Fluorescent Labeling of Newborn Dentate Granule Cells in GAD67-GFP Transgenic Mice: A Genetic Tool for the Study of Adult Neurogenesis

Abstract

Neurogenesis in the adult hippocampus is an important form of structural plasticity in the brain. Here we report a line of BAC transgenic mice (GAD67-GFP mice) that selectively and transitorily express GFP in newborn dentate granule cells of the adult hippocampus. These GFP+ cells show a high degree of colocalization with BrdU-labeled nuclei one week after BrdU injection and express the newborn neuron marker doublecortin and PSA-NCAM. Compared to mature dentate granule cells, these newborn neurons show immature morphological features: dendritic beading, fewer dendritic branches and spines. These GFP+ newborn neurons also show immature electrophysiological properties: higher input resistance, more depolarized resting membrane potentials, small and non-typical action potentials. The bright labeling of newborn neurons with GFP makes it possible to visualize the details of dendrites, which reach the outer edge of the molecular layer, and their axon (mossy fiber) terminals, which project to the CA3 region where they form synaptic boutons. GFP expression covers the whole developmental stage of newborn neurons, beginning within the first week of cell division and disappearing as newborn neurons mature, about 4 weeks postmitotic. Thus, the GAD67-GFP transgenic mice provide a useful genetic tool for studying the development and regulation of newborn dentate granule cells.

Presynaptic But Not Postsynaptic GABA Signaling at Unitary Mossy Fiber Synapses

“…This observation is consistent with a recent report using another GAD67-GFP mouse strain (Zhao et al, 2010). Although a majority of MFTs already express calbindin and not GAD67, a significant proportion of GCs still undergo differentiation and maturation after birth in rodents (Altman and Bayer, 1990).…”

Adult neurogenesis in the mammalian hippocampus: Why the dentate gyrus?

“…Other identification strategies include transgenic mice where certain promoters direct expression of fluorescent proteins in abGCs at specific phases of their development, e.g., POMC-eGFP up to 2 -3 wk (Overstreet et al 2004) and GAD-67-GFP up to 3 -4 wk (Zhao et al 2010b), or where inducible Cre-recombinase expression in neural stem cells combined with a Flx-stop fluorescent reporter gene can be used to label abGCs born after induction (Dranovsky et al 2011).…”

Viral and Transgenic Reporters and Genetic Analysis of Adult Neurogenesis

“…However, GAD67-labeled cells also have sparsely spiny dendrites that extend through the molecular layer, and they display a range of synaptic and electrophysiological properties. GAD67-labeled immature GCs are highly colabeled with BrdU between 7 and 21 d after BrdU administration, suggesting they overlap with the early developmental stage identified in POMC-GFP transgenics but also label new GCs at a more mature stage (Zhao et al 2010). The intrinsic and synaptic properties of immature GCs in another line of GAD67-GFP mice have also been well characterized (Cabezas et al 2013).…”

“…But, recently, BAC transgenic mice expressing GFP under the control of GAD67 regulatory elements revealed robust expression in immature dentate GC (Zhao et al 2010). Similar to many other GAD67-based reporter lines, GABAergic interneurons in other brain regions express GFP, but in this line hippocampal interneurons are unlabeled.…”

“…Similar to many other GAD67-based reporter lines, GABAergic interneurons in other brain regions express GFP, but in this line hippocampal interneurons are unlabeled. Interestingly, the transient expression of GFP in immature GCs appears to result from the low level of GAD67 expressed in developing GC mossy fibers (Sandler and Smith 1991;Sloviter et al 1996;Zhao et al 2010). GAD67-labeled GCs share several features with POMClabeled GCs, including overlap with immature markers like DCX and PSA-NCAM, as well as immature morphology.…”