Neonatal Pial Surface Electroporation

Abstract

Over the past several years the pial surface has been identified as a germinal niche of importance during embryonic, perinatal and adult neuroand gliogenesis, including after injury. However, methods for genetically interrogating these progenitor populations and tracking their lineages had been limited owing to a lack of specificity or time consuming production of viruses. Thus, progress in this region has been relatively slow with only a handful of investigations of this location. Electroporation has been used for over a decade to study neural stem cell properties in the embryo, and more recently in the postnatal brain. Here we describe an efficient, rapid, and simple technique for the genetic manipulation of pial surface progenitors based on an adapted electroporation approach. Pial surface electroporation allows for facile genetic labeling and manipulation of these progenitors, thus representing a time-saving and economical approach for studying these cells. Video LinkThe video component of this article can be found at

Sparse postnatal labeling and quantification of superficial cortical cell synapses in the mouse neocortex

“…The electroporation protocol is adapted from previously published protocols. 2 , 3 , 4 , 5 Load the micro-syringe with plasmid DNA solution. Note: To easily load the micro-syringe, put a drop of plasmid DNA solution on a piece of parafilm and tap it with the back of the micro-syringe; DNA solution will slowly enter by capillarity.…”

“…To quantify the density of SST + presynaptic boutons, we first perform pial-surface electroporation in the somatosensory cortex of neonatal mice. 2 , 3 , 4 , 5 This strategy allows for mosaic labeling of a handful of superficial cortical postmitotic SST + cells. Specifically, we electroporate SST Cre/+ pups at postnatal day (P) 1–2 with a CRE-dependent plasmid carrying a tdTomato and the mouse presynaptic protein synaptophysin (mSyp) fused with an enhanced green fluorescent protein (EGFP): pCALNL-tdTomato-2A-mSyp::EGFP (Addgene #191213).…”

Tetracycline‐Inducible and Reversible Stable Gene Expression in Human iPSC‐Derived Neural Progenitors and in the Postnatal Mouse Brain

“…Other methods of gene delivery to other regions of the brain, such as pial surface electroporation or conventional cloning the response and transactivator plasmids into viral vectors may also lend well to the use of the system (Braun et al, 2013; Levy et al, 2014). Specifically, therapeutic approaches to the CNS may favor viral-mediated delivery approaches (Chtarto et al, 2016).…”

“…Post‐natal delivery methods of the pB‐tet‐GOI system are not limited to electroporation of the lateral ventricle. Alternative methods of gene delivery to other regions of the brain, such as pial surface electroporation or cloning the response and transactivator plasmids into viral vectors may also lend well to the use of the system (Braun, Machado, & Jessberger, ; Levy, Molina, Danielpour, & Breunig, ). Specifically, therapeutic approaches to the CNS may favor viral‐mediated delivery approaches (Chtarto et al., ).…”

Lost highway(s): barriers to postnatal cortical neurogenesis and implications for brain repair

“…In humans, evidence exists for the local generation of interneurons in the cortical ventricular and subventricular zone (VZ and SVZ), respectively during embryogenesis, but this remains contentious (Letinic et al, 2002 ; Hansen et al, 2013 ; Radonjić et al, 2014 ). Moreover, there is evidence that interneuron progenitors can proliferate extensively during their migration, including in the cortex and, in addition, the tail end of this migrating progenitor population can be observed postnatally (Costa et al, 2007 ; Inta et al, 2008 ; Breunig et al, 2012 ; Levy et al, 2014 ).…”