Comparison of three DREADD agonists acting on Gq-DREADDs in the ventral tegmental area to alter locomotor activity in tyrosine hydroxylase:Cre male and female rats.

RATIONALE: Despite the increasingly pervasive use of chemogenetic tools in preclinical neuroscience research, the in vivo pharmacology of DREADD agonists remains poorly understood. The pharmacological effects of any ligand acting at receptors, engineered or endogenous, are influenced by numerous factors including potency, time course, and receptor selectivity. Thus, rigorous comparison of the potency and time course of available DREADD ligands may provide an empirical foundation for ligand selection. OBJECTIVES: Compare the behavioral pharmacology of three different DREADD ligands clozapine-N-oxide (CNO), compound 21 (C21), and deschloroclozapine (DCZ) in a locomotor activity assay in tyrosine hydroxylase:cre recombinase (TH:Cre) male and female rats. METHODS: Locomotor activity in nine adult TH:Cre Sprague-Dawley rats (5 female, 4 male) was monitored for two hours following administration of d-amphetamine (vehicle, 0.1-3.2 mg/kg, IP), DCZ (vehicle, 0.32-320 µg/kg, IP), CNO (vehicle, 0.32-10 mg/kg), and C21 (vehicle, 0.1-3.2 mg/kg, IP). Behavioral sessions were conducted twice per week prior to and starting three weeks after bilateral intra-VTA hM3Dq DREADD virus injection. RESULTS: d-Amphetamine significantly increased locomotor activity pre- and post-DREADD virus injection. DCZ, CNO, and C21 did not alter locomotor activity pre-DREADD virus injection. There was no significant effect of DCZ, CNO, and C21 on locomotor activity post-DREADD virus injection; however, large individual differences in both behavioral response and receptor expression were observed. CONCLUSIONS: Large individual variability was observed in both DREADD agonist behavioral effects and receptor expression. These results suggest further basic research would facilitate the utility of these chemogenetic tools for behavioral neuroscience research.

Protein kinetics of superoxide dismutase-1 in familial and sporadic amyotrophic lateral sclerosis.

OBJECTIVE: Accumulation of misfolded superoxide dismutase-1 (SOD1) is a pathological hallmark of SOD1-related amyotrophic lateral sclerosis (ALS) and is observed in sporadic ALS where its role in pathogenesis is controversial. Understanding in vivo protein kinetics may clarify how SOD1 influences neurodegeneration and inform optimal dosing for therapies that lower SOD1 transcripts. METHODS: We employed stable isotope labeling paired with mass spectrometry to evaluate in vivo protein kinetics and concentration of soluble SOD1 in cerebrospinal fluid (CSF) of SOD1 mutation carriers, sporadic ALS participants and controls. A deaminated SOD1 peptide, SDGPVKV, that correlates with protein stability was also measured. RESULTS: In participants with heterozygous SOD1A5V mutations, known to cause rapidly progressive ALS, mutant SOD1 protein exhibited ~twofold faster turnover and ~ 16-fold lower concentration compared to wild-type SOD1 protein. SDGPVKV levels were increased in SOD1A5V carriers relative to controls. Thus, SOD1 mutations impact protein kinetics and stability. We applied this approach to sporadic ALS participants and found that SOD1 turnover, concentration, and SDGPVKV levels are not significantly different compared to controls. INTERPRETATION: These results highlight the ability of stable isotope labeling approaches and peptide deamidation to discern the influence of disease mutations on protein kinetics and stability and support implementation of this method to optimize clinical trial design of gene and molecular therapies for neurological disorders. TRIAL REGISTRATION: Clinicaltrials.gov: NCT03449212.

Preclinical Characterization and Development on NAQ as a Mu Opioid Receptor Partial Agonist for Opioid Use Disorder Treatment.

Mu opioid receptor (MOR) selective antagonists and partial agonists have clinical utility for the treatment of opioid use disorders (OUDs). However, the development of many has suffered due to their poor pharmacokinetic properties and/or rapid metabolism. Our recent efforts to identify MOR modulators have provided 17-cyclopropylmethyl-3,14ß-dihydroxy-4,5α-epoxy-6α-(isoquinoline-3-carboxamido)morphinan (NAQ), a low-efficacy partial agonist, that showed sub-nanomolar binding affinity to the MOR (K i 0.6 nM) with selectivity over the delta opioid receptor (δ/µ 241) and the kappa opioid receptor (κ/µ 48). Its potent inhibition of the analgesic effect of morphine (AD50 0.46 mg/kg) and precipitation of significantly less withdrawal symptoms even at 100-fold greater dose than naloxone represents a promising molecule for further development as a novel OUD therapeutic agent. Therefore, further in vitro and in vivo characterization of its pharmacokinetics and pharmacodynamics properties was conducted to fully understand its pharmaceutical profile. NAQ showed favorable in vitro ADMET properties and no off-target binding to several classes of GPCRs, enzymes, and ion channels. Following intravenous administration, 1 mg/kg dose of NAQ showed a similar in vivo pharmacokinetic profile to naloxone; however, orally administered 10 mg/kg NAQ demonstrated significantly improved oral bioavailability over both naloxone and naltrexone. Abuse liability assessment of NAQ in rats demonstrated that NAQ functioned as a less potent reinforcer than heroin. Chronic 5 day NAQ pretreatment decreased heroin self-administration in a heroin-vs-food choice procedure similar to the clinically used MOR partial agonist buprenorphine. Taken together, these studies provide evidence supporting NAQ as a promising lead to develop novel OUD therapeutics.

Reinforcing effects of fentanyl and sufentanil aerosol puffs in rats.

RATIONALE: Rapidly evolving e-cigarette technology developed for self-administering nicotine aerosol has the potential to be utilized to self-administer other aerosolized drugs of abuse. Rodent models which mirror characteristics of human e-cigarette use are necessary to explore the degree to which this may be a public health concern. OBJECTIVES: Our goal was to develop a highly translational model of discrete nose-only aerosol puff drug delivery to explore the reinforcing effects of fentanyl and sufentanil aerosols in rats. METHODS: Male and female Sprague-Dawley rats were trained to perform a multiple schedule FR1 lever-press, 4-s (second) nose hold operant during which the subject's orofacial areas were exposed to drug-free glycerol/propylene glycol aerosol produced by a commercial e-cigarette at a power setting of 18 watts. Each completed 4-s drug-free vehicle aerosol exposure resulted in a 3-s presentation of a 0.1-ml dipper of sweetened milk solution. After training, rats were then allowed to self-administer 4-s nose-only puffs of fentanyl (100-6000 µg/ml) or sufentanil (30-500 µg/ml) aerosol in the absence of paired milk dipper reinforcers. RESULTS: All 31 rats learned the lever-press/nose-poke multiple schedule for milk dippers alone and 25 accepted exposure to 4 s of 18 watts of drug-free vehicle aerosol when paired with milk dipper presentations. In the absence of paired milk dipper presentations, fentanyl aerosol puffs at concentrations of 1000 and 3000 µg/ml as well as 100 µg/ml puffs of sufentanil served as reinforcers compared to both air puffs and drug-free vehicle aerosol puffs. There were no significant differences between males and females in number of fentanyl or sufentanil puffs self-administered. CONCLUSIONS: Discrete nose-only puffs of two potent opioids under exposure conditions comparable to puff durations in human e-cigarette users serve as reinforcers in rats. This outcome suggests that under appropriate conditions e-cigarettes might be a potential alternative delivery mechanism for illicit opioids.

SARS-CoV-2 PCR and antibody testing for an entire rural community: methods and feasibility of high-throughput testing procedures.

BACKGROUND: Early in the pandemic, inadequate SARS-CoV-2 testing limited understanding of transmission. Chief among barriers to large-scale testing was unknown feasibility, particularly in non-urban areas. Our objective was to report methods of high-volume, comprehensive SARS-CoV-2 testing, offering one model to augment disease surveillance in a rural community. METHODS: A community-university partnership created an operational site used to test most residents of Bolinas, California regardless of symptoms in 4 days (April 20th - April 23rd, 2020). Prior to testing, key preparatory elements included community mobilization, pre-registration, volunteer recruitment, and data management. On day of testing, participants were directed to a testing lane after site entry. An administrator viewed the lane-specific queue and pre-prepared test kits, linked to participants' records. Medical personnel performed sample collection, which included finger prick with blood collection to run laboratory-based antibody testing and respiratory specimen collection for polymerase chain reaction (PCR). RESULTS: Using this 4-lane model, 1,840 participants were tested in 4 days. A median of 57 participants (IQR 47-67) were tested hourly. The fewest participants were tested on day 1 (n = 338 participants), an intentionally lower volume day, increasing to n = 571 participants on day 4. The number of testing teams was also increased to two per lane to allow simultaneous testing of multiple participants on days 2-4. Consistent staffing on all days helped optimize proficiency, and strong community partnership was essential from planning through execution. CONCLUSIONS: High-volume ascertainment of SARS-CoV-2 prevalence by PCR and antibody testing was feasible when conducted in a community-led, drive-through model in a non-urban area.

SARS-CoV-2 PCR and antibody testing for an entire rural community: methods and feasibility of high-throughput testing procedures.

High-volume, community-wide ascertainment of SARS-CoV-2 prevalence by PCR and antibody testing was successfully performed using a community-led, drive-through model with strong operational support, well-trained testing units, and an effective technical platform.

Pharmacological validation of a translational model of cocaine use disorder: Effects of d-amphetamine maintenance on choice between intravenous cocaine and a nondrug alternative in humans and rhesus monkeys.

Drug self-administration procedures are the gold standard for laboratory research to study mechanisms of drug use disorders and evaluate candidate medications. However, preclinical-to-clinical translation has been hampered by a lack of coordination. To address this limitation, we previously developed homologous intravenous (IV) cocaine choice self-administration procedures in rhesus monkeys and humans, and then demonstrated their functional equivalence. The present studies sought to determine the sensitivity of these procedures to d-amphetamine maintenance. Three (N = 3) rhesus monkeys with histories of cocaine self-administration and 16 (N = 16) humans with cocaine use disorder completed the studies. Monkeys were maintained on IV d-amphetamine (0, 0.019, 0.037 and 0.074 mg/kg/h), and then completed 7 sessions during each condition in which they completed 9 choice trials to receive 0.14 mg/kg/injection IV cocaine (corresponding to 10 mg/70 kg in humans) or 10 food pellets under independent, concurrent progressive-ratio schedules. Humans were maintained on oral extended release d-amphetamine (0, 30 and 60 mg/day, corresponding to the lowest 3 doses in monkeys) and participated in 12 sessions in which they chose money ($6.00) or IV cocaine (0, 3, 10 and 30 mg/70 kg). Blood samples were taken to compare d-amphetamine plasma levels across species. In monkeys and humans, d-amphetamine reduced the number of cocaine choices and produced comparable blood levels at equivalent daily doses. d-Amphetamine had similar efficacy, though lower potency, at reducing choice for an equivalent cocaine dose in monkeys relative to humans. These coordinated studies support the utility of these procedures as a translational model for cocaine use disorder. (PsycInfo Database Record (c) 2020 APA, all rights reserved).

NurOwn, phase 2, randomized, clinical trial in patients with ALS: Safety, clinical, and biomarker results.

OBJECTIVE: To determine the safety and efficacy of mesenchymal stem cell (MSC)-neurotrophic factor (NTF) cells (NurOwn®, autologous bone marrow-derived MSCs, induced to secrete NTFs) delivered by combined intrathecal and intramuscular administration to participants with amyotrophic lateral sclerosis (ALS) in a phase 2 randomized controlled trial. METHODS: The study enrolled 48 participants randomized 3:1 (treatment: placebo). After a 3-month pretransplant period, participants received 1 dose of MSC-NTF cells (n = 36) or placebo (n = 12) and were followed for 6 months. CSF was collected before and 2 weeks after transplantation. RESULTS: The study met its primary safety endpoint. The rate of disease progression (Revised ALS Functional Rating Scale [ALSFRS-R] slope change) in the overall study population was similar in treated and placebo participants. In a prespecified rapid progressor subgroup (n = 21), rate of disease progression was improved at early time points (p < 0.05). To address heterogeneity, a responder analysis showed that a higher proportion of treated participants experienced ≥1.5 points/month ALSFRS-R slope improvement compared to placebo at all time points, and was significant in rapid progressors at 4 and 12 weeks (p = 0.004 and 0.046, respectively). CSF neurotrophic factors increased and CSF inflammatory biomarkers decreased in treated participants (p < 0.05) post-transplantation. CSF monocyte chemoattractant protein-1 levels correlated with ALSFRS-R slope improvement up to 24 weeks (p < 0.05). CONCLUSION: A single-dose transplantation of MSC-NTF cells is safe and demonstrated early promising signs of efficacy. This establishes a clear path forward for a multidose randomized clinical trial of intrathecal autologous MSC-NTF cell transplantation in ALS. CLASSIFICATION OF EVIDENCE: This phase II study provides Class I evidence.

Adjusted cost analysis of video televisits for the care of people with amyotrophic lateral sclerosis.

INTRODUCTION: We previously reported our amyotrophic lateral sclerosis (ALS) video televisit experience. Here we report on video televisit versus in-clinic costs, adjusting for perceived medical usefulness (MU). METHODS: We take the patient-perspective and a focused institutional-perspective. Costs are adjusted for patient/caregiver and physician perceptions of visit MU. The base-case reflects our outpatient ALS practice. RESULTS: In the base-case, from the patient perspective, in-clinic visits cost $1,116 and video televisits cost $89 ($119 after MU-adjustment). From the institutional perspective, clinic visits cost $799, and video televisits cost $354 ($472 after MU-adjustment). Adjusted cost-savings per televisit are $997 (patient) and $327 (institution). Sensitivity analyses on 5 variables accounted for uncertainty in base-case assumptions. CONCLUSIONS: Video televisits provide marked adjusted cost-savings for patients and institutions. Adjusted costs are sensitive to perceived MU of video televisits. Future research should explore the ability of video televisits to reduce healthcare resource usage. Muscle Nerve 60: 147-154, 2019.

Additive and subadditive antiallodynic interactions between µ-opioid agonists and N-methyl D-aspartate antagonists in male rhesus monkeys.

µ-Opioid agonists are clinically effective analgesics, but also produce undesirable effects such as sedation and abuse potential that limit their clinical utility. Glutamatergic systems also modulate nociception and N-methyl D-aspartate (NMDA) receptor antagonists have been proposed as one useful adjunct to enhance the therapeutic effects and/or attenuate the undesirable effects of µ-opioid agonists. Whether NMDA antagonists enhance the antiallodynic effects of µ-agonists in preclinical models of thermal hypersensitivity (i.e. capsaicin-induced thermal allodynia) are unknown. The present study determined the behavioral effects of racemic ketamine, (+)-MK-801, (-)-nalbuphine, and (-)-oxycodone alone and in fixed proportion mixtures in assays of capsaicin-induced thermal allodynia and schedule-controlled responding in rhesus monkeys. Ketamine, nalbuphine, and oxycodone produced dose-dependent antiallodynia. MK-801 was inactive up to doses that produced undesirable effects. Ketamine, but not MK-801, enhanced the potency of µ-agonists to decrease rates of operant responding. Ketamine and nalbuphine interactions were additive in both procedures. Ketamine and oxycodone interactions were additive or subadditive depending on the mixture. Furthermore, oxycodone and MK-801 interactions were subadditive on antiallodynia and additive on rate suppression. These results do not support the broad clinical utility of NMDA receptor antagonists as adjuncts to µ-opioid agonists for thermal allodynic pain states.

Development of a translational model to screen medications for cocaine use disorder I: Choice between cocaine and food in rhesus monkeys.

BACKGROUND: Homologous cocaine self-administration procedures in laboratory animals and humans may facilitate translational research for medications development to treat cocaine dependence. This study, therefore, sought to establish choice between cocaine and an alternative reinforcer in rhesus monkeys responding under a procedure back-translated from previous human studies and homologous to a human laboratory procedure described in a companion paper. METHODS: Four rhesus monkeys with chronic indwelling intravenous catheters had access to cocaine injections (0, 0.043, 0.14, or 0.43mg/kg/injection) and food (0, 1, 3, or 10 1g banana-flavored food pellets). During daily 5h sessions, a single cocaine dose and a single food-reinforcer magnitude were available in 10 30-min trials. During the initial "sample" trial, the available cocaine and food reinforcer were delivered non-contingently. During each of the subsequent nine "choice" trials, responding could produce either the cocaine or food reinforcer under an independent concurrent progressive-ratio schedule. RESULTS: Preference was governed by the cocaine dose and food-reinforcer magnitude, and increasing cocaine doses produced dose-dependent increases in cocaine choice at all food-reinforcer magnitudes. Effects of the candidate medication lisdexamfetamine (0.32-3.2mg/kg/day) were then examined on choice between 0.14mg/kg/injection cocaine and 10 pellets. Under baseline conditions, this reinforcer pair maintained an average of approximately 6 cocaine and 3 food choices. Lisdexamfetamine dose-dependently decreased cocaine choice in all monkeys, but food choice was not significantly altered. CONCLUSIONS: These results support utility of this procedure in rhesus monkeys as one component of a platform for translational research on medications development to treat cocaine use disorder.

Context-dependent GluN2B-selective inhibitors of NMDA receptor function are neuroprotective with minimal side effects.

Stroke remains a significant problem despite decades of work on neuroprotective strategies. NMDA receptor (NMDAR) antagonists are neuroprotective in preclinical models, but have been clinically unsuccessful, in part due to side effects. Here we describe a prototypical GluN2B-selective antagonist with an IC50 value that is 10-fold more potent at acidic pH 6.9 associated with ischemic tissue compared to pH 7.6, a value close to the pH in healthy brain tissue. This should maximize neuroprotection in ischemic tissue while minimizing on-target side effects associated with NMDAR blockade in noninjured brain regions. We have determined the mechanism underlying pH-dependent inhibition and demonstrate the utility of this approach in vivo. We also identify dicarboxylate dimers as a novel proton sensor in proteins. These results provide insight into the molecular basis of pH-dependent neuroprotective NMDAR block, which could be beneficial in a wide range of neurological insults associated with tissue acidification.

Pharmacological classification of the abuse-related discriminative stimulus effects of trichloroethylene vapor.

Inhalants are distinguished as a class primarily based upon a shared route of administration. Grouping inhalants according to their abuse-related in vivo pharmacological effects using the drug discrimination procedure has the potential to provide a more relevant classification scheme to the research and treatment community. Mice were trained to differentiate the introceptive effects of the trichloroethylene vapor from air using an operant procedure. Trichloroethylene is a chlorinated hydrocarbon solvent once used as an anesthetic as well as in glues and other consumer products. It is now primarily employed as a metal degreaser. We found that the stimulus effects of trichloroethylene were similar to those of other chlorinated hydrocarbon vapors, the aromatic hydrocarbon toluene and the vapor anesthetics methoxyflurane and isoflurane. The stimulus effects of trichloroethylene overlapped with those of the barbiturate methohexital, to a lesser extent the benzodiazepine midazolam and to ethanol. NMDA antagonists, the kappa opioid agonist U50,488 and the mixed 5-HT agonist mCPP largely failed to substitute for trichloroethylene. These data suggest that stimulus effects of chlorinated hydrocarbon vapors are mediated at least partially by GABAA receptor positive modulatory effects.

GLYX-13, a NMDA receptor glycine-site functional partial agonist, induces antidepressant-like effects without ketamine-like side effects.

Recent human clinical studies with the NMDA receptor (NMDAR) antagonist ketamine have revealed profound and long-lasting antidepressant effects with rapid onset in several clinical trials, but antidepressant effects were preceded by dissociative side effects. Here we show that GLYX-13, a novel NMDAR glycine-site functional partial agonist, produces an antidepressant-like effect in the Porsolt, novelty induced hypophagia, and learned helplessness tests in rats without exhibiting substance abuse-related, gating, and sedative side effects of ketamine in the drug discrimination, conditioned place preference, pre-pulse inhibition and open-field tests. Like ketamine, the GLYX-13-induced antidepressant-like effects required AMPA/kainate receptor activation, as evidenced by the ability of NBQX to abolish the antidepressant-like effect. Both GLYX-13 and ketamine persistently (24 h) enhanced the induction of long-term potentiation of synaptic transmission and the magnitude of NMDAR-NR2B conductance at rat Schaffer collateral-CA1 synapses in vitro. Cell surface biotinylation studies showed that both GLYX-13 and ketamine led to increases in both NR2B and GluR1 protein levels, as measured by Western analysis, whereas no changes were seen in mRNA expression (microarray and qRT-PCR). GLYX-13, unlike ketamine, produced its antidepressant-like effect when injected directly into the medial prefrontal cortex (MPFC). These results suggest that GLYX-13 produces an antidepressant-like effect without the side effects seen with ketamine at least in part by directly modulating NR2B-containing NMDARs in the MPFC. Furthermore, the enhancement of 'metaplasticity' by both GLYX-13 and ketamine may help explain the long-lasting antidepressant effects of these NMDAR modulators. GLYX-13 is currently in a Phase II clinical development program for treatment-resistant depression.

Benzodiazepine-like discriminative stimulus effects of toluene vapor.

In vitro studies show that the abused inhalant toluene affects a number of ligand-gated ion channels.The two most consistently implicated of these are γ-aminobutyric acid type A(GABAA) receptors which are positively modulated by toluene and N-methyl-D-aspartate(NMDA) receptors which are negatively modulated by toluene. Behavioral studies also suggest an interaction of toluene with GABAA and/or NMDA receptors but it is unclear if these receptors underlie the abuse-related intoxicating effects of toluene. Seventeen B6SJLF1/J mice were trained using a two-choice operant drug discrimination procedure to discriminate 10 min of exposure to 2000 ppm toluene vapor from 10 min of exposure to air. The discrimination was acquired in a mean of 65 training sessions. The stimulus effects of 2000 ppm toluene vapor were exposure concentration-dependent but rapidly diminished following the cessation of vapor exposure. The stimulus effects of toluene generalized to the chlorinated hydrocarbon vapor perchloroethylene but not 1,1,2-trichloroethane nor the volatile anesthetic isoflurane. The competitive NMDA antagonist CGS-19755, the uncompetitive antagonist dizocilpine and the glycine-site antagonist L701,324 all failed to substitute for toluene. The classical nonselective benzodiazepines midazolam and chlordiazepoxide produced toluene-like stimulus effects but the alpha 1 subunit preferring positive GABAA modulator zaleplon failed to substitute for toluene. The barbiturates pentobarbital and methohexital and the GABAA positive modulator neurosteroid allopregnanolone did not substitute for toluene. These data suggest that the stimulus effects of toluene may be at least partially mediated by benzodiazepine-like positive allosteric modulation of GABAA receptors containing alpha 2, 3 or 5 subunits.

GABAA-positive modulator selective discriminative stimulus effects of 1,1,1-trichloroethane vapor.

BACKGROUND: The abuse-related behavioral effects of inhalant vapors are poorly understood but probably involve multiple neurotransmitter receptor mechanisms. The present study examined the receptor systems responsible for transducing the discriminative stimulus of the abused chlorinated hydrocarbon 1,1,1-trichloroethane (TCE) in mice. METHODS: Thirty mice were trained to discriminate 10 min of 12,000 ppm TCE vapor exposure from air using an operant procedure. Substitution tests were then conduced with positive GABA(A) receptor modulators and/or NMDA receptor antagonists. RESULTS: The nonselective benzodiazepines midazolam and diazepam produced 62% and 61% and the barbiturate pentobarbital produced 68% TCE-lever selection. Zaleplon, an alpha1 subunit-preferring positive GABA(A) receptor benzodiazepine-site positive modulator resulted in 29% TCE-lever selection. The direct extrasynaptic GABA(A) agonist gaboxodol (THIP) and the GABA reuptake inhibitor tiagabine failed to substitute for TCE. No substitution was elicited by a competitive (CGS-19755), noncompetitive (dizocilpine) or glycine-site (L701,324) NMDA antagonist. The mixed benzodiazepine/noncompetitive NMDA antagonist anesthetic Telazol and the anticonvulsant valproic acid exhibited low levels of partial substitution for TCE (38% and 39%, respectively). Ethanol and nitrous oxide failed to substitute for TCE. CONCLUSIONS: The results suggest that the discriminative stimulus effects of TCE are fairly selectively mediated by positive modulation of GABA(A) receptors. The failure of gaboxadol to substitute and the poor substitution by zaleplon suggests that extrasynaptic GABA(A) receptors as well as GABA(A) receptors containing alpha1 subunits and are not involved in transducing the discriminative stimulus of TCE. Studies with additional GABA(A) benzodiazepine-site positive modulators will be necessary to confirm and extend these findings.

Locomotor activity changes in female adolescent and adult rats during repeated treatment with a cannabinoid or club drug.

Adolescents and young adults of both sexes are the primary consumers of "club" drugs; yet, most of the mechanistic preclinical research in this area has been performed in adult male rodents. The purpose of this study was to evaluate the acute and repeated effects of drugs that are commonly abused by adolescents in female adolescent and adult rats in a rodent model of behavioral sensitization. During two five-day periods separated by a two-day break, rats were injected daily with saline or with one of the following drugs: cocaine (7 or 15 mg/kg), ketamine (3 or 10 mg/kg), 3,4-methylenedioxymethamphetamine (MDMA) (3, 10, or 30 mg/kg), or Δ(9)-tetrahydrocannabinol (THC) (0.03, 0.1, 0.3 or 1 mg/kg) and their locomotor activity was measured. Cocaine increased activity across days in both age groups. Whereas ketamine produced progressive increases in activity with repeated administration in rats of both ages, MDMA increased, and then decreased, activity in the chronic dosing regimen in female adolescents only. Tolerance to the initial stimulatory effects of low doses of THC was observed at both ages. The results with THC are similar to those obtained for male rats tested under identical conditions in a previous study; however, in contrast with the present results in females, male adolescent rats in the previous study failed to develop behavioral sensitization to ketamine. Together, these results suggest that age and sex strongly influence the progressive adaptive changes that occur with repeated administration of some, but not all, of these commonly abused substances.

Characterization of the antinociceptive effects of the individual isomers of methadone after acute and chronic administrations.

Methadone is a long-acting opioid used in the treatment of various pain states and substitution therapy in heroin addiction. Extensive behavioral characterization has been carried out using the racemate, but limited investigation has been performed with the individual isomers. The L-isomer is a potent opioid agonist, whereas the D-isomer has weak µ-opioid activity and has also been shown to possess N-methyl-D-aspartate antagonist properties in vitro. The acute antinociceptive effects of the isomers were evaluated in rats using a warm-water, tail-withdrawal procedure at two stimulus intensities (50° and 55°C). Increasing dose ratios of D-methadone to L-methadone were administered chronically to determine the ability of the D-isomer to modulate antinociceptive tolerance to the L-isomer. Acutely, both L-methadone (0.1-5.6 mg/kg, subcutaneously) and D-methadone (3.0-56.0 mg/kg, subcutaneously) produced antinociception, although the efficacy of the D-isomer was limited at 55°C. These effects were dose dependently blocked by naltrexone (0.01-1.0 mg/kg, subcutaneously). Administered chronically, D-methadone (1.7-10 mg/kg, subcutaneously) dose dependently blocked tolerance development to the L-isomer (1.7 mg/kg, subcutaneously). These findings support the antinociceptive effects of the isomers being opioid receptor mediated with the L-isomer functioning as a full-efficacy agonist, whereas the D-isomer seems to have lower efficacy. The ability of nonracemic doses of the D-isomer to prevent tolerance development to the L-isomer may be attributed to partial µ-agonist activity; however, N-methyl-D-aspartate antagonist activity cannot be discounted.

GABA(A) positive modulator and NMDA antagonist-like discriminative stimulus effects of isoflurane vapor in mice.

RATIONALE: Several neurotransmitter systems have been hypothesized to be involved in the in vivo effects of volatile anesthetics. Drug discrimination may represent a novel procedure to explore the neurochemical systems underlying the sub-anesthetic behavioral effects of these compounds. OBJECTIVES: The purpose of the present study was to examine the contribution of GABA(A) and NMDA receptors to the discriminative stimulus effects of a behaviorally active sub-anesthetic concentration of isoflurane vapor. METHODS: Sixteen B6SJLF1/J mice were trained to discriminate 10 min of exposure to 6,000 ppm isoflurane vapor from air. Substitution tests were conducted with volatile anesthetics, abused vapors, GABA(A) positive modulators, NMDA antagonists, and nitrous oxide. RESULTS: The volatile anesthetics, enflurane and halothane as well as the abused vapors toluene and 1,1,1-trichloroethane fully substituted for isoflurane. The GABA(A) positive modulators, pentobarbital, midazolam, and zaleplon but not the direct GABA(A) agonist, muscimol, produced high levels of partial substitution for isoflurane. The anticonvulsant, valproic acid fully substituted for isoflurane but a second, tiagabine, did not substitute. The competitive NMDA antagonist, CGS-19755, fully and the non-competitive NMDA antagonist, dizocilpine, partially substituted for isoflurane. The glycine-site NMDA antagonist, L-701,324 did not substitute for isoflurane. Gamma-hydroxybutric acid and nitrous oxide gas also failed to substitute for isoflurane. CONCLUSIONS: The discriminative stimulus effects of sub-anesthetic concentrations of isoflurane vapor are shared by other vapor anesthetics and abused inhalants. The discriminative stimulus effects of isoflurane vapor appear to be mediated by both positive allosteric modulation of GABA(A) receptors as well as antagonism of NMDA receptors.