Remote control of induced dopaminergic neurons in parkinsonian rats
Maria Teresa Dell’Anno, … , Alexander Dityatev, Vania Broccoli
Maria Teresa Dell’Anno, … , Alexander Dityatev, Vania Broccoli
Published June 17, 2014
Citation Information: J Clin Invest. 2014;124(7):3215-3229. https://doi.org/10.1172/JCI74664.
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Research Article

Remote control of induced dopaminergic neurons in parkinsonian rats

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.

Authors

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

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Figure 3

WGA synaptic tracing of iDA neurons in vitro and in vivo.

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WGA synaptic tracing of iDA neurons in vitro and in vivo. (A) iDA neuron...
(A) iDA neurons were infected with WGA-IRES-mCherry lentivirus 1 day after sorting and then replated on mouse MGE primary cell culture for in vitro analysis. (B) iDA neurons showed dense arborization after 21 DIV and were able to establish functional synaptic contacts with mouse primary cells. (C–F) Staining for WGA clearly revealed WGA+GFP– cells, demonstrating that WGA from infected iDA neurons was uptaken from surrounding GAD+ neural cells through functional synapses. (G) iDA neurons expressing WGA were also transplanted into the striatum of immunodepressed mice (n = 4), and animals were kept alive for 5 weeks. (H–K) Immunohistochemistry shows WGA+GFP– cells in the host tissue. These cells were also positive for NeuN, as shown in triple immunostaining, thus proving the ability of iDA neurons to establish synaptic connections with neighboring neurons after grafting. Scale bars: 60 μm (B), 20 μm (C–F), 200 μm (G), 20 μm (H–K). B–K are representative images from 3 independent experiments performed on 4 mice per group.