A comparison of the physicochemical and biological properties of mirtazapine and mianserin.

Although the chemical structures of the antidepressants mirtazapine and mianserin are closely related there are considerable differences in their biological properties. To find an explanation of this, various physicochemical properties of mirtazapine and mianserin were measured or calculated. Isosteric replacement of CH in mianserin by N in mirtazapine has profound effects on physicochemical properties. The charge distributions as indicated by NMR and calculated by semi-empirical quantum mechanics differ, not only for the changed aromatic A-ring (as expected), but also in other regions of the molecule. The N5 atom in particular, which is conjugated to the changed aromatic ring, is less negatively charged in mirtazapine than in mianserin. Consequently the oxidation potential of mirtazapine is significantly higher than that of mianserin. Another result of this difference in charge distribution is that the (calculated) dipole-moment vectors of the compounds are oriented roughly perpendicular to each other. The dipole moment of mirtazapine is, moreover, three times larger than that of mianserin; mirtazapine is, therefore, more polar than mianserin and this is reflected in a lower retention index. Finally, the basicity of mirtazapine, expressed as the pKa value, is slightly but significantly lower than that of mianserin. The observed differences between the physicochemical properties of mirtazapine and mianserin result in different interactions of these two antidepressants with macromolecules, such as receptors, transporters and metabolizing enzymes; this might explain the differences observed in pharmacological activity and metabolic and kinetic behaviour, that is, the reduced affinity for the alpha 1-adrenoceptor and negligible noradrenaline reuptake of mirtazapine compared with mianserin.

Novel receptor site involved in enhancement of stimulus-induced acetylcholine, dopamine, and serotonin release.

The cognitive enhancer DuP 996 [3,3-bis(4-pyrindinylmethyl)-1-phenylindolin-2-one] and its structural analogs enhance the K(+)-stimulated release of acetylcholine, dopamine, and serotonin in brain slices, without effect on basal release. A novel receptor site labeled by [3H]DuP 996 has been identified. The [3H]DuP 996 binding site has a Kd of 19 nM and a Bmax of 102 fmol/mg of protein. Binding to this site is specific, saturable, reversible, and time, pH, and temperature dependent. Specific binding is decreased by treatment with trypsin and not affected by phospholipase C. Specific binding is inhibited by Ca2+ and increased by Mn2+ but not affected by Na+, K+, or Mg2+. The [3H]DuP 996 binding sites are heterogeneously distributed in brain, with striatum and hypothalamus having highest density and cerebellum lowest. The [3H]DuP 996 binding site does not belong to any known class of receptor site, because [3H]DuP 996 binding could not be displaced by a broad variety of standard pharmacological agents and neuropeptides. Physiological significance of this binding site is suggested by the excellent correlation between the binding affinity for this site and the potency to enhance K(+)-stimulated release of acetylcholine for a series of DuP 996 analogs. Ligands for this receptor site may have therapeutic potential for the treatment of cognitive deficits and neurodegenerative diseases.

A Parkinson-like neurologic deficit in primates is caused by a novel 4-substituted piperidine.

A neurologic deficit characterized by hypokinesia, postural flexion, and to a lesser extent, rigidity, tremor and myoclonus, has been observed in cynomolgus monkeys following administration of 1-methyl-4-(1-methylpyrrol-2-yl)-4-piperidinol (MMPP), a novel 4-substituted piperidine. The syndrome, similar to that described for 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), developed within 3-7 days after oral or i.v. dosing, and was accompanied by lesions in the substantia nigra. The behavioral syndrome was seen to a lesser extent in dogs but not in rats. MMPP contains a hydroxyl group on the 4-position of the pyridine ring; the corresponding dehydration product was inactive.

Response changes after repeated low apomorphine: dopamine autoreceptor desensitization or learning?

Repeated injection of rats with low doses of apomorphine (APO), which selectively interact with dopamine (DA) autoreceptors, caused a change in yawning responses that suggests initial low-APO-induced desensitization of DA autoreceptors, followed by a long-lasting rebound hypersensitivity. Repeated treatment with low APO followed by open-field testing, however, yielded totally different results. APO accelerated intrasession response decrement and upon repeated administration enhanced the intersession response decrement. Both for yawning and open-field behavior, the response change after the second dose of APO was only evident when the first as well as the second APO injection were followed by exposure of the rat to the same test situation. These results indicate that response changes after repeated treatment with low APO are not due to a simple DA-agonist-induced change in receptor sensitivity but that drug experience combined with environmental influences play a decisive role.

Dopamine uptake by rat striatal synaptosomes: time- and temperature-dependent decay and protection by dithiothreitol and dopamine.

The uptake of [3H]dopamine (DA) into rat striatal synaptosomes in the presence of a monoamine oxidase inhibitor was studied using a filtration technique. After a 10-min preincubation period, a fast initial uptake of [3H]DA was seen. Uptake reached a maximum after 4 min of incubation. If incubation was continued for more than 7 min, a gradual decrease in synaptosomal [3H]DA levels was found. Uptake was dependent on preincubation time; initial uptake velocity and maximal uptake decreased irreversibly with increasing preincubation periods. Moreover, the capacity of the synaptosomes to retain the [3H]DA during longer incubation times was progressively affected. The decrease in initial uptake activity was due to a decrease in the Vmax of the transport system. Dithiothreitol (2.8 mM) protected synaptosomal uptake activity against deterioration at 37 degrees C. Also, DA itself (10(-7)M) stabilized the uptake mechanism if added to the suspension before preincubation was started. Since [3H]DA uptake observed after loading the synaptosomes with labeled DA was similar to the uptake seen if the synaptosomes were not previously loaded with DA, it was concluded that under these conditions synaptosomal DA is completely exchangeable with exogenous substrate. Prolonged storage of the synaptosomes at 0 degree C also resulted in a time-dependent decrease in uptake activity (t1/2 = 116 min). The addition of unlabeled DA or dithiothreitol to the suspension did not affect instability at 0 degree C.

Interaction between dopamine influx and efflux in rat striatal synaptosomes.

Exchange release has previously been proposed as the mechanism by which dopamine is transported over the synaptosomal membrane. An increase in carrier-mediated dopamine release should therefore result in an enhanced rate of synaptosomal dopamine uptake. In order to test this hypothesis, rat striatal synaptosomes were incubated with 14C-dopamine until equilibrium. Then, the 14C-dopamine concentration in the medium was reduced in order to elicit various rates of net dopamine efflux, while influx was monitored using tracer amounts of 3H-dopamine. Dopamine release was concentration dependent and saturable and thus may be carrier-mediated. In contrast to that expected for a mechanism of exchange release, dopamine influx was depressed as compared to equilibrium conditions. Furthermore, nomifensine, a specific inhibitor of dopamine influx, did not attenuate dopamine efflux. It is concluded that dopamine transport over the synaptosomal membrane may not be via a mechanism of strict exchange release.

Synaptosomal uptake studies on recombinant inbred mice; neurotransmitter interaction and behavioral correlates.

The uptake of noradrenaline, choline and GABA into whole brain synaptosomes from the mice inbred strains C57BL/6By, BALB/cBy, their reciprocal crosses and seven recombinant inbred strains derived from the F2 crosses was measured in order to study the relation between different neurotransmitter systems and their possible correlations with behavioral parameters. Analysis of variance demonstrated significant differences in both Vmax and Km for each substrate between the inbred strains studied. Only the Km for GABA uptake did not reach significance although a strong trend (P = 0.07) was apparent. For none of the uptake parameters clear strain distribution patterns were found. A positive phenotypic as well as genotypic correlation was found between the Vmax and Km for noradrenaline, choline and GABA uptake. A genetic analysis showed a positive correlation between the Vmax for noradrenaline and choline uptake. Since prior to these neurochemical studies, all subjects were tested in a variety of behavioral paradigms, phenotypic and genotypic correlations between behavioral and neurochemical parameters could be calculated. The most consistent correlations were found between noradrenaline and choline uptake and water consumption and the preference ratio as measured in a conditioned taste aversion test.

"On" and "off" responses of K+-induced synaptosomal proline release: involvement of the sodium pump.

The K+-induced release of the putative neurotransmitter proline in rat cerebral cortex synaptosomes was studied by measuring [5-3H]proline released upon exposure to high-K+ media. In contrast to previous experiments, K+ was not substituted for Na+, which was held constant. Under the resulting conditions of increased osmolarity, two peaks of proline release were obtained, one upon exposure to increased [K+], another upon restoring normal [K+]. The dependence of the latter peak on ionic composition of the medium, and its sensitivity to ouabain are discussed.

Memory consolidation in senescence: effects of CO2, amphetamine and morphine.

Alterations in memory storage processes that occur in senescence were investigated by challenging young and old female "small Wistar" rats with posttraining administration of CO2, amphetamine or morphine, and measuring retention performance. Neither duration of CO2 immersion, nor the time of CO2 immersion after training had a differential amnestic effect with age on retention of a one-trial, shock-motivated inhibitory avoidance task. These results indicate that the times during which memory is susceptible to disruption for old and young rats are similar. Challenge with drugs, however, did reveal age-related alterations in memory storage processes. Amphetamine attenuated CO2-induced amnesia in young rats, but had no effect in old rats. This could not be attributed to a general decline in response to amphetamine in old rats because amphetamine increased open field activity of both young and old animals. Morphine also had a differential effect on memory with age: it caused amnesia in old rats trained in a one-trial hot plate escape task, while having no effect on retention performance of young rats. Thus, the modulatory influence of catecholamine and opioid systems on memory processes is probably altered in senescence.

Presynaptic alpha-block and inhibition of noradrenaline and 5-hydroxytryptamine reuptake by a series of compounds related to mianserin.

A structure-activity relationship study was undertaken for a variety of structural analogues of the tetracyclic antidepressant mianserin. Presynaptic alpha-blocking activity in vitro was evaluated measuring the potentiation of depolarization-induced noradrenaline (NA) release from rat cerebral cortex slices. Inhibition of NA and 5-hydroxytryptamine reuptake was measured in rat hypothalamic or striatal synaptosomes, respectively. Presynaptic alpha-blockade was only found in molecules with an overall bent shape. Flat rigid molecules or flexible ones were not active. Six-membered, chair-formed D-rings (containing the -NCH3 moiety) appeared better than 5- or 7-membered ones. Heteroatom substitution, but not hydroxylation or methylation, of the bridge between the two aromatic rings left presynaptic alpha-blockade unaffected. N-Demethylation and aromatic methyl- or chlorine-substitution reduced presynaptic alpha-blockade. In pyridine ring-substituted analogues the localization of the heteroatom appeared to be crucial. 5-Hydroxytryptamine reuptake inhibitory activity was only found in desmethylmianserin. NA uptake inhibition was found in many mianserin analogues, especially those with an exocyclic-N(CH3)2 moiety. Structure activity relationships for NA reuptake inhibition differed from those for presynaptic alpha-blockade and were generally less stringent. For both properties simple additivity relationships appeared to be absent.

Detailed analysis of the effects of apomorphine and d-amphetamine on spontaneous locomotor behaviour of rats as measured in a TV-based, automated open-field system.

Rat open field behaviour was measured in a TV-based, automated system. Habituation was evident in saline-treated rats. Most variables measured declined over a 10 min period. Apomorphine affected rat open-field behaviour bimodally, i.e. low doses of apomorphine (0.02-0.08 mg/kg, s.c.) decreased most aspects of rat open-field behaviour, whereas at higher doses (0.2 and 0.5 mg/kg, s.c.) various aspects of open-field behaviour were stimulated. Rearing and average speed, however, were monotonically depressed. Amphetamine (1 and 2 mg/kg) stimulated most aspects of rats open-field behaviour, including rearing. Speed was not affected by amphetamine. Habituation was more pronounced after low doses of apomorphine than after saline treatment and was absent after high doses of apomorphine and after amphetamine. The results represent a detailed analysis of apomorphine and amphetamine effects on rat open-field behaviour and show that the distinct components of this behaviour are affected differentially.

Differences in presynaptic alpha-blockade, noradrenaline uptake inhibition, and potential antidepressant activity between (+)- and (-)mianserin.

The effect of racemic mianserin on K+-evoked tritium release from rat brain cortex slices previously incubated with 3H-L-noradrenaline was studied. Racemic mianserin (10(-9)--10(-5) M) increased stimulation-induced release dose-dependently. As methysergide, metiamide, and cyproheptadine failed to do so, it was concluded that this effect was probably not caused by the antihistamine or antiserotonin activity of racemic mianserin, but due to its alpha-adrenolytic effect. Evaluation of the effects of the enantiomers (+)(S)mianserin and (-)(R)mianserin showed that the alpha-adrenolytic effect resided in the (+)isomer, whereas the (-)isomer was inactive at a concentration of 10(-6) M. Inhibition of noradrenaline into rat hypothalamic synaptosomes also showed stereospecificity in that (+)mianserin was about 300-times more active than(-)mianserin. Inhibition of rat muricidal behavior, a test for potential antidepressant activity, showed a similar dissociation in the effects of the two enantiomers, in that (+)mianserin was active, whereas (-)mianserin was not.

Changes in rat brain norepinephrine levels and turnover after olfactory bulbectomy.

After bilateral olfactory bulbectomy in rats a significant increase of norepinephrine (NE) level in the hypothalamus was found. However, no difference was observed between hypothalamic NE turnover of bulbectomized and sham operated animals in the amygdaloid cortex the NE level was not affected by bulbectomy. In this area, however, the NE turnover appeared to be decreased after bulbectomy. The latter finding may be related to the deficits in passive avoidance behaviour as found in bulbectomized rats.