Abstract
Intrastriatal grafts of stem cell-derived dopamine (DA) neurons induce behavioral recovery in animal models of Parkinson's disease (PD), but how they functionally integrate in host neural circuitries is poorly understood. Here, Wnt5a-overexpressing neural stem cells derived from embryonic ventral mesencephalon of tyrosine hydroxylase-GFP transgenic mice were expanded as neurospheres and transplanted into organotypic cultures of wild type mouse striatum. Differentiated GFP-labeled DA neurons in the grafts exhibited mature neuronal properties, including spontaneous firing of action potentials, presence of post-synaptic currents, and functional expression of DA D2 autoreceptors. These properties resembled those recorded from identical cells in acute slices of intrastriatal grafts in the 6-hydroxy-DA-induced mouse PD model and from DA neurons in intact substantia nigra. Optogenetic activation or inhibition of grafted cells and host neurons using channelrhodopsin-2 (ChR2) and halorhodopsin (NpHR), respectively, revealed complex, bi-directional synaptic interactions between grafted cells and host neurons and extensive synaptic connectivity within the graft. Our data demonstrate for the first time using optogenetics that ectopically grafted stem cell-derived DA neurons become functionally integrated in the DA-denervated striatum. Further optogenetic dissection of the synaptic wiring between grafted and host neurons will be crucial to clarify the cellular and synaptic mechanisms underlying behavioral recovery as well as adverse effects following stem cell-based DA cell replacement strategies in PD.
Highlights
Generation of dopamine (DA) neurons for transplantation from stem cells of various sources has gained substantial interest for restorative therapy in Parkinson’s disease (PD) [1,2,3]
For the first time, optogenetic methods in combination with patch-clamp recordings to analyze the functional integration of stem cell-derived DA neurons grafted into an in vitro PD model
Our findings indicate that the organotypic hemisphere cultures used here, having relatively preserved cortico-striatal connections but severed DA input to the striatum, represent a suitable model for preliminary screening of the functional properties of stem cellderived DA neurons and their integration into host neural circuitries
Summary
Generation of dopamine (DA) neurons for transplantation from stem cells of various sources has gained substantial interest for restorative therapy in Parkinson’s disease (PD) [1,2,3]. The reversal of PD symptoms following intrastriatal implantation of such cells in animal models raises questions about the cellular and synaptic mechanisms responsible for functional recovery, the level of synaptic integration of the stem cell-derived DA neurons into host neural circuitries. With the exception of mouse embryonic stem (ES) cell-derived DA neurons [4,5], the extent of DA fiber outgrowth has been limited and the afferent inputs to the grafted DA cells have not been determined. It is not known to what extent stem cell-derived DA neurons establish efferent synaptic connections with host neurons or receive functional synaptic inputs from other grafted cells or host neurons
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