Brain Opioid Binding Research Articles

The sequel to COVID-19: the antithesis to life

The sequel to COVID-19: the antithesis to life

Brain opioid receptor binding in early abstinence from alcohol dependence and relationship to craving: An [11C]diprenorphine PET study

The importance of the opioid receptor system in substance dependence is increasingly recognised. We used PET with the non-selective tracer [11C]diprenorphine to examine opioid receptor binding in early abstinence from alcohol dependence and the relationship to craving. We recruited 11 alcohol dependent patients and 13 controls. Subjects underwent one [11C]diprenorphine PET scan in early abstinence from dependent alcohol use (approximately 2 weeks) and 2 months later if continuously abstinent. Global and regional [11C]diprenorphine volumes of distribution (VD) were increased in alcohol dependent patients compared with controls but did not reach significance. We demonstrated a correlation between global and regional [11C]diprenorphine VD and craving in alcohol dependent patients which persisted in the anterior cingulate cortex into extended abstinence. This confirms previous work showing increased opioid receptor availability in early abstinence from substances of abuse and correlation with craving suggesting that the opioid system plays a fundamental role in this phase of addiction.

Divided Doses for Methadone Maintenance

Divided Doses for Methadone Maintenance

In vivo rat brain opioid receptor binding of LY255582 assessed with a novel method using LC/MS/MS and the administration of three tracers simultaneously

LY255582 is a pan opioid selective receptor antagonist that has been shown to have high affinity for mu, delta, and kappa receptors in vitro. In order to better understand the in vivo opioid receptor selectivity of LY255582, we developed in vivo receptor occupancy assays in the rat for the opioid mu, kappa and delta receptors using the occupancy tracers naltrexone, GR103545 and naltriben respectively. Individual assays for each target were established and then a “triple tracer” assay was created where all three tracers were injected simultaneously, taking advantage of LC/MS/MS technology to selectively monitor brain tracer levels. This is the first report of a technique to concurrently measure receptor specific occupancy at three opioid receptors in the same animal. The opioid subtype selective antagonists cyprodime, JDTic and naltrindole were used to validate selectivity of the assay. Examination of LY255582 in dose-occupancy experiments demonstrated a relative order of potency of mu>kappa>delta, reproducing the previously reported order determined with in vitro binding.

Brain opioid receptor binding in early abstinence from opioid dependence

Although opioid receptor function in humans is clearly reduced during opioid dependence, what happens to the receptor in early abstinence is not understood. This study sought to examine changes in opioid receptor availability in early abstinence from opioid dependence. Ten people with opioid dependence who had completed in-patient detoxification and 20 healthy controls underwent [11C]-diprenorphine positron emission tomography. Clinical variables were assessed with structured questionnaires. Opioid receptor binding was characterised as the volume of distribution of [11C]-diprenorphine using a template of predefined brain volumes and an exploratory voxel-by-voxel analysis. Compared with controls, participants with opioid dependence had increased [11C]-diprenorphine binding in the whole brain and in 15 of the 21 a priori regions studied. This study suggests that opioid receptor binding is increased throughout the brain in early abstinence from dependent opioid use. These data complement the findings in cocaine and alcohol dependence.

Global Increase in Cortical Opioid Binding Potential with Aging: In Vivo Quantification with 11C diprenorphine PET

Background:Recent studies using positron emission tomography (PET) have shown an increase of μ‐opioid receptors with aging in the human brain. However, its physiological role and relationship between the other subtypes of opioid receptors is still obscure. We used a non‐selective radiotracer [11C]diprenorphine and PET to estimate the effects of aging on brain opioid binding. Methods:Seventeen normal male subjects were studied. Their ages ranged from 27 to 65 years (mean; 46 years). The data was analysed with global and regional opioid activities on images reflecting binding potential (Bmax/Kd) using statistical parametric mapping (SPM). Results:The main effect of age was shown in the global opioid binding potential (p=0.046). A post‐hoc test revealed that the binding potentials began to increase abruptly in those aged 40 and above. SPM found that the binding activity of brain opioid receptors significantly increased with age throughout the cerebral cortex except in the primary visual and sensory‐motor cortices where little specific binding occurs. There were no age‐related changes in subcortical structures except for a part of the putamen. Conclusion:The increase of opioid binding activities with aging probably depends on the density of μ‐opioid receptors, not K‐ nor δ‐ opioid receptors in association cortices. The detailed distribution shown by SPM suggests that this phenomenon may not be due to compensatory up‐regulation by the decrease in the endogenous opioid. The Increase of cortical opioid binding may contribute to the pathophysiology of depression and high risk of suicide attempts with advancing age.

Gender differences in opioid-mediated analgesia: animal and human studies.

Gender differences in opioid-mediated analgesia: animal and human studies.

Anti-idiotypic antibodies: a useful alternative for studying the biochemical expression of μ/δ opioid binding sites in mammalian brain

A polyclonal antiserum was produced against opioid binding sites using an anti-idiotypic approach whereby antibodies directed against the opioid agonist DSLET were used as immunogen. The anti-idiotypic antiserum recognized specific brain proteins with molecular masses of 76 ± 4, 73 ± 4 and 59 ± 3 kDa, respectively. The immunolabeling of these proteins was mainly inhibited by μ, δ opioid agonists and a general antagonist, naloxone. The inhibition of immunoprecipitation by opioid agonists and antagonist and the developmental expression of these immunoreactive proteins found to occur during brain ontogeny strongly suggest that these three proteins were μ, δ but not κ opioid binding sites. The anti-idiotypic antiserum both inhibits 3H-DADLE binding and mimics the inhibitory agonist effects on the stimulated cAMP level of NG 108-15 cells which expressed 5 opiate receptors. Numerous mammalian brain opioid binding sites were labeled, due to the fact that the binding site was the epitope recognized by the anti-idiotypic antibodies. From the numerous studies performed with a view to characterizing the specificity of the anti-idiotypic antibodies, it was strongly suggested that the anti-idiotypic antibodies specifically recognize μ/δ opioid binding sites and they can therefore be powerful tools for studying the biochemical expression of these opioid binding sites in mammalian brains.

Social isolation and social interaction alter regional brain opioid receptor binding in rats

Endogenous opioid systems have been implicated in the consequences of social isolation and in the regulation of social behavior, although their precise role is not clear. There is not much information on a possible locus in the brain at which opioids exert their effects on social behavior. In an effort to address this issue we analyzed regional opioidergic activity upon social isolation-induced social interaction using in vivo autoradiography. Animals were either socially isolated for 7 days or group housed, and tested singly or in a dyadic encounter. Subsequently, a tracer dose of [ 3H]diprenorphine was administered and in vivo autoradiographic analysis was performed. Seven days of social isolation caused changes in both social behavior (dyadic encounters) and non-social behavior (singly tested animals). Opioid receptor binding was increased in the medial prefrontal cortex and the parafascicular area in isolates, suggesting that social isolation may evoke an upregulation of opioid receptors in these areas. Social interaction increased opioid binding in the parafascicular area of non-isolated rats. In substantia nigra pars compacta and ventral tegmental area binding was increased upon social isolation, and social interaction decreased opioid binding in isolates, but these changes failed to reach significance. These observed local changes in opioid receptor binding suggest a role for opioid systems in discrete areas in the consequences of social isolation and the regulation of social behavior in rats.

Social play alters regional brain opioid receptor binding in juvenile rats

An in vivo autoradiographic procedure was employed to visualize local changes in brain opioid receptor occupancy in juvenile rats. This procedure is based on the assumption that released endogenous ligand will exclude exogenously applied tracer, in this case [ 3H]diprenorphine, from opioid receptors. Increases in availability of opioid peptides will then result in decreased opioid receptor binding. From behavioral studies there is ample evidence that opioid systems are involved in the regulation of social play behavior in juvenile rats. In the present study, changes in regional brain opioid activity as a result of social isolation-induced social play behavior were monitored. Twenty-one-day-old rats were socially isolated for 0, 3.5 or 24 h prior to testing, and tested alone or in a dyadic encounter. After behavioral testing, [ 3H]diprenorphine was administered and the brain was prepared for autoradiography. Social isolation caused increases in social behavior (dyadic encounters) but not in non-social behavior (singly tested animals). Modest differences in brain opioid receptor binding due to social isolation, social play behavior, or an interaction of the two, were found in claustrum, nucleus accumbens, globus pallidus, paraventricular and arcuate nuclei of the hypothalamus, and the dorsolateral and paratenial thalamic nuclei. These results support the notion that opioid systems are involved in the regulation of social play behavior. In addition, the observation of changes in opioid binding in areas involved in reward processes, adds evidence to the hypothesis that opioid systems are involved in the regulation of the rewarding aspects of social plah in juvenile rats.

Effects of ovarian hormones on brain opioid binding sites in castrated female rats.

These experiments were performed to analyze whether treatments of ovariectomized female rats with ovarian steroid regimens able to induce either an increase (positive feedback effect) or a decrease (negative feedback effect) of serum levels of luteinizing hormone (LH) have some impact on the characteristics of mu-opioid binding sites in circumscribed areas of the brain. The increase of serum levels of LH elicited by a treatment with estradiol benzoate (EB) plus progesterone (P; positive feedback effect) was accompanied by a significant decrease in the number of mu-binding sites in the hypothalamus and in the corpus striatum. The decrease in serum levels of LH induced by a treatment with EB alone (negative feedback effect) brought about a significant increase of the number of mu-binding sites in the thalamus and in the hippocampus. These results seem to suggest that the release of LH induced by EB plus P may involve a decrease of hypothalamic mu-binding sites. Apparently, the inhibitory effect on LH release exerted by EB alone does not involve any change of the density of these binding sites in the hypothalamus.

Effect of adrenal and sex hormones on opioid analgesia and opioid receptor regulation

The role of endocrine factors on opioid analgesia (antinociception) and opioid receptors was studied in male and female Swiss-Webster mice. Morphine was more potent in male than in female mice, although this difference appears to be due to greater availability of morphine to the brain in males. Saturation binding studies indicated that the density and affinity of brain μ- and σ-opioid binding sites were equivalent in males and females. Males and females were implanted SC with naltrexone (NTX) or placebo pellets for 8 days, and then the pellets were removed. This treatment increased the density of μ and σ binding sites in brain and increased the potency of morphine for both sexes, although the increase in antinociceptive effects for males was greater than for females. Adrenalectomy (ADX) in male mice increased the potency of morphine and methadone but did not alter the brain levels of either drug. ADX did not alter brain opioid binding of either μ or σ ligands. When male ADX and control mice were treated with NTX, the potency of morphine and brain opioid binding sites were increased equivalently in both groups. Gonadectomy (GDX) in male mice tended to decrease morphine potency, although this was not found to be a very reliable effect.When male GDX and control mice were implanted with NTX, brain opioid binding was increased similarly in both groups, although morphine potency was increased less in GDX mice. Overall, these studies show that sex differences and hormones of the adrenals and gonads in male mice do not alter brain opioid receptors. Furthermore, these factors do not play a role in NTX-induced opioid receptor upregulation. However, all these factors can modify the potency of opioid agonists.

Dissociation of opioid receptor upregulation and functional supersensitivity

Alterations in brain opioid binding and opioid pharmacodynamics following chronic (8-day) naltrexone (NTX) treatment were determined in pertussis toxin (PTX)-treated mice. Intrathecal (IT) and intracerebroventricular (ICV) PTX produced a time-dependent, long-lasting inhibition of morphine (SC) analgesia without modifying basal nociception. Inhibition was maximal 16 days following PTX treatment, and was still observed at 40 days. Relative to placebo controls, NTX treatment produced supersensitivity to morphine analgesia in all control mice and in mice pretreated with PTX 1 day before NTX. Supersensitivity was not observed in 7-day PTX-pretreated mice. [ 3H]D-Ala 2-D-Leu 5]enkephalin ([ 3H]DADLE) and [ 3H][D-Ala 2-MePhe 4-Gly(ol) 5]enkephalin ([ 3H]DAMGO) binding sites were increased by NTX treatment in saline- and PTX-pretreated groups. K Ds were unchanged. These results indicate that PTX does not alter opioid antagonist-induced receptor upregulation. However, PTX treatment can diminish morphine potency in upregulated and control mice. Therefore, opioid analgesia in control and upregulated mice appears to be mediated by receptors linked to a common PTX-sensitive G-protein. Furthermore, in 7-day PTX-pretreated mice, NTX increased binding sites without altering morphine potency, which suggests that new binding sites can appear without being functionally coupled.

Simultaneous development of opioid tolerance and opioid antagonist-induced receptor upregulation

Mice treated chronically with opioid antagonists have increased receptor density in brain and are supersensitive to the pharmacodynamic action of morphine. In the present study mice were implanted subcutaneously with naltrexone or placebo pellets for 8 days. During implantation mice received daily injections of morphine (100 or 250 mg/kg) or saline. Morphine analgesia was completely blocked in mice that were implanted with naltrexone at the low dose of morphine; while some analgesic action was observed at the higher dose. Mice implanted with placebo were analgesic following the daily morphine treatment. At the end of 8 days the pellets were removed and 24 h later some mice were tested for morphine analgesia while others were examined in binding studies. Naltrexone treatment increased [ 3H]naloxone, 3H[ d-Ala 2- d-Leu 5]enkephalin (DADLE) and 3H[ d-Ala 2,NMePhe 4,Gly-ol 5]enkephalin (DAGO) binding compared to controls and increased the analgesic potency of morphine. Daily treatment with morphine did not alter brain opioid binding or naltrexone-induced receptor upregulation. Mice injected daily with morphine were significantly less sensitive to morphine (tolerant) than their respective saline control group for both the placebo and the naltrexone-treated groups. However, naltrexone-treated mice were more sensitive to morphine than placebo controls regardless of whether they were injected daily with morphine or not. These results indicate that if naltrexone-induced opioid receptor upregulation occurs in the presence of repeated agonist administration, the new binding sites mediate tolerance via desensitization to morphine.

Cholecystokinin antagonists proglumide, lorglumide and benzotript, but not L-364,718, interact with brain opioid binding sites

It has been reported that proglumide and L-364,718 potentiate opioid-induced antinociception. However, their mode of action in pain modulation is still not understood. To evaluate a possible interaction with opioid receptors, we determined the affinities of the CCK antagonists proglumide, lorglumide, benzotript and L-364,718 on mu, delta and kappa binding sites, using guinea pig brain crude synaptosome preparations. These affinities were compared to that of the central CCK binding site, using rat brain slide-mounted sections. At 100uM, proglumide competed for 13% and 17% of mu and kappa binding sites, but did not interact with delta and CCK sites. At this concentration, lorglumide reduced mu, delta, kappa and CCK specific binding by 44%, 69%, 35% and 88%, whereas benzotript diminished it by 16%, 13%, 38% and 48%, respectively. L-364,718 did not interact with opioid receptors (assay limit of solubility, 10 μM) but had a high affinity for CCK binding sites (IC 50, 126nM). The lack of selectivity of proglumide, lorglumide and benzotript for CCK receptors suggests that their reported ability to potentiate morphine analgesia may be related to an interaction with opioid receptors.

Chronic opioid antagonist treatment: assessment of receptor upregulation

Changes in specific brain opioid binding and opioid pharmacodynamics were determined in mice treated with the opioid antagonist naltrexone (subcutaneously implanted pellets) for 8 days. Chronic opioid antagonist treatment increased the number of binding sites (upregulation) for [ 3H]naloxone (+55%) and [ 3H][D-Ala 2,D-Leu 5]enkephalin (+41%) but did not alter the affinity of the ligands, as determined in saturation studies. Displacement studies of [ 3H]naloxone by morphine also indicated that there was no change in morphine's affinity. In vivo estimation of naloxone affinity (pA 2), agreed with the in vitro results indicating that chronic naltrexone treatment did not alter naloxone affinity. Chronic naltrexone treatment (0.5, 1.0, 15.0 mg pellets) increased the analgesic potency of morphine (supersensitivity) in a dose-dependent manner, up to a maximal increase in relative potency of 1.8. However, in mice tested with the naltrexone pellets still implanted, the 15 mg naltrexone pellet was able to shift the dose-response function for morphine analgesia more than 300-fold. The lowest dose naltrexone pellet (0.5 mg), produced significant antagonism of morphine analgesia, but did not produce significant supersensitivity. Thus, supersensitivity and upregulation are not proportional to the degree of antagonism of opioid effects; and supersensitivity in the mouse is related to increased binding sites and not to changes in receptor affinity as determined by in vivo and in vitro methods.

Recovery from opioid receptor alkylation: social conflict analgesia and brain [ 3H]etorphine binding in β-chlornaltrexamine-treated mice

The effects of β-chlornaltrexamine (CNA, 5 mg/kg s.c.) on social conflict analgesia and brain opioid binding were investigated in mice at different times after the administration of the alkylating antagonist. The specific binding of [ 3H]etorphine to high-affinity binding sites and the stress-induced analgesia of attacked mice (50 bites) were prevented for 6 h after CNA administration. Stress-mediated inhibition of pain fully recovered within 3 days after CNA treatment. Brain opioid binding was still reduced to 45% at this time and reached control values 9 days after treatment.

Upregulation of opioid receptor subtypes correlates with potency changes of morphine and dadle

Chronic treatment with opioid antagonists increases the potency of opioid agonists and produces an increase in brain opioid binding sites. In the present study, 8 day treatment with naltrexone blocked morphine and DADLE analgesia for the entire treatment period and increased μ 1, μ 2 and δ opioid receptor binding sites in mouse brain. μ 1 and μ 2 binding were increased by 81 and 67%, respectively, while δ binding was increased by 31%. Consistent with these binding changes, the potency of ICV morphine to produce analgesia was increased by over 3-fold, while the potency of ICV DADLE was increased by only 1.7. These findings indicate that relative increases in opioid receptor subtypes agree with pharmacodynamic studies on potency changes of opioid agonists.

Interaction with lectin of kappa opioid binding sites solubilized from human placenta

Kappa opioid binding sites from human placenta, prelabeled with 3H-etorphine and solubilized, were retained on wheat germ agglutinin (WGA) agarose and specifically eluted with N-acetylglucosamine. No significant retention was found with other immobilized lectins, including Concanavalin A (Con A), soybean seed lectin (SBA), Pisum sativum lectin (PsA), Lens culinaris Medik. lectin (LcA), and Lathyrus tingitanus lectin (LtA). About 23% of applied kappa sites were specifically eluted from WGA agarose, less than half of the proportion of rat brain opioid binding sites eluted from the same lectin (55%). Receptors from placental extracts were compared with those from other tissues enriched in either kappa or mu sites. The proportion of applied kappa sites from guinea pig cerebellum eluted specifically from WGA agarose was 36%, whereas elution of binding sites from rat thalamus and rabbit cerebellum (enriched in mu sites) was at a level of 55%. This difference in the level of retention on and elution from WGA may reflect differences in the sugar composition of the glycoproteins of the two types of receptors. Succinylation of WGA abolished its ability to retain opiod binding sites, consistent with involvement of sialic acid. However, currently available evidence suggests that differences in retention on WGA between kappa and mu sites may be due to differences in either sialic acid or N-acetylglucosamine content or both.

Hippocampal and cortical opioid receptor binding: Changes related to the hibernation state

In vitro opioid receptor binding in the dorsal hippocampal formation and parietal cortex was surveyed in ground squirrels ( Citellus lateralis) in the contrasting physiological states of hibernation and euthermia (i.e. not hibernating). Computer-assisted autoradiographic analysis of coronal sections incubated with [ 3H]dihydromorphine (DMH; 4 nM) revealed statistically significant reductions in specific opioid binding associated with hibernation. In the dorsal hippocampal formation of hibernating animals, binding in the striatum radiatum of CA3, hilus of the dentate gyrus and molecular layer of the dentate gyrus exhibited decreases up to 34% compared to euthermic animals. The stratum radiatum of CA3 exhibited the smallest decrease overall. DHM binding in parietal cortex displayed significant hibernation-related reductions, although they were not uniformly observed across all lamine at the 3 different brain levels examined. These experiments present evidence of changes in brain opioid binding related to the mammalian state of hibernation. The results suggest that changes in opioid receptor binding during hibernation may contribute to the earlier reported 5 apparent failure of morphine physical dependence to develop during hibernation.