Optogenetic inhibition of chemically induced hypersynchronized bursting in mice.

Synchronized activity is common during various physiological operations but can culminate in seizures and consequently in epilepsy in pathological hyperexcitable conditions in the brain. Many types of seizures are not possible to control and impose significant disability for patients with epilepsy. Such intractable epilepsy cases are often associated with degeneration of inhibitory interneurons in the cortical areas resulting in impaired inhibitory drive onto the principal neurons. Recently emerging optogenetic technique has been proposed as an alternative approach to control such seizures but whether it may be effective in situations where inhibitory processes in the brain are compromised has not been addressed. Here we used pharmacological and optogenetic techniques to block inhibitory neurotransmission and induce epileptiform activity in vitro and in vivo. We demonstrate that NpHR-based optogenetic hyperpolarization and thereby inactivation of a principal neuronal population in the hippocampus is effectively attenuating seizure activity caused by disconnected network inhibition both in vitro and in vivo. Our data suggest that epileptiform activity in the hippocampus caused by impaired inhibition may be controlled by optogenetic silencing of principal neurons and potentially can be developed as an alternative treatment for epilepsy.

Neuronal activity dynamics in the dentate gyrus during early epileptogenesis.

Epileptogenesis is a complex process involving a multitude of changes at the molecular, cellular and network level. Previous studies have identified several key alterations contributing to epileptogenesis and the development of hyper-excitability in different animal models, but only a few have focused on the early stages of this process. For post status epilepticus (SE) temporal lobe epilepsy in particular, understanding network dynamics during the early phases might be crucial for developing accurate preventive treatments to block the development of chronic spontaneous seizures. In this study, we used a viral vector mediated approach to examine activity of neurons in the dentate gyrus of the hippocampus during early epileptogenesis. We find that while granule cells are active 8 h after SE and then gradually decrease their activity, Calretinin-positive mossy cells and Neuropeptide Y-positive GABAergic interneurons in the hilus show a delayed activation pattern starting at 24 and peaking at 48 h after SE. These data suggest that indirect inhibition of granule cells by mossy cells through recruitment of local GABAergic interneurons could be an important mechanisms of excitability control during early epileptogenesis, and contribute to our understanding of the complex role of these cells in normal and pathological conditions.

Transplantation of Human Stem Cell-Derived GABAergic Neurons into the Early Postnatal Mouse Hippocampus to Mitigate Neurodevelopmental Disorders.

A reduced number or dysfunction of inhibitory interneurons is a common contributor to neurodevelopmental disorders. Therefore, cell therapy using interneurons to replace or mitigate the effects of altered neuronal circuits is an attractive therapeutic avenue. To this end, more knowledge is needed about how human stem cell-derived GABAergic interneuron-like cells (hdINs) mature, integrate, and function over time in the host circuitry. Of particular importance in neurodevelopmental disorders is a better understanding of whether these processes in transplanted cells are affected by an evolving and maturing host brain. The present protocol describes a fast and highly efficient generation of hdINs from human embryonic stem cells based on the transgenic expression of the transcription factors Ascl1 and Dlx2. These neuronal precursors are transplanted unilaterally, after 7 days in vitro, to the hippocampus of neonatal 2-day-old mice. The transplanted neurons disperse in the ipsi- and contralateral hippocampus of a mouse model of cortical dysplasia-focal epilepsy syndrome and survive for up to 9 months after transplantation. This approach allows for investigating the cellular identity, integration, functionality, and therapeutic potential of transplanted interneurons over an extended time in developing healthy and diseased brains.

Dynamic interaction of local and transhemispheric networks is necessary for progressive intensification of hippocampal seizures.

The detailed mechanisms of progressive intensification of seizures often occurring in epilepsy are not well understood. Animal models of kindling, with progressive intensification of stimulation-induced seizures, have been previously used to investigate alterations in neuronal networks, but has been obscured by limited recording capabilities during electrical stimulations. Remote networks in kindling have been studied by physical deletions of the connected structures or pathways, inevitably leading to structural reorganisations and related adverse effects. We used optogenetics to circumvent the above-mentioned problems inherent to electrical kindling, and chemogenetics to temporarily inhibit rather than ablate the remote interconnected networks. Progressively intensifying afterdischarges (ADs) were induced by repetitive photoactivation of principal neurons in the hippocampus of anaesthetized transgenic mice expressing ChR2. This allowed, during the stimulation, to reveal dynamic increases in local field potentials (LFPs), which coincided with the start of AD intensification. Furthermore, chemogenetic functional inhibition of contralateral hippocampal neurons via hM4D(Gi) receptors abrogated AD progression. These findings demonstrate that, during repeated activation, local circuits undergo acute plastic changes with appearance of additional network discharges (LFPs), leading to transhemispheric recruitment of contralateral dentate gyrus, which seems to be necessary for progressive intensification of ADs.