Pretreatment with pertussis toxin blocks morphine- but not beta-endorphin-induced antinociception in the mouse

We have previously demonstrated that the antinociception induced by morphine and beta-endorphin given intracerebroventricularly (i.c.v.) is mediated by the stimulation of respective mu- and epsilon-opioid receptors. The effects of i.c.v. pretreatment with pertussis toxin on the antinociception induced by morphine and beta-endorphin given i.c.v. were studied in male ICR mice. Antinociception was assessed by the tail-flick and hot-plate tests. Pretreatment with pertussis toxin (0.5 microgram) given i.c.v. 96 h earlier blocks the antinociception induced by i.c.v. administered morphine in both tail-flick and hot-plate tests. The same pretreatment did not affect the antinociception induced by i.c.v. administered beta-endorphin. Our results indicate that morphine-, but not beta-endorphin-induced antinociception is mediated by pertussis toxin sensitive G-proteins.     

Met]enkephalin in the spinal cord is involved in the antinociception induced by intracerebroventricularly-administered etorphine in the mouse

We have recently reported that the antinociception induced by etorphine given i.c.v. is mediated in part by the stimulation of both mu- and epsilon-opioid receptors and the activation of both monoaminergic and opioidergic descending pain control systems. [Xu J. Y. et al. (1992) J. Pharmac. exp. Ther. 263, 246-252]. Since the opioid epsilon-receptor-mediated antinociception induced by beta-endorphin is mediated by the release of [Met]enkephalin and subsequent stimulation of delta-opioid receptors in the spinal cord, the present studies were designed to determine if beta-endorphin-like action is also involved in etorphine-induced antinociception. The tail-flick test was used to assess the antinociceptive response performed in male ICR mice. Etorphine at doses from 5 to 20 ng given i.c.v. produced a dose-dependent inhibition of the tail-flick response. The inhibition of the tail-flick response induced by etorphine given i.c.v. was antagonized by intrathecal pretreatment for 60 min with antiserum against [Met]enkephalin (10 microg), but not with antiserum against [Leu]enkephalin (10 microg) or dynorphin A (1-13) (10 microg). Desensitization of delta-opioid receptors in the spinal cord by intrathecal pretreatment with [Met]enkephalin (5 microg) for 60 min attenuated i.c.v. administered etorphine-induced tail-flick inhibition. However, intrathecal pretreatment with [Leu]enkephalin (5 microg) or dynorphin A (1-17) (0.1 microg) for 60 min did not attenuate i.c.v. administered etorphine-induced tail-flick inhibition. The results indicate that antinociception induced by etorphine given i.c.v. is mediated in part by the stimulation of the epsilon-opioid receptor at the supraspinal sites and by the release of [Met]enkephalin, which subsequently stimulates delta-opioid receptors in the spinal cord.     

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.     

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.     

Spinal morphine/clonidine antinociceptive synergism is regulated by protein kinase C, but not protein kinase A activity

When coadministered spinally, morphine and clonidine interact synergistically to produce antinociception. The mechanism for the synergism is unknown, but may depend on intracellular messenger systems. Agents that alter the activities of protein kinases alter antinociception produced by opioids, but their effects on clonidine-induced antinociception or the morphine/clonidine interaction are not known. In these studies, mice were pretreated intrathecally with inhibitors or activators of protein kinase C and cyclic AMP-dependent protein kinase (protein kinase A). Antinociceptive responses to intrathecally administered morphine, clonidine and morphine/clonidine combinations were then measured in the radiant heat tail flick test. Inhibition of protein kinase C activity with chelerythrine or calphostin C changed the morphine/clonidine interaction from synergistic to additive. Inhibition of protein kinase A activity with H-89 did not alter the morphine/clonidine interaction, it remained synergistic. Stimulation of protein kinase C activity with phorbol 12,13-dibutyrate attenuated morphine antinociception, but did not alter the synergistic interaction. Increasing spinal cyclic AMP concentrations with either forskolin or rolipram attenuated the antinociception produced by separately administered morphine and clonidine, but had no effect on the morphine/clonidine interaction. These results suggest that protein kinase C activity may regulate the interaction between spinal opioid and alpha-2 receptors, stimulated by morphine and clonidine.     

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.     

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.     

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.     

Repeated administration of opioids alters characteristics of membrane-bound phorbol ester binding in rat brain

Scatchard analysis of saturation binding data indicated that dissociation constant (KD) of [3H]phorbol 12,13-dibutyrate (PDB) binding to the membrane-bound protein kinase C of rat cortex and midbrain was significantly decreased following systemic repeated administration of morphine (mu-opioid receptor agonist) and butorphanol (mu/delta/kappa-mixed opioid receptor agonist). No change in the receptor density (Bmax) of [3H]PDB binding was found following repeated treatment with morphine and butorphanol. On the other hand, neither the Bmax nor KD values in pons/medulla were altered in these rats. These results suggest that systemic repeated opioid treatment, such as morphine and butorphanol leads to an increased affinity for phorbol ester binding to the membrane-bound protein kinase C in rat cortex and midbrain.     

Differences in the influence of AIN-purified and non-purified diets on the development of hypertension in SHR and DOCA-salt hypertensive rats

The effect of nicotine administered supraspinally on antinociception induced by supraspinally administered opioids was examined in ICR mice. The intracerebroventricular (i.c.v.) injection of nicotine alone at doses from 1 to 12 micrograms produced only a minimal inhibition of the tail-flick response. Morphine (0.2 micrograms), beta-endorphin (0.1 microgram), D-Pen2.5-enkephalin (DPDPE; 0.5 microgram), trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl] benzeocetamide (U50, 488H; 6 micrograms) caused only slight inhibition of the tail-flick response. Nicotine dose dependently enhanced inhibition of the tail-flick response induced by i.c.v. administered morphine (0.2 microgram) or beta-endorphin (0.1 microgram). The degree of enhancing effect of nicotine toward beta-endorphin-induced inhibition of the tail-flick response was greater than toward morphine-induced inhibition of the tail-flick response. However, i.c.v. administered nicotine at the same doses was not effective in enhancing the inhibition of the tail-flick response induced by DPDPE (0.5 microgram) or U50, 488H (6 micrograms) administered i.c.v. Our results suggest that stimulation of supraspinal nicotinic receptors may enhance antinociception induced by morphine (a mu-opioid receptor agonist) and beta-endorphin (an epsilon-opioid receptor agonist) administered supraspinally. However, the activation of nicotinic receptors at supraspinal sites may not be involved in enhancing the antinociception induced by DPDPE (a delta-opioid receptor agonist) or U50, 488H (a kappa-opioid receptor agonist) administered supraspinally.     

The nitric oxide/cyclic GMP system at the supraspinal site is involved in the development of acute morphine antinociceptive tolerance

The role of the supraspinal nitric oxide (NO)/cyclic GMP system in the development of acute morphine antinociceptive tolerance was investigated by use of the mouse 55 degrees C warm-water tail-flick test. A single intracerebroventricular (i.c.v.) pretreatment of mice with morphine (3 nmol, 140 min before testing) produced an acute antinociceptive tolerance to subsequent i.c.v. doses of morphine, as demonstrated by a 120-fold rightward shift of the morphine dose-response curve. When co-administered with morphine (140 min before testing), the NO synthase inhibitors: N-nitro-L-arginine methyl ester (L-NAME), 3-bromo-7-nitroindazole, 7-nitroindazole and NG-monomethyl-L-arginine, attenuated the development of morphine tolerance. All four NO synthase inhibitors completely blocked the rightward shift of the morphine dose-response curve caused by i.c.v. morphine pretreatment (3 nmol, 140 min before testing). This effect was partially antagonized by L-arginine, but not D-arginine, in a dose-dependent manner. Also, D-NAME did not block the development of tolerance. Like the NO synthase inhibitors, LY-83,583, a guanylyl cyclase inhibitor, blocked the development of tolerance, which suggests that NO acting through the cyclic GMP pathway is involved in the development of acute antinociceptive tolerance. The effects of increased NO production on acute morphine antinociceptive tolerance were also studied. When co-administered with morphine (140 min before testing), neither L-arginine (100 nmol) nor the NO donors, sodium nitroprusside (5 nmol) and isosorbide dinitrate (10 nmol), had any effect on the magnitude of morphine antinociceptive tolerance. These results suggest that NO, acting through the cyclic GMP pathway, mediates the development of acute antinociceptive tolerance, but that NO production does not alter the magnitude of antinociceptive tolerance.     

Dermorphin analog Tyr-D-Arg2-Phe-sarcosine-induced opioid analgesia and respiratory stimulation: the role of mu 1-receptors?

Tyr-D-Arg2-Phe-sarcosine4 (TAPS), a mu-selective tetrapeptide analog of dermorphin, induced sustained antinociception and stimulated ventilatory minute volume (MV) at the doses of 3 to 100 pmol i.c.v. The doses of 30 and 100 pmol i.c.v. induced catalepsy. The effect of TAPS on MV was in negative correlation with the dose and the maximal response was achieved by the lowest (3 pmol) dose (+63 +/- 23%, P < .05). Morphine, an agonist at both mu 1 and mu 2 sites, at a dose of 150 nmol i.c.v. (equianalgesic to 100 pmol of TAPS decreased the MV by 30%, due to a decrease in ventilatory tidal volume. The antinociceptive effect of TAPS was antagonized by naloxone and the mu 1 receptor antagonist, naloxonazine. Naloxonazine also attenuated the catalepsy produced by 100 pmol of TAPS i.c.v. and the respiratory stimulation produced by 3 pmol of TAPS i.c.v. Pretreatment with 30 pmol of TAPS antagonized the respiratory depression induced by the mu opioid agonist dermorphin (changes in MV after dermorphin alone at 1 or 3 nmol were -22 +/- 10% and -60 +/- 9% and, after pretreatment with TAPS, +44 +/- 11% and -18 +/- 5%, respectively). After combined pretreatment with naloxonazine and TAPS, 1 nmol of dermorphin had no significant effect on ventilation. In contrast, pretreatment with a low respiratory stimulant dose (10 pmol i.c.v.) of dermorphin did not modify the effect of 1 nmol of dermorphin. In conclusion, the antinociceptive, cataleptic and respiratory stimulant effects of TAPS appear to be a related to its agonist action at the mu 1 opioid receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

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.     

Amygdaloid–thalamic interactions mediate the antinociceptive action of morphine microinjected into the periaqueductal gray.

The bilateral administration of the serotonin receptor antagonist methysergide (2.5 μg, 5 μg, and 10 μg) into either the central nucleus of the amygdala (ACe) or nucleus parafascicularis thalami (nPf) produced dose-dependent inhibition of the antinociceptive action of ventrolateral periaqueductal gray (vPAG)-administered morphine. Unilateral administration of these doses of methysergide into either the ACe or nPf had no effect on morphine-induced antinociception. However, the combined unilateral administration of these doses of methysergide into the ACe and nPf produced dose-dependent inhibition of morphine antinociception that was identical to that observed after its bilateral administration into either site. This latter finding is interpreted as evidence that a functional interaction between the ACe and nPf supports the antinociceptive action of morphine administered into the vPAG. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Streptozotocin-induced diabetes selectively enhances antinociception mediated by delta 1- but not delta 2-opioid receptors

We assessed the effect of diabetes on antinociception produced by intracerebroventricular injection of delta-opioid receptor agonists [D-Pen2,5]enkephalin (DPDPE) and [D-Ala2]deltorphin II. The antinociceptive effect of DPDPE (10 nmol), administered i.c.v., was significantly greater in diabetic mice than in non-diabetic mice. The antinociceptive effect of i.c.v. DPDPE was significantly reduced in both diabetic and non-diabetic mice following pretreatment with 7-benzylidenenaltrexone (BNTX), a selective delta 1-opioid receptor antagonist, but not with naltriben (NTB), a selective delta 2-opioid receptor antagonist. There were no significant differences in the antinociceptive effect of [D-Ala2]deltorphin II (3 nmol, i.c.v.) in diabetic and non-diabetic mice. Furthermore, the antinociceptive effect of i.c.v. [D-Ala2]deltorphin II was significantly reduced in both diabetic and non-diabetic mice following pretreatment with NTB, but not with BNTX. In conclusion, mice with diabetes are selectively hyper-responsive to supraspinal delta 1-opioid receptor-mediated antinociception, but are normally responsive to activation of delta 2-opioid receptors.     

Possible mechanisms of salt-induced hypertension in Dahl salt-sensitive rats

The effects of acute and chronic administration of cocaine on the antinociception and tolerance to the antinociceptive actions of mu-(morphine), kappa-(U-50,488H), and delta-([D-Pen2,D-Pen5]enkephalin; DPDPE), opioid receptor agonists were determined in male Swiss-Webster mice. Intraperitoneal injection of 40 mg/kg of cocaine by itself produced weak antinociceptive response as measured by the tail-fick test but the lower doses were ineffective. Administration of morphine (10 mg/kg, SC), U-50,488H (25 mg/kg, IP) or DPDPE (10 microg/mouse, ICV) produced antinociception in mice. Cocaine (20 mg/kg) potentiated the antinociceptive action of morphine and DPDPE but had no effect on U-50,488H-induced antinociception. Administration of morphine (20 mg/kg, SC), U-50,488H (25 mg/kg, IP) or DPDPE (20 microg/mouse, ICV) twice a day for 4 days resulted in the development of tolerance to their antinociceptive actions. Tolerance to the antinociceptive actions of morphine and U-50,488H was inhibited by concurrent treatment with 20 or 40 mg/kg doses of cocaine; however, tolerance to the antinociceptive action of DPDPE was not modified by cocaine. It is concluded that cocaine selectively potentiates the antinociceptive action of mu- and delta- but not of the kappa-opioid receptor agonist. On the other hand, cocaine inhibits the development of tolerance to the antinociceptive actions of mu- and kappa- but not of delta-opioid receptor agonists in mice.     

Evidence for the existence of the beta-endorphin-sensitive "epsilon-opioid receptor" in the brain: the mechanisms of epsilon-mediated antinociception

Recently, mu-, delta- and kappa-opioid receptors have been cloned and relatively well-characterized. In addition to three major opioid receptor types, more extensive studies have suggested the possible existence of other opioid receptor types that can be classified as non-mu, non-delta and non-kappa. Based upon anatomical and binding studies in the brain, the sensitive site for an endogenous opioid peptide, beta-endorphin, has been postulated to account for the unique characteristics of the opioid receptor defined as a putative epsilon-opioid receptor. Many epsilon-opioid receptors are functionally coupled to G-proteins. The functional epsilon-opioid receptors in the brain are stimulated by bremazocine and etorphine as well as beta-endorphin, but not by selective mu-, delta- or kappa-opioid receptor agonists. Epsilon-opioid receptor agonists injected into the brain produce profound antinociception. The brain sites most sensitive to epsilon-agonist-induced antinociception are located in the caudal medial medulla such as the nucleus raphe obscures, nucleus raphe pallidus and the adjacent midline reticular formation. The stimulation of epsilon-opioid receptors in the brain facilitates the descending enkephalinergic pathway, which probably originates from the brainstem terminating at the spinal cord. The endogenous opioid Met-enkephalin, released in the spinal cord by activation of supraspinal epsilon-opioid receptors, stimulates spinal delta2-opioid receptors for the production of antinociception. It is noteworthy that the epsilon-opioid receptor-mediated pain control system is different from that of other opioid systems. Although there appears to be no epsilon-selective ligand currently available, these findings provide strong evidence for the existence of the putative epsilon-opioid receptor and its unique function in the brain.     

Protein kinase A maintains cellular tolerance to mu opioid receptor agonists in hypothalamic neurosecretory cells with chronic morphine treatment: convergence on a common pathway with estrogen in modulating mu opioid receptor/effector coupling

The present study examined protein kinase A (PKA) and protein kinase C (PKC) involvement in the maintenance of cellular tolerance to mu opioid receptor agonists resulting from chronic opiate exposure in neurosecretory cells of the hypothalamic arcuate nucleus (ARC). The possibility that the diminution of mu opioid receptor/effector coupling produced by acute 17beta-estradiol or chronic opiate exposures is mediated by a common kinase pathway also was investigated. Intracellular recordings were made in hypothalamic slices prepared from ovariectomized female guinea pigs. The mu opioid receptor agonist D-Ala2, N-Me-Phe4, Gly-ol5-enkephalin (DAMGO) produced dose-dependent hyperpolarizations of ARC neurons. Chronic morphine treatment for 4 days reduced DAMGO potency 2.5-fold with no change in the maximal response. This effect was mimicked by a 20-min bath application of the PKA activator cAMP, Sp-isomer, or the PKC activator phorbol-12,13-dibutyrate. A 30-min bath application of the broad-spectrum protein kinase inhibitor staurosporine completely abolished the reduced DAMGO potency seen in morphine-tolerant neurosecretory cells, including those immunopositive for gonadotropin-releasing hormone. The effect of staurosporine was mimicked by the PKA inhibitor cAMP, Rp-isomer, but not by the PKC inhibitor calphostin C. Finally, a 20-min bath application of 17beta-estradiol did not further reduce DAMGO potency in morphine-tolerant ARC neurons. Therefore, increased PKA activity maintains cellular tolerance to mu opioid receptor agonists in ARC neurosecretory cells caused by chronic morphine treatment. Furthermore, acute 17beta-estradiol and chronic opiate treatments attenuate mu opioid receptor-mediated responses via a common PKA pathway.     

Antinociception produced by an ascending spino-supraspinal pathway

Studies in mice and rats have shown that antinociception produced by intrathecal (i.t.) administration of opioids can be partially inhibited by intracerebroventricular (i.c.v.) administration of naloxone. In this study we tested the hypothesis that this inhibition by i.c.v. naloxone results from antagonism of supraspinal endogenous opioid-mediated antinociception produced by the action of i.t. opioids on an ascending antinociceptive pathway. In rats lightly anesthetized with urethane/alpha-chloralose, i.t. DAMGO, i.t. lidocaine, or spinal transection at T5-T6 all attenuated the trigeminal jaw opening reflex (JOR) (i.e., were antinociceptive), an effect that was antagonized in each case by i.c.v. naloxone. These findings support the suggestion that there exists a pathway that ascends from the spinal cord to a supraspinal site that tonically inhibits antinociception mediated by supraspinal opioids. When activity in this ascending pathway is suppressed (e.g., by i.t. opioids or local anesthetics or by spinal cord transection), antinociception mediated by supraspinal opioids is disinhibited. To determine the supraspinal site(s) at which endogenous opioid-dependent antinociception is evoked by i.t. opioids, we microinjected naloxone methiodide into several supraspinal sites. Microinjection of naloxone methiodide into nucleus accumbens but not into the rostral ventral medulla (RVM) or the periaqueductal gray matter (PAG) antagonized the suppression of the JOR produced by i.t. DAMGO or lidocaine. The possibility that this ascending pathway may represent a source of spinal input to mesolimbic circuitry involved in setting the state of sensorimotor reactivity to noxious stimuli is discussed.     

Effect of 7-nitroindazole on tolerance to morphine, U-50,488H and [D-Pen2, D-Pen5] enkephalin in mice

The effects of 7-nitroindazole (7-NI), an inhibitor of the neuronal nitric oxide synthase (nNOS) which does not increase blood pressure, on tolerance to the antinociceptive activity of mu-(morphine), kappa-(U-50,488H) and delta-([D-Pen2, D-Pen5]enkephalin, DPDPE) opioid receptor agonists were determined in mice. Male Swiss-Webster mice were made tolerant by twice daily injections of morphine (20 mg/kg, s.c.), U-50,488H (25 mg/kg, i.p.) or DPDPE (20 micrograms/mouse, i.c.v.) for 4 days. When tested on day 5, tolerance to their antinociceptive activity was evidenced by decreased response in chronic drug treated mice in comparison to vehicle-injected mice. Concurrent administration of 7-NI (20, 40 or 80 mg/kg, i.p.) with DPDPE did not modify the development of tolerance to the antinociceptive action of DPDPE. However, 7-NI (40 or 80 mg/kg, i.p.) inhibited the development of tolerance to the antinociceptive activity of morphine and U-50,488H but the lower dose of 7-NI (20 mg/kg, i.p.) was not effective. Chronic administration of 7-NI by itself did not modify the acute response to morphine, U-50,488H or DPDPE. It is concluded that a specific inhibitor of nNOS can inhibit tolerance to the antinociceptive activity of mu- and kappa- but not of delta-opioid receptor agonists in mice.