Mariena V. Mattson

Binding of β-carbolines and related agents at serotonin (5-HT2 and 5-HT1A), dopamine (D2) and benzodiazepine receptors

A large series of β-carbolines was examined for their ability to bind at [3H]agonist-labeled 5-HT2A serotonin receptors. Selected β-carbolines were also examined at 5-HT2C serotonin receptors, 5-HT1A serotonin receptors, dopamine D2 receptors, and benzodiazepine receptors. Indolealkylamines and phenylisopropylamines were also evaluated in some of these binding assays. The β-carbolines were found to bind with modest affinity at 5-HT2A receptors, and affinity was highly dependent upon the presence of ring substituents and ring saturation. The β-carbolines displayed little to no affinity for 5-HT1A serotonin receptors, dopamine D2 receptors and, with the exception of β-CCM, for benzodiazepine receptors. Examples of β-carbolines, indolealkylamines (i.e. N,N-dimethyltryptamine analogs), and phenylisopropylamines have been previously shown to produce common stimulus effects in animals trained to discriminate the phenylisopropylamine hallucinogen DOM (i.e. 1-(2,5-dimethoxy-4-methylphenyl)-2-aminopropane) from vehicle. Although the only common receptor population that might account for this action is 5-HT2A, on the basis of a lack of enhanced affinity for agonist-labeled 5-HT2A receptors, as well as on their lack of agonist action in the PI hydrolysis assay, it is difficult to conclude that the β-carbolines behave in a manner consistent with that of other classical hallucinogens.

A specific acylating agent for the (3H)phencyclidine receptors in rat brain

A derivative of phencyclidine (PCP, 1 in fig. 1) bearing an isothiocyanate moiety on the meta position of the aromatic ring (Metaphit, 3 in fig. 1) has been synthesized and identified as a rapid and specific site‐directed acylating agent of the [3H]phencyclidine binding site in rat brain homogenates. The percentage of sites irreversibly inactivated by Metaphit was found to be the same in the hippocampus and striatum and the remaining sites were unaffected by Metaphit treatment under any conditions, suggesting that at least two distinct binding sites are present. An isomeric isothiocyanate derivative did not irreversibly inhibit [3H]phencyclidine receptors, indicating structural specificity for Metaphit in the inhibition of these receptors. The availability of Metaphit should greatly facilitate study of the structure and function of the phen‐cyclidine receptors.

Stereospecific reversal of nitrous oxide analgesia by naloxone.

The opiate antagonist naloxone was found to block nitrous oxide analgesia in a stereospecific fashion. Using a modified hotplate test in mice, the (-)-enantiomer of naloxone (which has a KD of approximately 1 nM for opiate receptors) antagonized the analgesic actions of nitrous oxide in a dose-dependent (2.5-20 mg/kg) fashion. In contrast, the (+)-enantiomer (KD approximately 10,000 nM) had no effect on nitrous oxide analgesia at the highest dose tested (40 mg/kg). These data strongly suggest that nitrous oxide analgesia is mediated via opiate receptors and is consistent with the hypotheses that this effect occurs either through the release of endogenous opioids or by physical perturbation of the opiate receptors.

In vivo effects of two novel alkylating benzodiazepines, irazepine and kenazepine

Intracerebroventricular administration of the alkylating benzodiazepines irazepine or kenazepine (20 nmol) resulted in a complete protection against convulsant doses of pentylenetetrazole (PTZ) for at least one hour, and a statistically significant protection for at least two and four hours, respectively. In contrast, administration of the non-alkylating parent benzodiazepine Ro-7/1986 or diazepam (20-60 nmol) resulted in no detectable anticonvulsants effects at fifteen minutes post-injection, the earliest interval examined. These results suggest that alkylating benzodiazepines which bind to brain benzodiazepine receptors in a non-competitive (covalent) fashion in vitro may exert a long lasting anticonvulsants effect by a similar mechanism.

Phencyclidine inhibits epinephrine-stimulated platelet aggregation independently of high affinity N-methyl-D-aspartate (NMDA)-type glutamatereceptors.

The psychotomimetic analgesic phencyclidine (PCP), which binds to a high affinity site on the neuronal N-methyl-D-aspartate (NMDA)-sensitive glutamate receptor, has previously been found to bind to platelets with high affinity and to specifically delay the onset of epinephrine-stimulated platelet aggregation (Jamieson et al. (1992) Biochem. J. 285, 35-39). We have now shown that the rank order of binding affinities of 14 synthetic PCP analogs at the high affinity binding site on platelets does not parallel the rank order of their affinities in binding to rat brain membranes, indicating that the high affinity PCP binding sites in platelets is distinct from the neuronal NMDA receptor. The order of potency of six of these analogs in delaying the onset of epinephrine-stimulated platelet aggregation also did not parallel the rank order of their binding affinities for platelet or brain binding sites. These data indicate that the ability of PCP analogs to inhibit epinephrine-stimulated aggregation is not related to their ability to bind to the high affinity platelet PCP binding site. Furthermore, (+)MK-801, which binds to the same high affinity binding site in neurons as does PCP, failed to inhibit epinephrine-stimulated platelet aggregation, further suggesting that the site at which PCP acts in platelets is not related to the NMDA-type glutamate receptor. Further studies showed that 5-HT2 receptors and effects on platelet secretion are not involved in PCP-mediated inhibition of epinephrine-induced platelet aggregation.

Etoxadrol-meta-isothiocyanate: A potent, enantioselective, electrophilic affinity ligand for the phencyclidine-binding site

Etoxadrol‐meta‐isothiocyanate (2S,4S,6S‐2‐ethyl‐2‐(3‐isothiocyanatophenyl)‐2‐piperidyl)1,3‐dioxolane, 4a) has been synthesized and characterized as an irreversible ligand for the phencyclidine (PCP)‐binding site. It is the first chiral electrophilic affinity ligand for this site to have been described. This affinity ligand is based upon etoxadrol, a 1,3‐dioxolane known to have PCP‐like effects in vivo and in vitro. Etoxadrol‐meta‐isothiocyanate was found to be four–five times more potent in vitro than metaphit (1‐[1‐(3‐isothiocyanatophenyl)cyclohexyl]piperidine), the only previously known electrophilic affinity ligand for the PCP‐binding site. The binding was shown to be highly enantioselective for etoxadrol‐meta‐isothiocyanate (4a). The 2R,4R,6R‐enantiomer of 4a was essentially inactive. The ability of the 2S,4S,6S‐enantiomer (4a) to interact with the benzodiazepine, muscarinic, and mu opioid receptor systems was also examined, and it was found not to interact with these receptor systems. It seems likely that 4a will prove to be a valuable tool in the study of structure and function of the PCP‐binding site.

Zipeprol: preclinical assessment of abuse potential

Zipeprol was evaluated in a number of in vitro and in vivo assays predictive of stimulant, depressant, or opioid abuse potential. Zipeprol had affinity for mu and kappa opioid binding sites as well as sigma binding sites. However, it failed to exert opioid-like agonist actions in rodents, and did not attenuate withdrawal signs in morphine- or pentobarbital-dependent rats. Zipeprol did not substitute for either amphetamine or pentobarbital in drug discrimination assays in rhesus monkeys. On the other hand, it suppressed morphine withdrawal signs in rhesus monkeys in two assays, and it acted as a quadazocine-sensitive reinforcer in monkeys trained to self-inject alfentanil. Zipeprol also acted as a reinforcer in monkeys trained to self-inject methohexital. In a dose range of 10-18 mg/kg, zipeprol induced convulsions in monkeys. Zipeprol appears to have abuse potential and a novel spectrum of action involving both opioid and non-opioid effects.