Opioid control of the in vitro release of calcitonin gene-related peptide from primary afferent fibres projecting in the rat cervical cord

In vitro superfusion of slices from the dorsal half of the rat cervical enlargement allowed the measurement of spontaneous, K+ (30 mM)- and capsaicin (0.5 microM)-evoked release of calcitonin gene-related peptide-like immunoreactive material (CGRPLI). The greater part of this immunoreactive material originated in primary afferent fibres since dorsal rhizotomy from C4 to Th2 (8 days before sacrifice) resulted in a 85-90% decrease in CGRPLI release. CGRPLI outflow which persisted after dorsal rhizotomy could still be enhanced by K+-induced depolarization but was no longer sensitive to the stimulatory effect of 0.5 microM capsaicin. Both delta (DTLET, D-Pen2-D-Pen5-enkephalin) and mu (DAGO, PL 017) opioid receptor agonists reduced the K+ evoked release of CGRPLI from the dorsal half of the cervical enlargement. Morphine was also inhibitory but the selective K opioid agonist U 69593 was inactive. As expected from the involvement of delta and mu receptors, the selective opioid antagonist ICI 174864 and naloxone prevented the inhibitory effects of DTLET and DAGO, respectively. These data suggest that opioid-induced presynaptic inhibiton of CGRP-containing primary afferent fibres may be involved in the analgesic effect of intrathecally injected delta and mu opioid agonists in rats.     

Opioid control of the release of Met-enkephalin-like material from the rat spinal cord.

The possible control by opioids of the release of Met-enkephalin-like material (MELM) from the rat spinal cord was investigated in vitro and in vivo. Superfusion of slices of the dorsal zone of the lumbar enlargement with the mu selective agonists DAGO or PL 017 or the delta selective agonist DTLET produced a significant reduction in the K(+)-evoked MELM release from these tissues. These effects persisted in the presence ot tetrodotoxin, as expected from their mediation through presynaptically located opioid autoreceptors. Furthermore, the inhibitory effect of DAGO and PL 017, but not that of DTLET, was prevented by the preferential mu antagonist naloxone. Conversely, the effect of DTLET was prevented by the delta antagonist naltrindole but not by naloxone. In vivo experiments performed in halothane-anaesthetized rats have shown that the intrathecal perfusion of DAGO and DTLET significantly depressed the spontaneous MELM outflow from the whole spinal cord. In contrast to these mu and delta agonists, the kappa selective agonist U 50488 H did not affect the in vivo- and only slightly reduced (at a very high concentration: 50 microM) the in vitro-release of MELM from the rat spinal cord. These data indicate that both mu and delta opioid autoreceptors are involved in a local presynaptic autoinhibitory control of MELM release in the rat dorsal horn.     

Opioid delta-receptor involvement in supraspinal and spinal antinociception in mice

The possibility that the opioid delta-receptor mediates antinociception in tests where heat is the noxious stimulus was investigated using highly selective mu- and delta-agonist and -antagonists. Antinociceptive dose-response curves were constructed for mu ([D-Ala2,NMePhe4,Gly-ol]enkephalin, DAGO; morphine) and delta ([D-Pen2,D-Pen5]enkephalin, DPDPE)-agonists in the absence, and in the presence of the mu non-surmountable antagonist, beta-funaltrexamine (beta-FNA) or the delta-antagonist ICI 174,864 (N,N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH, where Aib is alpha-amino-isobutyric acid). Agonists and ICI 174,864 were given alone in the same intracerebroventricular (i.c.v.) or intrathecal (i.th.) injection to mice 20 min prior to testing in the warm-water (55 degrees C) tail-withdrawal test (+10 min for i.th. DPDPE); beta-FNA was given as a single i.c.v. or i.th. pretreatment dose (20 and 0.01 nM, respectively) 4 h prior to testing. I.c.v. pretreatment with beta-FNA resulted in a rightward displacement of the DAGO and morphine antinociceptive dose-response lines, but failed to displace the i.c.v. DPDPE curve. Similarly, i.th. pretreatment with beta-FNA displaced the i.th. morphine dose-response curve to the right without affecting the i.th. DPDPE antinociceptive dose-response line. ICI 174,864 (1 and 3 micrograms) produced a dose-related antagonism of i.c.v. or i.th. DPDPE, but did not alter the antinociceptive effects of DAGO or morphine given by the same routes. Co-administration of ICI 174,864 (3 micrograms) with i.c.v. morphine in beta-FNA pretreated (but not control) mice resulted in a further rightward displacement of the morphine dose-response line.(ABSTRACT TRUNCATED AT 250 WORDS)     

Opioids modulate cell division in the germinal zone of the late embryonic neocortex.

Opioid effects on cell division in the embryonic cerebral cortex were examined using two experimental approaches: (i) the presence of opioid receptors in the embryonic day 16 mouse neocortex was tested using immunohistochemical techniques; (ii) the values of the indices of [3H]thymidine pulse labelled cells and mitotic indices were estimated in the ventricular zone of the embryonic day 16 mouse neocortex 2.5, 4.5 and 8.5 h after administration to pregnant females of selected opioid receptor agonists or the opioid antagonist naloxone. The immunohistochemical study demonstrated that distinct subpopulations of the ventricular zone cells express mu, delta or kappa opioid receptors. Acute exposure of mouse embryos to mu, delta and kappa opioid receptor agonists or naloxone differentially affects the indices of [3H] thymidine pulse labelled cells and mitotic indices indicating changes in the cell cycle composition. Treatment with the mu opioid receptor agonist D-Ala2-MePhe4, Gly-ol5-enkephalin (DAGO), or the partially selective kappa opioid receptor agonist bremazocine, increased the [3H]thymidine labelling and mitotic indices. In contrast, the delta receptor agonist (D-Ser8)-leucine enkephalin-Thr (DSLET) produced a decrease in the labelled cell indices and mitotic indices. Naloxone provided a biphasic effect: a decrease in the values of labelled cell indices 2.5 h after naloxone administration, followed by an increase in the values of the indices at 4.5 and 8.5 h. These results suggest that the endogenous embryonic/maternal opioid systems are involved in the regulation of cell division in the ventricular zone of the late embryonic cortex.     

Opioid receptor subtypes in the rat spinal cord: electrophysiological studies with μ- and δ-opioid receptor agonists in the control of nociception

We have compared the ability of selective mu- and delta-opiate agonists to modulate nociceptive transmission at the level of the rat dorsal horn using electrophysiological approaches. Single-unit extracellular recordings were made from neurones in the lumbar dorsal horn of the intact rat under halothane anaesthesia. Neurones could be activated by both A- and C-fibre electrical stimulation (and by natural innocuous and noxious stimuli). Agonists were applied directly onto the cord in a volume of 50 microliters. The intrathecal administration of 3 agonists, Tyr-D-Ala-Gly-MePhe-Gly-ol (DAGO) (mu-selective) (2 X 10(-3)-10 nmol) Tyr-D-Thr-Gly-Phe-Leu-Thr (DTLET) (mu/delta) (7 X 10(-4)-70 nmol), and cyclic Tyr-D-Pen-Gly-Phe-D-Pen (DPDPE) (delta) (2 X 10(-2)-100 nmol) produced dose-dependent inhibitions of C-fibre-evoked neuronal activity whilst A-fibre activity was relatively unchanged. DAGO produced near-maximal inhibitions which could be completely reversed by naloxone (1.5 nmol) whilst DPDPE causes less marked inhibitions which could only be partially reversed by naloxone (1.5-13.5 nmol). DTLET produced effects intermediate to those of DAGO and DPDPE. The results suggest that both mu- and delta-opioid receptors can modulate the transmission of nociceptive information in the rat spinal cord.     

Comparison of the antinociceptive action of mu and delta opioid receptor ligands in the periaqueductal gray matter, medial and paramedial ventral medulla in the rat as studied by the microinjection technique

In rats stereotaxically implanted with microinjection cannula in either the periaqueductal gray matter (PAG) or the medial/paramedial medullary reticular formation (MRF), microinjection of morphine, sufentanil, D-Ala2-D-Leu5-enkephalin (DADL) or D-Ser2-Thr6-leucine enkephalin (DSTLE) produced dose-dependent elevations in the response latency on tail-flick and hot plate tests. These effects were reversed by naloxone administered by microinjection into the same intracerebral site. Both mu (morphine and sufentanil) and delta (DADL and DSTLE) opioid receptor ligands produced a maximal elevation in the supraspinally mediated hot plate response when administered into either the PAG or the MRF. Similarly, mu and delta receptor ligands produced maximum elevations in the spinally mediated tail-flick response when microinjected into the PAG. In contrast, delta, but not mu, receptor agonists produced a total blockade of the tail-flick response following administration into the MRF. Microinjection of mu (morphine) or delta (DADL) agonists into the PAG or the MRF also resulted in a naloxone-reversible inhibition of the visceral chemical evoked writhing response. These observations suggest that mu and delta opioid receptor linked systems within the MRF but not the PAG produce their antinociceptive effects by discriminable mechanisms with a differential action on spinopetal vs supraspinal modulatory systems.     

Inhibition of trigemino-hypoglossal reflex in rats by oxytocin is mediated by mu and kappa opioid receptors

Recent studies showed that oxytocin plays an important role in the modulation of pain at different levels of the central nervous system. The present study was undertaken to investigate the effect of oxytocin on trigemino-hypoglossal reflex in rats. With the experimental settings used in this study, we have demonstrated that oxytocin showed significant analgesic effect after intracerebroventricular administration in rats, as assayed by the amplitude of the retractory movements of the tongue after tooth pulp stimulation. Antinociceptive effect of oxytocin was inhibited by subsequent perfusion of cerebral ventricles with oxytocin antagonist, [deamino-Cys1-D-Tyr(OEt)2-Thr4-Orn8]-oxytocin, atosiban. An involvement of opioid system in the oxytocin-induced analgesia was studied after intracerebroventricular administration of different opioid antagonists: non-selective naloxone, mu-selective beta-funaltrexamine, delta-selective naltrindole, and kappa-selective nor-binaltorphimine. It was shown that inhibition of antinociceptive effects was mediated through mu and kappa opioid receptors, indicating that there is a synergy between oxytocin and opioid systems in transmitting and modulating pain stimuli. Co-administration of oxytocin and a mu-selective endogenous opioid ligand endomorphin-2 did not significantly increase the antinociceptive activity of endomorphin-2.     

III. Comparison of the antinociceptive action of Mu and delta opioid receptor ligands in the periaqueductal gray matter, medial and paramedial ventral medulla in the rat as studied by the microinjection technique

In rats stereotaxically implanted with microinjection cannula in either the periaqueductal gray matter (PAG) or the medial/paramedial medullary reticular formation (MRF), microinjection of morphine, sufentanil,d-Ala²-d-Leu⁵-enkephalin (DADL) ord-Ser²- Thr⁶-leucine enkephalin (DSTLE) produced dose-dependent elevations in the response latency on tail-flick and hot plate tests. These effects were reversed by naloxone administered by microinjection into the same intracerebral site. Both mu (morphine and sufentanil) and delta (DADL and DSTLE) opioid receptor ligands produced a maximal elevation in the supraspinally mediated hot plate response when administered into either the PAG or the MRF. Similarly, mu and delta receptor ligands produced maximum elevations in the spinally mediated tail-flick response when microinjected into the PAG. In contrast, delta, but not mu, receptor agonists produced a total blockade of the tail-flick response following administraion into the MRF. Microinjection of mu (morphine) or delta (DADL) agonists into the PAG or the MRF also resulted in a naloxone-reversible inhibition of the visceral chemical evoked writhing response. These observations suggest that mu and delta opioid receptor linked systems within the MRF but not the PAG produce their antinociceptive effects by discriminable mechanisms with a differential action on spinopetal vs supraspinal modulatory systems.     

MU and delta opioid receptors control differently the horizontal and vertical components of locomotor activity in mice

The present study investigated the role of mu and delta opioid receptors in the control of the horizontal and vertical components of locomotion. Mice received intracerebroventricularly (i.c.v.) enkephalin analogs specific for either the mu or delta opioid receptors. The administration of the specific mu agonist [D-Ala2-NMePhe4-Gly5(ol)] enkephalin (DAGO) induced a dose-dependent increase in horizontal activity and a decrease in vertical activity. The specific delta agonist [D-Pen2,D-Pen5] enkephalin (DPDPE) increased both components of motor activity. The opiate antagonist naltrexone reversed the effects of DAGO, but did not influence the effects of DPDPE on motor activity. The pretreatment with the delta opiate antagonist ICI 154, 129 completely reversed the effects of DPDPE on locomotion but antagonized only partially the effects of DAGO on locomotion. These results indicate that the two components of locomotor activity--horizontal and vertical activity--are modulated differently by the stimulation of mu or delta opioid receptors.     

Differential depressive action of two μ and δ opioid ligands on neuronal responses to noxious stimuli in the thalamic ventrobasal complex of rat

In the present investigation the effects of selective agonists for μ (Tyr- -Ala-Me-Phe-Gly-ol (DAGO)) and δ (Tyr- -Thr-Gly-Phe-Leu-Thr (DTLET)) opioid receptors on neuronal activities induced by noxious cutaneous stimuli in the rat ventrobasal (VB) thalamus were analyzed. The two agonists produced a clear depressive action on thermal as well as mechanical noxious stimuli. The depressive action of DTLET (3 mg/kg i.v.) was lower and of shorter duration than that of DAGO (2 mg/kg i.v.). However, this effect is unambiguously related to the selective stimulation of opioid receptors since a consistent effect was also observed for a dose as low as 1.5 mg/kg i.v. of DTLET. Moreover, DTLET effect needs a high concentration of naloxone (0.5 mg/kg i.v.) to be reversed, while DAGO effect is totally reversed with 0.1 mg/kg i.v.     

Microinjection of opioid antagonists into the substantia nigra reduces stress-induced eating in rats

Stress produced by pinching the tail has been shown to cause satiated animals to eat and to display oral stereotypies. Endogenous opioids and central dopamine systems have been implicated in the mediation of these effects. In order to test the possibility that the substantia nigra (SN) might be involved, the amount of food intake and gnawing produced by mild tail pinch were assessed following bilateral microinjections of opioid antagonists into the SN. Evaluations of nociceptive thresholds were also conducted using tail flick and hot plate tests. Eating induced by tail pinch was reduced by microinjections of the non-selective opioid antagonist naloxone (3, 10, 20 and 30 nmol) and by the mu-selective antagonist Cys2, Tyr3, Orn5, Pen7 Amide (CTOP) (1, 3 and 10 nmol). These effects on eating occurred in the absence of effects on gnawing. kappa- and delta-antagonists (10 nmol) had no effect on eating or gnawing. Naloxone did not alter either tail flick or hot-plate response latencies. The highest dose of CTOP increased response latency on the hot-plate test only. The results are interpreted as suggesting that the SN may be an important central site of action for opioid antagonists in reducing stress-induced eating. The possibility that the SN may be a central site mediating the effects of dopamine on this phenomenon is also discussed.     

Autoradiographic study of irreversible binding of [3H]beta-funaltrexamine to opioid receptors in the rat forebrain: comparison with mu and delta receptor distribution.

beta-Funaltrexamine (beta-FNA) is an irreversible mu antagonist and a reversible kappa agonist in in vivo and in vitro tests. However, whether it produces irreversible delta antagonism is controversial. In binding studies, it is clear that beta-FNA does not bind irreversibly (it does reversibly) to kappa receptors. Yet there is no consensus as to whether beta-FNA binds irreversibly to mu and/or delta receptors. In this study, irreversible binding of [3H]beta-FNA to opioid receptors was examined in rat forebrain sections in the presence of 200 mM NaCl and its distribution compared with those of mu and delta opioid receptors, labeled by [3H][D-Ala2,MePhe4,Gly-ol5]enkephalin ([3H]DAMGO) and [3H][D-Pen2,D-Pen5]enkephalin ([3H]DPDPE), respectively. Irreversible binding of [3H]beta-FNA was determined as the binding that remained following 5 washes at room temp. for 1, 5, 20, 20, and 20 min each. Non-specific binding was defined by including 10 microM naloxone, beta-chlornaltrexamine (beta-CNA), or beta-FNA in the incubation mixture. At 37 degrees C, specific irreversible binding of [3H]beta-FNA to opioid receptors reached a plateau at 10 nM in 60 min, and constituted 50-70% of total irreversible binding. Series of 4 sections of similar anatomical levels were labeled with [3H]DAMGO, [3H]beta-FNA, [3H]beta-FNA + 10 microM naloxone, beta-CNA, or beta-FNA, and [3H]DPDPE, resp., and exposed to [3H]-Ultrofilm. The distribution of [3H]beta-FNA (5 nM) irreversible labeling is very similar to that of [3H]DAMGO, i.e. patches and subcallosal streaks in caudate-putamen, patches in nucleus accumbens, dense labeling in thalamus, and more binding in the rostral than caudal striatum.(ABSTRACT TRUNCATED AT 250 WORDS)     

Substantia nigra as a site of origin of dopamine-dependent motor syndromes induced by stimulation of mu and delta opioid receptors

Opioid agonists having different affinity for delta and mu receptors were injected bilaterally in the substantia nigra (SN) of rats. The selective agonist of mu receptors N-MePhe3,-D-Pro4 morphiceptin (PLO 17) produced a stereotyped behavior characterized by stereotyped sniffing and gnawing antagonized by the irreversible antagonist of mu receptors beta-funaltrexamine. In contrast, bilateral intranigral injection of the selective delta agonist D-Pen2,D-Pen5 enkephalin (DPDPE) elicited dose-dependent exploratory behavior and rearing but failed to produce gnawing. The behavioral syndrome induced by DPDPE was significantly reduced by the selective delta antagonist ICI 174,864. Naloxine, a non-selective opioid antagonist, antagonized the effects of both compounds. SCH 23390 and haloperidol, two antagonists of dopaminergic D1 and D2 receptors, respectively, blocked the effects of PLO 17 and DPDPE. The results indicate that stimulation of specific opioid receptor types in the SN elicits specific behavioral syndromes and suggest that the SN might be the site of origin of certain items of the behavioral syndrome evoked by systemic opiates. These items might be mediated by activation of dopaminergic neurons of the ventral mesencephalon.     

DAGO ([D-Ala2,N-Me-Phe4,Gly-ol]enkephalin) specifically reverses apomorphine-induced increase in rearing and grooming behaviors in the mouse.

The effects of intracerebroventricular injections (10 microliters) of the mu-selective opioid peptide DAGO on apomorphine (0.1, 0.56, 1.0 and/or 3.0 mg/kg)-induced motor activity were investigated in the mouse using multi-dimensional behavioral analyses. A lower dose (0.1 mg/kg) of apomorphine failed to significantly affect motor activity, whilst higher doses (0.56, 1.0 and 3.0 mg/kg) of the drug produced a marked increase in linear locomotion, circling, rearing and/or grooming behaviors. DAGO (0.003 and 0.01 micrograms) did not significantly affect different behaviors. DAGO (0.01 micrograms) antagonized the apomorphine (1.0 mg/kg)-induced increase in behaviors such as rearing and grooming. However, DAGO (0.003 or 0.01 micrograms) did not affect behaviors induced by a 3.0 mg/kg dose of apomorphine. Furthermore, the effects of DAG]O on apomorphine-induced behaviors were fully reversed by treatment with the mu-selective alkylating agent beta-FNA (beta-funaltrexamine) (5.0 micrograms). These results suggest that mu opioid receptors play a principal role in the apomorphine-induced increase in rearing and grooming behaviors.     

A single restraint stress exposure potentiates analgesia induced by intrathecally administered DAGO

In rats, restraint exposure potentiates the magnitude and duration of analgesia following both the peripheral and intracerebroventricular administration of several opioid agonists as compared to non-stressed controls. It has been suggested that the site of action whereby restraint leads to potentiated opioid analgesia is located supraspinally. However, the possible contribution of spinal analgesic mechanisms also warrants investigation. Thus, the purpose of the present study was two-fold: (1) to determine whether a single exposure to restraint stress would result in the dose-dependent potentiation of analgesia following the intrathecal (i.t.) administration of the mu (mu)-receptor selective opioid agonist [D-Ala2,N-Me-Phe4,Gly5-ol]enkephalin (DAGO) and (2) to quantify the degree of analgesia in restrained vs. non-restrained rats using the tail-flick and hot-plate analgesic assays. Using rats implanted with chronic i.t. cannula, dose- and time-course curves were observed following the i.t. administration of DAGO. The results demonstrate that both the duration and magnitude of analgesia was significantly potentiated in restrained rats compared to non-restrained controls. Restraint-treated rats receiving 0.15-0.6 micrograms of DAGO i.t. showed 1.3-1.5-fold potentiation of analgesia in the tail-flick assay and a 2.3-5.6-fold potentiation using the hot-plate assay. Restraint immobilization potentiated the magnitude and duration of DAGO-induced analgesia administered by the i.t. route as measured by the tail-flick and hot-plate assays. These data suggest that spinal analgesic mechanisms significantly contribute to the enhanced analgesic potency of opioids in subjects exposed to restraint stress.     

Multiple opioid receptors mediate feeding elicited by mu and delta opioid receptor subtype agonists in the nucleus accumbens shell in rats

The nucleus accumbens, and particularly its shell region, is a critical site at which feeding responses can be elicited following direct administration of opiate drugs as well as micro-selective and delta-selective, but not kappa-selective opioid receptor subtype agonists. In contrast to observations of selective and receptor-specific opioid antagonist effects upon corresponding agonist-induced actions in analgesic studies, ventricular administration of opioid receptor subtype antagonists blocks feeding induced by multiple opioid receptor subtype agonists. The present study examined whether feeding responses elicited by either putative mu ([D-Ala(2), NMe-Phe(4), Gly-ol(5)]-enkephalin (DAMGO)), delta(1) ([D-Pen(2), D-Pen(5)]-enkephalin (DPDPE)) or delta(2) ([D-Ala(2), Glu(4)]-deltorphin (Deltorphin)) opioid receptor subtype agonists administered into the nucleus accumbens shell were altered by accumbens pretreatment with either selective mu (beta-funaltrexamine), mu(1) (naloxonazine), delta(1) ([D-Ala(2), Leu(5), Cys(6)]-enkephalin (DALCE)), delta(2) (naltrindole isothiocyanate) or kappa(1) (nor-binaltorphamine) opioid receptor subtype antagonists. Similar magnitudes and durations of feeding responses were elicited by bilateral accumbens administration of either DAMGO (2.5 microg), DPDPE (5 microg) or Deltorphin (5 microg). DAMGO-induced feeding in the nucleus accumbens shell was significantly reduced by accumbens pretreatment of mu, delta(1), delta(2) and kappa(1), but not mu(1) opioid receptor subtype antagonists. DPDPE-induced feeding in the accumbens was significantly reduced by accumbens pretreatment of mu, delta(1), delta(2) and kappa(1), but not mu(1) opioid receptor subtype antagonists. Deltorphin-induced feeding in the accumbens was largely unaffected by accumbens delta(2) antagonist pretreatment, and was significantly enhanced by accumbens mu or kappa(1) antagonist pretreatment. These data indicate different opioid pharmacological profiles for feeding induced by putative mu, delta(1) and delta(2) opioid agonists in the nucleus accumbens shell, as well as the participation of multiple opioid receptor subtypes in the elicitation and maintenance of feeding by these agonists in the nucleus accumbens shell.     

Acute delta- and kappa-opioid agonist pretreatment potentiates opioid antagonist-induced suppression of water consumption

The primary objective of this study was to determine whether pretreatment with kappa- and delta-opioid agonists potentiates naltrexone-induced suppression of water consumption following 24 h of deprivation. This study also examined the temporal effects of agonist-induced antinociception using the tail-flick and hot-plate tests. Adult male Sprague-Dawley rats were water deprived 20 h and then given an injection (s.c. or i.c.) of an opioid agonist or saline. Drugs included the mu-opioid agonists morphine and DAMGO ([d-Ala2,NMePhe4,Gly-ol5]-enkephalin), the kappa-opioid agonists spiradoline, bremazocine, and U69,593, and the delta-opioid agonists BW 373U86 and DPDPE ([D-Pen2, D-Pen5]-enkephalin). Three hours and forty-five minutes later, animals received a single dose of naltrexone (0.1–30 mg/kg, s.c.) or saline. Fifteen minutes later, animals were allowed free access to water for 30 min. For the tail-flick and hot-plate tests, animals were given a single injection of agonist and tested in both procedures every 30 min for up to 2 h, then hourly up to 6 h post-injection. Naltrexone dose-dependently suppressed fluid consumption 24 h after deprivation. The effects of naltrexone on drinking were potentiated following pretreatment with at least one dose of the agonists tested except BW 373U86. With the exception of BW 373U86, DAMGO, and DPDPE, all of the opioid agonists produced significant antinociception in the hot-plate test. Only BW 373U86 failed to have an antinociceptive effect in the tail-flick test. By 4 h after treatment, drug-induced antinociception had largely waned, suggesting the potentiation of naltrexone-induced drinking suppression was not a result of a direct interaction with the agonists. In conclusion, kappa-opioid and delta-opioid receptors appear to contribute to the manifestation of acute opioid dependence, albeit to a lesser degree than mu-opioid receptors.     

Feedback inhibition of met-enkephalin release from the rat spinal cord in vivo.

The possible existence of a feedback control by endogenous opioids of the spinal release of met-enkephalin-like material was assessed in vivo, in halothane-anesthetized rats whose intrathecal space was continuously perfused with an artificial cerebrospinal fluid supplemented with various opioid-related drugs. Both the intrathecal perfusion of the mu agonist D-Ala2-D-MePhe4-Gly-ol5-enkephalin (DAGO) (10 microM) and the delta agonist Tyr-D-Thr-Gly-Phe-Leu-Thr (DTLET) (10 microM) produced a significant inhibition of the spinal outflow of met-enkephalin-like material. The effect of DAGO, but not that of DTLET, could be prevented by naloxone (10 microM), and, conversely, the effect of DLTET, but not that of DAGO, was no longer observed in the presence of naltrindole (10 microM). Therefore naloxone and naltrindole acted as potent and selective mu and delta antagonists, respectively, when perfused at 10 microM in the intrathecal space of halothane-anesthetized rats. As expected from the lack of a tonic opioid control of spinal enkephalinergic neurones, neither naloxone nor naltrindole alone affected the spontaneous outflow of met-enkephalin-like material. However, naltrindole, but not naloxone, markedly increased the spinal overflow of met-enkephalin-like material due to intrathecal administration of either porcine calcitonin (10 microM) or the peptidase inhibitors thiorphan (10 microM) plus bestatin (20 microM). These data suggest that delta, but not mu, receptors are involved in a phasic opioid inhibitory control of the release of met-enkephalin-like material in the rat spinal cord.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mu-Selective Opioid Peptides Stimulate Prolactin Release in Lactating Rats

The fact that opiates elicit prolactin secretion is well known. However, we have recently discovered that morphine does not stimulate prolactin release in lactating rats. The physiological basis for this alteration in opiate sensitivity during lactation is not known. Since morphine-induced prolactin secretion in male rats is mediated via the mu opioid receptor subtype, one possible explanation is that mu receptors are down-regulated during lactation. To address this possibility, the effects of mu opioid peptides on prolactin secretion were examined in lactating rats. The presumed mu-selective peptides DAGO ([D-Ala², Me-Phe⁴, Gly-ol⁵]-enkephalin) and PLO-17 ([NMe-Phe³, D-Pro⁴]-morphiceptin) were administered to primiparous lactating rats and the resulting hormone responses measured. Both DAGO and PLO-17 caused a rapid and significant rise in plasma prolactin during lactation. The prolactin-releasing effects of both peptides were naloxone reversible, suggesting involvement of opioid receptors. Moreover, the DAGO-induced secretion of prolactin could be completely abolished by pretreatment with the irreversible mu antagonist β-funaltrexamine. In lactating rats, DAGO and PLO-17 were poor growth hormone-releasing agents, providing further evidence for the mu specificity of these peptides. These results imply that during lactation, as in other reproductive states, mu opioid receptor sites are positively coupled to the prolactin secretory mechanism. Thus, the previously observed inability of morphine to elicit prolactin release in lactating rats cannot be explained on the basis of down-regulation of mu opioid receptors.     

Interaction of beta-funaltrexamine with [3H]cycloFOXY binding in rat brain: further evidence that beta-FNA alkylates the opioid receptor complex.

beta-Funaltrexamine (beta-FNA) is an alkylating derivative of naltrexone. In addition to acting as an irreversible inhibitor of mu-receptor-mediated physiological effects, intracerebroventricular (i.c.v.) administration of beta-FNA to rat attenuates the ability of selective delta receptor antagonists and naloxone to reverse delta receptor-mediated effects. Moreover, recent work demonstrated that i.c.v. administration of beta-FNA alters the conformation of the opioid receptor complex, as inferred by a decrease in the Bmax of the lower affinity [3H][D-ala2,D-leu5]enkephalin binding site. Consistent with the decreased potency of naloxone as an inhibitor of delta receptor mediated effects, beta-FNA doubled the naloxone IC50 for displacing [3H][D-ala2,D-leu5]enkephalin from its lower affinity binding site. These data collectively support the hypothesis that the opioid receptor complex postulated to mediate mu-delta interactions in vivo is identical to the opioid receptor complex as defined by vitro ligand binding studies. A direct prediction of this hypothesis is that beta-FNA should increase the Kd of antagonists for the mu binding site (mu cx) of the receptor complex. The data reported in this paper demonstrate that beta-FNA doubled the IC50 of the potent narcotic antagonist, 6-desoxy-6 beta-fluoronaltrexone (cycloFOXY) for displacing [3H][D-ala2,D-leu5]enkephalin from its lower affinity binding site, and doubled the Kd of [3H]cycloFOXY for its mu binding site, providing additional data that the mu binding site labeled by [3H]cycloFOXY is the mu binding site of the opioid receptor complex. beta-FNA also altered the kappa binding site labeled by [3H]cycloFOXY, and when administered intrathecally to mice, beta-FNA produced a longlasting antinociception in the acetic acid writhing test.