Preclinical assessment of candidate analgesic drugs: recent advances and future challenges.

In analgesic drug development, preclinical procedures are widely used to assess drug effects on pain-related behaviors. These procedures share two principal components: 1) a manipulation intended to produce a pain-like state in the experimental subject and 2) measurement of behaviors presumably indicative of that pain state. Drugs can then be evaluated for their ability to attenuate pain-related behaviors. In the simplest procedures, the pain state is produced by delivery of an acute noxious stimulus (e.g., a warm thermal stimulus), and the primary dependent measures focus on withdrawal responses or other nocifensive behaviors that increase in rate, frequency, or intensity in response to the noxious stimulus. This approach has been refined in two ways. First, new methods have been developed to induce more clinically relevant pain states. In particular, pain requiring clinical intervention is often associated with inflammation or neuropathy, and novel procedures have emerged to model these conditions and their ability to produce hypersensitive pain states, such as allodynia and hyperalgesia. Second, studies are incorporating a broader array of pain-related behaviors as dependent measures. For example, pain not only stimulates nocifensive behaviors but also suppresses many adaptive behaviors, such as feeding or locomotion. Measures of pain-suppressed behaviors can provide new insights into the behavioral consequences of pain and the effects of candidate analgesics. In addition, functional magnetic resonance imaging has emerged as a noninvasive tool for investigating changes in neural activity associated with pain and analgesia. Integration of these complementary approaches may improve the predictive validity of analgesic drug development.

Delta opioid antagonist effects of buprenorphine in rhesus monkeys.

Buprenorphine is an opioid with high affinity for delta, mu and kappa opioid receptors. The delta receptor-mediated effects of buprenorphine have not been studied. Thus, the present study examined the delta receptor-mediated effects of buprenorphine in rhesus monkeys. assays of receptor binding and agonist-stimulated GTP S binding confirmed that buprenorphine had high affinity for, and low efficacy at, delta receptors. In an assay of schedule-controlled responding for food presentation in four monkeys, buprenorphine produced little effect alone, but it antagonized the effects of the delta agonist SNC80, the mu agonist morphine and the kappa agonist U50,488. Buprenorphine was approximately 30-fold less potent as a delta antagonist than as a mu or kappa antagonist. In three monkeys trained to discriminate SNC80 from saline, buprenorphine alone produced only saline-appropriate responding, and buprenorphine pretreatment antagonized the discriminative stimulus effects of SNC80. In a fourth monkey, buprenorphine produced a partial substitution for SNC80 that could be blocked by the delta-selective antagonist naltrindole but not by the mu-selective antagonist quadazocine. These results indicate that, in rhesus monkeys, buprenorphine has very low efficacy at delta receptors, and that buprenorphine produces delta receptor-mediated effects with lower potency than it produces mu or kappa receptor-mediated effects.

Studies of tolerance and dependence with the delta-opioid agonist SNC80 in rhesus monkeys responding under a schedule of food presentation.

Tolerance and dependence after acute or chronic administration of the selective delta-opioid agonist SNC80 were assessed in rhesus monkeys (Macaca mulatta) responding under a schedule of food presentation. SNC80 dose dependently decreased response rates. These effects waned after 5 h. When administered as an acute 24-h pretreatment, SNC80 (1.0-10.0 mg/kg) produced tolerance as evidenced by dose-dependent rightward shifts in the SNC80 dose-effect curve. Pretreatments of 3.2 or 10.0 mg/kg SNC80 increased the SNC80 ED50 by 4- or 25-fold, respectively. Tolerance to acute SNC80 was also time-dependent as evidenced by increased ED50 values when administered as a 5-h (14-fold), 24-h (25-fold), or 3-day (11-fold) pretreatment. The SNC80 dose-effect curve was similar to control after a 7-day pretreatment. The selective delta-antagonist naltrindole (1.0 mg/kg) partially blocked tolerance to acute SNC80. Chronic SNC80 (1.0-10.0 mg/kg/day) also produced dose-dependent rightward shifts in the SNC80 dose-effect curve. Chronic SNC80 was more effective than acute SNC80 in producing tolerance. Moreover, tolerance to chronic SNC80 waned more slowly than to acute SNC80. Acute or chronic SNC80 (10.0 mg/kg/day) also produced cross-tolerance to the rate-decreasing effects of other delta-agonists (SNC162 and SNC243A) but not to mu- (morphine) or kappa (U-50,488)-agonists. Changes in response rates or behavioral signs of withdrawal were not observed after the administration of opioid antagonists (i.e., naltrindole or naltrexone) in monkeys treated with SNC80. These data suggest that a pharmacologically selective tolerance develops to delta-agonists after both acute and chronic administration of SNC80 with little or no dependence.

Antinociceptive effects of delta-opioid agonists in Rhesus monkeys: effects on chemically induced thermal hypersensitivity.

The effects of SNC80 and other structurally related delta-opioid receptor agonists were assessed under conditions of chemically induced hypersensitivity to thermal stimuli in four rhesus monkeys. The shaved tail of each monkey was exposed to warm water (38, 42, 46, and 50 degrees C), and the tail-withdrawal latency from each temperature was recorded. The effects of drugs on the temperature that produced a 10-s tail-withdrawal latency (the T(10) value) were examined. Capsaicin (0.01-0.32 mg) injected into the tail of monkeys dose dependently decreased the T(10), indicating that capsaicin increased sensitivity to thermal stimuli. A dose of 0.1 mg of capsaicin decreased the T(10) from 48.0 to 42.1 degrees C (a -5.9 degrees C change) 15 min after injection. SNC80 (1.0-10.0 mg/kg s.c.) dose dependently blocked the capsaicin-induced decrease in the T(10), and 10.0 mg/kg SNC80 fully blocked the effects of capsaicin. The delta-selective antagonist naltrindole (0.1-1.0 mg/kg) dose dependently antagonized the effects of SNC80, whereas a mu-selective dose of the opioid antagonist quadazocine (0.1 mg/kg) did not. Two other delta-selective agonists, SNC162 (1.0-10.0 mg/kg) and SNC243A (1.0-10.0 mg/kg), also dose dependently blocked capsaicin-induced thermal hypersensitivity. In contrast, neither SNC67 (10.0 mg/kg), which is the (-)-enantiomer of SNC80, nor the nonsteroidal anti-inflammatory drug (NSAID) ketorolac (1.0-10.0 mg/kg) modified the effects of capsaicin. SNC80 was also effective in reversing thermal hypersensitivity induced by prostaglandin E(2) (0.0158 mg) and Freund's complete adjuvant (10% concentration). These findings suggest that delta-agonists have antinociceptive effects in primates under conditions of chemically induced thermal hypersensitivity and might be effective under a broader range of conditions than clinically available NSAIDs.

Chronic l-alpha-acetylmethadol (LAAM) in rhesus monkeys: tolerance and cross-tolerance to the antinociceptive, ventilatory, and rate-decreasing effects of opioids.

Although l-alpha-acetylmethadol (LAAM) is a maintenance treatment for opioid dependence, few studies have systematically assessed the behavioral effects of LAAM and other drugs in LAAM-treated subjects. In the current study, we assessed the ventilatory, antinociceptive, and rate-decreasing effects of drugs (s.c. except dynorphin, which was administered i.v.) in rhesus monkeys (n = 3 or 4) before and during chronic treatment with 1.0 mg/kg/12 h LAAM (s.c.). Minute volume (V(E)) was reduced to 62% of baseline during LAAM treatment and remained depressed after more than 10 months of LAAM treatment. A cumulative dose of 10.0 mg/kg morphine decreased V(E) to similar values under baseline (53%) and LAAM-treated (52%) conditions; however, larger doses of morphine (up to 56.0 mg/kg) could be administered safely to LAAM-treated monkeys. LAAM treatment produced dependence as evidenced by a 220% increase in V(E) after a dose of naltrexone (0.032 mg/kg) that did not modify ventilation under baseline conditions. Compared with baseline, LAAM treatment increased the ED(50) values for the rate-decreasing effects of nalbuphine, morphine, and alfentanil by 7-, 7-, and 2-fold, respectively, in monkeys responding under a fixed ratio 10 schedule of food presentation. Similarly, LAAM treatment increased ED(50) values for the antinociceptive effects of morphine and alfentanil by 5- and 3-fold, respectively. LAAM treatment also increased the ED(50) values for the antinociceptive effects of the kappa-agonist enadoline by 5-fold and not those of U-50,488. That tolerance developed differentially to the ventilatory, rate, and antinociceptive effects of mu-agonists in LAAM-treated monkeys suggests that cross-tolerance might not be a safe therapeutic approach for the treatment of some opioid abusers.

The discriminative stimulus effects of morphine in pigeons responding under a progressive ratio schedule of food presentation.

The purpose of this study was to determine whether progressive ratio (PR) schedules might provide additional information, as compared with fixed ratio or fixed interval schedules, on pharmacologic features of discriminative stimuli (e.g. stimulus intensity). Five pigeons discriminated between 5.6 mg/kg morphine and saline under an arithmetic PR5 schedule of food presentation. The final ratio before pigeons either stopped responding for 5 min or switched responding from the selected to the non-selected key was designated as the last completed ratio (LCR). Pigeons responded 6.8% on the drug key following saline and 96.4% on the drug key following 5.6 mg/kg morphine. The average LCR value for saline was not significantly different from the average LCR value for morphine. A larger dose of morphine (10.0 mg/kg) increased the LCR value and significantly decreased rates of responding. Smaller doses of morphine (0.32 and 1.0 mg/kg) occasioned primarily saline-appropriate responding and decreased LCR values. Buprenorphine substituted for morphine and significantly increased LCR values, whereas nalbuphine produced only partial (20-80%) morphine-key responding and significantly decreased LCR. Cocaine did not substitute for morphine or modify LCR compared with saline control. Together, these results suggest that the stimulus effects of micro-opioids vary on a dimension (e.g. intensity) that can be quantified using PR schedules.

Effects of the long-acting monoamine reuptake inhibitor indatraline on cocaine self-administration in rhesus monkeys.

Cocaine is a nonselective monoamine reuptake inhibitor that is widely abused. Useful pharmacotherapies for cocaine dependence may include substitution medications that produce cocaine-like effects but have a slower onset and longer duration of action. Accordingly, the present study examined the effects of the long-acting, nonselective monoamine reuptake inhibitor indatraline in assays of cocaine discrimination and cocaine self-administration that have been used to evaluate other candidate treatment medications. In rhesus monkeys trained to discriminate cocaine (0.4 mg/kg) from saline, indatraline (0.1-1.0 mg/kg) produced a dose- and time-dependent substitution for cocaine. The effects of 1.0 mg/kg indatraline peaked after 30 min and lasted up to 24 h. In monkeys trained to self-administer 0.032 mg/kg/injection cocaine and food pellets during alternating daily sessions of cocaine and food availability, indatraline (0.0032-0.032 mg/kg/injection) maintained lower rates of responding than cocaine. Repeated treatments with indatraline (0.1-0.56 mg/kg/day) for 7 days produced dose-dependent and sustained decreases in cocaine self-administration across a broad range of cocaine doses (0.0032-0.1 mg/kg/injection), and the highest dose of indatraline (0.56 mg/kg/day) nearly eliminated cocaine-maintained responding. However, doses of indatraline that decreased cocaine self-administration also usually decreased rates of food-maintained responding and produced behavioral stereotypies and trends toward weight loss and mild anemia. These findings suggest that although indatraline may decrease cocaine-taking behavior in rhesus monkeys, it also produces undesirable side effects that may limit its clinical utility in the treatment of cocaine dependence.

Discriminative stimulus effects of the nonpeptidic delta-opioid agonist SNC80 in rhesus monkeys.

Five rhesus monkeys were trained to discriminate the nonpeptidic, delta-opioid agonist SNC80 (0.32 mg/kg i.m.) from saline by using a food-reinforced drug-discrimination procedure. Cumulative doses of SNC80 produced a dose-dependent increase in SNC80-appropriate responding and a dose-dependent decrease in response rate. In time-course studies, peak effects of the training dose of SNC80 were observed after 15 min, and these effects diminished over 240 min. In substitution studies, other piperazinyl benzamide delta agonists (SNC86, SNC162, and SNC243A) substituted for SNC80 with relative potencies similar those of SNC80. However, SNC67, the (-)-enantiomer of SNC80, did not occasion SNC80-appropriate responding up to a dose (32.0 mg/kg) that produced convulsions in one monkey. The mu agonists morphine and fentanyl and the kappa agonists U-50,488 and enadoline failed to substitute for SNC80 up to doses that eliminated responding. Two nonopioids (the N-methyl-D-aspartate antagonist ketamine and the monoamine reuptake inhibitor cocaine) also produced primarily saline-appropriate responding. Both the discriminative stimulus and rate-decreasing effects of SNC80 were antagonized by the delta-selective antagonist naltrindole (0.01-1.0 mg/kg) but not by doses of the opioid antagonist quadazocine (0.1-1.0 mg/kg) that block the effects of mu and kappa agonists. These data suggest that the discriminative stimulus effects of SNC80 are mediated by delta-opioid receptors and that the discriminative stimulus effects of delta opioids in primates can be differentiated from the effects of other opioid and nonopioid compounds.

Chronic l-alpha acetylmethadol in rhesus monkeys: discriminative stimulus and other behavioral measures of dependence and withdrawal.

This study characterized discriminative stimulus and other effects of naltrexone in rhesus monkeys treated daily with the long-acting opioid l-alpha acetylmethadol (LAAM). An initial dose-finding study assessed the rate-decreasing effects of naltrexone in three monkeys receiving LAAM daily (0.32-1.78 mg/kg); subsequently, these monkeys and a fourth received 1.0 mg/kg/12 hr of LAAM although discriminating between naltrexone and saline. Responding occurred on the saline lever after the administration of LAAM, whereas >90% drug-lever responding occurred after the administration of 0.1 mg/kg of naltrexone that also elicited signs of withdrawal. Naloxone and quadazocine, but not morphine, nalbuphine or ketamine, substituted for naltrexone. Morphine and nalbuphine shifted the naltrexone dose-effect curve to the right. Compared to precipitated withdrawal, deprivation-induced withdrawal occasioned less naltrexone-lever responding and fewer observable signs of withdrawal. Maximal naltrexone-level responding occurred 24 to 48 hr after the discontinuation of LAAM treatment; the frequency of other withdrawal signs also peaked 24 to 48 hr after the discontinuation of LAAM. Partial naltrexone-lever responding occurred for up to 10 days after discontinuation of LAAM treatment; 4 and 8 days after the discontinuation of LAAM treatment, 0.1 mg/kg of naltrexone did no further increase naltrexone-lever responding or withdrawal signs suggesting that less-then-maximal naltrexone-lever responding was not due to long-lasting effects of LAAM or its metabolites. The discriminative stimuli that are associated with LAAM deprivation might be different from the stimuli associated with either training condition. This study is the first antagonist discrimination in non-humans primates treated chronically with LAAM and the results indicate that the naltrexone stimulus is related to opioid withdrawal.

Pharmacological profile of a deuterium-substituted mirfentanil derivative, OHM10579, in rhesus monkeys.

The discriminative-stimulus, respiratory, and antinociceptive effects of OHM10579, an isotopic isomer of mirfentanil, were characterized in rhesus monkeys. In monkeys discriminating nalbuphine, 0.32 mg/kg of OHM10579 partially substituted for nalbuphine. In monkeys treated daily with 3.2 mg/kg of morphine and discriminating 0.01 mg/kg of naltrexone, 0.32 mg/kg of OHM10579 substituted for naltrexone. In morphine-abstinent monkeys, morphine reversed naltrexone-lever responding, an effect attenuated by OHM10579. The shift to the right in the morphine dose-effect curve was greater 2 h after 0.32 mg/kg of OHM10579 compared to 0.32 mg/kg of mirfentanil, indicating that OHM10579 has a longer duration of action than mirfentanil. In a warm-water tail-withdrawal procedure, 10 and 17.8 mg/kg of OHM10579 had antinociceptive effects that were not antagonized by naltrexone. Morphine decreased breathing in air to 48%, whereas the maximal decrease with OHM10579 was to 75% of control. OHM10579 attenuated hyperventilation induced by 5% CO2 and partially antagonized the respiratory-depressant effects of morphine. OHM10579 can be classified as a low-efficacy mu-opioid agonist with some nonopioid actions. These results indicate that the pharmacology of the mirfentanil isotope OHM10579 is similar to that of mirfentanil, but that OHM10579 might have a longer duration of action.

Studies on benzodiazepines and opioids administered alone and in combination in rhesus monkeys: ventilation and drug discrimination.

Benzodiazepines and opioids are co-administered recreationally as well as clinically; in the current study, the ventilatory-depressant and discriminative stimulus effects of several benzodiazepines and opioids were examined alone and in combination in order to evaluate any interaction between agonists from these pharmacological classes. The benzodiazepines alprazolam, diazepam, flunitrazepam, lorazepam, midazolam and triazolam and the opioids morphine and fentanyl decreased ventilation (V(E)) in monkeys breathing either air or 5% CO2 in air, although decreases in ventilation produced by opioids were greater in magnitude than decreases produced by benzodiazepines. Flumazenil antagonized the ventilatory-depressant effects of flunitrazepam and triazolam and not those of fentanyl; naltrexone antagonized the ventilatory-depressant effects of fentanyl and not those of flunitrazepam or triazolam. Interactions between the ventilatory-depressant effects of agonists from the two classes were less than additive. In monkeys receiving 3.2 mg/kg per day of morphine and discriminating 0.01 mg/kg naltrexone, neither flunitrazepam nor triazolam substituted for naltrexone; in morphine-deprived monkeys, morphine, and not flunitrazepam or triazolam, reversed naltrexone-lever responding. Moreover, benzodiazepines did not modify the discriminative stimulus effects of naltrexone in morphine-treated monkeys or of morphine in morphine-deprived monkeys. In contrast to studies showing synergism between benzodiazepines and opioids, the current study suggests that, under some conditions, combinations of these drugs can be administered without enhancing the ventilatory-depressant effects of either class of drugs or the discriminative stimulus effects of opioids.

Behavioral effects and binding affinities of the fentanyl derivative OHM3507.

Several fentanyl derivatives have been reported to have novel pharmacologies that might be exploited for developing alternate approaches to the treatment of pain. The purpose of the current series of studies was to evaluate OHM3507, a novel fentanyl derivative reported to have an unusual pharmacological profile in nonprimate species. Similar to several other fentanyl derivatives with clinical potential, OHM3507 had the highest affinity (IC50 = 10 nM) for mu ([3H]D-Ala2,N-Me-Phe4,Gly5-OH-labeled) receptors with 6- and 176-fold lower affinity for delta ([3H]D-Pen2-D-Pen5-labeled), and kappa ([3H]ethylketocyclazocine-labeled) receptors, respectively. In rhesus monkeys, OHM3507 shared discriminative stimulus effects with morphine, increased tail-withdrawal latencies in a warm-water procedure of antinociception, decreased ventilation in monkeys breathing normal air or 5% CO2, and failed to modify accuracy on acquisition and performance tasks up to doses that decreased rates of food-maintained responding. The opioid antagonists naltrexone and naltrindole antagonized the behavioral effects of OHM3507 in a manner that was consistent with mu-receptor mediation. Although OHM3507 appeared to have low efficacy opioid actions in nonprimate species, results from the current studies clearly show this compound to have strong, fentanyl-like mu agonist actions in rhesus monkeys. These results provide another example of the sometimes poor predictability in the behavioral pharmacology of fentanyl derivatives among species, in this case between monkeys and rats, mice and rabbits, and demonstrates the need for evaluating new drugs under a broad range of conditions to increase the probability of identifying novel compounds that can be used to treat pain.

Discriminative stimulus effects of l-alpha-acetylmethadol (LAAM), buprenorphine and methadone in morphine-treated rhesus monkeys.

The discriminative stimulus effects of l-alpha-acetylmethadol (LAAM), l-alpha-acetylnormethadol (nor-LAAM), l-alpha-acetyldinormethadol (dinor-LAAM), buprenorphine and methadone were investigated in morphine-treated (3.2 mg/kg/day) rhesus monkeys (n = 3-6) discriminating between saline and naltrexone (0.01 mg/kg) and responding under a fixed ratio (FR) schedule of stimulus-shock termination. Monkeys responded on the naltrexone lever after either the administration of 0.01 mg/kg of naltrexone or the substitution of saline for the daily dose of morphine (i.e., 27-hr morphine deprived). Morphine dose-dependently reversed naltrexone lever responding in morphine-deprived monkeys. Methadone, LAAM, nor-LAAM and dinor-LAAM had morphine-like discriminative stimulus effects in all monkeys, whereas, buprenorphine had naltrexone-like discriminative stimulus effects in three monkeys and morphine-like effects in two monkeys; 24 hr after administration, buprenorphine antagonized the effects of morphine in the former and antagonized the effects of naltrexone in the latter. The agonist and antagonist effects of buprenorphine persisted for more than 6 days. The relative duration of action was: buprenorphine > LAAM > nor-LAAM = methadone = dinor-LAAM = morphine. That buprenorphine had markedly different discriminative stimulus effects in monkeys treated identically with morphine is likely due to the low efficacy of buprenorphine and emphasizes the difficulty in predicting the behavioral effects of buprenorphine in opioid-dependent individuals. The considerably longer duration of LAAM, than either nor-LAAM or dinor-LAAM, indicates that the rate of metabolite formation is important for the long duration of LAAM and further suggests that variations in metabolic activity among individuals might result in differences in the behavioral effects of LAAM.

Discriminative stimulus effects on enadoline in pigeons.

The discriminative stimulus effects of enadoline were characterized in pigeons responding under a fixed-ratio 20 schedule of food presentation and discriminating between intramuscular injections of the kappa opioid agonist enadoline and saline. Cumulative doses of enadoline dose-dependently increased drug-key responding with the training dose of enadoline (0.178 mg/kg) producing > or = 90% drug key responding (% DR). In time course studies, doses of enadoline larger than 0.32 mg/kg produced > or = 90% DR for more than 40 min. Naltrexone antagonized both the discriminative stimulus and the rate-decreasing effects of enadoline (pA2 = 6.79 and 6.73, respectively); in some pigeons, naltrexone produced an unsurmountable antagonism of the enadoline discriminative stimulus. Substitution tests using the kappa agonists U-50,488, spiradoline, U-69,593 and ethylketocyclazocine resulted in > or = 90% DR in most, but not all, pigeons; at larger doses, all compounds markedly decreased response rates. Up to rate-decreasing doses, nalorphine, dynorphin A(1-13) amide (DYN), nalbuphine, butorphanol, morphine and ketamine failed to occasion > or = 90% DR; nalbuphine, nalorphine, butorphanol, but not DYN, antagonized the discriminative stimulus and the rate-decreasing effects of enadoline. This study established stimulus control with enadoline in pigeons and results from substitution studies in these pigeons support the view that the enadoline discriminative stimulus is mediated by kappa opioid receptors; these results further demonstrate that nalbuphine and butorphanol have kappa antagonist actions in pigeons. The negative results obtained with DYN are in contrast to previous demonstrations of kappa agonist effects for DYN and might provide support for the hypothesized importance of nonopioid systems in the effects of this peptide.

[Attitude to the criteria of death and kidney transplantation of the relatives of kidney donors]. / Holdningen til dødskriterium og nyretransplantation hos nyredonorers pårørende.

Fifty near relatives of cadaver kidney donors in the County of Ringkøbing were questioned about how they experienced information concerning the brain death of the donor and the request for permission to remove the kidneys for transplantation. This was undertaken as an interview investigation. In addition, they were questioned as to whether they subsequently experienced doubt about the decision and about their attitude to the criteria for death. It was found that approximately half were aware of the seriousness of the condition before they were told by a doctor but that 35% would have desired more detailed information. Emphasis is placed, in addition, on improved conditions for a dignified parting from the decreased, not information by telephone and that the time for decision making was too short. The reasons for accepting removal of the kidneys were either knowledge of the donor's attitude or consideration of the recipient. Only three donors had drawn up donor testaments. Only one of the relatives regretted the decision while 20% had experienced doubts but the great majority would have liked to discuss the subject with others later. 63% accept the criteria for brain death and 14% the criteria for heart death but only 9% have altered their attitude as a result of the current debate.

Mental function in elderly men after surgery during epidural analgesia.

To evaluate whether there is a difference in mental function after general anaesthesia and epidural analgesia, a homogeneous group of 40 elderly men (age between 60 and 80) undergoing transurethral prostatectomy was studied. The study was prospective, randomised and double blind. Patients with all types of complications believed to impair mental function were excluded. Long-term, short-term, verbal and visual memory were tested preoperatively, and 4 days, and 3 weeks postoperatively. In conclusion, we found a significant and equal decline in test performance on the fourth postoperative day. Three weeks postoperatively, however, both groups had returned to or exceeded preoperative levels of performance.