Vigilance-controlled quantified EEG in safety pharmacology.

The vigilance-controlled quantified EEG can be used either as a safety, or as a discovery pharmacology procedure. Put strategically into the preclinical development process of a drug, it can be useful for making the decision about the future research direction to be taken. The experimental approach described in this unit is based on the rat EEG. Even though there are considerable differences in function and structure between human and rat brain, the EEG response to psychoactive drugs and convulsants is similar in the two species. Thus, the rat EEG is generally a reliable predictor for human CNS drug effects.

Chronic mild stress-induced anhedonia model of depression; sleep abnormalities and curative effects of electroshock treatment.

A core symptom of human depressive disorder is anhedonia, the loss of interest or pleasure in daily activities. Anhedonia, measured as subsensitivity to reward, can be induced in rats by a regimen of repeated, mild, unpredictable stressors. Here, the hedonic state of rats was assessed using an intracranial self-stimulation (ICSS) procedure. The ICSS frequency threshold was determined before, during and after a period of exposure to the stress regimens. After 13 days of repeated mild stress, the ICSS threshold was significantly increased, suggesting a gradual decrease of sensitivity to reward. This anhedonic state lasted throughout the stress period. When stressed anhedonic animals were given electroshock treatment, the stress-induced increase in ICSS threshold was rapidly and completely reversed. Moreover, biological markers of human depression such as reduced latency to the first REM sleep episode or increased time spent in REM sleep were also found in electroencephalographic recordings of chronically stressed animals. These sleep abnormalities were observed beginning in the second week of a three-week stress regimen and progressively disappeared after termination of stress. In conclusion, these data provide further evidence supporting stress-induced anhedonia in rats as a unique animal model of human depression combining convergent elements of biological, etiological, symptomatological and therapeutic validity.

Benzodiazepines promote the intermediate stage at the expense of paradoxical sleep in the rat.

The effects of diazepam, a long half-life benzodiazepine, midazolam and triazolam, two with short half-life, on the transitional stage between deep slow wave sleep and paradoxical sleep were studied in Wistar and WAG/Rij rats. This intermediate stage is characterized by the unusual association of cortical spindles and low frequency hippocampal theta rhythm. The main result was extension of the intermediate stage at the expense of paradoxical sleep by diazepam and triazolam by influencing only the duration of the intermediate stage and both the onset and maintenance of paradoxical sleep. Midazolam increased both intermediate stage and paradoxical sleep. Several differences in the qualitative modulation of the stage characteristics and between rat strains were found. In regard to the possible peculiar physiological significance of the intermediate stage, we conclude that benzodiazepines promote a transient pharmacological cerveau isolé-like stage during sleep in rats.

Neurochemical and behavioral evidence that Ro 41-9067 is a selective presynaptic dopamine receptor agonist.

(-)-2,3,4a,5,6,10b-Hexahydro-7-hydroxy-2-methylbenzo(f)-quinoline 4(1H)-ethanol (Ro 41-9067) was compared with apomorphine, 2-amino-6-allyl-5,6,7,8-tetrahydro-4H-thiazolo-[4,5-d]-azepine (B-HT 920), lisuride and other dopamine (DA) receptor agonists in a series of tests designed to characterize its pharmacological activity on DA receptors. In vitro binding studies indicated that Ro 41-9067 bound selectively to DA D2 vs. D1 receptors. It also had a moderate affinity for serotonin1A and alpha-2 adrenergic receptors. Ro 41-9067 exhibited a marked agonistic component for the presynaptic DA autoreceptors. Indeed, it caused a dose-related reduction in locomotor activity over a wide dose range and prolonged periods of observation without stimulating locomotor activity, reflecting postsynaptic DA receptor activation, even at the highest doses. Ro 41-9067 inhibited the gamma-butyrolactone-induced increase in I-dopa accumulation in the rat striatum, an effect sensitive to haloperiodol. Ro 41-9067 inhibited K(+)-induced [3H]DA release and significantly reduced the striatal contents of the DA metabolites, dihydroxyphenilacetic acid and homovanillic acid. Furthermore, the compound counteracted stereotyped behavior and locomotor stimulation induced by amphetamine. Finally, Ro 41-9067 did not appear to act on postsynaptic D2 receptors because it, similarly to B-HT 920 but differently from bromocriptine, quinpirole and lisuride, did not change basal or forskolin-stimulated adenylate cyclase activity. Only at very high concentrations Ro 41-9067 increased adenylate cyclase activity, this effect being due to a D1 agonistic component.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochemistry and pharmacology of moclobemide, a prototype RIMA.

RIMA is a term for reversible inhibitors of monoamine oxidase (MAO) with preference for MAO-A; moclobemide is a prototype of this new class of antidepressants and is a highly selective inhibitor of MAO-A in vitro. This inhibition is reversible by dialysis in vitro, which accounts for the dose-dependent duration of in vivo enzyme inhibition of 12-24 h. Moclobemide increases the content of serotonin, noradrenaline and dopamine in the brain, and decreases that of their deaminated metabolites. Its biochemical, neurological and behavioural effects indicate that it increases the extracellular concentration of the classic monoamine neurotransmitters/neuromodulators - in particular 5-HT. Potentiation of the cardiovascular effects of tyramine is less pronounced after taking moclobemide than after irreversible MAO-A inhibitors. Understanding of the physiological role of MAO and of the events that link inhibition of the enzyme with modulation of neuronal activities in the CNS remains incomplete. A major physiological role of intraneuronal MAO is to keep cytosolic amine concentration very low, to enable the neuronal monoamine carriers to produce a net inward transport of monoamines, and thereby to act as the first step in the termination of action of extracellular monoamines. MAO is likely to have a similar function in non-monoaminergic cells with respect to the monoamine carriers they contain. In addition to the classic monoamines, "trace" amines may become functionally active after MAO inhibition. An alternative role for MAO is that of a scavenger, preventing natural substrates from accumulating in monoaminergic neurons and interacting with storage, release, uptake and receptor function of monoamines.

Sleep and EEG spectra in the rabbit under baseline conditions and following sleep deprivation.

The 24-hr sleep-wake distribution and power spectra of the electroencephalogram were determined in rabbits that had been implanted with cortical and hippocampal electrodes. A diurnal preference for sleep was observed. The spectral power density in nonrapid eye movement sleep (NREM sleep) of the cortex showed a decreasing trend in most frequencies within the 12-hr light period. In the 12-hr dim period no clear trend was present. Most hippocampal EEG frequencies decreased in NREM sleep in the first two hours of the light period, and thereafter stayed on a constant level. Sleep deprivation elicited the following changes: a prolonged increase of NREM sleep and a short increase of REM sleep; in the cortex, an increase of slow-wave activity (SWA; power density in the 0.25-2.0 Hz frequency band) in NREM sleep, which declined in the course of recovery; an enhancement of slow-wave (1.25-3 Hz) and theta (6.25-7 Hz) activity in REM sleep. The hippocampus showed an increase in NREM sleep power density in almost all frequencies. In REM sleep the hippocampus exhibited an increase in power density in the 6.25-7 Hz and 12.25-13 Hz bands, whereas in the 7.25-8 Hz band the values were below baseline. The results show that SWA in NREM sleep and theta activity in REM sleep are enhanced by sleep deprivation, as has been observed in other mammalian species. The EEG changes in the hippocampus resembled those in the cortex.

Sleep and EEG slow-wave activity in the domestic cat: effect of sleep deprivation.

The 24-h sleep-wake distribution, the vigilance states and the time course of EEG slow-wave activity was investigated in 8 adult domestic cats individually maintained in isolation under 14-h light (06.00-20.00 h)/10-h dim conditions. The frontal EEG and motor activity were continuously recorded for 22 h (during the daily cleaning and food-replenishing period between 07.00 and 09.00 h the cats were only observed). Sleep was prevented for 14 h (07.00-21.00 h) by playing with the animals. Recovery from sleep deprivation was recorded for the remaining 10 h of the dim period. Non-rapid eye movement (non-REM) sleep, REM sleep and waking were uniformly distributed over the light and dim period, with the exception of the 2-h feeding period where the cats were always awake. EEG power density in the delta band (0.75-4.5 Hz; slow-wave activity) was computed in non-REM sleep. The values in the light period were higher than in the dim period. In neither of the lighting periods was a trend observed. After sleep deprivation a small increase of non-REM sleep and REM sleep was present. EEG slow-wave activity was initially enhanced and then declined progressively. We conclude that despite the small circadian difference in the sleep-wake pattern observed in our cats, sleep homeostasis is similar to that observed in other mammalian species.

Pharmacological profile of moclobemide, a short-acting and reversible inhibitor of monoamine oxidase type A.

The novel antidepressant moclobemide is a reversible inhibitor of monoamine oxidase (MAO), preferentially of type A. Moclomide was active in three animal models considered predictive for antidepressant activity: 1) it prevented dose-dependently akinesia and blepharospasm induced in mice and rats by Ro 4-1284, a short-acting amine releasing agent. Prevention of akinesia by moclobemide also depended upon the dose of Ro 4-1284. For comparison also, effects of cimoxatone, harmaline, tranylcypromine and clorgyline are presented: 2) in cats, it selectively and dose-dependently suppressed rapid eye movement sleep without disturbing the sleep-wakefulness cycle; and 3) in the behavioral despair test in mice, it decreased the immobility score to a similar degree as amitriptyline or imipramine. In addition, moclobemide potentiated 5-hydroxytryptophan-induced stereotypies in rats with a potency similar to cimoxatone and with a duration of action of less than 24 hr. Moclobemide had almost no effect on the spontaneous behavior in mice, rats, cats and monkeys. Only in higher doses, marginal sedation and slight impairment in motor performance were seen. Moclobemide did not prevent pilcarpine-induced salivation in mice, demonstrating the absence of anticholinergic activity. Blood pressure and heart rate of freely moving, spontaneously hypertensive rats were only slightly decreased for less than 3 hr. Moclobemide moderately potentiated the pressor effect of p.o. tyramine in rats. In conclusion, the reversible MAO inhibitor moclobemide is active in animal models sensitive to all major drugs used in the treatment of depression. In contrast to imipramine-like antidepressants, it lacks anticholinergic activity and it differs from classic MAO inhibitors by potentiating only weakly the pressor effect of p.o. tyramine.

L-cycloserine: behavioural and biochemical effects after single and repeated administration to mice, rats and cats.

L-Cycloserine dose-dependently inhibited the activity of gamma-aminobutyric acid (GABA)-transaminase (GABA-T) and elevated the level of GABA in whole mouse brain with a peak effect 3-4 hr after a single intraperitoneal injection. At a dose (30 mg/kg) which elevated the level of GABA almost 4-fold, L-cycloserine moderately increased the content of alanine and slightly reduced that of aspartate, glutamate and glycine in the brain. L-Cycloserine (10-30 mg/kg, p.o. or i.p.) prevented tonic seizures induced by 3-mercaptopropionic acid (3-MPA) and audiogenic seizures in DBA/2 mice, without affecting those evoked by pentylenetetrazol, bicuculline and electroshock. Similarly small doses of L-cycloserine reduced the level of cGMP in the cerebellum of rats, prevented its elevation by 3-MPA and attenuated the hypothalamically-elicited rage reaction in cats. Larger doses of L-cycloserine (greater than 30-100 mg/kg) impaired the performance of mice in the rotarod, chimney and horizontal wire tests, and reduced spontaneous locomotor activity of rats. Upon repeated administration the inhibitory effect of L-cycloserine on the activity of GABA-T and on seizures elicited by 3-MPA in mice increased. In contrast, the depressant action of L-cycloserine on motor performance and locomotion declined in subchronically-treated mice and rats. The levels of amino acids in brain after repeated administration did not differ markedly from those in acutely-treated mice. It is suggested that small doses of L-cycloserine, probably by increasing GABAergic inhibition, reduce hyperexcitability in the brain in acute- and subchronically-treated animals. Larger doses of L-cycloserine, possibly by inducing multiple neurochemical changes, evoke central depressant effects which diminish during subchronic treatment.

Serotonin involvement in lisuride-induced mounting and in sleep.

Mounting behavior of rats induced by several drugs, such as e.g. p-CPA, 5,7-dihydroxytryptamine, L-DOPA or lisuride, appears to result from a combined decrease of 5-hydroxytryptamine (5-HT) and increase of dopamine (DA) neurotransmission. In this paper, lisuride-induced mounting is proposed as a behavioral model for detecting pharmacologically active drugs that interact with monoaminergic mechanisms, e.g. 5-HT reuptake blockers and type A monoamine oxidase inhibitors. With regard to sleep, 5-HT appears to sustain a relevant part in controlling the sleep-wakefulness cycle. However, other transmitters or neuromodulators (catecholamines, oligopeptides etc.) may also be involved in sleep mechanisms.

Some pharmacological effects of delta-sleep-inducing peptide (DSIP).

The synthetic nonapeptide DSIP was studied in rabbits and cats under normal conditions and under conditions of disturbed sleep. In other experiments, the effect of the oligopeptide on withdrawal jumping provoked by naloxone in morphine-dependent mice was studied. In rabbits, DSIP at 25 micrograms X kg-1 i.v. and 1 mg X kg-1 s.c. augmented spindle-dominated, light nonREM sleep and prevented hyposomnia after a stressful situation. In cats, 25 micrograms X kg-1 i.v. and 100 micrograms X kg-1 s.c. preferentially augmented REM sleep and abolished the sleep suppressant effect of morphine. In morphine-dependent mice, 25.5 micrograms X kg-1 i.v. as well as doses beyond 85 micrograms X kg-1 s.c. attenuated naloxone-induced withdrawal jumping. In most experimental situations, indications for bell-shaped dose-response curves of DSIP were found.

Effects of oral and intravenous midazolam, triazolam and flunitrazepam on the sleep-wakefulness cycle of rabbits.

The effect of the new, short-acting benzodiazepine, midazolam as well as that of triazolam and flunitrazepam on the sleep of rabbits was recorded for 6 h. Midazolam at 1 mg kg-1 i.v. augmented both rapid eye movement sleep (REMS) and non-REM sleep (NREMS) only in the first half of the observation period. At 10 mg kg-1 i.v., NREMS was further increased in the first and, to a lesser degree, in the second 3-h period, while REMS was suppressed. Both doses were less effective orally than intravenously. Qualitatively, the effect of triazolam 0.01 and 0.1 mg kg-1 i.v. was very similar to that of the corresponding low and high intravenous doses of midazolam, except that the high dose of triazolam had a prolonged effect on total sleep time. Like midazolam, triazolam was substantially less effective orally than intravenously. Flunitrazepam at 0.1 and 1 mg kg-1 i.v. produced almost the same effects as midazolam and triazolam at the respective low and high intravenous doses, but had a longer duration of action. In contrast to midazolam and triazolam, flunitrazepam was almost as active orally as intravenously.

Electrophysiological studies on the specific benzodiazepine antagonist Ro 15-1788.

This is an electrophysiological study in cats and rats of the imidazobenzodiazepinone derivative, Ro 15-1788, the first representative of specific benzodiazepine antagonists. (1) In unanaesthetized spinal cats, 1-10 mg kg-1 Ro 15-1788 i.v. did not affect segmental dorsal root potentials (DRPs), polysynaptic ventral root reflexes (VRRs), Renshaw cell responses to antidromic ventral root volleys and spontaneous gamma-motoneurone activity. However, at 1 mg kg-1 i.v., it antagonized the enhancement of DRPs as well as the depression of polysynaptic VRRs, Renshaw cell discharges and gamma-motoneurone activity induced by meclonazepam (0.1 mg kg-1 i.v.), diazepam (0.3 mg kg-1 i.v.) or zopiclone (1 mg kg-1 i.v.). The same dose of Ro 15-1788 failed to reduce similar effects of phenobarbital (10 mg kg-1 i.v.) on spinal cord activities. (2)In unanaesthetized "encéphale isole" rats, 3 mg kg-1 Ro 15-1788 i.v. abolished the decrease induced by 5 mg kg-1 midazolam i.v. of spontaneous multiunit activity (MUA) in the substantia nigra pars compacta, nucleus raphé dorsalis, nucleus locus coeruleus and the CAl area of the hippocampus dorsalis, but not the decrease produced by 10mg kg-1 pentobarbital i.v. Ro 15-1788 (12mg kg-1 i.v.) by itself did not affect MUA in the substantia nigra, but slightly depressed MUA in the other 3 areas. (3) In intact immobilized rats, the increase of power induced by 1 mg kg-1 flunitrazepam i.v. in the 0.5-48 Hz range of the electrocorticogram as well as in the 0.5-8 Hz, 8-32 Hz and 32-48 Hz frequency bands was transiently abolished by 5 mg kg-1 Ro 15-1788 i.v. (4) In unrestrained cats, 5 mg kg-1 Ro 15-1788 i.p. had no effect on the electrical threshold for eliciting a rage reaction evoked by electric hypothalamic stimulation, but abolished the threshold increase caused by 1 mg kg-1 diazepam i.p. These results are in line with biochemical and behavioural findings and support the selective antagonism by Ro 15-1788 of central effects of benzodiazepines through an interaction at benzodiazepine receptors.

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.

Effects of intracerebroventricularly injected "nerveside" on free behaviour and EEG activity of rats. A pilot study.

"Nerveside", a phosphopeptide extracted from ox and dog brains by Toh, was injected into one lateral ventricle of four rats previously prepared for the injection as well as for recordings of the EEG of the dorsal hippocampus (field CA1) and sensorimotor cortex. "Nerveside" in doses of 150 and 300 micrograms/animal induced a transient stereotyped behaviour and longer lasting alterations of the EEG activity and vigilance pattern. Wakefulness was increased, slow wave sleep and REM-sleep were diminished. The vigilance pattern was characterized by rapid shifts between all three vigilance states. The effects of "nerveside" were visible for about three hours.

[Pharmacological investigations with the combination sulfamoxole/trimethoprim, a new broadspectrum chemotherapeutic agent (author's transl)]. / Pharmakologische Untersuchungen zur Verträglichkeit der Kombination Sulfamoxol/Trimethoprim (CN 3123), eines neuen Breitbandchemotherapeutikums

Investigations were conducted with the combination of N1-(4,5-dimethyl-2-oxazolyl)-sulfanilamide (sulfamoxole) and 2,4-diamino-5-(3,4,5-trimethoxy-benzyl)-pyrimidine (trimethoprim) (CN 3123, Nevin, Supristol) in a dose ratio of 5:1, with respect to pharmacological activity and possible side effects. The effects obtained with the combination CN 3123 were compared with those of the single substances. In a dose range comparable to that as used in clinical treatment, there were no effects on cardiovascular or respiratory functions, on functions of autonomic and central nervous system, on contractility of smooth muscles and on data of clinical chemistry such as urine and electrolyte excretion, blood sugar, blood coagulation and liver function tests. Doses which are 5 to 10 times higher than the initial dose or 10 to 20 times higher than the maintenance dose used in man caused an increase of urine and sodium excretion without influencing potassium and chloride output. There were no signs of sedation as alteration of motility or EEG patterns, but in mice and rats there was an increase in both duration and depth of anaesthesia caused by barbiturates or ether. Only in a dose range 30 to 40 times higher than the initial dose for man there were some slight alterations with respect to cardiovascular system and liver function tests. In vitro, with high concentrations of CN 3123 there was a weak, unspecific spasmolytic effect on the isolated ureter and an increase in the refractory period of the guinea pig atrium. There were no hints that the side effects seen with separate administration of high or very high doses of sulfamoxole or trimethoprim were increased or poteniated by their simultaneous administration. Slight side effects in animals were only observed with doses exceeding the tenfold of the doses for therapeutic use in men. Therefore, the therapeutic range of CN 3123 seems to be more than adequate.