Peptide mimetics of thyrotropin-releasing hormone based on a cyclohexane framework: design, synthesis, and cognition-enhancing properties.

The design and synthesis of peptide mimetics of thyrotropin-releasing hormone (TRH) in which the peptide backbone is entirely replaced by a cyclohexane framework are described. The cis-1,3,5-trisubstituted ring was expected to permit key pharmacophoric groups to adopt conformations consistent with proposed bioactive conformations of the peptide. Compounds were synthesized by a stereoselective synthesis starting from L-glutamic acid. In a behavioral model of cognition in which TRH is active, the mimetics are potent, active compounds, exhibiting oral activity. One analog (26, (1S,3R,5(2S),5S)-5-[[5-oxo-1-(phenylmethyl)-2-pyrrolidinyl]-methyl]-5- [(1H-imidazol-5-yl)methyl]cyclohexaneacetamide) was radiolabeled for binding studies and evaluated in other binding assays and pharmacological tests. Competition binding of 26 vs [3H]MeTRH to rat brain slices suggests a two-site model for ligand binding with IC50's of 1 microM and 3 mM. Direct binding of [3H]-26 shows a biphasic curve with IC50's of 80 and 49 microM, respectively. Further studies would be needed to establish a link between the novel binding site(s) and the behavioral activity of 26 and TRH analogs.

Narrow temporal therapeutic window for NMDA antagonist protection against focal cerebral ischaemia.

N-methyl-d-aspartate (NMDA) receptor antagonists have been shown to protect against focal cerebral ischaemia when administered either before or soon after the onset of ischaemia. However, the precise therapeutic window for protection using these drugs remains to be defined. We studied dextrorphan administration delayed for 2 or 4 h after transient middle cerebral focal ischaemia in a rabbit model. With a 2h delay, the mid (12.5 mg kg-1 h-1) and high doses (17.5 mg kg-1 h-1) provided significant cortical neuroprotection (50% and 58% reduction, respectively), and the low dose (7.5 mg kg-1 h-1) protected against ischaemic damage in the basal ganglia (52% reduction). Animals having steady-state serum dextrorphan concentrations greater than 2000 ng ml-1 showed 50% cortical neuroprotection for the 2-h-delay group. No significant neuroprotection was seen in the 4-h-delay group, and the 4 h delay animals with dextrorphan levels greater than 2000 ng ml-1 had more severe ischaemic oedema than the saline controls. These results suggest a narrow temporal therapeutic window for neuroprotection, where delivery of drug delayed by 2 h was efficacious but treatment at 4 h after ischaemia onset was not beneficial and possibly harmful. These findings may have important implications for the treatment of clinical stroke.

AF102B, a novel M1 agonist, enhanced spatial learning in C57BL/10 mice with a long duration of action.

Orally administered AF102B, a selective muscarinic M1 cholinergic agonist, improved spatial learning in C57BL/10 mice in the Morris water maze. In four experiments in which all drug-treated animals received only one single administration of AF102B, improvement of acquisition depended on two factors: pretreatment time (tp) and dose. When a standard tp of 1 h was used, AF102B exhibited a U-shaped dose-response curve that is characteristic of many nootropic agents: learning was significantly improved by dose levels ranging from 0.1 to 1 mg/kg p.o. When the tp was extended out to as long as 8 days, two new effects emerged: (a) 1 mg/kg, the dose that had been the peak active dose at 1 h, exhibited a biphasic time course of action, being active at 1 h or at all tp intervals from 3 h to 5 days, but not at 1.5 h; (b) 0.03 mg/kg, a dose that had been inactive at a tp of 1 h, was active at all tp intervals from 3 h to 5 days, but not at shorter (1 and 2 h) or longer (6-8 days) tp intervals. In another experiment, animals received 0.03 mg/kg for 1-5 consecutive days: this dose level was active if the tp interval between the last dose and the learning session was 24-120 h, but not if it was only 1 h. Thus AF102B enhanced cognition in mice with a longer duration of action than reported for traditional muscarinic agonists.

Cephalosporin 3'-quinolone esters with a dual mode of action.

According to the generally accepted mechanism by which bacterial enzymes react with cephalosporins, opening of the beta-lactam ring can lead to the expulsion of a 3'-substituent. A series of dual-action cephalosporins was prepared in which antibacterial quinolones were linked to the cephalosporin 3'-position through an ester bond in the expectation that, in addition to exerting their own beta-lactam activity, these cephalosporins would act as prodrugs for the second antibacterial agent. Compared to parent cephalosporins in which the 3'-substituent was acetoxy, the bifunctional cephalosporins exhibited a broadened antibacterial spectrum, suggesting that a dual mode of action may indeed be operative.

Senile plaques in aged squirrel monkeys.

Aged squirrel monkeys develop senile plaques in the brain that are similar to those occurring in aged rhesus monkeys and aged humans. These plaques consist of abnormal, swollen neurites around an amyloid core. In whole-hemisphere coronal sections through the level of the rostral temporal lobe, plaques are present in temporal cortex, amygdala, hippocampal formation and, occasionally, in other cortical regions. In more rostral sections through the frontal lobe, plaques are most common in orbitofrontal and frontal opercular cortical regions. In immunocytochemical preparations, some neurites show immunoreactivity with antibodies directed against phosphorylated neurofilaments and neuropeptide Y. Thus, plaques in these New World primates are similar in distribution and composition to those occurring in aged Old World primates.

Acylmorphinans. A novel class of potent analgesic agents.

A series of novel 2- and 3-acylmorphinans (8-14) was synthesized in our search for a potent analgesic agent with low addiction potential. The compounds were evaluated for antinociceptive potency and receptor binding affinity. Among these compounds, the levorotatory 3-acetyl-N-(cyclopropylmethyl)morphinan (12) was found to be an orally active analgesic, comparable in potency to morphine (1), yet only weakly able to substitute for morphine (1) in morphine-dependent rats.

Potential synthetic codeine substitutes: (-)-3-O-aryl-N-methylmorphinans.

A series of novel O-aryl-N-methylmorphinans (7-19) were synthesized by the Ullmann reaction from levorphanol (4) in our search for a synthetic codeine (2) substitute with reduced addition liability. The compounds were evaluated for antinociceptive potency and receptor binding affinity. Among these compounds, (-)-3-phenoxy-N-methylmorphinan (7) is an orally active analgesic comparable in potency to codeine (2), which exhibits decreased physical dependence liability and longer duration of action.

2-benzazepines. 5. Synthesis of pyrimido[5,4-d][2]benzazepines and their evaluation as anxiolytic agents.

A series of 5H-pyrimido[5,4-d][2]benzazepines has been synthesized, starting from the corresponding 2-benzazepin-5-ones, and evaluated as potential anxiolytic agents. Selected compounds from this series show a pharmacological profile of action different than that of diazepam. They are more potent than diazepam in the anti-pentylenetetrazole test and in the [3H]diazepam binding assay, yet show less activity in the inclined screen test. A pharmacological data profile is given for 9-chloro-7-(2-chlorophenyl)-5H-pyrimido[5,4-d] [2]benzazepine (7c). The structure-activity relationships of these potential anxiolytic agents are discussed.

Pharmacodynamic studies with (-)-3-phenoxy-N-methylmorphinan in rats.

Analgesia and brain and plasma concentrations of (-)-3-phenoxy-N-methylmorphinan (PMM) and its metabolites were determined in rats administered 50 mg/kg of 3H-labeled PMM p.o., an approximate ED50. Unchanged PMM and two active metabolites, levorphanol and a different phenol, p-hydroxylated on the 3-phenoxy group (pOH-PMM), were present in brain at concentrations greater than in plasma. Analgesia was observed from 1 to 6 hr and was associated with brain concentrations of 400-1400 ng/g of PMM, 190-300 ng/g of pOH-PMM, and 16-27 ng/g of levorphanol. The presence of 58% of the administered dose as unchanged PMM in the gastrointestinal tract at 6 hr may reflect slow absorption and explain the persisting brain concentrations of PMM and its metabolites as well as the prolonged analgesia. Analgesia may have been due to the presence in brain of only PMM, pOH-PMM or levorphanol, or to the combined activity of two or three of these substances. Administration of the approximate ED50 of 3H-labeled levorphanol (0.1 mg/kg, s.c., or 6 mg/kg, p.o.) resulted in brain levorphanol concentrations (11-18 ng/g) close to those observed when PMM was administered p.o. at 50 mg/kg. After administration of an approximate subcutaneous ED50 of [3H]pOH-PMM of 24 mg/kg, the brains contained pOH-PMM (1500-4100 ng/g) and levorphanol (60-100 ng/g); these levorphanol concentrations were higher than those found after administration of the approximate ED50 of PMM or levorphanol. The findings indicate that brain levorphanol concentrations resulting from administration of PMM or pOH-PMM to rats may account for the analgesic activity observed, i.e. that PMM and pOH-PMM may act as prodrugs for levorphanol

Pharmacology of midazolam.

8-Chloro-6-(2-fluorophenyl)-1-methyl-4H-imidazo[1,5-a][1,4]benzodiazepine (midazolam, Ro 21-3981, Dormicum) is an imidazobenzodiazepine whose salts are soluble and stable in aqueous solution. It has a quick onset and, due to rapid metabolic inactivation, a rather short duration of action in all species studied. Midazolam has a similar pharmacologic potency and broad therapeutic range as diazepam. It produces all the characteristic effects of the benzodiazepine class, i.e., anticonvulsant, anxiolytic, sleep-inducing, muscle relaxant, and "sedative" effects. The magnitude of the anticonflict effect of midazolam is smaller than that of diazepam in rats and squirrel monkeys, probably because a more pronounced sedative component interferes with the increase of punished responses. In rodents, surgical anaesthesia is not attained with midazolam alone even in high i.v. doses, whereas this state is obtained in monkeys. The drug potentiates the effect of various central depressant agents. Midazolam is virtually free of effects on the cardiovascular system in conscious animals and produces only slight decreases in cardiac performance in dogs anaesthetized with barbiturates. No direct effects of the drugs on autonomic functions were found, however, stress-induced autonomic disturbances are prevented, probably by an effect on central regulatory systems. All animal data suggest the usefulness of midazolam as a sleep-inducer and i.v. anaesthetic of rapid onset and short duration.

Mechanism of action of the benzodiazepines: behavioral aspect.

The mechanism of action of benzodiazepines is considered from a behavioral pharmacology perspective, particularly with respect to methods that involve suppression of responding, such as the punishment-conflict model. Four biochemical hypotheses of current interrest are reviewed. 1) Several lines of evidence suggest that the benzodiazepine receptor in brain is related to the antianxiety actions of benzodiazepines, including a high correlation between potency in the binding assay and potency in the test, and some novel nonbenzodiazepine compounds that bind to the receptor and are also active in the conflict tests. 2) The evidence is mixed concerning whether inosine or hypoxanthine may be endogenous ligands for the benzodiazepine receptor. Unlike benzodiazepines, neither inosine nor hypoxanthine antagonizes convulsions induced by pentylenetetraxol in the rat. 3) Much research indicates that gamma-aminobutyric acid (GABA) mediates various electrophysiological and biochemical actions of benzodiazepines. Several studies have also found that picrotoxin or bicuculline block the behavior effects of benzodiazepines, although some conflicting results have been reported. Most available findings suggest that GABA agonists, e.g., muscimol, do not exhibit the antianxiety profile of the benzodiazepines. 4) Additional evidence has accumulated to support the hypothesis that the behavioral actions of the benzodiazepines are mediated by serotonin, possibly with the involvement of GABA.

Conflict behavior in the squirrel monkey: effects of chlordiazepoxide, diazepam and N-desmethyldiazepam.

Dose-response profiles were determined for chlordiazepoxide, diazepam and N-desmethyldiazepam in a squirrel monkey punishment (conflict) procedure. The monkeys were trained to lever press under a food-maintained concurrent schedule consisting of an unpunished 6-minute variable interval (VI) schedule, and a 1.5-minute VI schedule, on which responses were punished intermittently (24 response variable ratio) with electric footshocks. The three benzodiazepines effectively increased responding that had been suppressed by punishment; they had inverted U-shaped dose-effect curves. The minimum effective doses for increasing punished responding were: diazepam less than or equal to 0.31 mg/kg p.o.; N-desmethyldiazepam = chlordiazepoxide = 0.62 mg/kg. As a model to assess potential antianxiety activity, this procedure possessed excellent sensitivity and reliability. The following observations were also made. 1) During initial training, as shock intensity was increased and punished responding became suppressed, some monkeys exhibited an increase in unpunished response rates. This may have represented "positive behavioral contrast," but response rate changes were associated with changes in the amount of time the monkeys allocated to each schedule. 2) At certain dose levels, all three compounds exerted antipunishment effects 24 hours after administration. 3) As was reported previously for rats, when the monkeys had no previous drug experience ("drug-naive") they were more sensitive to the depressant effects of the benzodiazepines. With repeated administration, there was a reduction in this sedation and a concomitant increase in the antipunishment effect. This phenomenon was dose- and animal-dependent.

Reinforcement schedules and extrapolations to humans from animals in behavioral pharmacology.

Behavior controlled by various schedules of reinforcement is useful for characterizing drugs as well as for analyzing the mechanisms of action of their effects on behavior. Conditioned avoidance techniques have been useful for studying neuroleptics and for predicting their clinical antipsychotic acitivity; the possible involvement of dopaminergic mechanisms in the effect of neurolpetics on avoidance behavior is discussed. Tricyclic antidepressant agents have been studied in assays involving interactions with other agents, such as cocaine, amphetamine and tetrabenazine. One type of operant behavior, Sidman avoidance, has been used as particularly sensitive assay for such drug interactions. Another schedule, in which "observing" responses in pigeons are measured. seems to provide a method for studying antidepressants without involving drug interaction phenomena. For tricyclic compounds, facilitation of observing responses and weak potency of conditioned avoidance inhibition constitute a pharmacological profile that seems to have some predictive value for clinical imipramine-like antidepressant activity. "Conflict (punishment) schedules have been useful for predicting antianxiety activity in man. Although the degree of anticonflict effect observed is consistent with Dew's rate dependency hypothesis, this principle does not fully account for the observed drug effects. In the conflict model, the actions of benzodiazepines differ in drug-naive versus drug-experienced animals. Experiments with parachlorophenylalnine have not yet provided clear support for the postulated role of serotonin in related phenomena.

Behavioral analysis of the effects and mechanisms of action of benzodiazepines.

Conflict behavior is a powerful tool to reveal relevant pharmacologic correlates of the therapeutically desirable properties of benzodiazepine antianxiety compounds. The predictability of its clinical effectiveness in psychoneuroses is very high, and important quantitative and qualitative differences between compounds in this chemical class can be shown using such a behavioral technique. It is important to take these differences into account when studying biochemical correlates or mechanisms of action within the benzodiazepine class. Conflict behavior has been used to evaluate several biochemical hypotheses concerning the mechanism of action for the antianxiety properties of benzodiazepines. These studies found that inhibition of cyclic AMP phosphodiesterase did not seem to be a relevant factor. Similarly, experiments with AOAA did not provide evidence to support the involvement of GABA with benzodiazepines' antianxiety properties, nor did glycine receptor affinities correlate significantly with anticonflict effects. Thus, while it is reasonable at present to associate either GABA or glycine with the muscle-relaxant or anticonvulsant properties of benzodiazepines, no support was provided in the conflict procedure for their involvement in the anxiolytic effects. Partial support was provided for the proposal that serotonin is involved in the benzodiazepines' anxiolytic activity. This was determined in studies with the serotonin antagonists cinanserin and methysergide, which had anti-conflict activity, as well as in studies of monoamine turnover after initial chlordiazepoxide treatments to previously undrugged, conflict-trained rats.