Effects of the synthetic cannabinoid receptor agonist JWH-018 on abuse-related effects of opioids in rhesus monkeys.

Opioid abuse remains a public health crisis despite a tremendous outpouring of resources to address the problem. One factor that might complicate this issue is polydrug abuse. While cannabis is increasingly available due to legalization by states, phytocannabinoids do not appear to alter the abuse-related effects of opioids. Synthetic cannabinoids, which are not pharmacologically identical to phytocannabinoids, are also increasingly available, and differences among cannabinoids might affect their interactions with opioids. This study assessed the impact of one synthetic cannabinoid, JWH-018, on the effects of two µ opioid receptor agonists using two procedures that address different aspects of abuse. First, four monkeys could choose to self-administer the opioid remifentanil alone (0.32 µg/kg/infusion) or a mixture containing 0.32 µg/kg/infusion remifentanil and JWH-018 (1-10 µg/kg/infusion). On separate occasions, monkeys could choose between remifentanil available alone or combined with 100 µg/kg/infusion cocaine. While monkeys chose the remifentanil/cocaine mixture over remifentanil alone, they responded equally for remifentanil alone and the remifentanil/JWH-018 mixture. The ability of JWH-018 to reinstate extinguished responding previously maintained by heroin was examined in four other monkeys. When presented with drug-associated stimuli, heroin, but not JWH-018, reinstated responding, and when combined, JWH-018 did not increase the potency of heroin. While opioids and synthetic cannabinoids, including JWH-018, are abused, these results indicate that JWH-018 does not modify the behavioral effects of opioids in monkeys in a manner that would predict greater abuse liability of cannabinoid/opioid mixtures, a result that is consistent with a growing literature on mixtures of opioids and phytocannabinoids.

Effects of noncontingent ethanol, DHEA, and pregnanolone administration on ethanol self-administration in outbred female rats.

Previous research from this laboratory demonstrated that male outbred rats (Long-Evans) can be trained to prefer ethanol (10% v/v) over water during 30-min home-cage sessions and that higher ethanol concentrations (18-32% v/v) can serve as a reinforcer under various operant schedules. Further, we have shown that two neurosteroids, dehydroepiandrosterone (DHEA) and pregnanolone, can readily decrease ethanol self-administration in males. The present study used the same procedures in an attempt to systematically replicate the previous findings in female outbred rats. Rats were first trained to self-administer ethanol in the home cage using a saccharin-fading procedure. Subsequently, a two-bottle preference test was initiated by substituting different ethanol concentrations after subjects reliably consumed 10% ethanol alone. Water was always available during this phase. Next, subjects were transitioned to a fixed-ratio 10 (FR-10) schedule of reinforcement with 0.1 mL of ethanol (18% v/v) serving as the reinforcer so that a concentration-effect curve could be established. Upon completion, subjects were transitioned to an FR-10 FR-20 multiple schedule of ethanol (32% v/v) and food reinforcement to determine whether noncontingent ethanol, DHEA, and pregnanolone could selectively decrease ethanol intake. Not surprisingly, female subjects preferentially consumed ethanol over water at concentrations of 3.2-18% (v/v) during the home-cage procedure, and significantly increased the mean dose of ethanol consumed and blood ethanol concentration (BEC). Similarly, increasing concentrations under an FR-10 schedule significantly increased the dose of ethanol presented and BEC compared to control (water). Finally, under the multiple schedule, noncontingent injections of ethanol (0.32-1.8 g/kg), DHEA (10-100 mg/kg), and pregnanolone (1.8-32 mg/kg) dose-dependently decreased food- and ethanol-maintained responding and the dose of ethanol presented. BEC was significantly decreased by the neurosteroids, but increased by ethanol due to its noncontingent administration. Together, these data replicate only a subset of the data previously obtained in males, suggesting there are sex differences particularly with respect to the effects of DHEA and pregnanolone.

Ventilatory-depressant effects of opioids alone and in combination with cannabinoids in rhesus monkeys.

Pain is a serious health problem that is commonly treated with opioids, although the doses of opioids needed to treat pain are often similar to those that decrease respiration. Combining opioids with drugs that relieve pain through non-opioid mechanisms can decrease the doses of opioids needed for analgesia, resulting in an improved therapeutic window, but only if the doses of opioids that decrease respiration are not similarly decreased. Using small doses of opioids to treat pain has the potential to reduce the number of overdoses and deaths. This study investigated whether the cannabinoid receptor agonists Δ9-tetrahydrocannabinol (Δ9-THC) and CP 55,940 modify the ventilatory-depressant effects of morphine and fentanyl in three monkeys. Ventilatory parameters, including minute volume (VE), were monitored with a head plethysmograph. When given alone, morphine (0.032 - 10 mg/kg) and fentanyl (0.00032 - 0.1 mg/kg) dose dependently decreased VE. Doses of Δ9-THC (1 mg/kg) and CP 55,940 (0.01 mg/kg) that enhance the potency of opioids to produce antinociception modestly decreased ventilation when given alone but did not significantly change morphine or fentanyl dose-effect curves. A larger dose of CP 55,940 (0.032 mg/kg) shifted the fentanyl dose-effect curve downward in two monkeys, without significantly changing the morphine dose-effect curve. In summary, cannabinoid receptor agonists, which increase the potency of opioids to produce antinociception, did not increase their potency to depress ventilation. Thus, the therapeutic window is greater for opioids when they are combined with cannabinoid receptor agonists, indicating a possible advantage for these drug mixtures in treating pain.

Preference for an Opioid/Benzodiazepine Mixture over an Opioid Alone Using a Concurrent Choice Procedure in Rhesus Monkeys.

Increased abuse of opioids is contributing to an escalation in overdose deaths. Benzodiazepines are frequently abused with opioids, possibly because they increase the potency and/or effectiveness of opioids to produce reinforcing effects. This study used a concurrent-choice procedure to determine whether monkeys would choose to self-administer a mixture of the opioid remifentanil and the benzodiazepine midazolam over remifentanil alone. Initially, three monkeys could respond on one lever for saline and on a second lever for either remifentanil alone or midazolam alone. Thereafter, monkeys chose between a dose of remifentanil (0.32 µg/kg/infusion) that did not change and a dose of remifentanil that varied across sessions; for some sessions, midazolam was combined with varying doses of remifentanil. All monkeys received more infusions of remifentanil (0.0032-0.32 µg/kg/infusion) than saline, whereas only two monkeys responded more for midazolam than for saline. When 0.32 µg/kg/infusion remifentanil was available on one lever and a dose of remifentanil that varied across sessions (0.1-1 µg/kg/infusion) was available on the other lever, monkeys chose the larger dose. Combining 3.2 µg/kg/infusion midazolam with 0.32 µg/kg/infusion remifentanil increased responding for the mixture over 0.32 µg/kg/infusion remifentanil alone, although monkeys chose remifentanil alone over mixtures containing smaller doses of remifentanil. When 10 µg/kg/infusion midazolam was combined with 0.1 µg/kg/infusion remifentanil, monkeys chose the mixture over 0.32 µg/kg/infusion remifentanil alone. Thus, monkeys prefer some opioid/benzodiazepine mixtures to larger doses of the opioid alone, suggesting that opioid/benzodiazepine coabuse might be due to increased potency (and possibly effectiveness) of opioids to produce reinforcing effects.

Learning by subtraction: Hippocampal activity and effects of ethanol during the acquisition and performance of response sequences.

Learning is believed to be reflected in the activity of the hippocampus. However, neural correlates of learning have been difficult to characterize because hippocampal activity is integrated with ongoing behavior. To address this issue, male rats (n = 5) implanted with electrodes (n = 14) in the CA1 subfield responded during two tasks within a single test session. In one task, subjects acquired a new 3-response sequence (acquisition), whereas in the other task, subjects completed a well-rehearsed 3-response sequence (performance). Both tasks though could be completed using an identical response topography and used the same sensory stimuli and schedule of reinforcement. More important, comparing neural patterns during sequence acquisition to those during sequence performance allows for a subtractive approach whereby activity associated with learning could potentially be dissociated from the activity associated with ongoing behavior. At sites where CA1 activity was closely associated with behavior, the patterns of activity were differentially modulated by key position and the serial position of a response within the schedule of reinforcement. Temporal shifts between peak activity and responding on particular keys also occurred during sequence acquisition, but not during sequence performance. Ethanol disrupted CA1 activity while producing rate-decreasing effects in both tasks and error-increasing effects that were more selective for sequence acquisition than sequence performance. Ethanol also produced alterations in the magnitude of modulations and temporal pattern of CA1 activity, although these effects were not selective for sequence acquisition. Similar to ethanol, hippocampal micro-stimulation decreased response rate in both tasks and selectively increased the percentage of errors during sequence acquisition, and provided a more direct demonstration of hippocampal involvement during sequence acquisition. Together, these results strongly support the notion that ethanol disrupts sequence acquisition by disrupting hippocampal activity and that the hippocampus is necessary for the conditioned associations required for sequence acquisition.

Chronic Δ9-Tetrahydrocannabinol during Adolescence Differentially Modulates Striatal CB1 Receptor Expression and the Acute and Chronic Effects on Learning in Adult Rats.

The purpose of this study was to determine whether chronic administration of Δ(9)-tetrahydrocannabinol (THC) during adolescence would (1) modify any sex-specific effects of THC on learning and (2) affect the development of tolerance to THC as an adult. Male and female rats received daily injections of saline or 5.6 mg/kg of THC from postnatal day 35-75, yielding four groups (female/saline, female/THC, male/saline, and male/THC). Rats were then trained on a procedure that assayed both learning and performance behavior and administered 0.32-18 mg/kg of THC acutely as adults (experiment 1). THC produced rate-decreasing and error-increasing effects in both sexes; however, female rats were more sensitive than male rats were to the rate-decreasing effects. Rats were then chronically administered 10 mg/kg of THC (experiment 2). Rats that received THC during adolescence developed tolerance to the rate-decreasing effects more slowly and less completely than did rats that received saline; in addition, females developed tolerance to the error-increasing effects of THC slower than males did. Western blot analysis of brain tissue indicated long-term changes in hippocampal and striatal cannabinoid type-1 receptor (CB1R) levels despite levels that were indistinguishable immediately after chronic treatment during adolescence. Striatal CB1R levels were increased in adult rats that received THC during adolescence; hippocampal CB1R levels varied by sex. In summary, female rats were more sensitive than male rats were to the acute and chronic effects of THC, and chronic administration of THC during adolescence produced long-term changes in CB1R levels that correlated with decreased tolerance development to the rate-decreasing effects of THC.

Hormonal status and age differentially affect tolerance to the disruptive effects of delta-9-tetrahydrocannabinol (Δ(9)-THC) on learning in female rats.

The effects of hormone status and age on the development of tolerance to Δ(9)-THC were assessed in sham-operated (intact) or ovariectomized (OVX) female rats that received either intraperitoneal saline or 5.6 mg/kg of Δ(9)-THC daily from postnatal day (PD) 75-180 (early adulthood onward) or PD 35-140 (adolescence onward). During this time, the four groups for each age (i.e., intact/saline, intact/THC, OVX/saline, and OVX/THC) were trained in a learning and performance procedure and dose-effect curves were established for Δ(9)-THC (0.56-56 mg/kg) and the cannabinoid type-1 receptor (CB1R) antagonist rimonabant (0.32-10 mg/kg). Despite the persistence of small rate-decreasing and error-increasing effects in intact and OVX females from both ages during chronic Δ(9)-THC, all of the Δ(9)-THC groups developed tolerance. However, the magnitude of tolerance, as well as the effect of hormone status, varied with the age at which chronic Δ(9)-THC was initiated. There was no evidence of dependence in any of the groups. Hippocampal protein expression of CB1R, AHA1 (a co-chaperone of CB1R) and HSP90ß (a molecular chaperone modulated by AHA-1) was affected more by OVX than chronic Δ(9)-THC; striatal protein expression was not consistently affected by either manipulation. Hippocampal brain-derived neurotrophic factor expression varied with age, hormone status, and chronic treatment. Thus, hormonal status differentially affects the development of tolerance to the disruptive effects of delta-9-tetrahydrocannabinol (Δ(9)-THC) on learning and performance behavior in adolescent, but not adult, female rats. These factors and their interactions also differentially affect cannabinoid signaling proteins in the hippocampus and striatum, and ultimately, neural plasticity.

Comparison of dehydroepiandrosterone (DHEA) and pregnanolone with existing pharmacotherapies for alcohol abuse on ethanol- and food-maintained responding in male rats.

The present study compared two putative pharmacotherapies for alcohol abuse and dependence, dehydroepiandrosterone (DHEA) and pregnanolone, with two Food and Drug Administration (FDA)-approved pharmacotherapies, naltrexone and acamprosate. Experiment 1 assessed the effects of different doses of DHEA, pregnanolone, naltrexone, and acamprosate on both ethanol- and food-maintained responding under a multiple fixed-ratio (FR)-10 FR-20 schedule, respectively. Experiment 2 assessed the effects of different mean intervals of food presentation on responding for ethanol under a FR-10 variable-interval (VI) schedule, whereas Experiment 3 assessed the effects of a single dose of each drug under a FR-10 VI-80 schedule. In Experiment 1, all four drugs dose-dependently decreased response rate for both food and ethanol, although differences in the rate-decreasing effects were apparent among the drugs. DHEA and pregnanolone decreased ethanol-maintained responding more potently than food-maintained responding, whereas the reverse was true for naltrexone. Acamprosate decreased responding for both reinforcers with equal potency. In Experiment 2, different mean intervals of food presentation significantly affected the number of food reinforcers obtained per session; however, changes in the number of food reinforcements did not significantly affect responding for ethanol. Under the FR-10 VI-80 schedule in Experiment 3, only naltrexone significantly decreased both the dose of alcohol presented and blood ethanol concentration (BEC). Acamprosate and pregnanolone had no significant effects on any of the dependent measures, whereas DHEA significantly decreased BEC, but did not significantly decrease response rate or the dose presented. In summary, DHEA and pregnanolone decreased ethanol-maintained responding more potently than food-maintained responding under a multiple FR-10 FR-20 schedule, and were more selective for decreasing ethanol self-administration than either naltrexone or acamprosate under that schedule. Experiment 2 showed that ethanol intake was relatively independent of the interval of reinforcement in the food-maintained component, and Experiment 3 showed that naltrexone was the most effective drug at the doses tested when the interval for food reinforcement was low and maintained under a variable-interval schedule.

Estradiol administration to ovariectomized rats potentiates mephedrone-induced disruptions of nonspatial learning.

Mephedrone (4-methylmethcathinone) has been found in several over-the-counter products that are abused by humans, but very little is known about its behavioral effects and abuse liability. The present study examined the effects of mephedrone (1-10 mg/kg) on learning in female rats, as well as its interaction with the ovarian hormone estradiol. More specifically, female rats were trained to respond under a multiple schedule of repeated acquisition and performance of response sequences and then ovariectomized. Following ovariectomy, mephedrone dose-effect curves were obtained during periods of 17ß-estradiol administration and periods without estradiol administration. Unlike mephedrone, which was administered acutely (i.p.) before the experimental sessions, 17ß-estradiol was administered via subcutaneous Silastic capsules containing 25% 17ß-estradiol and 75% cholesterol. In general, mephedrone produced dose-dependent rate-decreasing and error-increasing effects in the acquisition and performance components of the schedule in all subjects. However, when estradiol was present, three of the four rats were more sensitive to the rate-decreasing effects of mephedrone, and all of the subjects were more sensitive to its error-increasing effects. These data indicate that estradiol can potentiate the disruptive effects of mephedrone on both the acquisition and performance of complex behavior in female rats.

Comparison of the behavioral and cardiovascular effects of mephedrone with other drugs of abuse in rats.

RATIONALE: Exceedingly little experimental research exists on the popular recreational drug mephedrone (4-methylmethcathinone) despite clinical reports concerning its behavioral and cardiovascular toxicity. OBJECTIVES: To characterize mephedrone preclinically by examining its capacity to (1) serve as a discriminative stimulus, (2) disrupt the acquisition of response sequences, and (3) disrupt mean arterial pressure (MAP) and heart rate (HR). METHODS AND RESULTS: In one group of subjects that reliably discriminated 3.2 mg/kg of mephedrone from saline (n = 9), substitution tests indicated that stimulants (cocaine, MDMA, and methamphetamine) more closely approximated the mephedrone discriminative stimulus than non-stimulants (fenfluramine, morphine, and phencyclidine), although none fully substituted. In a second group (n = 6), mephedrone (0.56-10 mg/kg, i.p.) dose-dependently decreased response rate and increased errors in both components of a procedure in which subjects either acquired a new response sequence each session (repeated acquisition) or completed the same response sequence each session (performance). Finally, in a third group (n = 12), radio telemetry probes were used to measure the changes in MAP and HR elicited by mephedrone and then compared them to a known stimulant, methamphetamine. In these studies, mephedrone (0.01-9 mg/kg, i.v.) elicited increases in MAP and HR that were very similar to those elicited by methamphetamine (0.01-9 mg/kg, i.v.). The tachycardia and pressor responses to mephedrone (3 mg/kg) were blocked by the ß-blocker atenolol (1 mg/kg, i.v.) and the α1, α2-blocker phentolamine (3 mg/kg, i.v.), respectively. CONCLUSIONS: Mephedrone produces behavioral and cardiovascular responses that are similar to other stimulants; however, differences from the classical stimulants were also apparent.