Behavioral Battery for Testing Candidate Analgesics in Mice. II. Effects of Endocannabinoid Catabolic Enzyme Inhibitors and ∆9-Tetrahydrocannabinol.

Enhanced signaling of the endocannabinoid (eCB) system through inhibition of the catabolic enzymes monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) has received increasing interest for development of candidate analgesics. This study compared effects of MAGL and FAAH inhibitors with effects of ∆9-tetrahydrocannabinol (THC) using a battery of pain-stimulated, pain-depressed, and pain-independent behaviors in male and female mice. Intraperitoneal injection of dilute lactic acid (IP acid) served as an acute visceral noxious stimulus to stimulate two behaviors (stretching, facial grimace) and depress two behaviors (rearing, nesting). Nesting and locomotion were also assessed in the absence of IP acid as pain-independent behaviors. THC and a spectrum of six eCB catabolic enzyme inhibitors ranging from MAGL- to FAAH-selective were assessed for effectiveness to alleviate pain-related behaviors at doses that did not alter pain-independent behaviors. The MAGL-selective inhibitor MJN110 produced the most effective antinociceptive profile, with 1.0 mg/kg alleviating IP acid effects on stretching, grimace, and nesting without altering pain-independent behaviors. MJN110 effects on IP acid-depressed nesting had a slow onset and long duration (40 minutes to 6 hours), were blocked by rimonabant, and tended to be greater in females. As inhibitors increased in FAAH selectivity, antinociceptive effectiveness decreased. PF3845, the most FAAH-selective inhibitor, produced no antinociception up to doses that disrupted locomotion. THC decreased IP acid-stimulated stretching and grimace at doses that did not alter pain-independent behaviors; however, it did not alleviate IP acid-induced depression of rearing or nesting. These results support further consideration of MAGL-selective inhibitors as candidate analgesics for acute inflammatory pain. SIGNIFICANCE STATEMENT: This study characterized a spectrum of endocannabinoid catabolic enzyme inhibitors ranging in selectivity from monoacylglycerol lipase-selective to fatty acid amide hydrolase-selective in a battery of pain-stimulated, pain-depressed, and pain-independent behaviors previously pharmacologically characterized in a companion paper. This battery provides a method for prioritizing candidate analgesics by effectiveness to alleviate pain-related behaviors at doses that do not alter pain-independent behaviors, with inclusion of pain-depressed behaviors increasing translational validity and decreasing susceptibility to motor-depressant false positives.

Behavioral Battery for Testing Candidate Analgesics in Mice. I. Validation with Positive and Negative Controls.

This study evaluated a battery of pain-stimulated, pain-depressed, and pain-independent behaviors for preclinical pharmacological assessment of candidate analgesics in mice. Intraperitoneal injection of dilute lactic acid (IP acid) served as an acute visceral noxious stimulus to produce four pain-related behaviors in male and female ICR mice: stimulation of 1) stretching, 2) facial grimace, 3) depression of rearing, and 4) depression of nesting. Additionally, nesting and locomotion in the absence of the noxious stimulus were used to assess pain-independent drug effects. These six behaviors were used to compare effects of two mechanistically distinct but clinically effective positive controls (ketoprofen and oxycodone) and two negative controls that are not clinically approved as analgesics but produce either general motor depression (diazepam) or motor stimulation (amphetamine). We predicted that analgesics would alleviate all IP acid effects at doses that did not alter pain-independent behaviors, whereas negative controls would not. Consistent with this prediction, ketoprofen (0.1-32 mg/kg) produced the expected analgesic profile, whereas oxycodone (0.32-3.2 mg/kg) alleviated all IP acid effects except depression of rearing at doses lower than those that altered pain-independent behaviors. For the negative controls, diazepam (1-10 mg/kg) failed to block IP acid-induced depression of either rearing or nesting and only decreased IP acid-stimulated behaviors at doses that also decreased pain-independent behaviors. Amphetamine (0.32-3.2 mg/kg) alleviated all IP acid effects but only at doses that also stimulated locomotion. These results support utility of this model as a framework to evaluate candidate-analgesic effects in a battery of complementary pain-stimulated, pain-depressed, and pain-independent behavioral endpoints. SIGNIFICANCE STATEMENT: Preclinical assays of pain and analgesia often yield false-positive effects with candidate analgesics. This study used two positive-control analgesics (ketoprofen, oxycodone) and two active negative controls (diazepam, amphetamine) to validate a strategy for distinguishing analgesics from nonanalgesics by profiling drug effects in a battery of complementary pain-stimulated, pain-depressed, and pain-independent behaviors in male and female mice.

Manipulating Pharmacodynamic Efficacy with Agonist + Antagonist Mixtures: In Vitro and In Vivo Studies with Opioids and Cannabinoids.

Pharmacodynamic efficacy of drugs to activate their receptors is a key determinant of drug effects, and intermediate-efficacy agonists are often useful clinically because they retain sufficient efficacy to produce therapeutically desirable effects while minimizing undesirable effects. Molecular mechanisms of efficacy are not well understood, so rational drug design to control efficacy is not yet possible; however, receptor theory predicts that fixed-proportion mixtures of an agonist and antagonist for a given receptor can be adjusted to precisely control net efficacy of the mixture in activating that receptor. Moreover, the agonist proportion required to produce different effects provides a quantitative scale for comparing efficacy requirements across those effects. To test this hypothesis, the present study evaluated effectiveness of fixed-proportion agonist/antagonist mixtures to produce in vitro and in vivo effects mediated by µ-opioid receptors (MOR) and cannabinoid type 1 receptors (CB1R). Mixtures of 1) the MOR agonist fentanyl and antagonist naltrexone and 2) the CB1R agonist CP55,940 and antagonist/inverse agonist rimonabant were evaluated in an in vitro assay of ligand-stimulated guanosine 5'-O-(3-[35S]thio)triphosphate binding and an in vivo assay of thermal nociception in mice. For both agonist/antagonist pairs in both assays, increasing agonist proportions produced graded increases in maximal mixture effects, and lower agonist proportions were sufficient to produce in vivo than in vitro effects. These findings support the utility of agonist-antagonist mixtures as a strategy to control net efficacy of receptor activation and to quantify and compare efficacy requirements across a range of in vitro and in vivo endpoints. SIGNIFICANCE STATEMENT: Manipulation of agonist proportion in agonist/antagonist mixtures governs net mixture efficacy at the target receptor. Parameters of agonist/antagonist mixture effects can provide a quantitative metric for comparison of efficacy requirements across a wide range of conditions.

Repeated Morphine Produces Sensitization to Reward and Tolerance to Antiallodynia in Male and Female Rats with Chemotherapy-Induced Neuropathy.

Paclitaxel is a cancer chemotherapy drug with adverse effects that include chemotherapy-induced neuropathic pain (CINP) as well as depression of behavior and mood. In the clinical setting, opioids are often used concurrently with or after chemotherapy to treat pain related to the cancer or CINP, but repeated opioid exposure can also increase the risk of opioid abuse. In this study, male and female Sprague-Dawley rats were used to test the hypothesis that repeated 3.2-mg/kg doses of morphine would induce tolerance to its antinociceptive effects in a mechanical sensitivity assay and increased expression of its abuse-related rewarding effects in an assay of intracranial self-stimulation (ICSS). Three weeks after four injections of vehicle or 2.0 mg/kg of paclitaxel, the initial morphine dose-effect curves were determined in both assays. Subsequently, rats were treated with 3.2 mg/kg per day morphine for 6 days. On the final day of testing, morphine dose-effect curves were redetermined in both assays. On initial exposure, morphine produced dose-dependent antiallodynia in the assay of mechanical sensitivity, but it produced little or no rewarding effects in the assay of ICSS. After 6 days of repeated treatment, morphine antiallodynia decreased, and morphine reward increased. Females exhibited greater morphine reward on initial exposure than males, but repeated morphine eliminated this sex difference. These results suggest that repeated morphine may produce tolerance to therapeutically beneficial analgesic effects of morphine but increased sensitivity to abuse-related rewarding effects of morphine in subjects treated with paclitaxel.

Apparent CB1 Receptor Rimonabant Affinity Estimates: Combination with THC and Synthetic Cannabinoids in the Mouse In Vivo Triad Model.

Synthetic cannabinoids (SCs) represent an emerging class of abused drugs associated with psychiatric complications and other substantial health risks. These ligands are largely sold over the internet for human consumption, presumably because of their high cannabinoid 1 receptor (CB1R) affinity and their potency in eliciting pharmacological effects similar to Δ9-tetrahydrocannabinol (THC), as well as circumventing laws illegalizing this plant. Factors potentially contributing to the increased prevalence of SC abuse and related hospitalizations, such as increased CB1R efficacy and non-CB1R targets, highlight the need for quantitative pharmacological analyses to determine receptor mediation of the pharmacological effects of cannabinoids. Accordingly, the present study used pA2 and pKB analyses for quantitative determination of CB1R mediation in which we utilized the CB1R-selective inverse agonist/antagonist rimonabant to elicit rightward shifts in the dose-response curves of five SCs (i.e., A-834,735D; WIN55,212-2; CP55,950; JWH-073; and CP47,497) and THC in producing common cannabimimetic effects (i.e., catalepsy, antinociception, and hypothermia). The results revealed overall similarity of pA2 and pKB values for these compounds and suggest that CB1Rs, and not other pharmacological targets, largely mediated the central pharmacological effects of SCs. More generally, affinity estimation offers a powerful pharmacological approach to assess potential receptor heterogeneity subserving in vivo pharmacological effects of SCs.

Stratification of Cannabinoid 1 Receptor (CB1R) Agonist Efficacy: Manipulation of CB1R Density through Use of Transgenic Mice Reveals Congruence between In Vivo and In Vitro Assays.

Synthetic cannabinoids (SCs) are an emerging class of abused drugs that differ from each other and the phytocannabinoid ∆9-tetrahydrocannabinol (THC) in their safety and cannabinoid-1 receptor (CB1R) pharmacology. As efficacy represents a critical parameter to understanding drug action, the present study investigated this metric by assessing in vivo and in vitro actions of THC, two well-characterized SCs (WIN55,212-2 and CP55,940), and three abused SCs (JWH-073, CP47,497, and A-834,735-D) in CB1 (+/+), (+/-), and (-/-) mice. All drugs produced maximal cannabimimetic in vivo effects (catalepsy, hypothermia, antinociception) in CB1 (+/+) mice, but these actions were essentially eliminated in CB1 (-/-) mice, indicating a CB1R mechanism of action. CB1R efficacy was inferred by comparing potencies between CB1 (+/+) and (+/-) mice [+/+ ED50 /+/- ED50], the latter of which has a 50% reduction of CB1Rs (i.e., decreased receptor reserve). Notably, CB1 (+/-) mice displayed profound rightward and downward shifts in the antinociception and hypothermia dose-response curves of low-efficacy compared with high-efficacy cannabinoids. In vitro efficacy, quantified using agonist-stimulated [35S]GTPγS binding in spinal cord tissue, significantly correlated with the relative efficacies of antinociception (r = 0.87) and hypothermia (r = 0.94) in CB1 (+/-) mice relative to CB1 (+/+) mice. Conversely, drug potencies for cataleptic effects did not differ between these genotypes and did not correlate with the in vitro efficacy measure. These results suggest that evaluation of antinociception and hypothermia in CB1 transgenic mice offers a useful in vivo approach to determine CB1R selectivity and efficacy of emerging SCs, which shows strong congruence with in vitro efficacy.

Opioid-like adverse effects of tianeptine in male rats and mice.

RATIONALE: Tianeptine is a mu-opioid receptor (MOR) agonist with increasing reports of abuse in human populations. Preclinical data regarding the abuse potential and other opioid-like adverse effects of tianeptine at supratherapeutic doses are sparse. OBJECTIVES: The present study evaluated tianeptine in a rat model of abuse potential assessment and in mouse models of motor, gastrointestinal, and respiratory adverse effects. METHODS: Abuse potential was assessed in adult male Sprague-Dawley rats using an intracranial self-stimulation (ICSS) procedure to determine effects of acute and repeated tianeptine on responding for electrical brain stimulation. Male ICR mice were used to determine the effects of tianeptine in assays of locomotor behavior and gastrointestinal motility. Male Swiss-Webster mice were monitored for respiratory changes using whole-body plethysmography. RESULTS: In rats, acute tianeptine produced weak and delayed evidence for abuse-related ICSS facilitation at an intermediate dose (10 mg/kg, IP) and pronounced, naltrexone-preventable ICSS depression at a higher dose (32 mg/kg, IP). Repeated 7-day tianeptine (10 and 32 mg/kg/day, IP) produced no increase in abuse-related ICSS facilitation, only modest tolerance to ICSS depression, and no evidence of physical dependence. In mice, tianeptine produced dose-dependent, naltrexone-preventable locomotor activation. Tianeptine (100 mg/kg, SC) also significantly inhibited gastrointestinal motility and produced naloxone-reversible respiratory depression. CONCLUSIONS: Tianeptine presents as a MOR agonist with resistance to tolerance and dependence in our ICSS assay in rats, and it has lower abuse potential by this metric than many commonly abused opioids. Nonetheless, tianeptine produces MOR agonist-like acute adverse effects that include motor impairment, constipation, and respiratory depression.

Effects of repeated treatment with monoamine-transporter-inhibitor antidepressants on pain-related depression of intracranial self-stimulation in rats.

RATIONALE: Synaptic neurotransmission with dopamine (DA), norepinephrine (NE), and serotonin (5-HT) is terminated primarily by reuptake into presynaptic terminals via the DA, NE, and 5-HT transporters (DAT/NET/SERT, respectively). Monoamine transporter inhibitors constitute one class of drugs used to treat both depression and pain, and therapeutic effects by these compounds often require repeated treatment for days or weeks. OBJECTIVES: The present study compared antinociceptive effects produced by repeated treatment with monoamine transporter inhibitors in a preclinical assay of pain-related depression of positively reinforced operant responding. METHODS: Adult Sprague-Dawley rats equipped with microelectrodes targeting a brain-reward area responded for pulses of electrical brain stimulation in an intracranial self-stimulation (ICSS) procedure. Intraperitoneal injection of dilute lactic acid served as a noxious stimulus that repeatedly depressed ICSS and also produced weight loss during 7 days of repeated acid administration. RESULTS: Acid-induced depression of both ICSS and body weight were completely blocked by repeated pretreatment with the nonsteroidal anti-inflammatory drug ketorolac. The DAT-selective inhibitor bupropion also fully blocked acid-induced ICSS depression and weight loss throughout all 7 days of treatment. The NET-selective inhibitor nortriptyline and the SERT-selective inhibitor citalopram were generally less effective, but both drugs blocked acid-induced ICSS depression by the end of the 7-day treatment. Acid-induced depression of ICSS and body weight were not blocked by the kappa opioid receptor (KOR) agonist U69593 or the KOR antagonist norbinaltorphimine. CONCLUSIONS: These results support effectiveness of bupropion to alleviate signs of pain-related behavioral depression in rats and further suggest that nortriptyline and citalopram produce significant but less reliable effects.

Selective suppression of cocaine- versus food-maintained responding by monoamine releasers in rhesus monkeys: benzylpiperazine, (+)phenmetrazine, and 4-benzylpiperidine.

Monoamine releasers constitute one class of drugs currently under investigation as potential agonist medications for the treatment of cocaine dependence. The efficacy and safety of monoamine releasers as candidate medications may be influenced in part by their relative potency to release dopamine and serotonin, and we reported previously that releasers with approximately 30-fold selectivity for dopamine versus serotonin release may be especially promising. The present study examined the effects of the releasers benzylpiperazine, (+)phenmetrazine, and 4-benzylpiperidine, which have 20- to 48-fold selectivity in vitro for releasing dopamine versus serotonin. In an assay of cocaine discrimination, rhesus monkeys were trained to discriminate 0.4 mg/kg i.m. cocaine from saline in a two-key, food-reinforced procedure. Each of the releasers produced a dose- and time-dependent substitution for cocaine. 4-Benzylpiperidine had the most rapid onset and shortest duration of action. Phenmetrazine and benzylpiperazine had slower onsets and longer durations of action. In an assay of cocaine self-administration, rhesus monkeys were trained to respond for cocaine injections and food pellets under a second order schedule. Treatment for 7 days with each of the releasers produced a dose-dependent and selective reduction in self-administration of cocaine (0.01 mg/kg/injection). The most selective effects were produced by phenmetrazine. Phenmetrazine also produced a downward shift in the cocaine self-administration dose effect curve, virtually eliminating responding maintained by a 30-fold range of cocaine doses (0.0032-0.1 mg/kg/injection) while having only small and transient effects on food-maintained responding. These findings support the potential utility of dopamine-selective releasers as candidate treatments for cocaine dependence.

Effects of the monoamine uptake inhibitors RTI-112 and RTI-113 on cocaine- and food-maintained responding in rhesus monkeys.

Cocaine blocks uptake of the monoamines dopamine, serotonin and norepinephrine, and monoamine uptake inhibitors constitute one class of drugs under consideration as candidate "agonist" medications for the treatment of cocaine abuse and dependence. The pharmacological selectivity of monoamine uptake inhibitors to block uptake of dopamine, serotonin and norepinephrine is one factor that may influence the efficacy and/or safety of these compounds as drug abuse treatment medications. To address this issue, the present study compared the effects of 7-day treatment with a non-selective monoamine uptake inhibitor (RTI-112) and a dopamine-selective uptake inhibitor (RTI-113) on cocaine- and food-maintained responding in rhesus monkeys. Monkeys (N=3) were trained to respond for cocaine injections (0.01 mg/kg/inj) and food pellets under a second-order schedule [FR2(VR16:S)] during alternating daily components of cocaine and food availability. Both RTI-112 (0.0032-0.01 mg/kg/hr) and RTI-113 (0.01-0.056 mg/kg/h) produced dose-dependent, sustained and nearly complete elimination of cocaine self-administration. However, for both drugs, the potency to reduce cocaine self-administration was similar to the potency to reduce food-maintained responding. These findings do not support the hypothesis that pharmacological selectivity to block dopamine uptake is associated with behavioral selectivity to decrease cocaine- vs. food-maintained responding in rhesus monkeys.

Effects of repeated treatment with methcathinone, mephedrone, and fenfluramine on intracranial self-stimulation in rats.

RATIONALE: Synthetic cathinones constitute a class of abused drugs that can act at dopamine, norepinephrine, and serotonin transporters (DAT, NET, and SERT, respectively). Intracranial self-stimulation (ICSS) is a preclinical procedure that can be used to evaluate abuse potential of drugs, and prior studies have indicated that abuse-related ICSS effects of monoamine-transporter substrates, including some synthetic cathinones, are positively correlated with drug selectivity for DAT vs. SERT. Abuse potential of drugs can also be influenced by regimens of repeated drug exposure, but the role of repeated exposure on abuse-related ICSS effects of synthetic cathinones has not been examined. OBJECTIVES: This study used ICSS to evaluate effects of repeated treatment with the DAT>SERT substrate methcathinone, the DAT

Sex differences in abuse-related neurochemical and behavioral effects of 3,4-methylenedioxymethamphetamine (MDMA) in rats.

3,4-Methylenedioxymethamphetamine (MDMA) is a substrate for dopamine (DA), norepinephrine and serotonin (5HT) transporters that produces greater pharmacological effects on certain endpoints in females than males in both clinical and rodent preclinical studies. To evaluate potential for sex differences in abuse-related MDMA effects, the present study compared MDMA effects on intracranial self-stimulation (ICSS) and on in vivo microdialysis measurements of DA or 5HT in the nucleus accumbens (NAc) in female and male Sprague-Dawley rats. For ICSS studies, electrodes were implanted in the medial forebrain bundle and rats trained to press for electrical stimulation over a range of frequencies (56-158Hz, 0.05 log increments) under a fixed-ratio 1 schedule, and the potency (0.32-3.2mg/kg, 10min pretreatment) and time course (3.2. mg/kg, 10-180min pretreatment) of MDMA effects were determined. For in vivo microdialysis, rats were implanted with bilateral guide cannulae targeting the NAc, and the time course of MDMA effects (1.0-3.2mg/kg, 0-180min) on DA and 5HT was determined. MDMA produced qualitatively similar effects in both sexes on ICSS (both increases in low ICSS rates maintained by low brain-stimulation frequencies and decreases in high ICSS rates maintained by high brain-stimulation frequencies) and microdialysis (increases in both DA and 5HT). The duration and peak levels of both abuse-related ICSS facilitation and increases in NAc DA were longer in females. MDMA was also more potent to increase 5HT in females. These results provide evidence for heightened sensitivity of females to abuse-related behavioral and neurochemical effects of MDMA in rats.

Effects of the noncompetitive N-methyl-d-aspartate receptor antagonists ketamine and MK-801 on pain-stimulated and pain-depressed behaviour in rats.

BACKGROUND: Pain is a significant public health concern, and current pharmacological treatments have problematic side effects and limited effectiveness. N-methyl-d-aspartate (NMDA) glutamate receptor antagonists have emerged as one class of candidate treatments for pain because of the significant contribution of glutamate signalling in nociceptive processing. METHODS: This study compared effects of the NMDA receptor antagonists ketamine and MK-801 in assays of pain-stimulated and pain-depressed behaviour in rats. The nonsteroidal anti-inflammatory drug ketoprofen was examined for comparison as a positive control. Intraperitoneal injection of dilute acid served as an acute visceral noxious stimulus to stimulate a stretching response or depress intracranial self-stimulation (ICSS) in male Sprague-Dawley rats. RESULTS: Ketamine (1.0-10.0 mg/kg) blocked acid-stimulated stretching but failed to block acid-induced depression of ICSS, whereas MK-801 (0.01-0.1 mg/kg) blocked both acid-stimulated stretching and acid-induced depression of ICSS. These doses of ketamine and MK-801 did not alter control ICSS in the absence of the noxious stimulus; however, higher doses of ketamine (10 mg/kg) and MK-801 (0.32 mg/kg) depressed all behaviour. Ketoprofen (1.0 mg/kg) blocked both acid-induced stimulation of stretching and depression of ICSS without altering control ICSS. CONCLUSION: These results support further consideration of NMDA receptor antagonists as analgesics; however, some NMDA receptor antagonists are more efficacious at attenuating pain-depressed behaviours. WHAT DOES THIS STUDY ADD: NMDA receptor antagonists produce dissociable effects on pain-depressed behaviour. Provides evidence that pain-depressed behaviours should be considered and evaluated when determining the antinociceptive effects of NMDA receptor antagonists.

Steric parameters, molecular modeling and hydropathic interaction analysis of the pharmacology of para-substituted methcathinone analogues.

BACKGROUND AND PURPOSE: There is growing concern over the abuse of certain psychostimulant methcathinone (MCAT) analogues. This study extends an initial quantitative structure-activity relationship (QSAR) investigation that demonstrated important steric considerations of seven 4- (or para-)substituted analogues of MCAT. Specifically, the steric character (Taft's steric ES ) of the 4-position substituent affected in vitro potency to induce monoamine release via dopamine and 5-HT transporters (DAT and SERT) and in vivo modulation of intracranial self-stimulation (ICSS). Here, we have assessed the effects of other steric properties of the 4-position substituents. EXPERIMENTAL APPROACH: Definitive steric parameters that more explicitly focus on the volume, width and length of the MCAT 4-position substituents were assessed. In addition, homology models of human DAT and human SERT based upon the crystallized Drosophila DAT were constructed and docking studies were performed, followed by hydropathic interaction (HINT) analysis of the docking results. KEY RESULTS: The potency of seven MCAT analogues at DAT was negatively correlated with the volume and maximal width of their 4-position substituents, whereas potency at SERT increased as substituent volume and length increased. SERT/DAT selectivity, as well as abuse-related drug effects in the ICSS procedure, also correlated with the same parameters. Docking solutions offered a means of visualizing these findings. CONCLUSIONS AND IMPLICATIONS: These results suggest that steric aspects of the 4-position substituents of MCAT analogues are key determinants of their action and selectivity, and that the hydrophobic nature of these substituents is involved in their potency at SERT.

Stereoselective Actions of Methylenedioxypyrovalerone (MDPV) To Inhibit Dopamine and Norepinephrine Transporters and Facilitate Intracranial Self-Stimulation in Rats.

The designer stimulant methylenedioxypyrovalerone (MDPV) is a potent reuptake inhibitor at transporters for dopamine (DAT) and norepinephrine (NET) that produces a constellation of abuse-related behavioral effects. MDPV possesses a chiral center, and the abused formulation of the drug is a racemic mixture, but no data are available on the pharmacology of its isomers. Here, the individual optical isomers of MDPV were prepared and examined with respect to their neurochemical actions on neurotransmitter reuptake and behavioral effects in an assay of intracranial self-stimulation (ICSS) in rats. In assays of DAT uptake inhibition, S(+)MDPV (EC50 = 2.13 nM) was more potent than either (±)MDPV (EC50 = 4.85 nM) or R(-)MDPV (EC50 = 382.80 nM); the three drugs were less potent at NET uptake inhibition, with the same rank order of potency. Neither racemic MDPV nor its optical isomers inhibited the reuptake of serotonin at concentrations up to 10 µM. S(+)MDPV produced an abuse-related and dose-dependent facilitation of ICSS, and the potency of S(+)MDPV (significant facilitation at doses ≥ 0.1 mg/kg) was greater than that of the racemate (significant facilitation at doses ≥ 0.32 mg/kg). R(-)MDPV failed to alter ICSS at doses up to 100 times greater than the lowest effective dose of S(+)MDPV. The results indicate that abuse-related neurochemical and behavioral effects of racemic MDPV reside primarily with its S(+) isomer.

Quantitative structure-activity relationship analysis of the pharmacology of para-substituted methcathinone analogues.

BACKGROUND AND PURPOSE: Methcathinone (MCAT) is a potent monoamine releaser and parent compound to emerging drugs of abuse including mephedrone (4-CH3 MCAT), the para-methyl analogue of MCAT. This study examined quantitative structure-activity relationships (QSAR) for MCAT and six para-substituted MCAT analogues on (a) in vitro potency to promote monoamine release via dopamine and serotonin transporters (DAT and SERT, respectively), and (b) in vivo modulation of intracranial self-stimulation (ICSS), a behavioural procedure used to evaluate abuse potential. Neurochemical and behavioural effects were correlated with steric (Es ), electronic (σp ) and lipophilic (πp ) parameters of the para substituents. EXPERIMENTAL APPROACH: For neurochemical studies, drug effects on monoamine release through DAT and SERT were evaluated in rat brain synaptosomes. For behavioural studies, drug effects were tested in male Sprague-Dawley rats implanted with electrodes targeting the medial forebrain bundle and trained to lever-press for electrical brain stimulation. KEY RESULTS: MCAT and all six para-substituted analogues increased monoamine release via DAT and SERT and dose- and time-dependently modulated ICSS. In vitro selectivity for DAT versus SERT correlated with in vivo efficacy to produce abuse-related ICSS facilitation. In addition, the Es values of the para substituents correlated with both selectivity for DAT versus SERT and magnitude of ICSS facilitation. CONCLUSIONS AND IMPLICATIONS: Selectivity for DAT versus SERT in vitro is a key determinant of abuse-related ICSS facilitation by these MCAT analogues, and steric aspects of the para substituent of the MCAT scaffold (indicated by Es ) are key determinants of this selectivity.

Intravenous self-administration studies with l-deprenyl (selegiline) in monkeys.

l-Deprenyl and its stereoisomer d-deprenyl did not maintain intravenous self-administration behavior in rhesus monkeys. In contrast, l-methamphetamine, the major metabolite of l-deprenyl, as well as the baseline drug, cocaine, maintained high rates of intravenous self-administration behavior. Treatment with l-deprenyl doses up to 1.0 mg/kg before self-administration sessions failed to alter self-administration of either cocaine or l-methamphetamine. Thus l-deprenyl did not appear to have cocaine- or methamphetamine-like reinforcing properties in monkeys and was ineffective in altering established patterns of psychomotor-stimulant self-administration behavior. These results support clinical findings that despite long-term use of l-deprenyl for the treatment of Parkinson's disease by large numbers of patients, no instances of abuse have been documented. l-Deprenyl has recently been suggested as a potential medication for the treatment of various types of drug abuse, including cocaine abuse, but its failure to produce selective effects in decreasing cocaine or methamphetamine self-administration behavior in the present experiments makes such an application seem unlikely.

Preclinical evaluation of pharmacotherapies for treatment of cocaine and opioid abuse using drug self-administration procedures.

Drug abuse is a major public health problem, and the relationship between intravenous drug abuse and AIDS underscores the need for more effective treatment medications. Animal models of drug self-administration are useful to systematically evaluate new treatment medications and predict clinical efficacy. This review summarizes the status of preclinical evaluations of medications for treatment of cocaine and opiate abuse. The basic drug self-administration methodology and the rationale for experimental designs and outcome criteria are described. Studies of the effects of dopamine or opioid receptor agonists and antagonists as well as medications used clinically for other indications on drug self-administration are critically examined. Where possible, the degree of concordance between clinical and preclinical studies of drug abuse treatment medications is discussed. We conclude that drug self-administration models are valuable for preclinical assessment of medication efficacy, and we recommend some strategies to further improve evaluation procedures. The discovery of more effective medications for substance abuse treatment should be facilitated by recent advances in behavioral science, pharmacology, neurobiology and medicinal chemistry.

Effects of indatraline and buprenorphine on self-administration of speedball combinations of cocaine and heroin by rhesus monkeys.

The simultaneous intravenous (i.v.) administration of heroin and cocaine, called a "speedball," is often reported clinically, and identification of effective pharmacotherapies is a continuing challenge. We hypothesized that treatment with combinations of a dopamine reuptake inhibitor, indatraline, and a mu partial agonist, buprenorphine, might reduce speedball self-administration by rhesus monkeys more effectively than either drug alone. Speedballs (0.01 mg/kg/inj cocaine + 0.0032 mg/kg/inj heroin) and food (1 g banana pellets) were available in four daily sessions on a second-order schedule of reinforcement [fixed ratio (FR)4; variable ratio (VR)16:S]. Monkeys were treated for 10 days with saline or ascending dose combinations of indatraline (0.001-0.032 mg/kg/day) and buprenorphine (0.00032-0.01 mg/kg/day). Two combinations of indatraline (0.32 and 0.56 mg/kg/day) + buprenorphine (0.10 and 0.18 mg/kg/day) significantly reduced speedball self-administration in comparison to the saline treatment baseline (p <.01-.001), whereas the same doses of each compound alone had no significant effect on speedball-maintained responding. Daily treatment with 0.56 mg/kg/day indatraline + 0.18 mg/kg/day buprenorphine produced a significant downward shift in the speedball dose-effect curve (p <.01) and transient changes in food-maintained responding. These findings suggest that medication mixtures designed to target both the stimulant and opioid component of the speedball combination may be an effective approach to polydrug abuse treatment.