Antagonism of the discriminative stimulus effects of the kappa-opioid agonist spiradoline.

This study compared the potency of mixed-action opioids (i.e., agonist-antagonists) and pure antagonists to block the discriminative stimulus effects of spiradoline, a kappa-opioid agonist. Rats were trained to discriminate between 3.0 mg/kg spiradoline and saline (SC) in a two-choice discrete-trial procedure. Graded doses of test drugs were administered in combination with 3.0 mg/kg spiradoline and the dose that reduced selection of the spiradoline-appropriate choice lever by 50% (AD50) was calculated. Ten drugs blocked the discriminative effects of spiradoline in an orderly dose-dependent manner. They spanned a 150-fold potency range from diprenorphine (5 times as potent as naloxone) to nalbuphine (0.03 times as potent as naloxone). Antagonism was stereoselective: 0.1-1.0 mg/kg (-)-N-allylnormetazocine (NANM) reduced spiradoline-appropriate responding by 50% whereas 3.0 mg/kg (+)-NANM did not. (-)-Pentazocine (0.1-10 mg/kg) and butorphanol (0.1-3.0 mg/kg) also did not block the discriminative effects of spiradoline. Antagonism was surmountable when graded doses of spiradoline were tested with a fixed dose of diprenorphine, naloxone, or levallorphan. Apparent pKB values derived from the interactions between those three drugs and spiradoline were in accord with relative antagonist potencies based upon the AD50s. Because the potency of a competitive antagonist is determined by its receptor affinity, the relative potencies of mixed-action opioids and pure antagonists in blocking the discriminative stimulus effects of spiradoline can provide an estimate of the relative in vivo affinities of these drugs for the kappa-opioid receptor.

Assessing spiradoline-like discriminative effects of DuP 747: influence of route of administration.

DuP 747, trans-3,4-dichloro-N-methyl-N-[2-(pyrrolidin-1-yl)-1,2,3,4- tetrahydronaphthalen-1-yl]benzeacetamide methanesfonate, is a recently synthesized analgesic drug that binds with high affinity and selectivity to the kappa-opioid receptor. In order to determine if DuP 747 has kappa-like discriminative effects it was tested for stimulus generalization in rats trained to discriminate between SC injections of saline and 3.0 mg/kg of spiradoline, a potent kappa-opioid agonist. A range of drug doses was administered by each of several routes 30 min before a test session. Spiradoline occasioned orderly dose-dependent increases in spiradoline-appropriate lever selection after SC or IP administration, with ED50s of 0.65 and 1.75 mg/kg, respectively. In contrast, DuP 747 (1.0-30 mg/kg) occasioned little spiradoline-appropriate lever selection when administered SC, but was generalized from spiradoline partially when administered IP (ED50 = 5.9 mg/kg) or PO (ED50 = 59 mg/kg). The 5-hydroxy-desmethoxy metabolite of DuP 747, administered SC (0.3-10 mg/kg), occasioned selection of the saline-appropriate lever only. That DuP 747 had little spiradoline-like activity after SC administration suggests that a metabolite of DuP 747 was responsible for the spiradoline-appropriate responding that followed IP and PO administration of the drug, apparently a metabolite other than 5-hydroxy-desmethoxy-DuP 747.

Monoamine systems in the discriminative effects of spiradoline, a kappa-opioid agonist.

The results of studies on mice indicate that the antinociceptive effects of kappa-opioid agonists are due, in part, to activation of the 5-HT2 type of serotonin receptor. One objective of this study was to determine if the discriminative effects of spiradoline, a kappa-opioid agonist, are mediated by 5-HT2 receptors in rats also. A second objective was to confirm findings that dopamine receptor antagonists produce spiradoline-like discriminative effects (Ohno et al., 1992). Rats were trained to discriminate between spiradoline (3.0 mg/kg) and saline in a discrete-trial avoidance/escape procedure. In subsequent tests of stimulus generalization, the discriminative effects of spiradoline were not mimicked by fenfluramine (0.3-10 mg/kg) or fluoxetine (1.0-10 mg/kg), drugs that enhance serotonergically mediated neurotransmission, nor were they blocked by the 5-HT2 antagonists pirenperone (0.01-1.0 mg/kg) and ketanserin (0.1-10 mg/kg), or potentiated by fluoxetine pretreatment. Neither the dopamine receptor antagonists haloperidol (0.01-0.3 mg/kg) and sulpiride (3.0-100 mg/kg) nor the agonists apomorphine (0.03-0.3 mg/kg) and d-amphetamine (0.1-3.0 mg/kg) engendered spiradoline-like discriminative effects. These results demonstrate further the pharmacological specificity of the discriminative effects of spiradoline, but provide no evidence for mediation by serotonergic or dopaminergic systems.

Interactions between U-50,488H and sigma receptor antagonists: EEG, EEG power spectral and behavioral correlates.

I.v. administration of U-50,488H after i.v. saline pretreatment produced occasional high-voltage EEG slow-wave bursts that were associated with relatively small increases in spectral power in the 2.5-7.5 Hz band as a spectral peak and with behavioral incidents of sedation, ataxia, ptosis, straub tail, hunching of the back, and backing-up. After pretreatment with the sigma antagonists rimcazole and DuPont Merck S-7389-4, U-50,488H administration produced significantly larger increases in absolute EEG spectral power, both over the 1-50 Hz range and in the 2.5-5.0 and 5.0-7.5 Hz bands, than after saline pretreatment; rimcazole pretreatment eliminated U-50,488H-induced incidents of ataxia, ptosis, hunching of the back and backing-up. In summary, effects of U-50,488H on EEG, EEG power spectra and behavior may reflect interactions between kappa opioid and sigma (non-opioid) receptor-effector systems.

DuP 734 [1-(cyclopropylmethyl)-4-(2'(4''-fluorophenyl)-2'-oxoethyl)- piperidine HBr], a sigma and 5-hydroxytryptamine2 receptor antagonist: receptor-binding, electrophysiological and neuropharmacological profiles.

It has been suggested that sigma receptor antagonists may be useful as antipsychotic drugs and that 5-hydroxytryptamine (5-HT2) receptor antagonists produce improvements of the negative symptoms of schizophrenia. [1-(Cyclopropylmethyl)-4-(2'-(4''-fluorophenyl)-2'- oxoethyl)-piperidine HBr] (DuP 734) is a novel compound with high affinity for the sigma (Ki = 10 nM) and 5-HT2 (Ki = 15 nM) receptors, but low affinity for dopamine receptors (Ki > 1000 nM) as well as 33 other receptors, ion channels and second messenger systems in vitro. DuP 734 did not inhibit the synaptosomal uptake of dopamine, 5-HT or norepinephrine. Oral administration of DuP 734 potently blocked 5-hydroxy-L-trytophan (5-HTP)-induced head twitch in the rat (ED50 = 6.5 mumol/kg), indicating 5-HT2 antagonist activity. Extracellular single-unit recording studies demonstrated that DuP 734 antagonized the effect of the selective sigma ligand (+)-3-(3-hydroxyphenyl-N-(1-propyl) piperidine [(+)-3-PPP] on dopamine neuronal activity in the substantia nigra of the rat with an ED90 of 3.6 mumol/kg i.v. The sigma receptor agonists (+)-SKF 10,047 and phencyclidine both elicited rotational behavior in rats with unilateral lesion of the substantia nigra. The rotational behavior induced by either (+)-SKF 10,047 or phencyclidine was dose-dependently antagonized by DuP 734 with oral ED50 of 8.7 and 19.6 mumol/kg, respectively. The 5-HT2 receptor antagonist ICI 169,369, even at high doses (up to 33 mumol/kg, s.c.), did not antagonize the rotational behavior induced by (+)-SKF 10,047.(ABSTRACT TRUNCATED AT 250 WORDS)

Discriminative stimulus effects of spiradoline, a kappa-opioid agonist.

This study represents the initial step in assessing the discriminative effects of spiradoline (U62,066), a potent congener of the selective kappa-opioid agonist, U50,488. Separate groups of rats were trained to discriminate between SC injections of saline and either 3.0 mg/kg spiradoline or 3.0 mg/kg morphine in a discrete-trial shock-avoidance/escape procedure. Spiradoline-trained rats generalized completely to U50,488 (ED50S for spiradoline and U50,488 were 0.66 and 8.71 mg/kg, respectively), but selected the choice lever appropriate for saline in generalization tests with graded doses of morphine, phencyclidine, and agonist-antagonist opioids with varying degrees of kappa activity, ethylketocyclazocine, nalorphine, and butorphanol. Morphine-trained rats did not generalize to spiradoline. Naltrexone (0.01 or 0.1 mg/kg) blocked surmountably the discriminative effects of both spiradoline and morphine, but was approximately 10-fold less potent against spiradoline. These results indicate that the discriminative effects of spiradoline are mediated by kappa-opioid receptors; meaningful mu-opioid and PCP/sigma components of action were not in evidence. The potency and apparent pharmacological selectivity of spiradoline suggest the potential value of this drug for studying kappa-opioid-mediated stimulus control of behavior.

Electrophysiological effects of selective sigma-receptor agonists, antagonists, and the selective phencyclidine receptor agonist MK-801 on midbrain dopamine neurons.

Extracellular single unit recording techniques were used to study the effects of selective sigma-receptor agonist [(+)-3-PPP, (+)-pentazocine, and DTG] and selective sigma-receptor antagonists (BMY 14802 and Rimcazole) on dopamine neurons of the substantia nigra. Intravenous (IV) administration of sigma agonists decreased, whereas IV administration of the sigma antagonist BMY-14802 increased the firing rate of dopamine neurons. The other sigma antagonist Rimcazole produced inconsistent changes in dopamine unit activity. These data, in conjunction with anatomic data suggesting sigma receptor localization on dopamine neurons in the substantia nigra (Gundlach et al: J Neurosci 6:1757-1770, 1986; Graybiel et al: Soc Neurosci Abstr 13:28, 1987) demonstrate a relationship of the sigma receptor with the dopamine system and further suggest a model system to study agonist-antagonist interactions of sigma ligands. The selective phencyclidine (PCP) agonist MK-801 was equipotent to PCP in regard to stimulatory properties on dopamine neurons. However, the relative potencies do not correspond to their relative binding affinities, suggesting that non-PCP-receptor properties may mediate this effect.

Selective sigma receptor agonist and antagonist affect dopamine neuronal activity.

Extracellular recording techniques were used to study the effects of the selective sigma receptor agonist (+)-[3H]3-(3-hydroxyphenyl)-N-(1-propyl)piperidine ((+)-3-PPP) and selective sigma receptor antagonist BMY 14802 on dopamine (DA) neurons of the substantia nigra. Intravenous administration of (+)-3-PPP produced a dose-dependent inhibition of DA neuron firing rate. Complete inhibition of DA neurons produced by (+)-3-PPP could be completely reversed by administration of BMY 14802. Also, pretreatment with BMY 14802 shifted the (+)-3-PPP dose response curve to the right. These data demonstrate a relationship of the sigma receptor with the dopamine system and further suggest a model system to study agonist/antagonist interactions of sigma ligands.

Biochemical, behavioral, and electrophysiologic actions of the selective sigma receptor ligand (+)-pentazocine.

Research on the sigma receptor, a binding site associated with drug-induced psychotomimetic behaviors, has been hampered because most sigma agonists also interact with the phencyclidine (PCP) receptor. (+)-Pentazocine, a human psychotogen, is a selective sigma receptor ligand. To demonstrate sigma receptor activities, we studied the behavioral and electrophysiologic actions for (+)-pentazocine. In the behavioral drug discrimination procedure in which rats were trained to discriminate between 2.0 mg/kg (5.59 mumol/kg) (+)-pentazocine and saline, (+)-pentazocine produced dose-related increases in the percentage of trials completed on the (+)-pentazocine lever. At a dose of 1.0 mg/kg (3.29 mumol/kg) (+)-N-allylnormetazocine generalized completely to (+)-pentazocine. By contrast, PCP only partially generalized. In the visual evoked potential test, these compounds produced a significant dose-dependent slowing of the N2 latency. This response was prevented by haloperidol pretreatment. These results demonstrate pharmacologic actions for the selective sigma receptor ligand (+)-pentazocine and suggest some overlapping pharmacologic properties of the sigma and PCP receptor sites despite differences in central nervous system distribution.

Discriminative stimulus properties of a sigma receptor agonist in the rat: role of monoamine systems.

The role of the dopaminergic, noradrenergic and serotonergic systems in the production of the discriminative stimulus (DS) properties of the sigma receptor agonist (+)-N-allylnormetazocine [(+)-NANM/(+)-SKF 10,047] was measured in rats. The partial generalization of the noradrenergic agonist clonidine with (+)-NANM and partial antagonism of (+)-NANM by yohimbine suggest that the noradrenergic system may play a role in (+)-NANM DS. Activation of the dopaminergic system by apomorphine did not produce (+)-NANM DS however, haloperidol, which binds to the sigma receptor with high affinity, antagonized (+)-NANM DS. These data suggest that the haloperidol antagonism of (+)-NANM DS is a function of antagonism at the sigma receptor rather than antagonism at the dopamine receptor. Since yohimbine does not bind to the sigma receptor, yohimbine antagonizes (+)-NANM by a mechanism different from that of haloperidol. There was no evidence to suggest serotonergic mediation of (+)-NANM DS.

Discriminative stimulus properties of (+)-N-allylnormetazocine in the rat: correlations with (+)-N-allylnormetazocine and phencyclidine receptor binding.

Recent studies have identified a stereospecific (+)-NANM binding site that binds psychotomimetic opioids and phencyclidine (PCP) but has a distribution in brain different from the PCP binding site. Since (+)-NANM has no opioid activity and (-)-NANM has opioid activity, rats were trained to discriminate (+)-NANM from saline in order to develop an ability to distinguish the (+)-NANM cues from other opioid agonist and antagonist activities. Cyclazocine, PCP, and ketamine all produced (+)-NANM-like stimuli in a dose-dependent manner. Behaviorally, cyclazocine and PCP are equipotent to (+)-NANM whereas ketamine is 6.7 times less potent than (+)-NANM. Pentazocine had the highest affinity for the (+)-[3H]NANM binding site, yet did not produce (+)-NANM-like discriminative stimuli. By contrast, ketamine had the lowest binding affinity for the (+)-[3H]NANM binding site and did produce (+)-NANM-like discriminative stimuli. Drug discrimination potencies relative to (+)-NANM were not predictive of relative binding affinities at (+)-NANM or PCP binding sites, although there was a trend toward a stronger correlation with the PCP binding site. Therefore, the discriminative stimulus properties of (+)-NANM cannot be explained by pharmacologic actions at either (+)-NANM or PCP binding sites alone, and may involve concurrent actions at both sites.

(+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine [(+)-3-PPP] but not (-)-3-PPP produces (+)-N-allylnormetazocine-like (SKF 10,047) discriminative stimuli.

In rats trained to discriminate the prototypic sigma receptor agonist, (+)-N-Allylnormetazocine [(+)-N-Allylnormetazocine [(+)-NANM/SKF 10,047], from saline, the (+)- but not the (-)-isomer of 3-(3-hydroxyphenyl)-N-(1-propyl)piperidine (3-PPP) produced (+)-NANM-like discriminative stimuli. (+)-3-PPP binds stereo selectively to the (+)-NANM binding site, but not to the phencyclidine binding site. Additionally, phencyclidine was found to produce (+)-NANM-like discriminative stimuli. Although the 3-PPP isomers were shown to produce changes in central dopaminergic activity (Hjorth et al. Life Sci 37, 673, 1985), the discriminative stimulus properties of (+)-3-PPP are apparently not mediated via the dopaminergic system. This hypothesis is supported by the fact that apomorphine did not produce (+)-NANM-like discriminative stimuli. These stimuli are thus non-dopaminergic and may be due to the (+)-3-PPP actions at the sigma binding site. However, it is possible that (+)-NANM, PCP, and (+)-3-PPP may have common non-sigma pharmacologic properties that account for the similar discriminative stimulus properties of these compounds.

Antinociceptive profiles of mu and kappa opioid agonists in a rat tooth pulp stimulation procedure.

The purpose of the study was to develop a tooth pulp stimulation procedure in the awake, freely moving rat and to then quantitatively assess the analgesic effects of compounds reported to act on mu and/or kappa receptors. The mu receptor agonists produce a biphasic (primary and secondary slope) dose-response curve (DRC) whereas kappa agonist and mixed agonist/antagonist analgesics produce single-slope DRCs. The primary slopes calculated from the mu agonist biphasic DRC are steeper than the slopes calculated for the other types of opioid analgesics. The rank order of analgesic potency for the mu agonist analgesics is oxymorphone greater than morphine = methadone greater than meperidine. The rank order analgesic potency for the kappa agonist analgesics is tifluadom greater than ethylketocyclazocine greater than U50488H and for the mixed agonist/antagonist analgesics, butorphanol greater than nalbuphine greater than pentazocine. The nonsteroidal anti-inflammatory drugs, aspirin and zomepirac, are also effective analgesics in this test procedure; but the DRC slopes for these compounds are lower than all opioid analgesics tested. The opioid antagonist naloxone produces no significant changes in threshold responses. Naloxone did reverse the threshold increases produced by morphine and ethylketocyclazocine but not aspirin. This study demonstrates that the electrical stimulation of the tooth pulp in the rat can be used as an assay for evaluation of opioid and nonopioid analgesics. When minimal effective dose values of each analgesic are plotted as a function of the clinical analgesic dose a high correlation is observed.

Caudate unit activity in freely moving cats: effects of phasic auditory and visual stimuli.

Quantitative analyses of the single unit responses to sensory stimuli were made in caudate nucleus neurons in the unanesthetized freely moving cat. Over half of the cells were responsive to repetitive presentation of simple sensory stimuli. Typically, neurons gave similar responses to both auditory (click) and visual (flash) stimuli. This study confirms previous reports of the polysensory response characteristics of caudate nucleus neurons, and extends these observations to the freely moving animal.

Antinociceptive profiles of opioid peptide agonists in a rat tooth pulp stimulation procedure.

The analgesic activity of the prototypic opioid peptides for the mu (D-Ala2-Me-Phen4-Gly-ol5-enkephalin [DAGO]) kappa (Dynorphin 1-13), delta (D-Ala2-D-Leu5-enkephalin [DADLE]), or epsilon (beta-endorphin) receptor was assessed in a rat tooth pulp stimulation procedure. All opioid peptides tested and the opioid alkaloid U50, 488H (kappa receptor agonist) significantly elevated response thresholds. The rank order of potency based on the Minimum Effective Dose values was beta-endorphin greater than DAGO = dynorphin A (1-13) amide greater than DADLE greater than dynorphin A (1-13) greater than U50,488H. Based on absolute magnitude, the rank order of dose response slopes was DAGO greater than U50,488H greater than dynorphin A (1-13) amide greater than beta-endorphin greater than DADLE. Dynorphin A (1-13) produced the shallowest dose response slope and the magnitude of response threshold was the lowest for all compounds tested. Finally, the general conclusion that mu agonists are effective against noxious stimuli derived from thermal, chemical, and mechanical is extended by our data to include electrical sources derived from tooth pulp stimulation; kappa agonists are effective against noxious stimuli derived from chemical, mechanical, and electrical sources (tooth pulp stimulation) and delta agonists are effective analgesics against thermal, chemical and electrical stimuli (tooth pulp stimulation).

Chloral hydrate anesthesia alters the responsiveness of central serotonergic neurons in the cat.

The influence of chloral hydrate anesthesia on the spontaneous activity and responsiveness of serotonergic neurons was examined by administering chloral hydrate (300 mg/kg, i.p.) to freely moving cats from which serotonergic unit activity in the dorsal raphe nucleus (DRN) was being recorded. Although chloral hydrate administration produced a surgical level of anesthesia within 15 min following injection, it produced only a small decrease (approximately 20%) in the spontaneous activity of DRN serotonergic neurons. In contrast, the responsiveness of these same neurons was greatly altered by chloral hydrate administration. By examining the same neuron before and after chloral hydrate injection, it was found that chloral hydrate anesthesia completely abolished the excitatory responses of DRN serotonergic neurons to auditory and visual stimuli, as well as their excitatory response to electrical stimulation of the gigantocellular tegmental field (FTG) in the pontine reticular formation. On the other hand, the inhibition of serotonergic neuron firing resulting from systemic administration of WB 4101 (1.0 mg/kg, i.p.), a selective alpha 1 adrenergic receptor antagonist, was greatly potentiated by chloral hydrate anesthesia. Therefore, these data indicate that chloral hydrate anesthesia produces profound changes in the physiological and pharmacological responses of central serotonergic neurons which are not predictable by examination of spontaneous activity alone. Furthermore, as discussed, it it not clear to what extent these confounding influences might generalize to other anesthetized or immobilized preparations. Thus, beyond the obvious advantage which allows for the study of relationships between neuronal activity and behavior, single unit studies conducted in awake, freely moving animals also may be of greater value for basic physiological and pharmacological studies.

Raphe unit activity in freely moving cats is altered by manipulations of central but not peripheral motor systems.

Single unit activity of serotonergic neurons in the dorsal raphe nucleus of freely moving cats was recorded in experiments which manipulated central or peripheral motor systems. Unilateral microinjections of the cholinomimetic agent, carbachol, into the pontine tegmentum, produced muscle atonia. During these periods of drug-induced atonia, the activity of serotonergic neurons was reduced 97% below pre-drug baseline rates. In experiments where microinjections of carbachol did not produce muscle atonia, no significant change occurred in serotonergic unit discharge rate. Muscle tonus was also altered by systemic injections of mephenesin, a centrally acting muscle relaxant. A low dose of mephenesin (50 mg/kg) produced mild atonia which was correlated with a 16% reduction in serotonergic neuron discharge rate relative to pre-drug baseline. A higher dose of mephenesin (150 mg/kg) produced complete atonia, during which serotonergic unit activity was reduced by 68% below baseline firing rate. To distinguish between centrally and peripherally induced atonia, we injected either succinylcholine or dantrolene, systemically. These are both drugs whose muscle relaxant properties are known to be mediated by peripheral mechanisms. In neither case was a change in serotonergic unit discharge rate seen following drug-induced atonia. These data demonstrate that manipulation of central, but not peripheral, motor systems can profoundly affect the activity of serotonergic neurons of the dorsal raphe nucleus. Alternate hypotheses are also discussed.

Response of dopaminergic neurons in cat to auditory stimuli presented across the sleep-waking cycle.

The response of dopaminergic neurons of the substantia nigra pars compacta to auditory clicks continuously presented across the sleep-wake cycle was studied in cats. The initial excitatory followed by inhibitory response to the click which occurred during quiet waking diminished as the cat progressed into slow-wave sleep and was absent during REM sleep. Upon awakening from REM sleep, dopamine neurons once again displayed an excitatory/inhibitory response to the clicks, implying that the decrease across the sleep-wake cycle was not attributable to long-term habituation.

Dopaminergic unit activity in freely moving cats: lack of relationship to feeding, satiety, and glucose injections.

The activity of dopaminergic neurons in the substantia nigra was recorded from freely moving cats during feeding and satiety, and following injections of glucose. At no time during feeding or in the postprandial satiety period was there a significant increase or decrease in firing rate of these neurons relative to baseline. Additionally, no change in firing rate was observed following injections of glucose (300, 500 and 1000 mg/kg) or glucose in combination with insulin (300 mg/kg glucose and 0.8 units/kg insulin).

Behavioral correlates of dopaminergic unit activity in freely moving cats.

Single unit activity of dopaminergic neurons in the substantia nigra was recorded in freely moving cats under a variety of conditions. These neurons displayed their highest discharge rate during active waking (3.68 +/- 0.30 spikes/s), which was 20% greater than their discharge rate during quiet waking (3.07 +/- 0.20). Although these cells fired somewhat faster during active waking, their activity displayed no correlation with phasic EMG changes, and, in general, their activity showed little relationship to overt behavioral changes. As the cat progressed from quiet waking through slow-wave sleep and REM sleep there was no significant change in either the rate or pattern of firing of dopaminergic neurons. In addition, no correlation was observed between the activity of these neurons and either sleep spindles or PGO waves. These neurons did respond, however, to the repeated presentation of a click or light flash with excitation followed by inhibition, with no evidence of habituation. One of the most impressive changes in dopaminergic unit activity was a large decrease in association with orienting responses. This was seen in over 50% of the cells in which this relationship was examined. As the behavioral orientation habituated with repeated stimulus presentation, so did the associated dopaminergic unit suppression. In conclusion, dopaminergic neurons maintain a remarkably constant rate and pattern of firing across a variety of behaviors and states. However, this stability can be dramatically altered under special circumstances, such as during and following orienting responses.