Optogenetic Interrogation of ChR2-Expressing GABAergic Interneurons After Transplantation into the Mouse Brain

Hippocampal transplants of fetal GABAergic progenitors regulate adult neurogenesis in mice with temporal lobe epilepsy

“…Embryonic day 13.5 embryos were obtained by timed breeding of VGAT-ChR2-YFP adult male mice (stock number: 014548, The Jackson Labs) to wild-type females ( n = 6; C57Bl/6NHsd, Envigo). VGAT-ChR2-YFP + embryos were identified under fluorescent illumination and the MGEs were isolated, as described ( Arshad et al, 2021 ). Following trypsin digestion and trituration to obtain a single-cell suspension, the cells were re-suspended at a concentration of 1 × 10 5 cells/μL in transplantation media (Leibovitz’s L-15 (Gibco, 11415–064), supplemented with caspase inhibitor (Promega G7231), B27 (Gibco, 17504–044), fibroblast growth factor (Sigma, F0291), and epidermal growth factor (Invitrogen, 53,303–018) ( Henderson et al, 2014 ).…”

“…Injections of systemic pilocarpine have become a common laboratory method for models of TLE in inbred strains of mice, due to the relative ease of inducing status epilepticus (SE) using repeated subthreshold injections of the drug. This method leads to the reliable development of spontaneous, recurrent seizures in mice ( Ahmed Juvale and Che Has, 2020 ; Arshad and Naegele, 2020 ; Arshad et al, 2021 ). Pilocarpine-induced SE also results in aberrant neurogenesis ( Parent et al, 1997 ; Parent et al, 2006 ; Jessberger and Parent, 2015 ; Danzer, 2018 ; Sasaki-Takahashi et al, 2020 ) and loss of GABAergic interneurons ( Thind et al, 2010 ; Buckmaster et al, 2017 ).…”

“…Indeed, patch-clamp electrophysiological recordings in brain slices from TLE mice with transplants showed increased spontaneous inhibitory postsynaptic currents in granule cells ( Henderson et al, 2014 ). Transplants containing Channel Rhodopsin 2 (ChR2)-expressing GABAergic neurons formed extensive inhibitory synaptic contacts onto adult-born granule cells and, when stimulated with blue light, the transplanted interneurons induced strong inhibitory postsynaptic currents in adult-born granule cells in the hippocampus ( Gupta et al, 2019 ; Arshad et al, 2021 ). Despite mounting evidence that engraftment of developing GABAergic progenitors enhances inhibitory networks and suppresses seizures in TLE, the underlying mechanisms for this therapeutic effect are not well understood.…”

Hippocampal Transplants of Fetal GABAergic Progenitors Regulate Adult Neurogenesis in Mice with Temporal Lobe Epilepsy

“…Embryonic day 13.5 embryos were obtained by timed breeding of VGAT-ChR2-YFP adult male mice (stock number: 014548, The Jackson Labs) to wild-type females (C57Bl/6NHsd, Envigo). VGAT-ChR2-YFP+ embryos were removed and MGEs from each YFP+ embryo were isolated as described (Arshad et al, 2021). Following trypsin digestion and trituration to obtain a single-cell suspension, cells were re-suspended at a concentration of 1 × 10 5 cells/μl in transplantation media (Leibovitz’s L-15 (Gibco, 11415-064), supplemented with caspase inhibitor (Promega G7231), B27 (Gibco, 17504-044), fibroblast growth factor (Sigma, F0291), and epidermal growth factor (Invitrogen, 53303-018) (Henderson et al, 2014).…”

“…Indeed, patch-clamp electrophysiological recordings in brain slices from mice with TLE showed increased spontaneous inhibitory postsynaptic currents in granule cells in slices with transplants (Henderson et al, 2014). The transplanted cells form extensive synaptic contacts associated with post-synaptic gephyrin puncta in the host brains and optogenetic stimulation of the transplanted cells induces strong inhibitory postsynaptic currents (Henderson et al, 2014; Gupta et al, 2019; Arshad et al, 2021). Despite mounting evidence that enhancing inhibitory synaptic transmission in the hippocampus or reducing adult neurogenesis are effective approaches for reducing seizures in rodent models of TLE, little is known about the underlying mechanisms for these therapeutic effects.…”