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6. L-Glutamate Biosensor for In Vitro Investigations: Application in Brain Extracts.

Author

Razumiene, Julija, Leo, Damiana, Gureviciene, Vidute, Ratautas, Dalius, Gaidukevic, Justina, and Sakinyte-Urbikiene, Ieva

Subjects
BIOSENSORS, MALACHITE green, GRAPHENE oxide, HYDROGEN peroxide, NEUROLOGICAL disorders, DYES & dyeing
Abstract

Investigations of L-glutamate release in living organisms can help to identify novel L-glutamate-related pathophysiological pathways, since abnormal transmission of L-glutamate can cause many neurological diseases. For the first time, a nitrogen-modified graphene oxide (GO) sample (RGO) is prepared through a simple and facile one-pot hydrothermal reduction of GO in the presence of 20 wt.% of the dye malachite green and is used for amperometric biosensing. The biosensor demonstrates adequate stability and is easy to prepare and calibrate. The biosensor detects the current generated during the electrooxidation of hydrogen peroxide released in the L-glutamate that is converted to the alpha-ketoglutarate catalyzed by L-glutamate oxidase. The biosensor consists of a semipermeable membrane, with L-glutamate oxidase (EC 1.4.3.11) immobilized in albumin and RGO and the working Pt electrode. First, the basic version of the L-glutamate biosensor is examined in PBS to investigate its sensitivity, reliability, and stability. To demonstrate the applicability of the L-glutamate biosensor in the analysis of complex real samples, quantification of L-glutamate in bovine brain extract is performed and the accuracy of the biosensor is confirmed by alternative methods. The enhanced version of the L-glutamate biosensor is applied for L-glutamate release investigations in a newly developed strain of rats (DAT-knockout, DAT-KO). [ABSTRACT FROM AUTHOR]

Published
2023
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8. Dopamine Transporter Knockout Rats Show Impaired Wellbeing in a Multimodal Severity Assessment Approach

Author

Mallien, Anne Stephanie, primary, Becker, Laura, additional, Pfeiffer, Natascha, additional, Terraneo, Federica, additional, Hahn, Melissa, additional, Middelman, Anthonieke, additional, Palme, Rupert, additional, Creutzberg, Kerstin Camile, additional, Begni, Veronica, additional, Riva, Marco Andrea, additional, Leo, Damiana, additional, Potschka, Heidrun, additional, Fumagalli, Fabio, additional, Homberg, Judith R., additional, and Gass, Peter, additional

Published
2022
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11. Disruption of the PDZ domain–binding motif of the dopamine transporter uniquely alters nanoscale distribution, dopamine homeostasis, and reward motivation

Author

Sørensen, Gunnar, primary, Rickhag, Mattias, additional, Leo, Damiana, additional, Lycas, Matthew D., additional, Ridderstrøm, Pernille Herrstedt, additional, Weikop, Pia, additional, Lilja, Jamila H., additional, Rifes, Pedro, additional, Herborg, Freja, additional, Woldbye, David, additional, Wörtwein, Gitta, additional, Gainetdinov, Raul R., additional, Fink-Jensen, Anders, additional, and Gether, Ulrik, additional

Published
2021
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12. Remote control of induced dopaminergic neurons in parkinsonian rats

Author

Dell'Anno, Maria Teresa, Caiazzo, Massimiliano, Leo, Damiana, Dvoretskova, Elena, Medrihan, Lucian, Colasante, Gaia, Giannelli, Serena, Theka, Ilda, Russo, Giovanni, Mus, Liudmila, Pezzoli, Gianni, Gainetdinov, Raul R., Benfenati, Fabio, Taverna, Stefano, Dityatev, Alexander, and Broccoli, Vania

Subjects
Neurons -- Physiological aspects, Medical research, Medicine, Experimental, Cellular therapy -- Research, Dopaminergic mechanisms -- Physiological aspects, Parkinson's disease -- Care and treatment, Health care industry
Abstract

Direct lineage reprogramming through genetic-based strategies enables the conversion of differentiated somatic cells into functional neurons and distinct neuronal subtypes. Induced dopaminergic (iDA) neurons can be generated by direct conversion of skin fibroblasts; however, their in vivo phenotypic and functional properties remain incompletely understood, leaving their impact on Parkinson's disease (PD) cell therapy and modeling uncertain. Here, we determined that iDA neurons retain a transgene- independent stable phenotype in culture and in animal models. Furthermore, transplanted iDA neurons functionally integrated into host neuronal tissue, exhibiting electrically excitable membranes, synaptic currents, dopamine release, and substantial reduction of motor symptoms in a PD animal model. Neuronal cell replacement approaches will benefit from a system that allows the activity of transplanted neurons to be controlled remotely and enables modulation depending on the physiological needs of the recipient; therefore, we adapted a DREADD (designer receptor exclusively activated by designer drug) technology for remote and real- time control of grafted iDA neuronal activity in living animals. Remote DREADD-dependent iDA neuron activation markedly enhanced the beneficial effects in transplanted PD animals. These data suggest that iDA neurons have therapeutic potential as a cell replacement approach for PD and highlight the applicability of pharmacogenetics for enhancing cellular signaling in reprogrammed cell-based approaches., Introduction The differentiated cell state has been traditionally considered irreversible and insensitive to epigenetic modifications. Nevertheless, in contrast with this classical view, accumulating evidence indicates that cell identity relies on [...]

Published
2014
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14. Disruption of the PDZ domain-binding motif of the dopamine transporter uniquely alters nanoscale distribution, dopamine homeostasis, and reward motivation

Author

Sørensen, Gunnar, Rickhag, Mattias, Leo, Damiana, Lycas, Matthew D., Ridderstrøm, Pernille Herrstedt, Weikop, Pia, Lilja, Jamila H., Rifes, Pedro, Herborg, Freja, Woldbye, David, Wörtwein, Gitta, Gainetdinov, Raul R., Fink-Jensen, Anders, Gether, Ulrik, Sørensen, Gunnar, Rickhag, Mattias, Leo, Damiana, Lycas, Matthew D., Ridderstrøm, Pernille Herrstedt, Weikop, Pia, Lilja, Jamila H., Rifes, Pedro, Herborg, Freja, Woldbye, David, Wörtwein, Gitta, Gainetdinov, Raul R., Fink-Jensen, Anders, and Gether, Ulrik

Abstract

The dopamine (DA) transporter (DAT) is part of a presynaptic multiprotein network involving interactions with scaffold proteins via its C-terminal PDZ domain-binding sequence. Using a mouse model expressing DAT with mutated PDZ-binding sequence (DAT-AAA), we previously demonstrated the importance of this binding sequence for striatal expression of DAT. Here, we show by application of direct stochastic reconstruction microscopy not only that the striatal level of transporter is reduced in DAT-AAA mice but also that the nanoscale distribution of this transporter is altered with a higher propensity of DAT-AAA to localize to irregular nanodomains in dopaminergic terminals. In parallel, we observe mesostriatal DA adaptations and changes in DA-related behaviors distinct from those seen in other genetic DAT mouse models. DA levels in the striatum are reduced to ∼45% of that of WT, accompanied by elevated DA turnover. Nonetheless, fast-scan cyclic voltammetry recordings on striatal slices reveal a larger amplitude and prolonged clearance rate of evoked DA release in DAT-AAA mice compared with WT mice. Autoradiography and radioligand binding show reduced DA D2 receptor levels, whereas immunohistochemistry and autoradiography show unchanged DA D1 receptor levels. In behavioral experiments, we observe enhanced self-administration of liquid food under both a fixed ratio of one and progressive ratio schedule of reinforcement but a reduction compared with WT when using cocaine as reinforcer. In summary, our data demonstrate how disruption of PDZ domain interactions causes changes in DAT expression and its nanoscopic distribution that in turn alter DA clearance dynamics and related behaviors.

Published
2021

15. Direct generation of functional dopaminergic neurons from mouse and human fibroblasts

Author

Caiazzo, Massimiliano, Dell'Anno, Maria Teresa, Dvoretskova, Elena, Lazarevic, Dejan, Taverna, Stefano, Leo, Damiana, Sotnikova, Tatyana D., Menegon, Andrea, Roncaglia, Paola, Colciago, Giorgia, Russo, Giovanni, Carninci, Piero, Pezzoli, Gianni, Gainetdinov, Raul R., Gustincich, Stefano, Dityatev, Alexander, and Broccoli, Vania

Subjects
Neurons -- Physiological aspects -- Health aspects, Dopamine -- Physiological aspects -- Health aspects, Fibroblasts -- Physiological aspects -- Health aspects, Parkinson's disease -- Care and treatment -- Physiological aspects, Environmental issues, Science and technology, Zoology and wildlife conservation
Abstract

Transplantation of dopaminergic neurons can potentially improve the clinical outcome of Parkinson's disease, a neurological disorder resulting from degeneration of mesencephalic dopaminergic neurons (1,2). In particular, transplantation of embryonic-stem-cellderived dopaminergic neurons has been shown to be efficient in restoring motor symptoms in conditions of dopamine deficiency (3,4). However, the use of pluripotent-derived cells might lead to the development of tumours if not properly controlled (5). Here we identified a minimal set of three transcription factors--Mash1 (also known as Ascll), Nurr1 (also known as Nr4a2) and Lmx1a--that are able to generate directly functional dopaminergic neurons from mouse and human fibroblasts without reverting to a progenitor cell stage. Induced dopaminergic (iDA) cells release dopamine and show spontaneous electrical activity organized in regular spikes consistent with the pacemaker activity featured by brain dopaminergic neurons. The three factors were able to elicit dopaminergic neuronal conversion in prenatal and adult fibroblasts from healthy donors and Parkinson's disease patients. Direct generation of iDA cells from somatic cells might have significant implications for understanding critical processes for neuronal development, invitrodisease modelling and cell replacement therapies., Seminal studies have demonstrated that functional neurons can be generated independently of stem cells by direct cell conversion through genetics-based approaches (6). More recently, in a set of elegant experiments, [...]

Published
2011
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17. Remote control of induced dopaminergic neurons in parkinsonian rats

Author

DellʼAnno, Maria Teresa, Caiazzo, Massimiliano, Leo, Damiana, Dvoretskova, Elena, Medrihan, Lucian, Colasante, Gaia, Giannelli, Serena, Theka, Ilda, Russo, Giovanni, Mus, Liudmila, Pezzoli, Gianni, Gainetdinov, Raul R., Benfenati, Fabio, Taverna, Stefano, Dityatev, Alexander, and Broccoli, Vania

Published
2014
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19. Disrupted PDZ-domain binding motif of the dopamine transporter uniquely alters nanoscale distribution, dopamine homeostasis and reward motivation

Author

Sørensen, Gunnar, primary, Rickhag, Mattias, additional, Leo, Damiana, additional, Lycas, Matthew D., additional, Ridderstrøm, Pernille Herrstedt, additional, Weikop, Pia, additional, Herborg, Freja, additional, Woldbye, David, additional, Wörtwein, Gitta, additional, Gainetdinov, Raul R., additional, Fink-Jensen, Anders, additional, and Gether, Ulrik, additional

Published
2021
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22. Trace Amine-Associated Receptor 5 Provides Olfactory Input Into Limbic Brain Areas and Modulates Emotional Behaviors and Serotonin Transmission

Author

Espinoza, Stefano, primary, Sukhanov, Ilya, additional, Efimova, Evgeniya V., additional, Kozlova, Alena, additional, Antonova, Kristina A., additional, Illiano, Placido, additional, Leo, Damiana, additional, Merkulyeva, Natalia, additional, Kalinina, Daria, additional, Musienko, Pavel, additional, Rocchi, Anna, additional, Mus, Liudmila, additional, Sotnikova, Tatiana D., additional, and Gainetdinov, Raul R., additional

Published
2020
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27. Presynaptic action of neurotensin on dopamine release through inhibition of D2 receptor function

Author

Trudeau Louis-Eric, Leo Damiana, Martel Philippe, and Fawaz Charbel S

Subjects
Neurosciences. Biological psychiatry. Neuropsychiatry, RC321-571, Neurophysiology and neuropsychology, QP351-495
Abstract

Abstract Background Neurotensin (NT) is known to act on dopamine (DA) neurons at the somatodendritic level to regulate cell firing and secondarily enhance DA release. In addition, anatomical and indirect physiological data suggest the presence of NT receptors at the terminal level. However, a clear demonstration of the mechanism of action of NT on dopaminergic axon terminals is lacking. We hypothesize that NT acts to increase DA release by inhibiting the function of terminal D2 autoreceptors. To test this hypothesis, we used fast-scan cyclic voltammetry (FCV) to monitor in real time the axonal release of DA in the nucleus accumbens (NAcc). Results DA release was evoked by single electrical pulses and pulse trains (10 Hz, 30 pulses). Under these two stimulation conditions, we evaluated the characteristics of DA D2 autoreceptors and the presynaptic action of NT in the NAcc shell and shell/core border region. The selective agonist of D2 autoreceptors, quinpirole (1 μM), inhibited DA overflow evoked by both single and train pulses. In sharp contrast, the selective D2 receptor antagonist, sulpiride (5 μM), strongly enhanced DA release triggered by pulse trains, without any effect on DA release elicited by single pulses, thus confirming previous observations. We then determined the effect of NT (8–13) (100 nM) and found that although it failed to increase DA release evoked by single pulses, it strongly enhanced DA release evoked by pulse trains that lead to prolonged DA release and engage D2 autoreceptors. In addition, initial blockade of D2 autoreceptors by sulpiride considerably inhibited further facilitation of DA release generated by NT (8–13). Conclusion Taken together, these data suggest that NT enhances DA release principally by inhibiting the function of terminal D2 autoreceptors and not by more direct mechanisms such as facilitation of terminal calcium influx.

Published
2009
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28. In Vivo Amphetamine Action is Contingent on αCaMKII

Author

Steinkellner, Thomas, Mus, Liudmilla, Eisenrauch, Birgit, Constantinescu, Andreea, Leo, Damiana, Konrad, Lisa, Rickhag, Mattias, Sørensen, Gunnar, Efimova, Evgenia V, Kong, Eryan, Willeit, Matthäus, Sotnikova, Tatyana D, Kudlacek, Oliver, Gether, Ulrik, Freissmuth, Michael, Pollak, Daniela D, Gainetdinov, Raul R, and Sitte, Harald H

Subjects
Male, Mice, Knockout, Dextroamphetamine, Dopamine, Membrane Proteins, Hyperkinesis, Corpus Striatum, Receptors, Dopamine, Mice, Inbred C57BL, Reward, Space Perception, Conditioning, Psychological, Animals, Original Article, Central Nervous System Stimulants, Calcium-Calmodulin-Dependent Protein Kinase Type 2, Disks Large Homolog 4 Protein, Guanylate Kinases, Locomotion
Abstract

Addiction to psychostimulants (ie, amphetamines and cocaine) imposes a major socioeconomic burden. Prevention and treatment represent unmet medical needs, which may be addressed, if the mechanisms underlying psychostimulant action are understood. Cocaine acts as a blocker at the transporters for dopamine (DAT), serotonin (SERT), and norepinephrine (NET), but amphetamines are substrates that do not only block the uptake of monoamines but also induce substrate efflux by promoting reverse transport. Reverse transport has been a focus of research for decades but its mechanistic basis still remains enigmatic. Recently, transporter-interacting proteins were found to regulate amphetamine-triggered reverse transport: calmodulin kinase IIα (αCaMKII) is a prominent example, because it binds the carboxyl terminus of DAT, phosphorylates its amino terminus, and supports amphetamine-induced substrate efflux in vitro. Here, we investigated whether, in vivo, the action of amphetamine was contingent on the presence of αCaMKII by recording the behavioral and neurochemical effects of amphetamine. Measurement of dopamine efflux in the dorsal striatum by microdialysis revealed that amphetamine induced less dopamine efflux in mice lacking αCaMKII. Consistent with this observation, the acute locomotor responses to amphetamine were also significantly blunted in αCaMKII-deficient mice. In addition, while the rewarding properties of amphetamine were preserved in αCaMKII-deficient mice, their behavioral sensitization to amphetamine was markedly reduced. Our findings demonstrate that amphetamine requires the presence of αCaMKII to elicit a full-fledged effect on DAT in vivo: αCaMKII does not only support acute amphetamine-induced dopamine efflux but is also important in shaping the chronic response to amphetamine.

Published
2014

31. Pronounced Hyperactivity, Cognitive Dysfunctions, and BDNF Dysregulation in Dopamine Transporter Knock-out Rats

Author

Leo, Damiana, primary, Sukhanov, Ilya, additional, Zoratto, Francesca, additional, Illiano, Placido, additional, Caffino, Lucia, additional, Sanna, Fabrizio, additional, Messa, Giulia, additional, Emanuele, Marco, additional, Esposito, Alessandro, additional, Dorofeikova, Mariia, additional, Budygin, Evgeny A., additional, Mus, Liudmila, additional, Efimova, Evgenia V., additional, Niello, Marco, additional, Espinoza, Stefano, additional, Sotnikova, Tatyana D., additional, Hoener, Marius C., additional, Laviola, Giovanni, additional, Fumagalli, Fabio, additional, Adriani, Walter, additional, and Gainetdinov, Raul R., additional

Published
2018
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33. Role of Dopamine D2/D3 Receptors in Development, Plasticity, and Neuroprotection in Human iPSC-Derived Midbrain Dopaminergic Neurons

Author

Bono, Federica, primary, Savoia, Paola, additional, Guglielmi, Adele, additional, Gennarelli, Massimo, additional, Piovani, Giovanna, additional, Sigala, Sandra, additional, Leo, Damiana, additional, Espinoza, Stefano, additional, Gainetdinov, Raul R., additional, Devoto, Paola, additional, Spano, PierFranco, additional, Missale, Cristina, additional, and Fiorentini, Chiara, additional

Published
2017
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35. Rapid Conversion of Fibroblasts into Functional Forebrain GABAergic Interneurons by Direct Genetic Reprogramming

Author

Colasante, Gaia, primary, Lignani, Gabriele, additional, Rubio, Alicia, additional, Medrihan, Lucian, additional, Yekhlef, Latefa, additional, Sessa, Alessandro, additional, Massimino, Luca, additional, Giannelli, Serena G., additional, Sacchetti, Silvio, additional, Caiazzo, Massimiliano, additional, Leo, Damiana, additional, Alexopoulou, Dimitra, additional, Dell’Anno, Maria Teresa, additional, Ciabatti, Ernesto, additional, Orlando, Marta, additional, Studer, Michele, additional, Dahl, Andreas, additional, Gainetdinov, Raul R., additional, Taverna, Stefano, additional, Benfenati, Fabio, additional, and Broccoli, Vania, additional

Published
2015
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37. TAAR1 Modulates Cortical Glutamate NMDA Receptor Function

Author

Espinoza, Stefano, primary, Lignani, Gabriele, additional, Caffino, Lucia, additional, Maggi, Silvia, additional, Sukhanov, Ilya, additional, Leo, Damiana, additional, Mus, Liudmila, additional, Emanuele, Marco, additional, Ronzitti, Giuseppe, additional, Harmeier, Anja, additional, Medrihan, Lucian, additional, Sotnikova, Tatyana D, additional, Chieregatti, Evelina, additional, Hoener, Marius C, additional, Benfenati, Fabio, additional, Tucci, Valter, additional, Fumagalli, Fabio, additional, and Gainetdinov, Raul R, additional

Published
2015
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38. A Caenorhabditis elegans model to study dopamine transporter deficiency syndrome.

Author

Illiano, Placido, Lanzo, Ambra, Leo, Damiana, Paglione, Maria, Zampi, Giuseppina, Gainetdinov, Raul R., Di Schiavi, Elia, and Bolam, Paul

Subjects
CAENORHABDITIS elegans, NEUROBIOLOGY, GENETIC mutation, GENETIC disorders, GENE knockout
Abstract

Dopamine transporter deficiency syndrome ( DTDS) is a novel autosomal recessive disorder caused by mutations in the dopamine transporter ( DAT), which leads to the partial or total loss of function of the protein. DTDS is a pharmacoresistant syndrome and very little is known about its neurobiology, in part due to the lack of relevant animal models. The objective of this study was to establish the first animal model for DTDS with strong construct validity, using Caenorhabditis elegans, and to investigate the in vivo role played by DTDS-related mutations found in human DAT ( hDAT). We took advantage of a C. elegans knockout for the hDAT orthologue, ce dat-1, to obtain genetically humanized animals bearing hDAT, in the wild type and in two mutated forms (399delG and 941C>T), in a null background. In C. elegans transgenic animals expressing the human wild-type form, we observed a rescue of the knockout phenotype, as assessed using two well-established paradigms, known to be regulated by the endogenous uptake of dopamine or 6-hydroxydopamine (6- OHDA) by DAT. The less severe mutation (941C>T) was able to partially rescue only one of the knockout phenotypes, whereas the 399delG mutation impaired DAT function in both phenotypic paradigms. Our in vivo phenotypic findings demonstrate a functional conservation between human and nematode DAT and validate previous in vitro indications of the loss of function of hDAT in carriers of DTDS-related mutations. Taken together, these observations establish C. elegans as a novel animal model for fast and inexpensive screening of hDAT mutations in functional and in vivo tests. [ABSTRACT FROM AUTHOR]

Published
2017
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39. Remote control of induced dopaminergic neurons in parkinsonian rats

Author

Dell’Anno, Maria Teresa, primary, Caiazzo, Massimiliano, additional, Leo, Damiana, additional, Dvoretskova, Elena, additional, Medrihan, Lucian, additional, Colasante, Gaia, additional, Giannelli, Serena, additional, Theka, Ilda, additional, Russo, Giovanni, additional, Mus, Liudmila, additional, Pezzoli, Gianni, additional, Gainetdinov, Raul R., additional, Benfenati, Fabio, additional, Taverna, Stefano, additional, Dityatev, Alexander, additional, and Broccoli, Vania, additional

Published
2014
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41. Social withdrawal and gambling-like profile after lentiviral manipulation of DAT expression in the rat accumbens

Author

Adriani, Walter, Boyer, Frederic, Leo, Damiana, Canese, Rossella, Podo, Franca, Perrone-Capano, Carla, Dreyer, Jean-Luc, Laviola, Giovanni, Adriani, Walter, Boyer, Frederic, Leo, Damiana, Canese, Rossella, Podo, Franca, Perrone-Capano, Carla, Dreyer, Jean-Luc, and Laviola, Giovanni

Abstract

Dysfunction of brain dopamine transporter (DAT) has been associated with sensation seeking and impulse-control disorders. We recently generated a new animal model by stereotaxical inoculation of lentiviral vectors, which allowed localized intra-accumbal delivery of modulators for DAT gene: GFP (green fluorescent protein) control, silencers (Sil), a regulatable enhancer (DAT+), or both (DAT+Sil). Wistar male rats were followed both for socio-emotional profiles and for propensity to seek risky, uncertain rewards. Elevated anxiety and affiliation towards an unfamiliar partner emerged in Sil rats. Interestingly, in DAT+Sil rats (and Sil rats to a lesser extent) levels of playful social interaction were markedly reduced compared to controls. These DAT+Sil rats displayed a marked ‘gambling-like’ profile (i.e. preference for a large/uncertain over a small/sure reward), which disappeared upon doxycycline-induced switch-off onto DAT enhancer, but consistently reappeared with doxycycline removal. MRI-guided ¹H-MRS (at 4.7 T) examinations in vivo (under anaesthesia) revealed changes in the bioenergetic metabolites (phosphocreatine and total creatine) for DAT+Sil rats, indicating a functional up-regulation of dorsal striatum (Str) and conversely a down-regulation of ventral striatum (i.e. nucleus accumbens, NAc). A combined profile of (1) enhanced proneness to gambling and (2) strong social withdrawal is thus associated with altered DAT-induced balance within forebrain dopamine systems. In fact, risk of developing a gambling-prone, social-avoidant psychopathology might be associated with (1) dominant semi-automatic strategies and/or habits, developed within Str circuits, and (2) reduced NAc function, with poorer feedback adjustment on decisions by aversive experiences.

Published
2010

42. Rapid Generation of Functional Dopaminergic Neurons From Human Induced Pluripotent Stem Cells Through a Single-Step Procedure Using Cell Lineage Transcription Factors

Author

Theka, Ilda, primary, Caiazzo, Massimiliano, additional, Dvoretskova, Elena, additional, Leo, Damiana, additional, Ungaro, Federica, additional, Curreli, Sebastiano, additional, Managò, Francesca, additional, Dell'Anno, Maria Teresa, additional, Pezzoli, Gianni, additional, Gainetdinov, Raul R., additional, Dityatev, Alexander, additional, and Broccoli, Vania, additional

Published
2013
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