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 4

Functional integration of transplanted iDA neurons.

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Functional integration of transplanted iDA neurons. (A and B) Immunohist...
(A and B) Immunohistochemistry of iDA neurons grafted into immunodepressed mice for electrophysiological recordings. (C–E) High magnification of a biocytin-filled reprogrammed neuron immunostained for GFP. (F) Voltage-dependent Na+ (negative-going) and K+ (positive-going) currents in response to 10-mV steps (starting from a Vh of –70 mV) in voltage-clamp mode. Inset shows a magnified view of the first portion of the traces. (G) Voltage responses of an iDA neuron to hyperpolarizing and depolarizing current steps in current-clamp mode. (H) Continuous AP firing was induced by a slow suprathreshold current ramp (~4 pA/second). (I) Assessment of FFN511+ synaptic terminals on transplanted TH-GFP+ iDA neuronal fibers in brain slices. (J) Upper panel: sEPSCs recorded at a Vh of –70 mV were blocked by the extracellular application of 5 μM NBQX. Lower panel: sIPSCs recorded at a Vh of 0 mV were blocked by the extracellular application of 10 μM gabazine. Insets show averaged EPSCs and IPSCs, respectively, in an expanded time scale. Scale bars: 600 μm (A), 150 μm (B), 50 μm (C–E), 100 μm (I). A–J are representative images of 3 independent experiments performed on 3 mice and 6 cells (F–J).