Nerve growth factor ß polypeptide (NGFB) genetic variability: association with the methadone dose required for effective maintenance treatment.

Opioid addiction is a chronic disease with high genetic contribution and a large inter-individual variability in therapeutic response. The goal of this study was to identify pharmacodynamic factors that modulate methadone dose requirement. The neurotrophin family is involved in neural plasticity, learning, memory and behavior and deregulated neural plasticity may underlie the pathophysiology of drug addiction. Brain-derived neurotrophic factor (BDNF) was shown to affect the response to methadone maintenance treatment. This study explores the effects of polymorphisms in the nerve growth factor (ß polypeptide) gene, NGFB, on the methadone doses required for successful maintenance treatment for heroin addiction. Genotypes of 14 NGFB polymorphisms were analyzed for association with the stabilizing methadone dose in 72 former severe heroin addicts with no major co-medications. There was significant difference in methadone doses required by subjects with different genotypes of the NGFB intronic single-nucleotide polymorphism rs2239622 (P=0.0002). These results may have clinical importance.

Integrated internist - addiction medicine - hepatology model for hepatitis C management for individuals on methadone maintenance.

Despite a high prevalence of hepatitis C virus (HCV) among drug users, HCV evaluation and treatment acceptance are extremely low among these patients when referred from drug treatment facilities for HCV management. We sought to increase HCV treatment effectiveness among patients from a methadone maintenance treatment program (MMTP) by maintaining continuity of care. We developed, instituted and retrospectively assessed the effectiveness of an integrated, co-localized care model in which an internist-addiction medicine specialist from MMTP was embedded in the hepatitis clinic. Methadone maintenance treatment program patients were referred, evaluated by the internist and hepatologist in hepatitis clinic and provided HCV treatment with integration between both sites. Of 401 evaluated patients, anti-HCV antibody was detected in 257, 86% of whom were older than 40 years. Hepatitis C virus RNA levels were measured in 222 patients, 65 of whom were aviremic. Of 157 patients with detectable HCV RNA, 125 were eligible for referral to the hepatitis clinic, 76 (61%) of whom accepted and adhered with the referral. Men engaged in MMTP <36 months were significantly less likely to be seen in hepatitis clinic than men in MMTP more than 36 months (odds ratio = 7.7; 95% confidence interval 2.6-22.9) or women. We evaluated liver histology in 63 patients, and 83% had moderate to advanced liver disease. Twenty-four patients initiated treatment with 19 completing and 13 (54%) achieving sustained response. In conclusion, integrated care between the MMTP and the hepatitis clinic improves adherence with HCV evaluation and treatment compared to standard referral practices.

Aticaprant (Clinically Developed Kappa-Opioid Receptor Antagonist) Combined With Naltrexone Prevents Alcohol "Relapse" Drinking.

Alcohol relapse is the treatment target for medications development for alcohol dependence. Aticaprant, a selective and short-acting kappa-opioid receptor (KOR) antagonist, has recently been under development for new clinical implications (depression or anhedonia). Recent studies have also found that aticaprant reduces alcohol intake and prevents stress- triggered alcohol seeking in rodents via a KOR-mediated mechanism. Here, we further investigated whether aticaprant alone or in combination with naltrexone (mu-opioid receptor [MOR] antagonist) altered alcohol relapse-like drinking using a mouse alcohol deprivation effect (ADE) paradigm to mimic the relapse episodes in human alcoholics. A long-acting and selective KOR antagonist nor-BNI was used as a reference compound for the effects of the KOR antagonism on the ADE. After 3-week intermittent-access alcohol drinking (two-bottle choice, 24-h access every other day), male and female mice displayed excessive alcohol intake and then pronounced ADE after 1-week abstinence. Aticaprant alone decreased alcohol ADE in a dose- dependent manner (1-3 mg/kg) in both males and females. Aticaprant at a lower dose (0.3 mg/kg) than the effective one (3 mg/kg) combined with a low dose of naltrexone (1 mg/kg) reduced the ADE in both sexes, and the combination was effective after a multi-dosing regimen (5 daily injections during the abstinence) without development of tolerance, suggesting synergistic effects of the combination. In contrast, nor-BNI alone or with naltrexone had no effect on the ADE in either sex. Our present study suggests that a combination of clinically developed, short-acting KOR antagonist aticaprant with low-dose naltrexone has therapeutic potential in alcohol "relapse" treatment.

Association of polymorphisms of the cannabinoid receptor (CNR1) and fatty acid amide hydrolase (FAAH) genes with heroin addiction: impact of long repeats of CNR1.

Alterations in expression of a cannabinoid receptor (CNR1, CB1), and of fatty acid amide hydrolase (FAAH) that degrades endogenous ligands of CB1, may contribute to the development of addiction. The 385C>A in the FAAH gene and six polymorphisms of CNR1 were genotyped in former heroin addicts and control subjects (247 Caucasians, 161 Hispanics, 179 African Americans and 19 Asians). In Caucasians, long repeats (>or=14) of 18087-18131(TAA)(8-17) were associated with heroin addiction (P=0.0102). Across three ethnicities combined, a highly significant association of long repeats with heroin addiction was found (z=3.322, P=0.0009). Point-wise significant associations of allele 1359A (P=0.006) and genotype 1359AA (P=0.034) with protection from heroin addiction were found in Caucasians. Also in Caucasians, the genotype pattern, 1359G>A and -6274A>T, was significantly associated with heroin addiction experiment wise (P=0.0244). No association of FAAH 385C>A with heroin addiction was found in any group studied.

Bidirectional translational research: Progress in understanding addictive diseases.

The focus of this review is primarily on recent developments in bidirectional translational research on the addictions, within the Laboratory of the Biology of Addictive Diseases at The Rockefeller University. This review is subdivided into major interacting aspects, including (a) Investigation of neurobiological and molecular adaptations (e.g., in genes for the opioid receptors or endogenous neuropeptides) in response to cocaine or opiates, administered under laboratory conditions modeling chronic patterns of human self-exposure (e.g., chronic escalating "binge"). (b) The impact of such drug exposure on the hypothalamic-pituitary-adrenal (HPA) axis and interacting neuropeptidergic systems (e.g., opioid, orexin and vasopressin). (c) Molecular genetic association studies using candidate gene and whole genome approaches, to define particular systems involved in vulnerability to develop specific addictions, and response to pharmacotherapy. (d) Neuroendocrine challenge studies in normal volunteers and current addictive disease patients along with former addicts in treatment, to investigate differential pharmacodynamics and responsiveness of molecular targets, in particular those also investigated in the experimental and molecular genetic approaches as described above.

Mu opioid receptor knockdown in the substantia nigra/ventral tegmental area by synthetic small interfering RNA blocks the rewarding and locomotor effects of heroin.

Mu opioid receptors (MOP-r) play an important role in the rewarding and locomotor stimulatory effects of heroin. The aim of the current study was to determine whether infusion of small interfering RNAs (siRNA) targeting MOP-r into the midbrain could knock down MOP-r mRNA and affect heroin-induced locomotor activity or heroin-induced conditioned place preference. Ten-week-old male C57BL/6J mice were surgically implanted bilaterally with guide cannulae directed between the substantia nigra and ventral tegmental area. After 4 days' recovery, mice were infused bilaterally with siRNAs that target the MOP-r (2 mMx0.75 microl/side/day for 3 days) or control siRNA. Seven days after the last infusion, a procedure for conditioned place preference was begun with four heroin (3 mg/kg i.p.) administration sessions alternating with four saline sessions. While heroin induced an increase in locomotor activity in all groups, siRNAs targeting specific regions of MOP-r significantly attenuated this effect. Of particular interest, mice infused with specific siRNAs targeting the MOP-r failed to develop and express conditioned place preference to heroin, or showed a significantly attenuated preference. These alterations in reward-related behaviors are likely due to the reduction in MOP-r mRNA and protein, shown in separate studies by in situ hybridization and autoradiography using the same MOP-r- siRNA infusions. Taken together, these studies demonstrate the utility of siRNA in the neurobiological study of specific components of the reward system and should contribute to the study of other complex behaviors.

Genotype patterns that contribute to increased risk for or protection from developing heroin addiction.

A genome-wide association study was conducted using microarray technology to identify genes that may be associated with the vulnerability to develop heroin addiction, using DNA from 104 individual former severe heroin addicts (meeting Federal criteria for methadone maintenance) and 101 individual control subjects, all Caucasian. Using separate analyses for autosomal and X chromosomal variants, we found that the strongest associations of allele frequency with heroin addiction were with the autosomal variants rs965972, located in the Unigene cluster Hs.147755 (experiment-wise q=0.053), and rs1986513 (q=0.187). The three variants exhibiting the strongest association with heroin addiction by genotype frequency were rs1714984, located in an intron of the gene for the transcription factor myocardin (P=0.000022), rs965972 (P=0.000080) and rs1867898 (P=0.000284). One genotype pattern (AG-TT-GG) was found to be significantly associated with developing heroin addiction (odds ratio (OR)=6.25) and explained 27% of the population attributable risk for heroin addiction in this cohort. Another genotype pattern (GG-CT-GG) of these variants was found to be significantly associated with protection from developing heroin addiction (OR=0.13), and lacking this genotype pattern explained 83% of the population attributable risk for developing heroin addiction. Evidence was found for involvement of five genes in heroin addiction, the genes coding for the mu opioid receptor, the metabotropic receptors mGluR6 and mGluR8, nuclear receptor NR4A2 and cryptochrome 1 (photolyase-like). This approach has identified several new genes potentially associated with heroin addiction and has confirmed the role of OPRM1 in this disease.

Current status of opioid addiction treatment and related preclinical research.

Opioid use disorders (OUDs) are diseases of the brain with behavioral, psychological, neurobiological, and medical manifestations. Vulnerability to OUDs can be affected by factors such as genetic background, environment, stress, and prolonged exposure to µ-opioid agonists for analgesia. Two standard-of-care maintenance medications, methadone and buprenorphine-naloxone, have a long-term positive influence on health of persons with opioid addiction. Buprenorphine and another medication, naltrexone, have also been approved for administration as monthly depot injections. However, neither medication is used as widely as needed, due largely to stigma, insufficient medical education or training, inadequate resources, and inadequate access to treatment. Ongoing directions in the field include (i) personalized approaches leveraging genetic factors for prediction of OUD vulnerability and prognosis, or for targeted pharmacotherapy, and (ii) development of novel analgesic medicines with new neurobiological targets with reduced abuse potential, reduced toxicity, and improved effectiveness, especially for chronic pain states other than cancer pain.

Effects of cocaine place conditioning, chronic escalating-dose "binge" pattern cocaine administration and acute withdrawal on orexin/hypocretin and preprodynorphin gene expressions in lateral hypothalamus of Fischer and Sprague-Dawley rats.

Recent evidence suggests an important role for hypothalamic orexins/hypocretins in modulation of drug reward and addiction-like behaviors in rodents. Our recent study has shown that the aversive state of arousal during acute morphine withdrawal is associated with increased orexin gene expression in lateral hypothalamus (LH) of Fischer 344 (F344) inbred rats, with no change in the expression of preprodynorphin (ppDyn), a gene co-expressed with LH orexin. Therefore, we determined whether orexin and ppDyn mRNA levels in LH or medial hypothalamus (including perifornical and dorsomedial areas) of F344 or Sprague-Dawley (SD) outbred rats, are altered following: 1) cocaine (10 mg/kg, i.p.) conditioned place preference (CPP); 2) chronic (14 days) cocaine exposure using both "binge" pattern administration in steady-dose (45 mg/kg/day) and escalating-dose (45-90 mg/kg/day) regimens; and 3) acute (1 day) and chronic (14 days) withdrawal from cocaine with opioid receptor antagonist naloxone treatment (1 mg/kg). We found that orexin mRNA levels were decreased after cocaine place conditioning in the LH of SD rats. A decreased LH orexin mRNA level was also observed after chronic escalating-dose cocaine (but not CPP pattern regimen without conditioning, or steady-dose regimen) in both strains. In F344 rats only, acute withdrawal from chronic escalating-dose cocaine administration resulted in increases in both LH orexin and ppDyn mRNA levels, which were unaltered by naloxone or after chronic withdrawal. Our results suggest that (1) alteration of LH orexin gene expression is region-specific after cocaine place conditioning in SD rats and dose-dependent after chronic exposure in both strains; and (2) increased LH orexin and ppDyn gene expressions in F344 rats may contribute to negative affective states in cocaine withdrawal.

Steady-state methadone effect on generalized arousal in male and female mice.

Methadone is widely used in treatment of short-acting opiate addiction. The on-off effects of opioids have been documented to have profound differences from steady-state opioids. The authors hypothesize that opioids play important roles in either generalized arousal (GA) or aversive state of arousal during opioid withdrawal. Both male and female C57BL6 mice received steady-state methadone (SSM) through osmotic pumps at 10 or 20 mg/kg/day, and GA was measured in voluntary motor activity, sensory responsivity, and contextual fear conditioning. SSM did not have any effect on those GA behaviors in either sex. Females had higher activity and less fear conditioning than males. The effects of SSM on stress-responsive orexin gene expression in the lateral hypothalamus (LH) and medial hypothalamus (MH, including perifornical and dorsomedial areas) were measured after the behavioral tests. Females showed significantly lower basal LH (but not MH) orexin mRNA levels than males. A panel of GA stressors increased LH orexin mRNA levels in females only; these increases were blunted by SSM at 20 mg/kg. In summary, SSM had no effect on GA behaviors. In females, SSM blunted the GA stress-induced LH orexin gene expression.

Cocaine self-administration in mice is inversely related to phosphorylation at Thr34 (protein kinase A site) and Ser130 (kinase CK1 site) of DARPP-32.

The reinforcing effect of cocaine is associated with increases in dopamine in the striatum. The phosphoprotein DARPP-32 (dopamine- and cAMP-regulated phosphoprotein) has been shown to mediate the intracellular events after activation of dopamine receptors. DARPP-32 is phosphorylated at multiple sites by different protein kinases, but little is known about the functional role of these different sites. Cocaine self-administration and striatal levels of dopamine after acute "binge" cocaine administration were measured in separate lines of mice with alanine mutations introduced into DARPP-32 at either Thr34 (protein kinase A site, Thr34A), Thr75, (cyclin-dependent kinase 5 site, Thr75A), Ser97 (kinase CK2 site, Ser97A), or Ser130 (kinase CK1 site, Ser130A). Acquisition of stable cocaine self-administration required significantly more time in Thr34A-/- mice. Both Thr34A- and Ser130A-DARPP-32 mutant mice self-administered more cocaine than their respective wild-type controls. Also, cocaine-induced increases of dopamine in dorsal striatum were attenuated in the Thr34A- and Ser130A-DARPP-32 phosphomutant mice compared with wild-type mice. Notably, levels of P-Thr34- and P-Ser130-DARPP-32 were reduced after self-administration of cocaine in wild-type mice. Thus, phosphorylation states of Thr34- and Ser130-DARPP-32 play important roles in modulating the reinforcing effects of cocaine.

Structure and expression of the guinea pig preproenkephalin gene: site-specific cleavage in the 3' untranslated region yields truncated mRNA transcripts in specific brain regions.

We isolated the guinea pig preproenkephalin gene from a genomic library by hybridization to a rat cDNA probe. The entire nucleotide sequence of the gene was determined. Genomic Southern blot hybridization demonstrated that the gene exists in a single copy within the genome. On the basis of RNase protection transcript mapping and homology comparisons with known preproenkephalin sequences from other species and assuming a poly(A) tail length of 100 residues, we predicted an mRNA transcript of approximately 1,400 nucleotides encoded by three exons. Northern (RNA) blot analysis of total RNA from several brain regions showed high levels of preproenkephalin mRNA in the caudate putamen, nucleus accumbens, and hypothalamus, with detectable levels in the amygdala, ventral tegmental area, and central gray and also in the pituitary. Unexpectedly, in several brain regions, the mRNA appeared not only in the 1,400-nucleotide length but also in a shorter length of approximately 1,130 bases. Significant amounts of the shorter mRNA were found in the caudate putamen, nucleus accumbens, and amygdala. The longer, but not the shorter, transcripts from the caudate putamen were found to be polyadenylated, but the difference in size was not due solely to the presence of poly(A) tails. Northern gel analysis of total RNA from the caudate putamen with probes from each exon, together with RNase protection mapping of the 3' end of the mRNA demonstrated that the 1,400-base preproenkephalin mRNA transcripts are cleaved in a site-specific manner in some brain regions, yielding a 1,130-base transcript and a 165-base polyadenylated fragment derived from the terminal end of the 3' untranslated region of the mRNA. This cleavage may serve as a preliminary step in RNA degradation and provide a mechanism for control of preproenkephalin mRNA abundance through selective degradation.

Hypothalamic-specific proopiomelanocortin deficiency reduces alcohol drinking in male and female mice.

Opioid receptor antagonist naltrexone reduces alcohol consumption and relapse in both humans and rodents. This study investigated whether hypothalamic proopiomelanocortin (POMC) neurons (producing beta-endorphin and melanocortins) play a role in alcohol drinking behaviors. Both male and female mice with targeted deletion of two neuronal Pomc enhancers nPE1 and nPE2 (nPE-/-), resulting in hypothalamic-specific POMC deficiency, were studied in short-access (4-h/day) drinking-in-the-dark (DID, alcohol in one bottle, intermittent access (IA, 24-h cycles of alcohol access every other day, alcohol vs. water in a two-bottle choice) and alcohol deprivation effect (ADE) models. Wild-type nPE+/+ exposed to 1-week DID rapidly established stable alcohol drinking behavior with more intake in females, whereas nPE-/- mice of both sexes had less intake and less preference. Although nPE-/- showed less saccharin intake and preference than nPE+/+, there was no genotype difference in sucrose intake or preference in the DID paradigm. After 3-week IA, nPE+/+ gradually escalated to high alcohol intake and preference, with more intake in females, whereas nPE-/- showed less escalation. Pharmacological blockade of mu-opioid receptors with naltrexone reduced intake in nPE+/+ in a dose-dependent manner, but had blunted effects in nPE-/- of both sexes. When alcohol was presented again after 1-week abstinence from IA, nPE+/+ of both sexes displayed significant increases in alcohol intake (ADE or relapse-like drinking), with more pronounced ADE in females, whereas nPE-/- did not show ADE in either sex. Our results suggest that neuronal POMC is involved in modulation of alcohol 'binge' drinking, escalation and 'relapse', probably via hypothalamic-mediated mechanisms, with sex differences.

Reliability of salivary cortisol assessments in cocaine dependent individuals.

The aim of this study was to assess the reliability of salivary cortisol as a measure of hypothalmic-pituitary-adrenal (HPA) axis function in cocaine dependent individuals. Saliva and total plasma samples were collected from 49 abusers on 3 testing days in the morning, across eight time points per day. Significant associations between saliva and plasma cortisol were observed across all time points collapsed across 3 days in both men and women. These moderately significant correlations suggest that salivary measurements represent a stable, non-invasive and broad indicator of HPA axis functioning in cocaine dependent individuals.

Corticotropin-releasing factor decreases vasogenic brain edema.

We report the first series of studies comparing the anti-edematous effects of human corticotropin-releasing factor (hCRF) and dexamethasone in an experimental model of vasogenic peritumoral brain edema. Both hCRF and dexamethasone effectively decreased blood-brain barrier (BBB) permeability of intracerebral RG2 gliomas in rats as observed by contrast-enhanced T(1)-weighted magnetic resonance imaging. A decrease in the water content of tumor and peritumoral brain tissue was observed with proton-density magnetic resonance imaging and confirmed by direct wet/dry tissue measurements. The calculated ED(50) for hCRF was 59 micrograms/kg s.c. twice a day, and that for dexamethasone was 0.61 mg/kg i.m. twice a day; the hCRF:dexamethasone dose-potency ratio was 120:1 on a molar basis. The anti-edematous action of hCRF is not mediated by the release of adrenal corticosteroids. A direct action of hCRF on the tumor microvasculature results in restoration of BBB integrity and up-regulation of BBB-specific protein expression. The average survival time with chronic treatment was prolonged significantly in the hCRF-treated group (35 days) compared with the dexamethasone-treated group (28 days; P < 0.05) and the saline-treated control group (22 days; P < 0.0001). hCRF, as an alternative to corticosteroid therapy, may provide substantial benefits with respect to reducing the major side effects encountered with long-term, high-dose corticosteroid treatment.

Amygdalar vasopressin mRNA increases in acute cocaine withdrawal: evidence for opioid receptor modulation.

In humans, stress is recognized as a major factor contributing to relapse to drug abuse in abstinent individuals; drugs of abuse themselves or withdrawal from such drugs act as stressors. In the animals, evidence suggests that centrally released arginine vasopressin in both amygdala and hypothalamus plays an important role in stress-related anxiogenic behaviors. The stress responsive hypothalamic-pituitary-adrenal axis is under tonic inhibition via endogenous opioids, and cocaine withdrawal stimulates hypothalamic-pituitary-adrenal activity. The present studies were undertaken to determine whether: (1) 14-day (chronic) "binge" pattern cocaine administration (45 mg/kg/day) or its withdrawal for 3 h (acute), 1 day (subacute) or 10 days (chronic) alters arginine vasopressin mRNA levels in amygdala or hypothalamus; (2) the opioid receptor antagonist naloxone (1mg/kg) alters arginine vasopressin mRNA or hypothalamic-pituitary-adrenal hormonal responses in acute cocaine withdrawal; and (3) there are associated changes of mu opioid receptor or proopiomelanocortin mRNA levels. In amygdala, arginine vasopressin mRNA levels were unchanged after chronic "binge" cocaine, but were increased during acute cocaine withdrawal. Naloxone completely blocked this increase. Neither chronic cocaine nor its acute withdrawal altered amygdalar mu opioid receptor mRNA levels. The increase in amygdalar arginine vasopressin mRNA levels was still observed after subacute withdrawal, but not after chronic withdrawal. Although hypothalamic-pituitary-adrenal tolerance developed with chronic "binge" cocaine, there were modestly elevated plasma adrenocorticotropin hormone levels during acute withdrawal. While naloxone produced modest adrenocorticotropin hormone elevations in cocaine-naïve rats, naloxone failed to elicit an adrenocorticotropin hormone response in cocaine-withdrawn rats. In hypothalamus, neither chronic cocaine nor acute withdrawal altered arginine vasopressin, proopiomelanocortin or mu opioid receptor mRNA levels. These results show that: (1) opioid receptors mediate increased amygdalar arginine vasopressin gene expression during acute cocaine withdrawal, and (2) cocaine withdrawal renders the hypothalamic-pituitary-adrenal axis insensitive to naloxone. Our findings suggest a potential role for amygdalar arginine vasopressin in the aversive consequences of early cocaine withdrawal.

Persistent increases in rat hypothalamic POMC gene expression following chronic withdrawal from chronic "binge" pattern escalating-dose, but not steady-dose, cocaine.

Recent research suggests an involvement of pro-opiomelanocortin (POMC) gene products (e.g., beta-endorphin) in modulating cocaine-induced reward and addiction-like behaviors in rodents. In this study, we investigated whether chronic "binge" cocaine and its withdrawal altered POMC gene expression in the brain of rats. Male Fischer rats were treated with two different chronic (14-day) "binge" pattern cocaine administration regimens (three injections at 1-h intervals, i.p.): steady-dose (45mg/kg/day) and escalating-dose (90mg/kg on the last day). Although there was no POMC mRNA alteration after chronic steady-dose cocaine, a significant decrease in POMC mRNA levels in the hypothalamus was found after chronic escalating-dose cocaine. In contrast, after acute (1-day) withdrawal from chronic "binge" escalating-dose regimen, but not steady-dose regimen, there were increased hypothalamic POMC mRNA levels that persisted into 14days of protracted withdrawal. To study the role of the endogenous opioid systems in the cocaine withdrawal effects, we administered a single naloxone injection (1mg/kg) that caused elevated POMC mRNA levels observed 24h later in cocaine naïve rats, but it did not lead to further increases in cocaine-withdrawn rats. Our results suggest that during withdrawal from chronic "binge" escalating-dose cocaine: (1) there was a persistent increase in hypothalamic POMC gene expression; and (2) hyposensitivity of the POMC gene expression to naloxone indicates altered opioidergic tone at or above the hypothalamic level.

Self administration of oxycodone alters synaptic plasticity gene expression in the hippocampus differentially in male adolescent and adult mice.

Abuse and addiction to prescription opioids such as oxycodone (a short-acting Mu opioid receptor (MOP-r) agonist) in adolescence is a pressing public health issue. We have previously shown differences in oxycodone self-administration behaviors between adolescent and adult C57BL/6J mice and expression of striatal neurotransmitter receptor genes, in areas involved in reward. In this study, we aimed to determine whether oxycodone self-administration differentially affects genes regulating synaptic plasticity in the hippocampus of adolescent compared to adult mice, since the hippocampus may be involved in learning aspects associated with chronic drug self administration. Hippocampus was isolated for mRNA analysis from mice that had self administered oxycodone (0.25 mg/kg/infusion) 2h/day for 14 consecutive days or from yoked saline controls. Gene expression was analyzed with real-time polymerase chain reaction (PCR) using a commercially available "synaptic plasticity" PCR array containing 84 genes. We found that adolescent and adult control mice significantly differed in the expression of several genes in the absence of oxycodone exposure, including those coding for mitogen-activated protein kinase, calcium/calmodulin-dependent protein kinase II gamma subunit, glutamate receptor, ionotropic AMPA2 and metabotropic 5. Chronic oxycodone self administration increased proviral integration site 1 (Pim1) and thymoma viral proto-oncogene 1 mRNA levels compared to controls in both age groups. Both Pim1 and cadherin 2 mRNAs showed a significant combined effect of Drug Condition and Age × Drug Condition. Furthermore, the mRNA levels of both cadherin 2 and cAMP response element modulators showed an experiment-wise significant difference between oxycodone and saline control in adult but not in adolescent mice. Overall, this study demonstrates for the first time that chronic oxycodone self-administration differentially alters synaptic plasticity gene expression in the hippocampus of adolescent and adult mice.

Beta endorphin levels during heroin, methadone, buprenorphine, and naloxone challenges: preliminary findings.

Beta endorphin (BE) is a polypeptide agonist for the brain's endogenous opioid system. Levels of BE are elevated by opioid antagonists such as naloxone and depressed by short-acting agonists such as heroin and morphine; they become normalized during steady-state methadone. Buprenorphine (BUP) is a partial opioid agonist whose effects on BE levels were examined in six former heroin addicts and 14 methadone-maintained patients before and after being switched to sublingual BUP 2 mg daily for 1 month. In six former methadone-treated subjects BE levels also were measured after stopping BUP and after naloxone challenge. Levels of BE were not significantly lower in subjects started on BUP after stopping heroin (n = 6) (8.0 versus 8.1 ng/ml) or in subjects started on BUP after stopping methadone (n = 14) (11.6 vs 15.6 ng/ml). However, BE levels were lower on BUP than after naloxone challenge (n = 6) (7.0 versus 34.9 ng/ml). Levels of BE did not significantly change between the first 2 weeks ("early") and "later," although BE levels on methadone significantly correlated with BE levels on BUP in the "early" but not the "later" phase. The BE levels on BUP also did not differ from BE levels of unmedicated normals.

Methadone, monoamine oxidase, and depression: opioid distribution and acute effects on enzyme activity.

Narcotic withdrawal is often accompanied by an atypical depression with responds to resumption of narcotics. We hypothesized that methadone might exert its antidepressant effects through monoamine oxidase (MAO) inhibition. The current study examined 3H-methadone distribution in rat brain and effects on regional MAO activity with acute doses (2.5 mg/kg) which approximate those found during chronic methadone maintenance in man. Limbic areas (amygdala, basomedial hypothalamus, caudate-putamen, hippocampus, preoptic nucleus), as well as pituitary and liver were assayed for MAO activity and methadone concentration. MAO activities did not differ significantly in acute methadone or saline-treated cage-mates at 1 or 24 hr. The concentrations of methadone at 1 hr ranged between 17 and 223 ng/100 mg wet wt tissue in the preoptic nucleus and pituitary, respectively. No significant correlation was found between change in MAO activity (MAO methadone/MAO saline) and methadone concentration in any region at 1 or 24 hr. This study does not support the hypothesis that methadone acts as an antidepressant through MAO inhibition, at least not following acute administration of this exogenous opioid.