H. H. Keller

Immunoregulation mediated by the sympathetic nervous system.

Abstract A postulated immunoregulatory role for the autonomous nervous system was explored utilizing several in vivo and in vitro approaches. Local surgical denervation of the spleen in rats and general chemical sympathectomy by 6-hydroxydopamine combined with adrenalectomy yielded a similar removal of restraint expressed as enhancement in the number of PFC in response to immunization. Noradrenaline and the synthetic α-agonist clonidine which are, respectively, natural and artificial effector molecules of the sympathetic nervous system each strongly suppressed the in vitro induced immune response of murine spleen cells to SRBC. Further, radiometric-enzymatic assay of noradrenaline in the splenic pulp revealed a decrease in the content of this neurotransmitter just preceding the exponential phase of the immune response to SRBC (Days 3 and 4) in this site. Taken together, these findings point to a dynamic immunoregulatory relationship between the immune and sympathetic nervous system.

Benzodiazepine antagonist Ro 15-1788: binding characteristics and interaction with drug-induced changes in dopamine turnover and cerebellar cGMP levels.

Abstract: The recently discovered benzodiazepine antagonist Ro 15‐1788 was characterized in binding studies, and its potency and selectivity were determined in vivo by interaction with drug‐induced changes in dopamine turnover and cerebellar cGMP level. Ro 15‐1788 reduced [3H]flunitrazepam binding in the brain in vivo with a potency similar to that of diazepam and effectively inhibited [3H]diazepam binding in vitro (IC50= 2.3 ± 0.6 nmol/liter). [3H]Ro 15‐1788 bound to tissue fractions of rat cerebral cortex with an apparent dissociation constant (KD) of 1.0 ± 0.1 nmol/liter. The in vitro potency of various benzodiazepines in displacing [3H]Ro 15‐1788 from its binding site was of the same rank order as found previously in [3H]diazepam binding. Autoradiograms of [3H]Ro 15‐1788 binding in sections of rat cerebellum showed the same distribution of radioactivity as with [3H]flunitrazepam. The attenuating effect of diazepam on the chlorpromazine‐ or stress‐induced elevation of homovanillic acid in rat brain was antagonized by Ro 15‐1788. Among a series of compounds which either decreased or increased the rat cerebellar cGMP level, only the effect of benzodiazepine receptor ligands (diazepam, zopiclone, CL 218 872) was antagonized by Ro 15‐1788. Thus, Ro 15‐1788 is a selective benzodiazepine antagonist acting at the level of the benzodiazepine receptor in the central nervous system. Peripheral benzodiazepine binding sites in kidney and schistosomes were not affected by Ro 15‐1788.

Increase of 3-methoxy-4-hydroxyphenylethylene glycol in rat brain by neuroleptic drugs

Abstract The potency of various neuroleptic drugs in increasing the content of endogenous 3-methoxy-4-hydroxyphenyl-ethylene glycol (MOPEG) in rat brain decreased in the order methiothepin, haloperidol, clozapine, thioridazine, chlorpromazine, pimozide. The neuroleptics, except pimozide and chlorpromazine, also caused a slight to moderate diminution of the endogenous cerebral noradrenaline (NA). Based on these and earlier findings it is concluded that (a) changes in brain NA turnover induced by neuroleptics can be estimated, in a relatively simple way, by measuring cerebral MOPEG; (b) these drugs markedly differ in their ability to activate noradrenergic neurons; and (c) the activation of noradrenergic neurons by neuroleptics does not seem to parallel that of dopaminergic neurons.

Effects of clozapine on cerebral catecholaminergic neurone systems.

1 . Clozapine, a dibenzodiazepine derivative claimed to possess antipsychotic properties in man without producing extrapyramidal disorders, greatly increased the turnover of cerebral dopamine in the rat. 2 . The drug itself was virtually devoid of cataleptigenic activity in rats; however, it antagonized prochlorperazine‐induced catalepsy. 3 . It is proposed that clozapine causes a blockade of striatal dopamine receptors which is of the surmountable type in contrast to that produced by cataleptigenic neuroleptics. In addition, clozapine may also increase the turnover of cerebral noradrenaline.

From moclobemide to Ro 19-6327 and Ro 41-1049: the development of a new class of reversible, selective MAO-A and MAO-B inhibitors

This study describes the serendipitous discovery of moclobemide, a short-acting MAO-A inhibitor which is in an advanced stage of clinical development as an antidepressant. The short duration of action of this MAO inhibitor containing a morpholine ring moiety is due to the complete reversibility (probably by metabolism of the inhibitory molecular species) of MAO-A inhibition. Since moclobemide is much more effective in vivo than expected from its in vitro activity, investigations to identify a possible metabolite(s) more active as MAO-A inhibitor than the parent compound were carried out. The study of the MAO inhibitory characteristics of several known and putative moclobemide metabolites did not allow the identification of a potent MAO-A inhibitor but led to the discovery of Ro 16-6491, a potent MAO-B inhibitor of novel chemical structure. Systematic chemical modification of the aromatic ring system of Ro 16-6491 finally provided Ro 19-6327 and Ro 41-1049 which are highly selective and reversible inhibitors of MAO-B and MAO-A, respectively. Tritiated derivatives of Ro 19-6327 and Ro 41-1049 were used in binding studies to elucidate their mechanisms of action and to study their cellular distribution by quantitative enzyme radioautography.

The pharmacology of Parkinson's disease: Basic aspects and recent advances

Basic aspects and recent advances in the understanding of the pharmacological mechanism of action of the clinically most used antiparkinson drugs are reviewed. Recent human and animal biochemical investigations clearly confirm and extend previous findings indicating that benserazide is much more potent than carbidopa as peripheral decarboxylase inhibitor. L-DOPA in combination with benserazide or carbidopa constitutes the best available therapy for Parkinsons disease (PD). To reduce peaks and rapid fluctuations of L-DOPA plasma levels (possibly responsible for peak-dose dyskinesias and end-of-dose deterioration) a slow-release formulation of L-DOPA in combination with benserazide or with benserazide plus catechol-O-methyltransferase inhibitors should be developed. In parkinsonian patients under long-term L-DOPA therapy monoamine oxidase inhibitors type B (MAO-B) e.g. (−)-deprenyl and firect dopamine receptor agonists (bromocriptine, lisuride, pergolide etc.), due to their L-DOPA-sparing effects, alleviate in some cases L-DOPA-induced side-effects e.g. dyskinesias and on-off phenomena. However, since (−)-deprenylm, due to its metabolism to (−)methamphetamine and (−)amphetamine, seem to have indirect sympathomimetic activity, new selective MAO-B inhibitors devoid of indirect sympathomimetic effects should be tested clinically to assess the functional role of pure MAO-B inhibition in the therapy of PD. The auxiliary therapy with direct dopmaine receptor agonists of the D-2 subtype represents another valid approach which should be further investigated in order to find novel dopamine agonists, less expensive than bromocriptine and strictly selective for D-2 receptor sites.

Baclofen and γ-hydroxybutyrate: similar effects on cerebral dopamine neurones

Abstract Baclofen (20 mg/kg) caused an increase in the content of homovanillic acid (HVA) and dopamine (DA) in rat brain 2–3 h after drug injection without appreciable changes in the level of other monoamines and their main metabolites. Six and eight hours after baclofen, the content of HVA but not that of DA was reduced. Moreover, baclofen initially (20 min after injection) reduced, but later (105 min post drug) enhanced the accumulation of HVA induced by probenecid. The shortlasting (20 min) initial reduction of HVA elevation in probenecid-pretreated animals as well as the longlasting (6–8 h) decrease of HVA levels in rats injected with baclofen alone are interpreted to be due to a decreased release and metabolism of DA, probably as a consequence of the blockade of impulse flow in mesolimbic and nigro-striatal DA neurones. The increase in HVA and DA seen during the first few hours is thought to result from enhanced DA synthesis similar to that known for γ-hydroxybutyrate (GHB). This initial rise in HVA due to synthesis stimulation probably masked a reduction of HVA to be expected immediately after baclofen injection. The similarity between baclofen and GHB is stressed by the finding that baclofen counteracted the increase of HVA occuring after chlorpromazine and D-amphetamine but not that induced by the benzoquinolizine derivative, Ro 4-1284.

Short-acting novel MAO inhibitors: in vitro evidence for the reversibility of MAO inhibition by moclobemide and Ro 16-6491

SummaryThe inhibition of monoamine oxidase (MAO) in rat liver and brain by the short-acting MAO-A inhibitors moclobemide (Ro 11-1163 = p-chloro-N-[2-morpholinoethyl]benzamide) and brofaremine and by the short-acting MAO-13 inhibitors Ro 16-6491 (N-[2-aminoethyl]-p-chlorobenzamide) and almoxatone, administered p. o. at roughly equieffective doses 2 h before decapitation, was investigated for its reversibility under various in vitro conditions. MAO A activity in liver homogenates, inhibited by moclobemide (300 μmol/kg) to approx. 15% of control, time dependently recovered during 0.5 to 2 h of incubation at 37°C, irrespective of whether the homogenates were prepared and incubated in distilled water or Krebs-Ringer buffer (KRB). Dialysis of such homogenates for 4 h in distilled water at 37°C (but not at 13°C) led to a complete return of the MAO activity. In liver homogenates from rats pretreated with brofaremine (30 μmol/kg), dialysis for 4 h at 37°C against distilled water caused only little recovery of the MAO activity. Likewise, MAO-B inhibited by Ro 16–6491 (30 μmol/kg) to approx. 4% of control returned to almost control activity after 4 h of dialysis at 37°C, while inhibition induced by almoxatone (30 μmol/kg) was little or not reversed at all. In brain homogenates prepared in, and dialysed against, distilled water or KRB at 37°C (but not at 13°C), MAO-A inhibited by moclobemide (100–300 μmol/kg) to approx. 15% of control, partially (KRB) or almost completely (dist. water) returned to control activity after 4 h of dialysis. From rats pretreated with Ro 16–6491 (30 μmol/kg), MAO-B in brain homogenates prepared in KRB was reduced to 12% of control and returned to control value upon dialysis for 4 h in KRB at 37°C; in homogenates prepared in H2O, MAO-B was reduced to only 60% of control and completely recovered by dialysis against dest. water even at 13°C. In all of these conditions, recovery of the enzyme activity was small after brofaremine and almoxatone. Analogous results were obtained with brain slices (0.2 × 0.2 × 1.5 mm) in KRB at 37°C, whereby time dependent recovery of MAO activity during incubation was achieved, and superfusion was somewhat more effective than incubation in restoring enzyme activity. In the experiments with incubated or superfused brain slices, inhibition of MAO-A and -B by the irreversible inhibitors clorgyline and selegiline (l-deprenyl), resp., could not be reversed at all. Tyramine (0.3 mmol/l) clearly enhanced the recovery of MAO-A in KRB-prepared liver homogenates and brain slices of moclobemide-pretreated rats but not in brain slices of brofaremine- and clorgyline-pretreated rats. Thus, the reversibility of MAO inhibition in vitro could be convincingly demonstrated for moclobemide and Ro 16–6491 but not for the other novel, short-acting MAO inhibitors studied.

Spontaneous and drug-induced changes of cerebral dopamine turnover during postnatal development of rats

Abstract In the whole brain, the striatum and the brain without the striata of rats, the dopamine (DA) and the noradrenaline (NA) levels progressively increased from day 1 to day 60 after birth. Furthermore, in whole brain a continuous rise of the 3,4-dihydroxyphenylacetic acid (DOPAC) content was observed during the postnatal period. The homovanillic acid (HVA) concentration showed peak values at 12–18 days; thereafter it declined again until adult levels were reached at 30–60 days. The striatal HVA levels at 60 days were higher than those at day 4, whereas in the brain without striata the concentration of the acid was lower at day 60 than at day 4. Probenecid caused the same HVA increment in the brain at all ages. The haloperidol-induced increase of cerebral HVA was smaller in the early (1–12 days) than in the later period (18–60 days) of postnatal development. Apomorphine was progressively more effective in reducing the HVA levels from day 1 to day 60. It is concluded that the turnover of striatal DA is relatively high during the first days of the postnatal period and then progressively decreases until the adult rate is reached at about 60 days. This change may be connected with the maturation of the DA stores, the DA receptors and the feedback mechanism regulating DA synthesis. The decrease of the HVA levels in the extrastriatal brain parts between days 12 and 60 is possibly due to maturation of the noradrenergic system.

Effect of neuroleptics on endogenous norepinephrine in rat brain.

Abstract The neuroleptic drugs clozapine, thioridazine and methiothepin, but not chlorpromazine, markedly reduced the norepinephrine (NE) levels in the brain of rats kept normothermic. Clonidine prevented the NE lowering effect of clozapine. In animals with sectioned spinal cords, clozapine decreased the NE levels only in the segment cranial to the lesion whereas in normal animals the drug induced a similar NE decrease in the cranial and caudal part of the spinal cord. After pretreatment with the dopamine-β-hydroxylase inhibitor FLA 63 in a dose which did not markedly diminish the cerebral NE, chlorpromazine caused a small but significant decrease of this amine. It is concluded that, due to blockade of NE receptors and a subsequent feed-back activation of NE neurones, clozapine, thioridazine and methiothepin increase the release of NE to such a degree that the amine loss cannot be fully compensated by synthesis possibly as a consequence of an insufficient rate of β-hydroxylation of dopamine. In contrast, in the case of chlorpromazine which seems to be a relatively weak NE-receptor blocking agent, this compensation is still possible.