidenenaltrexone. Moreover, the regional distribution of [3H]naltriben in brains from CXB-7/BY (CXBK) mice, a strain that shows supraspinal delta1- but not delta2-opioid receptor agonist effects, was quite similar to that found for CD1 mice.     

The competence-related abilities of women criminal defendants

Delta9-tetrahydrocannabinol (delta9-THC) elicits antinociception in rodents through the central CB1 cannabinoid receptor subtype. In addition. Delta9-THC stimulates the release of dynorphin-related peptides leading to kappa-opioid spinal antinociception. In this work we describe the effect of a mixture of thiorphan (a neutral endopeptidase EC3.4.24.11 inhibitor) and bestatin (an aminopeptidase inhibitor), administered i.c.v., on the antinociceptive effect of peripherally administered delta9-THC in mice. As in the case of morphine or DAMGO ([D-Ala2.N-Me-Phe4,Gly-ol]enkephalin), a mu-selective opioid receptor agonist, the mixture of enkephalin-degrading enzyme inhibitors also enhanced the antinociceptive effect of delta9-THC. This effect was blocked by the CB1 cannabinoid receptor antagonist, SR-141,716-A, as well as by naloxone. The kappa-opioid receptor antagonist nor-binaltorphimine, administered i.t., also antagonized the effect of this combination. Similar results were obtained with the mu-opioid receptor antagonist beta-funaltrexamine after i.c.v. administration. These results demonstrate the involvement of both mu-opioid supraspinal and kappa-opioid spinal receptors in the interaction of both opioid and cannabinoid systems regulating nociception in mice.     

Logistic regression analysis of twin data: estimation of parameters of the multifactorial liability-threshold model

This study with the rat evaluated the contribution of omega-conotoxin GVIA-(omega-CgTx) and verapamil-sensitive Ca2+ channels in behavioural, antinociceptive and thermoregulatory responses to intracerebroventricular (i.c.v.) injection of [D-Ala2,NMePhe4,Gly-ol5]enkephalin (DAMGO), [D-Pen2,D-Pen5]enkephalin (DPDPE) and dynorphin A-(1-17), which are selective agonists for putative mu, delta and kappa-opioid receptors, respectively. The rats treated with omega-CgTx (8-32 pmol i.c.v.) showed transient, dose-dependent shaking behaviour, hyperalgesia and hypothermia which gradually disappeared within 4 h. The behaviour of the rats was normal by 24 h. Histological examination of brain sections showed morphological alterations of neurons in the hippocampus, medial-basal hypothalamus and pyriform cortex. antinociception, catalepsy and thermoregulatory responses elicited by DAMGO (0.4 and 2.0 nmol) were significantly prolonged and potentiated by verapamil (20 pmol i.c.v. 15 min before) or omega-CgTx (8 pmol 24 h before). Antinociception and hypothermia induced by DPDPE were antagonized by verapamil and omega-CgTx, whereas only omega-CgTx prevented the behavioural arousal observed after DPDPE. Similarly, hypothermia induced by dynorphin A-(1-17) (5.0 nmol) and by the kappa-opioid receptor agonist U50,488H (215 nmol) was antagonized by the two Ca2+ channel blockers but only omega-CgTx prevented the barrel rolling and bizarre postures caused by the opioid peptide.     

Modification of morphine-induced place preference by diabetes

The effects of diabetes on morphine-induced place preference in mice were examined. Morphine caused dose-related place preference in both diabetic and non-diabetic mice. This morphine-induced place preference in diabetic mice was greater than that in non-diabetic mice. The morphine (5 mg/kg)-induced place preference in both diabetic and non-diabetic mice was significantly antagonized by pretreatment with beta-funaltrexamine, a selective mu-opioid receptor antagonist, but not with naloxonazine, a selective mu1-opioid receptor antagonist. The morphine (5 mg/kg)-induced place preference in non-diabetic mice was attenuated by pretreatment with either naltriben, a selective delta2-opioid receptor antagonist, or 7-benzylidenenaltrexone. a selective delta1-opioid receptor antagonist. Moreover, the morphine (10 mg/kg)-induced place preference in non-diabetic mice was antagonized by pretreatment with 7-benzylidenenaltrexone (0.7 mg/kg). Although 7-benzylidenenaltrexone had no effect on the place preference induced by 5 mg/kg morphine in diabetic mice, it reduced the place preference induced by 3 mg/kg morphine. Furthermore, the morphine (5 mg/kg)-induced place preference in diabetic mice was significantly antagonized by co-pretreatment with beta-funaltrexamine (10 mg/kg) and 7-benzylidenenaltrexone (0.7 mg/kg). 2-Methyl-4a alpha-(3-hydroxyphenyl)- 1,2,3,4,4a,5,12,12a alpha-octahydroquinolino[2,3,3-g]isoquinoline (TAN-67), a non-peptide delta-opioid receptor agonist, produced place preference in diabetic, but not in non-diabetic mice. These results support the hypothesis that the morphine-induced place preference is mainly mediated through the activation of the mu2-opioid receptor. Furthermore, the enhancement of the morphine-induced place preference in diabetic mice may be due to the up-regulation of delta-opioid receptor-mediated functions.     

Pretreatment with protein kinase C activator phorbol 12,13-dibutyrate attenuates the antinociception induced by mu- but not epsilon-opioid receptor agonist in the mouse

The effects of pretreatment with a protein kinase C activator, phorbol 12,13-dibutyrate, on antinociception induced by i.c.v.-administered mu-opioid receptor agonist (D-Ala2, NMePhe4, Gly(ol)5) enkephalin (DAMGO) or morphine and epsilon-opioid receptor agonist beta-endorphin were studied in male ICR mice. The tail-flick responses were used for antinociceptive tests. I.c.v. pretreatment with phorbol 12,13-dibutyrate (50 pmol) for 30 or 60 but not 10 min attenuated antinociception induced by i.c.v.-administered DAMGO. I.c.v. pretreatment with phorbol 12,13-dibutyrate (10 and 50 pmol) for 60 min caused a dose-dependent attenuation of DAMGO (19.5 pmol)- or morphine (6.0 nmol)-induced antinociception. The dose-response curve for DAMGO-induced antinociception was shifted to the right by 7.3-fold by i.c.v. pretreatment with phorbol 12,13-dibutyrate (50 pmol) for 60 min. However, the i.c.v.-administered beta-endorphin-induced antinociception was not affected by the same pretreatment with phorbol 12,13-dibutyrate. The attenuation of i.c.v.-administered DAMGO- and morphine-induced antinociception by phorbol 12,13-dibutyrate was reversed by concomitant i.c.v. pretreatment with a selective protein kinase C inhibitor calphostin C. These results suggest that activation of protein kinase C by phorbol 12,13-dibutyrate leads to the desensitization of mu-, but not epsilon-opioid receptor-mediated antinociception. These findings also provide additional evidence for differential intracellular modulation on antinociceptive action of mu- and epsilon-opioid receptor agonists.     

In vivo injection of antisense oligodeoxynucleotides to G alpha subunits and supraspinal analgesia evoked by mu and delta opioid agonists

For 5 consecutive days repeated intracerebroventricular (i.c.v.) administration of antisense oligodeoxynucleotides (ODNs) to G alpha subunit mRNAs was used to impair the function of mouse Gi1, Gi2, Gi3 and Gx/z regulatory proteins. Decreases of 20 to 60% on the G alpha-like immunoreactivity could be observed in neural structures of mouse brain, an effect that was not produced by a random-sequence ODN used as a control. The ODN to Gi1 alpha subunits lacked effect on opioid-evoked analgesia. In mice injected with the ODN to Gi2 alpha subunits the antinociceptive activity of all the opioids studied appeared greatly impaired. The ODN to Gi3 alpha subunits reduced the effects of the selective agonists of delta opioid receptors, [D-Pen2,5]-enkephalin and [D-Ala2]deltorphin II. Conversely, the analgesia evoked by opioids binding mu opioid receptors, [D-Ala2, N-MePhe4,Gly-ol5]enkephalin and morphine, appeared consistently and significantly attenuated in mice injected with the ODN to Gx/z alpha. The effect of the neuropeptide beta-endorphine-(1-31) agonist at mu and delta receptors was also reduced by ODNs to Gi3 alpha or Gx/z alpha subunits. l.c.v. injection of antibodies directed to these G alpha subunits antagonized opioid-induced analgesia with a pattern similar to that observed for the ODNs. Thus, the mu and delta opiod receptors regulate different classes of G transducer proteins to mediate the analgesic effect of agonists. The in vivo antisense strategy and the use of specific antibodies to G alpha subunits gave comparable results, indicating that in the neural tissue the mRNAs and the G alpha subunits can be accessed by the corresponding ODNs and IgGs.     

Antinociceptive evaluation of 14 beta-(bromoacetamido)-7,8-dihydro- N(cyclopropylmethyl)-normorphinone in mice

This study evaluated the supraspinal opioid effects of 14 beta-(bromoacetamido)-7,8-dihydro-N(cyclopropylmethyl)-normorphinone+ ++ (N-CPM-H2BAMO) in the mouse acetic acid-induced writhing and tail-flick assays. In the writhing test, N-CPM-H2BAMO produced a time- and dose-dependent antinociception after i.c.v. administration, with a 50% antinociceptive response being obtained with 0.28 (0.19-0.39) nmol when given 10 min before testing. The antinociceptive effect of N-CPM-H2BAMO was antagonized in a dose-dependent manner by the kappa-selective opioid receptor antagonist, nor-binaltorphimine. In the mouse tail-flick assay, N-CPM-H2BAMO failed to produce any antinociception after i.c.v. administration. N-CPM-H2BAMO produced a dose-dependent antagonism of morphine-induced antinociception but not antinociception induced by the delta-opioid receptor agonist [D-Pen2,D-Pen5]enkephalin. Nor-binaltorphimine (0.3 nmol) at dose that completely antagonized N-CPM-H2BAMO-induced antinociception in the writhing assay did not prevent the antagonistic effect of N-CPM-H2BAMO on morphine-induced antinociception. Therefore, these data indicate that N-CPM-H2BAMO produces antinociception by acting at supraspinal kappa-opioid receptors in the writhing assay, and also acts as a mu-opioid receptor antagonist.     

Characterisation of integrons and antibiotic resistance genes in Danish multiresistant Salmonella enterica Typhimurium DT104

This study examined the contribution of spinal delta1 and delta2 opioid receptors to the antinociception produced by microinjection of L-glutamate in either the nucleus raphe magnus (NRM) or the nucleus reticularis gigantocellularis pars alpha (NGCp alpha) of the rat. Intrathecal (i.t.) pretreatment with 1 microg of 7-benzylidinenaltrexone (BNTX), a delta1 opioid receptor antagonist, did not antagonize the increase in tail flick latency (TFL) produced by microinjection of L-glutamate in either the NRM or the NGCp alpha. In contrast, i.t. pretreatment with 3 microg of naltriben (NTB), a delta2 opioid receptor antagonist, completely antagonized the increase in TFL evoked by microinjection of L-glutamate in the NRM, but did not antagonize the increase in TFL evoked from the NGCp alpha. These results suggest that the antinociception produced by activation of these bulbospinal pathways is predominantly mediated by spinal delta2 opioid receptors and that there is little, if any, contribution by spinal delta1 opioid receptors.