Directing astroglia from the cerebral cortex into subtype specific functional neurons

Astroglia from the postnatal cerebral cortex can be reprogrammed in
vitro to generate neurons following forced expression of neurogenic
transcription factors, thus opening new avenues towards a potential
use of endogenous astroglia for brain repair. However, in previous
attempts astroglia-derived neurons failed to establish functional
synapses, a severe limitation towards functional neurogenesis. It
remained therefore also unknown whether neurons derived from
reprogrammed astroglia could be directed towards distinct neuronal
subtype identities by selective expression of distinct neurogenic
fate determinants. Here we show that strong and persistent
expression of neurogenic fate determinants driven by
silencing-resistant retroviral vectors instructs astroglia from the
postnatal cortex in vitro to mature into fully functional,
synapse-forming neurons. Importantly, the neurotransmitter fate
choice of astroglia-derived neurons can be controlled by selective
expression of distinct neurogenic transcription factors: forced
expression of the dorsal telencephalic fate determinant
neurogenin-2 (Neurog2) directs cortical astroglia to generate
synapse-forming glutamatergic neurons; in contrast, the ventral
telencephalic fate determinant Dlx2 induces a GABAergic identity,
although the overall efficiency of Dlx2-mediated neuronal
reprogramming is much lower compared to Neurog2, suggesting that
cortical astroglia possess a higher competence to respond to the
dorsal telencephalic fate determinant. Interestingly, however,
reprogramming of astroglia towards the generation of GABAergic
neurons was greatly facilitated when the astroglial cells were
first expanded as neurosphere cells prior to transduction with
Dlx2. Importantly, this approach of expansion under neurosphere
conditions and subsequent reprogramming with distinct neurogenic
transcription factors can also be extended to reactive astroglia
isolated from the adult injured cerebral cortex, allowing for the
selective generation of glutamatergic or GABAergic neurons. These
data provide evidence that cortical astroglia can undergo a
conversion across cell lineages by forced expression of a single
neurogenic transcription factor, stably generating fully
differentiated neurons. Moreover, neuronal reprogramming of
astroglia is not restricted to postnatal stages but can also be
achieved from terminally differentiated astroglia of the adult
cerebral cortex following injury-induced reactivation.