beta-funaltrexamine and Morphine-Dependence

Tolerance to the pain-relieving effects of opiates limits their clinical use. Although morphine tolerance is associated with desensitization of mu-opioid receptors, the underlying cellular mechanisms are not understood. One problem with the desensitization hypothesis is that acute morphine does not readily desensitize mu-opioid receptors in many cell types. Given that neurons in the periaqueductal gray (PAG) contribute to morphine antinociception and tolerance, an understanding of desensitization in PAG neurons is particularly relevant. Opioid activity in the PAG can be monitored with activation of G-protein-mediated inwardly rectifying potassium (GIRK) currents. The present data show that opioids have a biphasic effect on GIRK currents in morphine tolerant rats. Opioid activation of GIRK currents is initially potentiated in morphine (EC(50)=281 nM) compared to saline (EC(50)=8.8 microM) pretreated rats as indicated by a leftward shift in the concentration-response curve for met-enkephalin (ME)-induced currents. These currents were inhibited by superfusion of the mu-opioid receptor antagonist beta-funaltrexamine (beta-FNA) suggesting that repeated morphine administration enhances agonist stimulation of mu-opioid receptor coupling to G-proteins. Although supersensitivity of mu-opioid receptors in the PAG is counterintuitive to the development of tolerance, peak GIRK currents from tolerant rats desensitized more than currents from saline pretreated rats (56% of peak current after 10 min compared to 15%, respectively). These data indicate that antinociceptive tolerance may be triggered by enhanced agonist potency resulting in increased desensitization of mu-opioid receptors.

Topics: Analgesics, Opioid; Animals; Brain Chemistry; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Tolerance; Enkephalin, Methionine; G Protein-Coupled Inwardly-Rectifying Potassium Channels; Male; Morphine Dependence; Naltrexone; Narcotic Antagonists; Periaqueductal Gray; Rats; Rats, Sprague-Dawley; Receptors, Opioid; Receptors, Opioid, mu

We have recently shown concurrent changes in behavioural responses and c-Fos protein expression in the central nervous system in both naive and morphine-dependent rats after systemic administration of the opioid antagonist naloxone. However, because naloxone acts on the three major types of opioid receptors, the present study aimed at determining, in the same animals, both changes in behaviour and c-Fos-like immunoreactivity after intravenous injection of selective opioid antagonists, such as mu (beta-funaltrexamine, 10 mg/kg), delta (naltrindole, 4 mg/kg) or kappa (nor-binaltorphimine, 5 mg/kg) opioid receptor antagonists, in naive or morphine-dependent rats. In a first experimental series, only beta-funaltrexamine increased c-Fos expression in the eight central nervous system structures examined, whereas no effect was seen after naltrindole or nor-binaltorphimine administration in naive rats. These results suggest a tonic activity in the endogenous opioid peptides acting on mu opioid receptors in normal rats. A second experimental series in morphine-dependent rats showed that beta-funaltrexamine had the highest potency in the induction of classical signs of morphine withdrawal syndrome, as well as the increase in c-Fos expression in the 22 central nervous system structures studied, suggesting a major role of mu opioid receptors in opioid dependence. However, our results also demonstrated that naltrindole and, to a lesser extent, nor-binaltorphimine were able to induce moderate signs of morphine withdrawal and relatively weak c-Fos protein expression in restricted central nervous system structures. Therefore, delta and kappa opioid receptors may also contribute slightly to opioid dependence.

Topics: Animals; Behavior, Animal; Cell Count; Central Nervous System; Gene Expression Regulation; Immunohistochemistry; Male; Morphine; Morphine Dependence; Naltrexone; Narcotic Antagonists; Narcotics; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Substance Withdrawal Syndrome

Two of the synthesized (-)-(1R,5R,9R)-N-homologues (N-but-3-enyl- and N-but-3-ynyl-5,9-dimethyl-2'-hydroxy-6,7-benzomorphan (9, 13)) were found to be about 20 times more potent than morphine in the mouse tail-flick assay (ED(50) = 0.05 mg/kg), and (-)-(1R,5R, 9R)-N-but-2-ynyl-5,9-dimethyl-2'-hydroxy-6,7-benzomorphan ((-)-(1R, 5R,9R)-N-but-2-ynylnormetazocine, 12) was about as potent as the opioid antagonist N-allylnormetazocine (AD(50) in the tail-flick vs morphine assay = 0.3 mg/kg). All of the homologues examined had higher affinity for the kappa-opioid receptor than the mu-receptor except (-)-N-but-2-ynyl-normetazocine (12), which had a kappa/mu ratio = 7.8 and a delta/mu ratio = 118. The (-)-N-2-cyanoethyl (3), -allyl (8), and -but-3-ynyl (13) analogues had good affinity (<10 nM) for delta-opioid receptors. Two homologues in the (+)-(1S,5S,9S)-normetazocine series, N-pent-4-enyl (24) and N-hex-5-enyl (25), were high-affinity and selective sigma(1)-ligands (K(i) = 2 nM, sigma(2)/sigma(1) = 1250, and 1 nM, sigma(2)/sigma(1) = 750, respectively); in contrast, N-allylnormetazocine (22) had relatively poor affinity at sigma(1), and its sigma(1)/sigma(2) ratio was <100.

Topics: Analgesics; Animals; Benzomorphans; Binding, Competitive; Cerebral Cortex; Ligands; Macaca mulatta; Mice; Morphine; Morphine Dependence; Narcotic Antagonists; Pain Measurement; Radioligand Assay; Receptors, Opioid, delta; Receptors, Opioid, kappa; Receptors, Opioid, mu; Stereoisomerism; Structure-Activity Relationship; Substance Withdrawal Syndrome

We examined the locomotor-enhancing action of mu-opioid receptor agonists, such as morphine and [D-Ala2, N-MePhe4, Gly-ol5]enkephalin (DAMGO), and physical dependence on morphine in diabetic and nondiabetic mice. Morphine (5-20 mg/kg, s.c.) and DAMGO (1-4 nmol, i.c.v.) had a dose-dependent locomotor-enhancing effect in both nondiabetic and diabetic mice. The locomotor-enhancing effects of morphine and DAMGO were significantly less in diabetic mice than in nondiabetic mice, and were significantly reduced after pretreatment with either beta-funaltrexamine (20 mg/kg, s.c.), a selective mu-opioid receptor antagonist, or naloxonazine (35 mg/kg, s.c.), a selective mu1-opioid receptor antagonist. Both diabetic and nondiabetic mice were chronically treated with morphine (8-45 mg/kg, s.c.) for 5 days. During this treatment, neither diabetic nor nondiabetic mice showed any signs of toxicity. After morphine treatment, withdrawal was precipitated by injection of naloxone (0.3-10 mg/kg, s.c.). Several withdrawal signs, such as weight loss, diarrhea, ptosis, jumping and body shakes, were observed after naloxone challenge in morphine-dependent nondiabetic mice. Although morphine-dependent diabetic mice showed greater weight loss than nondiabetic mice, the incidence of jumping and body shakes after naloxone challenge in diabetic mice were lower than that in nondiabetic mice. These results suggest that diabetic mice are selectively hyporesponsive to mu1-opioid receptor-mediated locomotor enhancement. Furthermore, diabetes may affect mu1-opioid receptor-mediated naloxone-precipitated signs of withdrawal from physical dependence on morphine.

Topics: Animals; Diabetes Mellitus, Experimental; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Male; Mice; Mice, Inbred ICR; Morphine; Morphine Dependence; Motor Activity; Naloxone; Naltrexone; Receptors, Opioid, mu; Streptozocin

Involvement of supraspinal and spinal mu opioid receptors in the development degree of morphine dependence was estimated by the ED50 values of naloxone (s.c.) required to precipitate withdrawal jumping and diarrhea 72 hr after morphine-pellet implantation. beta-FNA was administered 4 times (24 hr before, just before, 24 and 48 hr after morphine-pellet implantation) by the i.c.v. or i.t. route. beta-FNA (both i.c.v. and i.t. routes) significantly increased the ED50 values of naloxone for jumping and diarrhea and the increase in the ED50 value of naloxone for jumping was much greater than that for diarrhea. I.c.v. administered beta-FNA was more potent in increasing the ED50 value of naloxone for jumping than i.t. administered beta-FNA. On the other hand, i.c.v. administered beta-FNA was equipotent with i.t. administered beta-FNA in increasing the ED50 value of naloxone for diarrhea. These results suggest that both supraspinal and spinal mu opioid receptors are involved in the development of morphine dependence and that supraspinal mu receptors play a more important role than spinal mu receptors in withdrawal jumping which may reflect an excitation of the central nervous system whereas supraspinal and spinal mu receptors have similar importance in withdrawal diarrhea which may reflect an abnormal function of the autonomic nervous system.

Topics: Animals; Brain; Cerebral Ventricles; Diarrhea; Injections, Intraventricular; Injections, Spinal; Male; Mice; Morphine Dependence; Motor Activity; Naloxone; Naltrexone; Narcotic Antagonists; Receptors, Opioid, mu; Spinal Cord; Substance Withdrawal Syndrome

We have previously reported that chicken embryos injected with a single dose of methadone (Meth) on day 3, 7 or 11 of embryogenesis fail to show dependence on day 14, measured as a significant overshoot in motility above baseline after challenge with the opioid antagonist naloxone (Nx). Constant infusion of Meth from day 7 to 14 also failed to produce evidence of dependence on day 14. To address the question of whether the 14-day-old embryo is capable of expressing withdrawal, isobutylmethylxanthine (IBMX), a compound that produces quasi-opioid withdrawal, was injected directly into the embryo, resulting in a significant increase in motility. To determine whether the 14-day-old embryo could also express true opioid withdrawal, the embryos were injected with various doses of Meth or morphine (Morph), followed at different time intervals by injections of varying doses of Nx. A high dose of Morph followed 24 hours later by a low dose of Nx produced evidence of withdrawal, as did a low dose of Meth followed 1 hour later by a higher dose of Nx, U50488H, a selective kappa agonist, had no effect on motility in the 14-day-old embryo, suggesting that the decrease in motility seen after Meth was not mediated by a kappa receptor. Pretreatment with the irreversible mu antagonist, beta-funaltrexamine (B-FNA), blocked the decrease in motility seen after Meth and also prevented the overshoot in motility when Nx was given 1 hour post-Meth. We were also able to demonstrate dependence/withdrawal in the 12-day-old embryo, but higher doses of both Meth and Nx were required.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 1-Methyl-3-isobutylxanthine; 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Animals; Chick Embryo; Methadone; Morphine Dependence; Motor Activity; Naltrexone; Narcotic Antagonists; Opioid-Related Disorders; Pyrrolidines; Receptors, Opioid; Substance Withdrawal Syndrome

The specific, irreversible binding of [3H]beta-funaltrexamine (beta-FNA) to mu opioid receptors in slices from the corpus striatum and midbrain region containing the ventral tegmentum area (VTA) was accomplished by incubation with [3H]beta-FNA followed by a washing procedure with 1 microM unlabeled naltrexone in the washing medium. Regional distribution studies of the specific, irreversible binding of [3H]beta-FNA revealed that slices of striatum and midbrain were richest in mu opioid receptors. Slices of cerebral cortex and medulla-pons had much lower amounts and the cerebellum had virtually no mu opioid receptors. Examination of the kinetics of the specific, irreversible binding of [3H]beta-FNA revealed that although the maximum binding of beta-FNA to striatal and midbrain slices of control and morphine tolerant/dependent mice did not differ, the rate at which beta-FNA associated with the receptor was increased in the slices from morphine tolerant/dependent animals. The increase in association probably occurred at the initial recognition step before alkylation takes place and is attributable to an increase in affinity of mu opioid receptors for beta-FNA in both the striatal and midbrain regions of morphine tolerant/dependent mice. This finding supports and expands earlier conclusions that the affinity of mu opioid receptors for opioid antagonists is increased during the development of opiate tolerance and dependence in mice.

Topics: Animals; Brain Chemistry; Corpus Striatum; Drug Tolerance; In Vitro Techniques; Male; Mesencephalon; Mice; Morphine; Morphine Dependence; Naltrexone; Narcotic Antagonists; Stereoisomerism

The i.c.v. administration of opioid peptides having selectivity for the mu receptor (D-Ala2-NMePhe4-Gly5(ol)enkephalin and FK 33,824) produced effects on the locomotor activity of nondependent and morphine-dependent rats that differed both quantitatively and qualitatively from those effects produced by peptides having selectivity for the delta receptor (D-Ala2-D-Leu5enkephalin and metkephamid) and beta-endorphin, which has similar affinity for both receptors. Peptides selective for the mu receptor: had a biphasic effect on locomotor activity of nondependent rats, inducing an increase at low doses and an initial decrease followed by a later increase at higher doses and had an enhanced stimulant effect on locomotor activity with tolerance to the depressant effect in morphine-dependent rats. Peptides selective for the delta receptor and beta-endorphin: induced only a dose-related increase in the locomotor activity of nondependent rats and had effects on the locomotor activity of morphine-dependent rats that did not differ substantially from those in nondependent rats. Naltrexone (0.1 mg/kg s.c.) and beta-funaltrexamine (5.0 micrograms/rat i.c.v.), an irreversible antagonist, each blocked to a comparable extent the effects of D-Ala2-NMePhe4-Gly5(ol)enkephalin and DAla2-D-Leu5enkephalin on the locomotor activity of nondependent rats. Thus, effects of opioid peptides that act predominantly at mu or delta receptors on locomotor activity cannot be differentiated in nondependent rats by antagonists but can be differentiated in morphine-dependent rats. These results suggest that the depressant and stimulant effects of opioid peptides on locomotor activity are mediated by distinct neuronal sites.

Topics: Animals; beta-Endorphin; D-Ala(2),MePhe(4),Met(0)-ol-enkephalin; Dose-Response Relationship, Drug; Endorphins; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, Leucine; Enkephalin, Leucine-2-Alanine; Enkephalins; Male; Morphine Dependence; Motor Activity; Naltrexone; Rats; Rats, Inbred Strains; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, mu

The opioid mu receptor antagonist beta-funaltrexamine (beta-FNA) blocked the development of physical dependence in rats when infused simultaneously with morphine for 6 days. In addition, beta-FNA given s.c. 24 h prior to the initiation and on day 3 of a 6 day period of morphine infusion in rats reduced the development of physical dependence in a dose-dependent manner. In morphine-dependent rhesus monkeys, beta-FNA precipitated a prompt and long-lasting withdrawal, which was not reversed within 30 h by subsequent injections of morphine. In contrast, naloxone-induced withdrawal lasted approximately 90 min. These results provide further evidence that beta-FNA is a long-acting antagonist of the opioid mu receptor, and that this receptor has a major role in the development of morphine-induced physical dependence.

Topics: Animals; Body Weight; Dose-Response Relationship, Drug; Macaca mulatta; Male; Morphine Dependence; Naltrexone; Narcotic Antagonists; Rats; Rats, Inbred Strains; Species Specificity; Substance Withdrawal Syndrome; Time Factors

We previously demonstrated that the spinal cord is a primary site for the development of morphine-induced tolerance and dependence. In the current investigation, we have determined the significance of mu opioid receptors in the spinal cord in tolerance and dependence induced by systemically administered morphine. Rats surgically implanted with intrathecal (i.t.) catheters were injected i.t. with various doses of beta-funaltrexamine (beta-FNA), a specific irreversible mu opioid receptor antagonist. beta-FNA (i.t.) dose-dependently antagonized morphine-induced analgesia (i.p.) with approximately one-half the potency of beta-chlornaltrexamine, a nonselective irreversible opioid receptor antagonist. Rats injected with saline i.t. 24 h before implanting morphine pellets developed a significant degree of tolerance and dependence 72 h after morphine administration. Tolerance did not develop in similarly treated animals that received i.t. injections of 4.5 nmol beta-FNA. Signs of naloxone-precipitated withdrawal were also significantly antagonized in all instances, except weight loss, in animals pretreated with beta-FNA (i.t.). We conclude that i.t. injections of beta-FNA significantly antagonized the development of tolerance and dependence induced by systemically administered morphine. Therefore, our results indicate that mu opioid receptors within the spinal cord, and probably throughout the central nervous system, play a primary role in morphine-induced tolerance and dependence.

Topics: Analgesia; Animals; Body Weight; Drug Tolerance; Humans; Male; Morphine; Morphine Dependence; Naltrexone; Rats; Rats, Inbred Strains; Receptors, Opioid; Receptors, Opioid, mu; Spinal Cord