Properties of dopaminergic neurons in organotypic mesencephalic-striatal co-cultures - evidence for a facilitatory effect of dopamine on the glutamatergic input mediated by α-1 adrenergic receptors

Cucchiaroni, Maria Letizia; Freestone, Peter S.; Berretta, Nicola; Viscomi, Maria Teresa; Bisicchia, Elisa; Molinari, Marco; Bernardi, Giorgio; Lipski, Janusz; Mercuri, Nicola Biagio; Guatteo, Ezia

doi:10.1111/j.1460-9568.2011.07659.x

Cited by 11 publications

(8 citation statements)

References 76 publications

(104 reference statements)

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“…Therefore, α1-AR actions on DA neuron excitability follow synaptic activation induced by action potentials from presynaptic neurons. In accordance with our results, Cucchiaroni et al,(2011) reported that α1-AR activation increases glutamate release onto DA cells of mesencephalic-striatal co-cultures. The enhanced glutamate release was dependent on presynaptic action potentials and had no effect on the sensitivity of postsynaptic glutamate receptors in DA cells.…”

Section: Discussionsupporting

confidence: 93%

Activation of alpha1-adrenoceptors enhances glutamate release onto ventral tegmental area dopamine cells

2012

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The ventral tegmental area (VTA) plays an important role in reward and motivational processes that facilitate the development of drug addiction. Glutamatergic inputs into the VTA contribute to dopamine (DA) neuronal activation related to reward and response-initiating effects in drug abuse. Previous investigations indicate that alpha1-adrenoreceptors (α1-AR) are primarily localized at presynaptic elements in the ventral midbrain. Studies from several brain regions have shown that presynaptic α1-AR activation enhance glutamate release. Therefore, we hypothesized that glutamate released onto VTA-DA neurons is modulated by pre-synaptic α1-AR. Recordings were obtained from putative VTA-DA cells of male Sprague-Dawley rats (28–50 days postnatal) using voltage clamp techniques. Phenylephrine (10 µM) and methoxamine (80 µM), both α1-AR agonists, increased AMPA receptor-mediated excitatory postsynaptic currents (EPSCs) amplitude evoked by electrical stimulation of afferent fibers (p<0.05). This effect was blocked by the α1-AR antagonist prazosin (1 µM). Phenylephrine decreased the paired-pulse ratio and increased spontaneous EPSCs frequencies but not their amplitudes suggesting a presynaptic locus of action. No changes in miniature EPSCs (0.5 µM TTX) were observed after phenylephrine’s application which suggest that α1-AR effect was action potential dependent. Normal extra- and intracellular Ca2+ concentration seems necessary for the α1-AR effect since phenylephrine in low Ca2+ ACSF and depletion of intracellular Ca2+ stores with thapsigargin (10 µM) failed to increase the AMPA EPSCs amplitude . Chelerythrine (1 µM, PKC inhibitor) but not Rp-cAMPS (11 µM, PKA inhibitor) blocked the α1-AR activation effect on AMPA EPSCs, indicating that a PKC intracellular pathway is required. These results demonstrated that presynaptic α1-ARs activation modulates glutamatergic inputs that affect VTA-DA neurons excitability. α1-ARs action might be heterosynaptically localized at glutamatergic fibers terminating onto VTA-DA neurons. It is suggested that drug-induced changes in α1-AR could be part to the neuroadaptations occurring in the mesocorticolimbic circuitry during the addiction process.

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Section: Discussionsupporting

confidence: 93%

Activation of alpha1-adrenoceptors enhances glutamate release onto ventral tegmental area dopamine cells

2012

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“…Gruol and co-workers [39] reported that cells with little electrical activity had low resting [Ca 2+ ] i levels (∼50 nM); this may be consistent with our changes in [Ca 2+ ] i during differentiation. This developing spontaneous activity may also be consistent with the observation of spontaneous and burst firing potentials in dopaminergic neurons [42] , [43] , [44] , [45] . Ca 2+ signaling responses have not been studied widely in ESC-derived neuronal cultures before; apart from our previous work, a recent study by Malmersjo and co-workers [46] in human ESC-derived TH + neurons observed the development of Ca2+ responses in those cells (to, for example, KCl, Glut, or neurotensin stimulation) and concluded there were functional similarities to primary ventral midbrain dopaminergic neurons of mice.…”

Section: Discussionsupporting

confidence: 89%

In Vitro Maturation of Dopaminergic Neurons Derived from Mouse Embryonic Stem Cells: Implications for Transplantation

Watmuff¹,

Pouton

Haynes

2012

PLoS ONE

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The obvious motor symptoms of Parkinson's disease result from a loss of dopaminergic neurons from the substantia nigra. Embryonic stem cell-derived neural progenitor or precursor cells, adult neurons and fetal midbrain tissue have all been used to replace dying dopaminergic neurons. Transplanted cell survival is compromised by factors relating to the new environment, for example; hypoxia, mechanical trauma and excitatory amino acid toxicity. In this study we investigate, using live-cell fluorescence Ca2+ and Cl− imaging, the functional properties of catecholaminergic neurons as they mature. We also investigate whether GABA has the capacity to act as a neurotoxin early in the development of these neurons. From day 13 to day 21 of differentiation [Cl−]i progressively dropped in tyrosine hydroxylase positive (TH+) neurons from 56.0 (95% confidence interval, 55.1, 56.9) mM to 6.9 (6.8, 7.1) mM. At days 13 and 15 TH+ neurons responded to GABA (30 µM) with reductions in intracellular Cl− ([Cl−]i); from day 21 the majority of neurons responded to GABA (30 µM) with elevations of [Cl−]i. As [Cl−]i reduced, the ability of GABA (30 µM) to elevate intracellular Ca2+ ([Ca2+]i) did also. At day 13 of differentiation a three hour exposure to GABA (30 µM) or L-glutamate (30 µM) increased the number of midbrain dopaminergic (TH+ and Pitx3+) neurons labeled with the membrane-impermeable nuclear dye TOPRO-3. By day 23 cultures were resistant to the effects of both GABA and L-glutamate. We believe that neuronal susceptibility to amino acid excitotoxicity is dependent upon neuronal maturity, and this should be considered when isolating cells for transplantation studies.

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“…The remaining neurons had a much higher basal firing frequency, ranging between 4 and 11 Hz and characterized by mean values of 6.8 ± 1.4 and 6.4 ± 0.5 Hz, when measured with μG-SCD-MEAs and MEAs ( p > 0.1), respectively. This heterogeneity of responses can, most likely, be ascribed to the presence of distinct neuronal populations such as DA neurons from SN, GABAergic and DA neurons from the nearby ventral tegmental area (Berretta et al, 2010; Cucchiaroni et al, 2011). Optical images of GFP-TH + neurons that were cultured on μG-SCD-MEAs are provided in Figure 3D.…”

Section: Resultsmentioning

confidence: 99%

Quantal Release of Dopamine and Action Potential Firing Detected in Midbrain Neurons by Multifunctional Diamond-Based Microarrays

et al. 2019

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Micro-Graphitic Single Crystal Diamond Multi Electrode Arrays (μG-SCD-MEAs) have so far been used as amperometric sensors to detect catecholamines from chromaffin cells and adrenal gland slices. Besides having time resolution and sensitivity that are comparable with carbon fiber electrodes, that represent the gold standard for amperometry, μG-SCD-MEAs also have the advantages of simultaneous multisite detection, high biocompatibility and implementation of amperometric/potentiometric protocols, aimed at monitoring exocytotic events and neuronal excitability. In order to adapt diamond technology to record neuronal activity, the μG-SCD-MEAs in this work have been interfaced with cultured midbrain neurons to detect electrical activity as well as quantal release of dopamine (DA). μG-SCD-MEAs are based on graphitic sensing electrodes that are embedded into the diamond matrix and are fabricated using MeV ion beam lithography. Two geometries have been adopted, with 4 × 4 and 8 × 8 microelectrodes (20 μm × 3.5 μm exposed area, 200 μm spacing). In the amperometric configuration, the 4 × 4 μG-SCD-MEAs resolved quantal exocytosis from midbrain dopaminergic neurons. KCl-stimulated DA release occurred as amperometric spikes of 15 pA amplitude and 0.5 ms half-width, at a mean frequency of 0.4 Hz. When used as potentiometric multiarrays, the 8 × 8 μG-SCD-MEAs detected the spontaneous firing activity of midbrain neurons. Extracellularly recorded action potentials (APs) had mean amplitude of ∼-50 μV and occurred at a mean firing frequency of 0.7 Hz in 67% of neurons, while the remaining fired at 6.8 Hz. Comparable findings were observed using conventional MEAs (0.9 and 6.4 Hz, respectively). To test the reliability of potentiometric recordings with μG-SCD-MEAs, the D 2 -autoreceptor modulation of firing was investigated by applying levodopa ( L -DOPA, 20 μM), and comparing μG-SCD-MEAs, conventional MEAs and current-clamp recordings. In all cases, L -DOPA reduced the spontaneous spiking activity in most neurons by 70%, while the D 2 -antagonist sulpiride reversed this effect. Cell firing inhibition was generally associated with increased APs amplitude. A minority of neurons was either insensitive to, or potentiated by L -DOPA, suggesting that AP recordings originate from different midbrain neuronal subpopulations and reveal different modulatory pathways. Our data demonstrate, for the first time, that μG-SCD-MEAs are multi-functional biosensors suitable to resolve real-time DA release and AP firing in in vitro neuronal networks.

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Properties of dopaminergic neurons in organotypic mesencephalic-striatal co-cultures - evidence for a facilitatory effect of dopamine on the glutamatergic input mediated by α-1 adrenergic receptors

Cited by 11 publications

References 76 publications

Activation of alpha1-adrenoceptors enhances glutamate release onto ventral tegmental area dopamine cells

Activation of alpha1-adrenoceptors enhances glutamate release onto ventral tegmental area dopamine cells

In Vitro Maturation of Dopaminergic Neurons Derived from Mouse Embryonic Stem Cells: Implications for Transplantation

Quantal Release of Dopamine and Action Potential Firing Detected in Midbrain Neurons by Multifunctional Diamond-Based Microarrays

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