Neurologia Sperimentale

From using in vitro intracellular recordings from mesencephalic neurons and monoamine-depleted rats, we report that the functions of levodopa in the brain are greatly enhanced and prolonged by high doses of the monoamine oxidase (MAO) inhibitor deprenyl. Dopaminergic neurons were hyperpolarized and inhibited by levodopa application. These effects of levodopa were largely potentiated by pretreatment with nonselective doses of deprenyl. Furthermore, when locomotor aaivity induced by levodopa was examined on a rodent model of Parkinson's disease, pretreatment of the animals with nonselective doses of deprenyl caused an enhancement of the antiparkinsonian action of levodopa. The great increase in levodopa responses by deprenyl suggests a likely therapeutic use of this dopamine precursor with a higher dosage of the MA0 inhibitor, to reduce effectively the daily levodopa requirements in Parkinson's disease patients. Mercuri NB, Scarponi M, Federici M, Bonci A, Siniscalchi A, Bernardi G. Modification of levodopa responses by deprenyl (Selegiline): an electrophysiological and behavioral study in the rat relevant to Parkinson's disease.

The dopaminergic system constitutes the principal target of many psychostimulants, antidepressant, and antiparkinsonian drugs. The effects caused by these compounds are partly associated with an increased dopamine (DA) levels within the terminal areas of DA neurons and in the ventral midbrain. Therefore, several substances of abuse, antidepressants, and endogenous compounds (levodopa and trace amines [TAs]) regulate the activity of DA cells by activating D2 autoreceptors located on the terminals, soma, and dendrites. Considering our past and recent experimental studies on this issue, here we will briefly reexamine the mechanisms of action of several psychoactive drugs on DA neurons. In particular, we propose three different modalities by which the mesencephalic DA neurons can be regulated by drugs: amphetamine/TAs-like, cocaine-like, and levodopa-like. We, therefore, discuss the potential therapeutic and addictive properties of the psychoactive substances.

Background: Methylphenidate has been suggested to exert its therapeutic effect mainly by blocking the dopamine transporter. In spite of the importance of this interaction, no detailed information is available yet on its actions on single dopaminergic neurons. Methods: We examined the effects of methylphenidate on dopaminergic neurons using electrophysiological recordings from rat midbrain slices. Results: Methylphenidate inhibited spontaneous firing and caused a membrane hyperpolarization in current clamp or an outward current in voltage clamp. These effects were antagonized by the D 2 receptor antagonist sulpiride. An acute dopamine-depleting treatment of the slices with the dopa-decarboxylase inhibitor carbidopa significantly reduced the effects of methylphenidate. This drug potentiated, in a concentration-dependent manner, cellular responses to exogenous dopamine application. Conclusions: Our electrophysiological data are consistent with the hypothesis that methylphenidate inhibits dopamine transporter and suggest that the depression of firing is mediated by the release of newly synthesized dopamine which accumulates extracellularly due to inhibition of its reuptake.

Nutrition is the result of a complex interaction among environmental, homeostatic and rewardrelated processes. Accumulating evidence supports key roles for the dopaminergic neurons of the ventral midbrain in regulating feeding behaviour. For this reason, in the present study, we have investigated the electrophysiological effects of two centrally acting anorexic agents, fenfluramine and sibutramine, on these cells. Experimental approach: Rat midbrain slices were used to make intracellular recordings from dopaminergic neurons of the substantia nigra and the ventral tegmental area. Gamma-aminobutyric acid (GABA)-mediated synaptic transmission was assessed from the inhibitory postsynaptic potentials (IPSPs) mediated by GABAA and GABAB receptors. Key results: Fenfluramine and sibutramine reduced, concentration-dependently, the GABAB IPSPs, without affecting the GABAA-mediated potentials. This effect is presynaptic, as postsynaptic membrane responses induced by application of a GABAB receptor agonist, baclofen, were not affected by the two drugs. Furthermore, the selective 5-hydroxytriptamine 1B (5-HT1B) receptor antagonist, SB216641, blocked the reduction of GABAB IPSPs caused by fenfluramine and sibutramine, indicating that the receptor mediating this effect is 5-HT1B. Conclusions and implications: Two anorexic agents, fenfluramine and sibutramine, induced the activation of 5-HT1B receptors located on presynaptic GABAergic terminals, thus reducing the release of GABA. This action can alter the strength of synaptic afferents that modify the activity of dopaminergic neurons, inducing neuronal excitation. Our results reveal an additional mechanism of action for fenfluramine and sibutramine that might contribute to reducing food intake, by influencing the pleasurable and motor aspects of feeding behaviour.

Background and purpose: Although trace amines (TAs) are historically considered 'false neurotransmitters' on the basis of their ability to induce catecholamine release, there is evidence that they directly affect neuronal activity via TA receptors, ligand-gated receptor channels and/or s receptors. Here, we have investigated the effects of two TAs, tyramine (TYR) and b-phenylethylamine (b-PEA), on electrophysiological responses of substantia nigra pars compacta (SNpc) dopaminergic cells to the D2 receptor agonist, quinpirole. Experimental approach: Electrophysiological recordings of D2 receptor-activated G-protein-gated inward rectifier K + channel (GIRK) currents were performed on dopaminergic cells from midbrain slices of mice and on Xenopus oocytes expressing D2 receptors and GIRK channels. Key results: TYR and b-PEA reversibly reduced D2 receptor-activated GIRK currents in a concentration-dependent manner on SNpc neurones. The inhibitory effect of TAs was still present in transgenic mice with genetically deleted TA1 receptors and they could not be reproduced by the selective TA1 agonist, o-phenyl-3-iodotyramine (O-PIT). Pretreatment with antagonists of s1 and s2 receptors did not block TA-induced effects. In GTPgS-loaded neurones, the irreversibly-activated GIRK-current was still reversibly reduced by b-PEA. Moreover, b-PEA did not affect basal or dopamine-evoked GIRK-currents in Xenopus oocytes.

In order to study whether the membrane hyperpolarization and firing inhibition caused by dopamine and levodopa on rat midbrain dopamine cells are affected by the inhibition of brain catechol-O-methyl-transferase (COMT), intracellular electrophysiological recordings were made from these neurons maintained in vitro. Here we report that a treatment of the cerebral tissue with tolcapone, a central and peripheral inhibitor of COMT, does not change the membrane responses of midbrain dopamine neurons to dopamine and levodopa. The lack of modification of the dopaminergic effects by tolcapone suggests that the pharmacological inhibition of intracerebral COMT does not have detectable action on dopamine neurotransmission. Therefore, the therapeutic action of tolcapone in Parkinson's disease, might be dependent on the reduction of COMT activity in the extracerebral tissue.

A multidisciplinary approach was followed to investigate whether the opioid-like peptide nociceptin/orphanin FQ (N/OFQ) regulates the nigrostriatal dopaminergic pathway and motor behavior. Nigrostriatal dopaminergic cells, which express N/OFQ peptide (NOP) receptors, are located in the substantia nigra pars compacta and extend their dendrites in the substantia nigra pars reticulata, thereby modulating the basal ganglia output neurons.

In the present study, we demonstrate a direct role for D-aspartate in regulating hippocampal synaptic plasticity. These evidences were obtained using two different experimental strategies which enabled a nonphysiological increase of endogenous D-aspartate levels in the mouse hippocampus: a genetic approach based on the targeted deletion of Daspartate oxidase gene and another based on the oral administration of D-aspartate. Overall, our results indicate that increased D-aspartate content does not affect basal properties of synaptic transmission but enhances long-term potentiation in hippocampal slices from both genetic and pharmacological animal models. Besides electrophysiological data, behavioral analysis suggests that altered levels of D-aspartate in the hippocampus do not perturb basal spatial learning and memory abilities, but may selectively interfere with the dynamic NMDARdependent processes underlying cognitive flexibility.

Trace amines (TAs) are present in the central nervous system in which they up-regulate catecholamine release and are implicated in the pathogenesis of addiction, attention-deficit/hyperactivity disorder, Parkinson's disease, and schizophrenia. By using intracellular and patch-clamp recordings from dopaminergic cells in the rat midbrain slices, we report a depressant postsynaptic action of two TAs, ␤-phenylethylamine (␤-PEA) and tyramine (TYR) on the GABA B -mediated slow inhibitory postsynaptic potential and baclofen-activated outward currents. ␤-PEA and TYR activated G-proteins, interfering with the coupling between GABA B receptors and G-␤␥-gated inwardly rectifying potassium channels. This is the first demonstration that ␤-PEA and TYR depress inhibitory synaptic potentials in neurons of the central nervous system, supporting their emerging role as neuromodulators.

Background: The dopamine transporter (DAT) regulates the outflow of dopamine from synapses. Results: We present evidence that the DAT blockers cocaine and methylphenidate increase or reduce the release of DA in the striatum. Conclusion: The reducing effects on DA release are not dependent on a typical blockade of DAT. Significance: The paradoxical blunting of dopamine (DA) release could account for differential effects of psychostimulants.

The cellular mechanisms regulating dopamine (DA) release in the striatum have attracted much interest in recent years. By in vitro amperometric recordings in mouse striatal slices, we show that a brief (5 min) exposure to the metabotropic glutamate receptor agonist DHPG (50 μM) induces a profound depression of synaptic DA release, lasting over 1 h from DHPG washout. This long-term depression is sensitive to glycine, which preferentially inhibits local cholinergic interneurons, as well as to drugs acting on nicotinic acetylcholine receptors and to the pharmacological depletion of released acetylcholine. The same DHPG treatment induces a parallel long-lasting enhancement in the tonic firing of presumed striatal cholinergic interneurons, measured with multi-electrode array recordings. When DHPG is bilaterally infused in vivo in the mouse striatum, treated mice display an anxiety-like behavior. Our results demonstrate that metabotropic glutamate receptors stimulation gives rise to a pro...

Background-Homozygous or compound heterozygous mutations in the PTEN-induced kinase 1 (PINK1) gene are causative of autosomal recessive, early onset PD. Single heterozygous mutations have been repeatedly detected in a subset of patients as well as in non-affected subjects, and their significance has long been debated. Several neurophysiological studies from nonmanifesting PINK1 heterozygotes have shown the existence of neural plasticity abnormalities, indicating the presence of specific endophenotypic traits in the heterozygous state. Methods-In the present study, we performed a functional analysis of corticostriatal synaptic plasticity in heterozygous PINK1 knockout (PINK1 +/−) mice by a multidisciplinary approach. Results-We found that, despite a normal motor behavior, repetitive activation of cortical inputs to striatal neurons failed to induce long-term potentiation (LTP), whereas long-term depression (LTD) was normal. Although nigral dopaminergic neurons exhibited normal morphological and electrophysiological properties with normal responses to dopamine receptor activation, we measured a significantly lower dopamine release in the striatum of PINK1 +/− , compared to control mice, suggesting that a decrease in stimulus-evoked dopamine overflow acts as a major determinant for the LTP deficit. Accordingly, pharmacological agents capable of increasing the

Metabolic stress associated to mitochondrial dysfunction has been put forward as an important factor causing degeneration of mesencephalic dopamine-containing neurons in Parkinson's disease (PD). Here we overview how these neurons react to acute hypoxia or hypoglycemia, that are conditions of energy deprivation causing a reduced production of ATP by mitochondria. These neurons, which show a tonic firing discharge under normal condition, undergo into membrane hyperpolarization during hypoxia or hypoglycemia that silence their spontaneous activity. We outline the cellular mechanisms causing membrane hyperpolarization and the accompanied disturbances of intracellular calcium and sodium homeostasis. A better understanding of the changes occurring during transient energy deprivation might contribute to understand the physiopathology of these neurons that derives from mitochondrial dysfunction.

The presence of α-synuclein (α-syn) in Lewy bodies and Lewy neurites is an important characteristic of the neurodegenerative processes of substantia nigra pars compacta (SNpc) dopaminergic (DAergic) neurons in Parkinson's disease (PD) and other synucleinopathies. Here we report that Berlin-Druckrey rats carrying a spontaneous mutation in the 3′ untranslated region of α-syn mRNA (m/m rats) display a marked accumulation of α-syn in the mesencephalic area, striatum and frontal cortex, accompanied to severe dysfunctions in the dorsolateral striatum. Despite a small reduction in the number of SNpc and ventral tegmental area DAergic cells, the surviving dopaminergic neurons of the m/m rats do not show clearcut alterations of the spontaneous and evoked firing activity, DA responses and somatic amphetamine-induced firing inhibition. Interestingly, mutant DAergic neurons display diminished whole-cell Ih conductance and a reduced frequency of spontaneous excitatory synaptic currents. By contrast, m/m rats show a severe impairment of DA and glutamate release in the dorsolateral striatum, as revealed by amperometric measure of DA currents and by electrophysiological recordings of glutamatergic synaptic events in striatal medium spiny neurons. These functional impairments are paralleled by a decreased expression of the DA transporter and VGluT1 proteins in the same area. Thus, together with α-syn overload in the mesencephalic region, striatum and frontal cortex, the main functional alterations occur in the DAergic and glutamatergic terminals in the dorsal striatum of the m/m rats.

Alterations of the dopaminergic (DAergic) system are frequently reported in Alzheimer's disease (AD) patients and are commonly linked to cognitive and non-cognitive symptoms. However, the cause of DAergic system dysfunction in AD remains to be elucidated. We investigated alterations of the midbrain DAergic system in the Tg2576 mouse model of AD, overexpressing a mutated human amyloid precursor protein (APPswe). Here, we found an age-dependent DAergic neuron loss in the ventral tegmental area (VTA) at pre-plaque stages, although substantia nigra pars compacta (SNpc) DAergic neurons were intact. The selective VTA DAergic neuron degeneration results in lower DA outflow in the hippocampus and nucleus accumbens (NAc) shell. The progression of DAergic cell death correlates with impairments in CA1 synaptic plasticity, memory performance and food reward processing. We conclude that in this mouse model of AD, degeneration of VTA DAergic neurons at pre-plaque stages contributes to memory ...