Partial dopamine-agonistic and atypical neuroleptic properties of the amino-ergolines SDZ 208-911 and SDZ 208-912

The aminoergolines SDZ 208-911 [N-[(8-alpha)-2,6-dimethylergoline-8-yl]-2,2- dimethylpropanamide] and SDZ 208-912 [N-[8-alpha)-2-chloro-6-methylergoline-8-yl]- 2,2-dimethylpropanamide] exhibit nonclassical, neuroleptic-like properties in rodents. Thus, they are equipotent to haloperidol as inhibitors of apomorphine-induced gnawing behavior and conditioned avoidance responding, but are essentially devoid of cataleptogenic activity. In addition, they show high affinity for central D-2 receptors in vitro and elevate striatal homovanillic acid levels. In contrast to haloperidol, however, SDZ 208-911 and 208-912 strongly inhibit prolactin secretion and induce contralateral circling behavior in 6-hydroxydopamine-lesioned animals. These profiles are consistent with the drugs exhibiting varying degrees of partial agonistic activity at dopamine D-2 receptors, with SDZ 208-911 being considerably more agonistic than SDZ 208-912. Support for this contention stems from the ability of SDZ 208-911 to reduce the elevation of striatal L-dopa formation induced by gamma-butyrolactone, and SDZ 208-912's partial reversal of apomorphine's inhibitory action on gamma-butyrolactone activity. SDZ 208-911's effects are reduced after the partial alkylation of D-2 receptors with N-ethoxy-carbonyl-2-ethoxy-1,2-dihydroquinoline, confirming its partial agonistic properties. SDZ 208-911 and SDZ 208-912 could be effective against both the positive and negative symptoms of schizophrenia, while exhibiting a reduced incidence of dystonic and parkinsonian side-effects. In addition, their clinical testing might throw more light on the central dopaminergic status of schizophrenic subjects.     

Uptake and metabolism of catecholamines in rat brain synaptosomes: Studies on the contribution of monoamine oxidase

The uptake of catecholamines into rat brain synaptosomes was studied without suppressing MAO activity. Control experiments confirmed that in our purified synaptosomal preparations catecholamine metabolism was attributable to intrasynaptosomal MAO activity only. Thus, total uptake was defined as the sum of accumulated and metabolised amine. The following results were found. (1) Accumulation of dopamine (DA) as well as norepinephrine (NE) reached a saturation level after about 20 min. Deaminated products were formed in a linear fashion over time. However, NE metabolites constituted less than 20% of the total NE taken up at 20 min, while DA metabolites were an important part of the total DA taken up (44 and 77% at 2 and 20 min, respectively). (2) The distribution of deaminated metabolites between the intrasynaptosomal compartment and the incubation medium was consistent with the assumption that they were released from the synaptosomes by passive diffusion. (3) Kinetic investigations of active transport gave K_m values of 1 × 10^?7 M for DA uptake and 5 × 10^?7 M for NE uptake. For DA, but not for NE, results for maximal velocities were considerably higher when calculated on the basis of the sum of accumulation and metabolism, compared to accumulation alone. (4) Blocking MAO activity resulted in a reduced total DA uptake to approximately the same level of accumulation as in controls. Total NE uptake was not significantly affected by MAO inhibition. (5) In synaptosomes from reserpinized animals, a significant decrease of accumulation of both DA and NE was found. In the case of DA this reduction was largely compensated by increased MAO activity, while with NE the decrease of storage resulted in a lower total uptake of NE.It is concluded from these results that intrasynaptosomal MAO activity allows removal of DA from the extracellular space under conditions in which equilibrium between inward and outward fluxes would otherwise abolish net uptake of DA. With NE, however, MAO activity was too low to accomplish this function, and the fate of this amine subsequent to its uptake seems to depend mainly on intracellular storage.     

Alterations in opiate receptor function after chronic ethanol exposure

The dose-response curve for morphine-induced stimulation of striatal dopamine metabolism was shifted to the right in mice which had been withdrawn for 24 hours after chronic consumption of an ethanol-containing liquid diet. The apparent ED50 for morphine was increased by 33% in ethanol-treated mice. Concomitant with the shift in the dose-response curve, the affinity for dihydromorphine of the high-affinity caudate morphine receptor was decreased in ethanol-treated mice. The change in receptor properties after ethanol treatment included a decreased sensitivity of the receptor to the effects of sodium ion on morphine binding. The results suggest: 1) that the effect of morphine on dopamine metabolism in the mouse striatum is, at least in part, mediated by receptors that exhibit a high affinity for dihydromorphine: and 2) that ethanol treatment and withdrawal may induce specific changes in these particular opiate receptors.     

Studies on the subcellular localization of monoamine oxidase types A and B and its importance for the deamination of dopamine in the rat brain

The distribution of the MAO-forms A and B between intra- and extrasynaptosomal rat brain mitochondria was studied with the aid of their known substrate and inhibitor specificities. The activities with the selective substrates serotonin, PEA and benzylamine indicated that intrasynaptosomal mitochondria have about a 3.4-fold higher MAO A:MAO B ratio than extrasynaptosomal mitochondria. However, PEA was found to be a selective substrate for MAO B only at low concentrations (such as 5 × 10^?6M), whereas at higher concentrations (such as 10^?3M) it was a substrate for both forms of MAO. The different ratios of the two enzyme forms in the two mitochondrial populations were confirmed when the selective inhibitors clorgyline and deprenyl were used with dopamine or 10^?3M PEA. With these two amines, the ratios of MAO A: MAO B activities were 3–4.5 times higher in intrasynaptosomal than in extrasynaptosomal mitochondria. In particular, when the activity with dopamine was measured in intact synaptosomes, deamination being preceded by a specific uptake into these particles, the inhibitor sensitivities clearly showed that MAO activity was almost exclusively attributable to the A-form of the enzyme. Thus, mitochondria in the terminals of dopaminergic neurones have an even more pronounced enrichment in MAO A than the mitochondria obtained by osmotic lysis of a total brain synaptosomal preparation. It was also found that clorgyline and deprenyl have an inhibitory effect on the uptake of dopamine into nerve endings with IC₅₀ values in the range of 10^?5 to 10^?4M. These results are discussed in terms of possible physiological significancies of the properties and distribution of the two forms of MAO.     

Receptor activation involving positive allosteric modulation, unlike full agonism, does not result in GABAB receptor desensitization

Allosteric modulators act more physiologically than orthosteric ligands, targeting only endogenously activated receptors and not their whole population, which is why they are expected to produce less side effects and tolerance. To inspect the role of the positive allosteric modulator GS39783 in GABA_B receptor desensitization, we examined receptor function and cell surface expression in a recombinant GABA_B cell line and in primary neuronal cultures upon persistent treatments with GABA_B agonists, and combinations of agonists and GS39783. The potency of GABA to inhibit 7β-forskolin-induced cAMP formation in recombinant cells decreased after the exposure to a saturating GABA concentration, but not after a combination of a low GABA concentration and GS39783, that activated the receptor to the same extent. Concordantly, a significant decrease of cell surface receptors was found after GABA-induced desensitization, unlike after the combined treatment with GABA and GS39783. Similar observations regarding receptor function were found in primary neurons for baclofen-induced inhibition of spontaneous Ca²⁺ oscillations. However, the cell surface receptor density remained unaffected upon baclofen-induced desensitization in the primary neurons, possibly due to different mechanisms of desensitization in the neurons and the recombinant cell line. These findings indicate that the degree of occupancy of the orthosteric site determines desensitization rather than the degree of receptor activation. In summary, our results conform to predictions that positive allosteric modulators have less propensity for the development of tolerance due to receptor desensitization than classical agonists.     

Allosteric modulation of family C G-protein-coupled receptors: from molecular insights to therapeutic perspectives

Allosteric receptor modulation is an attractive concept in drug targeting because it offers important potential advantages over conventional orthosteric agonism or antagonism. Allosteric ligands modulate receptor function by binding to a site distinct from the recognition site for the endogenous agonist. They often have no effect on their own and therefore act only in conjunction with physiological receptor activation. This article reviews the current status of allosteric modulation at family C G-protein coupled receptors in the light of their specific structural features on the one hand and current concepts in receptor theory on the other hand. Family C G-protein-coupled receptors are characterized by a large extracellular domain containing the orthosteric agonist binding site known as the "venus flytrap module" because of its bilobal structure and the dynamics of its activation mechanism. Mutational analysis and chimeric constructs have revealed that allosteric modulators of the calcium-sensing, metabotropic glutamate and GABA(B) receptors bind to the seven transmembrane domain, through which they modify signal transduction after receptor activation. This is in contrast to taste-enhancing molecules, which bind to different parts of sweet and umami receptors. The complexity of interactions between orthosteric and allosteric ligands is revealed by a number of adequate biochemical and electrophysiological assay systems. Many allosteric family C GPCR modulators show in vivo efficacy in behavioral models for a variety of clinical indications. The positive allosteric calcium sensing receptor modulator cinacalcet is the first drug of this type to enter the market and therefore provides proof of principle in humans.     

A dual microtiter plate (192 sample) luminometer employing computer-aided single-photon imaging applicable to cellular luminescence and luminescence immunoassay

In spite of the increasing applications of luminescence measurements, no instrumentation is yet commercially available that permits sensitive luminescence measurements to be performed simultaneously on a large number of samples. Based on the principle of single-photon imaging, we describe here the performance of a computer-aided image luminometer with a capacity for two microtiter plates. Applications to the study of chemiluminescence by a B lymphocyte cell line, and to an IgE luminescence immunoassay are used to exemplify the capabilities of the system, which we have termed 'chemiluminescence multiwell analyser'.     

Biochemical and pharmacological effects of fluperlapine on noradrenaline and acetylcholine systems in some rodent,bovine and crustacean preparations

Summary Fluperlapine was compared with clozapine, chlorpromazine, haloperidol and imipramine regarding its effects on some cholinergic and noradrenergic animal systems.Fluperlapine and clozapine showed the most pronounced anticholinergic effects. Fluperlapine was equipotent with clozapine in displacing [³H]-QNB from muscarinic receptors of the calf cerebral cortex (IC₅₀ about 15 nM). In the mydriasis test in the mouse and in the crayfish hindgut bioassay the differences between fluperlapine and clozapine were small.Like the other antischizophrenic drugs tested, fluperlapine displayed a marked affinity for ₁-adrenoceptors (calf cerebral cortex: IC₅₀ about 10 nM) but a neglible affinity for ₂-adrenoceptors in the same tissue. Only clozapine showed a weak affinity for the latter receptor type.Fluperlapine was as effective as imipramine in antagonizing tetrabenazine-induced ptosis in the rat, the antiptotic effect remaining constant after up to ten daily drug administrations. Still, imipramine was stronger than fluperlapine as an inhibitor of the accumulation of [³H]-noradrenaline ([³H]-NA) in rat cerebral cortex slices. Fluperlapine's effects on the spontaneous and the electrically-induced release of [³H]-NA from rat cerebral cortex slices, with and without protriptyline, showed it to be an inhibitor of the reuptake of NA.The results indicate that the pharmacological profile of fluperlapine is similar to that of clozapine, with additional antidepressant properties.