The optogenetic approach to gain control over neuronal excitability both in vitro and in vivo has emerged as a fascinating scientific tool to explore neuronal networks, but it also opens possibilities for developing novel treatment strategies for neurologic conditions. We have explored whether such an optogenetic approach using the light-driven halorhodopsin chloride pump from Natronomonas pharaonis (NpHR), modified for mammalian CNS expression to hyperpolarize central neurons, may inhibit excessive hyperexcitability and epileptiform activity. We show that a lentiviral vector containing the NpHR gene under the calcium/calmodulin-dependent protein kinase IIα promoter transduces principal cells of the hippocampus and cortex and hyperpolarizes these cells, preventing generation of action potentials and epileptiform activity during optical stimulation. This study proves a principle, that selective hyperpolarization of principal cortical neurons by NpHR is sufficient to curtail paroxysmal activity in transduced neurons and can inhibit stimulation train-induced bursting in hippocampal organotypic slice cultures, which represents a model tissue of pharmacoresistant epilepsy. This study demonstrates that the optogenetic approach may prove useful for controlling epileptiform activity and opens a future perspective to develop it into a strategy to treat epilepsy.

Optogenetic techniques provide powerful tools for bidirectional control of neuronal activity and investigating alterations occurring in excitability disorders, such as epilepsy. In particular, the possibility to specifically activate by light-determined interneuron populations expressing channelrhodopsin-2 provides an unprecedented opportunity of exploring their contribution to physiological and pathological network activity. There are several subclasses of interneurons in cortical areas with different functional connectivity to the principal neurons (e.g., targeting their perisomatic or dendritic compartments). Therefore, one could optogenetically activate specific or a mixed population of interneurons and dissect their selective or concerted inhibitory action on principal cells. We chose to explore a conceptually novel strategy involving simultaneous activation of mixed populations of interneurons by optogenetics and study their impact on ongoing epileptiform activity in mouse acute hippocampal slices. Here we demonstrate that such approach results in a brief initial action potential discharge in CA3 pyramidal neurons, followed by prolonged suppression of ongoing epileptiform activity during light exposure. Such sequence of events was caused by massive light-induced release of GABA from ChR2-expressing interneurons. The inhibition of epileptiform activity was less pronounced if only parvalbumin- or somatostatin-expressing interneurons were activated by light. Our data suggest that global optogenetic activation of mixed interneuron populations is a more effective approach for development of novel therapeutic strategies for epilepsy, but the initial action potential generation in principal neurons needs to be taken in consideration.

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.

Gene therapy using recombinant adeno-associated viral vectors overexpressing neuropeptide Y in the hippocampus exerts seizure-suppressant effects in rodent epilepsy models and is currently considered for clinical application in patients with intractable mesial temporal lobe epilepsy. Seizure suppression by neuropeptide Y in the hippocampus is predominantly mediated by Y2 receptors, which, together with neuropeptide Y, are upregulated after seizures as a compensatory mechanism. To explore whether such upregulation could prevent seizures, we overexpressed Y2 receptors in the hippocampus using recombinant adeno-associated viral vectors. In two temporal lobe epilepsy models, electrical kindling and kainate-induced seizures, vector-based transduction of Y2 receptor complementary DNA in the hippocampus of adult rats exerted seizure-suppressant effects. Simultaneous overexpression of Y2 and neuropeptide Y had a more pronounced seizure-suppressant effect. These results demonstrate that overexpression of Y2 receptors (alone or in combination with neuropeptide Y) could be an alternative strategy for epilepsy treatment.