Santiago Ballaz

MDMA ('Ecstasy') enhances 5-HT1A receptor density and 8-OH-DPAT-induced hypothermia: blockade by drugs preventing 5-hydroxytryptamine depletion.

One week after a single administration of 3,4-methylenedioxymethamphetamine (MDMA HCI, 30 mg/kg i.p.), 5-HT1A receptor density was significantly increased by approximately 25-30% in the frontal cortex and hypothalamus of rats. The increased density correlated with the potentiation of the hypothermic response to the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT, 1 mg/kg s.c.). Hypothalamic 5-HT7 receptors, which also bind 8-OH-DPAT, were not changed, however, by MDMA. Fluoxetine (5 mg/kg s.c.), ketanserin (5 mg/kg s.c.) or haloperidol (2 mg/kg i.p.), given 15 min prior to MDMA, prevented the depletion of 5-hydroxytryptamine (5-HT) induced by MDMA and also blocked the effects of this neurotoxin on 5-HT1A receptor density and on 8-OH-DPAT-induced hypothermia. The protection afforded by drugs against 5-HT loss did not correlate, however, with the antagonism of the acute hyperthermic effect of MDMA. The present results indicate that drugs able to prevent or to attenuate MDMA-induced 5-HT loss also prevent the changes in 5-HT1A receptor density as well as the enhanced hypothermic response to the 5-HT1A receptor agonist 8-OH-DPAT in MDMA-treated rats.

Facilitation by 8-OH-DPAT of passive avoidance performance in rats after inactivation of 5-HT1A receptors

Pretraining administration of 8‐hydroxy‐2‐di‐n‐propylamino‐tetralin (8‐OH‐DPAT 0.1 mg kg−1), a 5‐HT1A receptor agonist, or buspirone (1 mg kg−1), a 5‐HT1A receptor partial agonist, markedly impaired passive avoidance retention in rats 24 h later. The effect of 8‐OH‐DPAT was prevented by the 5‐HT1A receptor antagonists, NAN‐190 and WAY‐100635, at doses without any intrinsic effect. N‐ethoxycarbonyl‐2‐ethoxy‐1,2‐dihydroquinoline (EEDQ 10 mg kg−1), an alkylating agent that inactivates different G‐protein coupled receptors, impaired retention performance when given 48 h pretraining. The disruptive effect of EEDQ was reversed by 8‐OH‐DPAT or buspirone, given 30 min before training. Non‐specific actions did not account for 8‐OH‐DPAT‐induced reversal of the EEDQ effect since no significant difference in locomotor activity or in pain threshold was found between rats receiving EEDQ or EEDQ+8‐OH‐DPAT. When NAN‐190 (1 mg kg−1) or WAY‐100635 (0.5 mg kg−1) were given before 8‐OH‐DPAT to EEDQ‐pretreated animals, the reversal by 8‐OH‐DPAT of EEDQ‐induced retention impairment was still more pronounced. However, no EEDQ reversal by 8‐OH‐DPAT was found when 5‐HT1A receptors were protected by WAY‐100635 (10 mg kg−1) 30 min before EEDQ. In the hippocampus of EEDQ‐treated rats, 5‐HT7 receptors were less inactivated than 5‐HT1A receptors and significant increases were found in 5‐HT1A but not in 5‐HT7 receptor mRNA levels. Ritanserin and methiothepin (10 mg kg−1 each), antagonists with higher affinity at 5‐HT7 than at 5‐HT1A receptors, prevented the retention impairment induced by EEDQ but did not significantly protect against 5‐HT7 receptor inactivation. The results indicate that the facilitatory effect of 8‐OH‐DPAT is not mediated through 5‐HT1A receptors and suggest that other 8‐OH‐DPAT‐sensitive receptors could be involved in the dual effect of 8‐OH‐DPAT on passive avoidance performance in rats.

Pharmacological evaluation of IQM-95,333, a highly selective CCKA receptor antagonist with anxiolytic-like activity in animal models

The pyridopyrimidine derivative IQM‐95,333 ((4aS,5R)‐2‐benzyl‐5‐[Nα‐tert‐butoxicarbonyl)L‐tryptophyl]amino‐1,3dioxoperhydropyrido[1,2‐c]pyrimidine), a new non‐peptide antagonist of cholecystokinin type A (CCKA) receptors, has been evaluated in vitro and in vivo in comparison with typical CCKA and CCKB receptor antagonists, such as devazepide, lorglumide, L‐365,260 and PD‐135,158. IQM‐95,333 displaced [3H]‐CCK‐8S binding to CCKA receptors from rat pancreas with a high potency in the nanomolar range. Conversely, the affinity of this new compound at brain CCKB receptors was negligible (IC50>10 μM). IQM‐95,333 was a more selective CCKA receptor ligand than devazepide and other CCKA receptor antagonists. Like devazepide, IQM‐95,333 was a more potent antagonist of CCK‐8S‐ than of CCK‐4‐induced contraction of the longitudinal muscle from guinea‐pig ileum, suggesting selective antagonism at CCKA receptors. IQM‐95,333 and devazepide were also potent inhibitors of CCK‐8S‐stimulated amylase release from isolated pancreatic acini, a CCKA receptor‐mediated effect. The drug concentrations required (IC50s around 20 nM) were higher than in binding studies to pancreas homogenates. Low doses (50–100 μg kg−1, i.p.) of IQM‐95,333 and devazepide, without any intrinsic effect on food intake or locomotion, blocked the hypophagia and the hypolocomotion induced by systemic administration of CCK‐8S, two effects associated with stimulation of peripheral CCKA receptors. IQM‐95,333 showed an anxiolytic‐like profile in the light/dark exploration test in mice over a wide dose range (10–5,000 μg kg−1). Typical CCKA and CCKB antagonists, devazepide and L‐365,260 respectively, were only effective within a more limited dose range. In a classical conflict paradigm for the study of anxiolytic drugs, the punished‐drinking test, IQM‐95,333, devazepide and L‐365,260 were effective within a narrow dose range. The dose‐response curve for the three drugs was biphasic, suggesting that other mechanisms are operative at higher doses. In conclusion, IQM‐95,333 is a potent and selective CCKA receptor antagonist both in vitro and in vivo with an anxiolytic‐like activity in two different animal models, which can only be attributed to blockade of this CCK receptor subtype.

Ascorbate prevents cell death from prolonged exposure to glutamate in an in vitro model of human dopaminergic neurons.

Ascorbate (vitamin C) is a nonenzymatic antioxidant highly concentrated in the brain. In addition to mediating redox balance, ascorbate is linked to glutamate neurotransmission in the striatum, where it renders neuroprotection against excessive glutamate stimulation. Oxidative stress and glutamatergic overactivity are key biochemical features accompanying the loss of dopaminergic neurons in the substantia nigra that characterizes Parkinsons disease (PD). At present, it is not clear whether antiglutamate agents and ascorbate might be neuroprotective agents for PD. Thus, we tested whether ascorbate can prevent cell death from prolonged exposure to glutamate using dopaminergic neurons of human origin. To this purpose, dopamine‐like neurons were obtained by differentiation of SH‐SY5Y cells and then cultured for 4 days without antioxidant (antiaging) protection to evaluate glutamate toxicity and ascorbate protection as a model system of potential factors contributing to dopaminergic neuron death in PD. Glutamate dose dependently induced toxicity in dopaminergic cells largely by the stimulation of AMPA and metabotropic receptors and to a lesser extent by N‐methyl‐D‐aspartate and kainate receptors. At relatively physiological levels of extracellular concentration, ascorbate protected cells against glutamate excitotoxicity. This neuroprotection apparently relies on the inhibition of oxidative stress, because ascorbate prevented the pro‐oxidant action of the scavenging molecule quercetin, which occurred over the course of prolonged exposure, as is also seen with glutamate. Our findings show the relevance of ascorbate as a neuroprotective agent and emphasize an often underappreciated role of oxidative stress in glutamate excitotoxicity. Occurrence of a glutamate–ascorbate link in dopaminergic neurons may explain previous contradictions regarding their putative role in PD.

Striatal interaction among dopamine, glutamate and ascorbate

Despite evidence suggesting the interaction among glutamate (GLU), dopamine (DA) and ascorbic acid (AA) in the striatum, their actions are often studied separately. Microdialysis was used here to quantify the extracellular interaction among GLU-DA-AA in the striatum of rats, an interaction which was compared with those studied in the substantia nigra (SN). Perfusion of GLU by reverse microdialysis increased DA and decreased 3,4-dihydroxyphenylacetic acid (DOPAC) in the extracellular medium of the striatum, but increased both DA and DOPAC in the SN. The increase of extracellular DA-concentration induced by the local DA-perfusion decreased the extracellular level of GLU and glutamine, an effect that, as suggested by the GLU and glutamine increase observed after the haloperidol administration, probably involves the D2 dopamine receptor. Local administration of AA increased the extracellular DA, decreased DOPAC and had no effect on GLU and glutamine. Present data suggest that, in the striatum, GLU-release inhibits DA-uptake, DA-release inhibits GLU-release, and AA-release prevents DA-oxidation increasing its extracellular diffusion. These effects were different in the SN where GLU probably promoted the DA-release instead of inhibiting the DA-uptake as presumably occurred in the striatum. Present data denote a marked GLU-DA-AA interaction in the striatum, which might be relevant for the pharmacological control of basal ganglia disorders.

Pseudopeptide CCK-4 analogues incorporating the Ψ[CH(CN)NH] peptide bond surrogate

Abstract The synthesis, binding to CCK receptors, and in vitro functional activity of pseudopeptide CCK-4 analogues incorporating the ( R ) or ( S ) Ψ[CH(CN)NH] peptide bond surrogate at the NIe 31 -Asp 32 or or Trp 30 -NIe 13 bonds are described. Z-TrpΨ[( S )CH(CN)NH]NIe-Asp-Phe-NH 2 retained the high CCK-B receptor binding affinity of Boc-[NIe 31 ]-CCK-4, and was a potent and selective CCK-B antagonist in the isolated guinea pig ileum.

CCK-4 restricted analogues containing a 3-oxoindolizidine skeleton

Abstract A series of CCK-4 restricted analogues, incorporating a 3-oxoindolizidine bicyclic lactam as conformational constraint, is described. The 8-(Boc-tryptophyl)amino-2-benzyl derivatives behave as weak CCK-A or CCK-B receptor antagonists. The main factors for selectivity are the configuration at C-2 position of the 3-oxoindolizidine ring and at the Trp residue.