A. Groppetti

Significance of dopamine metabolites in the evaluation of drugs acting on dopaminergic neurones

The effect of various drugs was studied on 3-methoxytyramine (3-MT) concentrations in rat striatum. The drugs were chosen for their ability to interfere with the dopaminergic system at different levels. Dopamine (DA) acidic metabolites, i.e. homovanillic acid (HVA) and 3,4-dihydroxyphenylacetic acid (DOPAC), were also measured. Changes of 3-MT, unlike those of DOPAC and HVA, seem to reflect the functional activity of dopaminergic neurons. In fact drugs believed to increase or decrease DA content in the synaptic cleft produce predictable changes of striatal 3-MT. Thus cocaine, nomifensine and d-amphetamine increase 3-MT concentrations while gamma-butyrolactone, alpha-methyltyrosine and apomorphine decrease it.

Role of dopamine in manganese neurotoxicity

Manganese chloride increased cell mortality when added to human fibroblast cultures. The toxicity of the metal was greatly enhanced by dopamine; this effect was antagonized by the presence in the culture medium of catalase and superoxide dismutase enzymes. Manganese chloride also caused a marked decrease of striatal dopamine concentrations when infused into rat substantia nigra. Manganese neurotoxicity was lowered by pretreating the animals with drugs that reduced striatal dopamine turnover rate. Administration of an antioxidant, such as vitamin E, also partially prevented striatal dopamine decline induced by intranigral manganese infusion. Therefore, the decreased availability or autoxidation of dopamine attenuated manganese neurotoxicity. These findings are in agreement with previous observations suggesting that manganese increases toxic products originating from dopamine catabolism.

Changes in specific activity of dopamine metabolites as evidence of a multiple compartmentation of dopamine in striatal neurons.

The functional status of dopaminergic nerve terminals has been studied with a method that allows the simultaneous determination of the specific activities of dopamine (DM), tyrosine (Tyr), 3‐methoxytyramine (3‐MT) and 3,4‐dihydroxyphenylacetic acid (DOPAC), after administration of [3H]tyrosine ([3H]Tyr).

Reduction of food intake by apomorphine: A pimozide-sensitive effect

brain 5-HT and 5-HIAA and preventing 5-HT accumulation after pargyline. Thus D-PCPA, like its L-isomer, inhibits 5-HT synthesis. As previously shown for the racemic compound (Koe & Weissman, 1966), the depletion of brain 5-HT and 5-HIAA induced by Dor L-PCPA was maximal after a long latency and persisted for several days. These results suggest that the inhibition of 5-HT synthesis by either isomer is an irreversible process and depends on the slow formation of an active metabolite. Since D-amino acids are not incorporated into proteins (Berg, 1959), the incorporation of D-PCPA into tryptophan hydroxylase is unlikely. A conversion of D-PCPA in vivo to L-PCPA is theoretically possible via its deamination by D-amino acid oxidase (Blaschko & Stiven, 1950) to p-chlorophenylpyruvic acid followed by transamination of the latter to L-PCPA (Spencer & Brock, 1962). However, this is difficult to reconcile with the fact D-PCPA decreases the level of 5-HT and 5-HIAA at the same rate, to the same extent, and for the same time as the L-isomer. These considerations lead to the conclusion that stereoisomerism is not essential for PCPA-induced inhibition of 5-HT synthesis.

Effect of aspirin on serotonin and met-enkephalin in brain: correlation with the antinociceptive activity of the drug

Intravenous administration of acetyl salicylate of lysine, a soluble salt of aspirin, reduced in rats the firing discharge of thalamic neurones, evoked by noxious stimuli. Concomitantly, concentrations of 5-hydroxyindole acetic acid increased, while those of met-enkephalin-like immuno-reactive derivatives were decreased in several areas of the brain. Similar electrophysiological and biochemical responses were obtained by administering tryptophan or 5-hydroxytryptophan plus carbidopa. The effect of aspirin on the evoked firing of the thalamic neurones was counteracted by pretreating the animals with metergoline. On the other hand, naloxone did not antagonize the inhibitory effect of aspirin and 5-hydroxytryptophan on pain-induced neuronal excitation. These data indicate that a serotonin-, but not a naloxone-sensitive opiate mechanism, may be relevant for aspirin-mediated antinociception.

Pertussis toxin inhibits morphine analgesia and prevents opiate dependence

Six days after intracerebroventricular pretreatment of rats with pertussis toxin (PTX 0.5 microgram/rat) there was a marked decrease in the antinociceptive effect of morphine, regardless of the route of opioid administration (into the periaqueductal gray matter, intrathecally or intraperitoneally) or the analgesic test used (tail flick and jaw opening reflex). PTX pretreatment also partially attenuated the naloxone-precipitated withdrawal syndrome in morphine-dependent rats, significantly reducing teeth chattering, rearing and grooming. These in vivo findings indicate that G-protein-dependent mechanisms are involved in morphine analgesia and dependence. The biochemical mechanism could be related to ADP ribosylation of Gi coupled to the adenylate cyclase system, but an interaction of PTX with other G-proteins linked to different second messengers or directly to ionic channels cannot be excluded.

A mass fragmentographic assay of 3-methoxytyramine in rat brain.

Abstract— A mass fragmentographic method for the measurement of 3‐methoxytyramine, a metabolite of dopamine formed after its neuronal release, is described. This method allows the assay of this dopamine metabolite in the picogram range, thus utilizing only few milligrams of brain areas containing dopaminergic neurons. Moreover, the method is simple and possesses the high specificity intrinsic to mass fragmentography.

Manganese neurotoxicity: Effects ofl-DOPA and pargyline treatments

Single, monolateral injection into rat substantia nigra of manganese chloride produced within two weeks from its administration a loss of dopamine in the striatum ipsilateral to the injected side. The effect was dose-dependent and was not extended to serotoninergic terminals present in this brain area, whose content in serotonin and 5-hydroxyindoleacetic acid was not affected. When L-DOPA + carbidopa or pargyline were given to these animals the decrease of striatal dopamine was more marked. Moreover, rats treated two weeks before with a dose of manganese chloride that produced a 70-80% drop in striatal dopamine concentrations, rotated ipsilaterally to the dopamine-depleted striatum when injected with apomorphine, suggesting that in these animals the stimulatory effects of apomorphine were more relevant in striatum where presynaptic dopaminergic neurons were not affected by manganese chloride. These data indicate that the alterations of dopaminergic postsynaptic receptors may be different in parkinsonian and in manganese-intoxicated patients and that current therapy used for Parkinsons disease could be a hazard in treating manganese poisoning.

Nefopam inhibits calcium influx, cGMP formation, and NMDA receptor-dependent neurotoxicity following activation of voltage sensitive calcium channels

Summary.Nefopam hydrochloride is a potent non sedative benzoxazocine analgesic that possesses a profile distinct from that of anti-inflammatory drugs. Previous evidence suggested a central action of nefopam but the detailed mechanism remains unclear. We have investigated the actions of nefopam on voltage sensitive calcium channels and calcium-mediated pathways. We found that nefopam prevented N-methyl-D-aspartate (NMDA)-mediated excitotoxicity following stimulation of L-type voltage sensitive calcium channels by the specific agonist BayK8644. Nefopam protection was concentration-dependent. 47 μM nefopam provided 50% protection while full neuroprotection was achieved at 100 μM nefopam. Neuroprotection was associated with a 73% reduction in the BayK8644-induced increase in intracellular calcium concentration. Nefopam also inhibited intracellular cGMP formation following BayK8644 in a concentration-dependent manner, 100 μM nefopam providing full inhibition of cGMP synthesis and 58 μM allowing 50% cGMP formation. Nefopam reduced NMDA receptor-mediated cGMP formation resulting from the release of glutamate following activation of channels by BayK8644. Finally, we also showed that nefopam effectively reduced cGMP formation following stimulation of cultures with domoic acid, while not providing neuroprotection against domoic acid. Thus, the novel action of nefopam we report here may be important both for its central analgesic effects and for its potential therapeutic use in neurological and neuropsychiatric disorders involving an excessive glutamate release.

Interactions between serotonergic and enkephalinergic neurons in rat striatum and hypothalamus

We report evidence for an interaction between serotonergic and enkephalinergic neurons in rat striatum and hypothalamus. The administration of drugs such as p-chlorophenylalanine (PCPA), 5,6-dihydroxytryptamine (5,6-DHT) and fenfluramine that lower the striatal and hypothalamic serotonin (5-HT) content caused an increase in Met-enkephalin-like immunoreactive material (ME-IR) in these brain areas. Moreover, chronic treatment with PCPA induced an increase in the number of striatal [3H][D]Ala2,Met5]enkephalinamide binding sites. These observations suggest that serotonergic neurons modulate the functional activity of enkephalinergic neurons in the rat striatum and hypothalamus. The significance of this interaction is discussed.