Changing the wiring pattern of L4 neurons (green cells, left) by ectopic Fezf2 expression (right). Reprinted from Nature Neuroscience1.

It is a Kafkaesque era for biology: under the right 'genetic spell', cells of a certain identity can wake up one morning metamorphosed into another. But there are still unanswered questions about the boundaries of this cellular plasticity.

Can, for example, one type of neuron be engineered into another in the living brain? Can a set of genes change a cell's characteristic morphology and firing pattern, and even rewire it into a different network?

Although it is known that manipulating gene expression in early neuronal progenitor cells can modify the identity of their progeny, it is not known whether neurons can be reprogrammed in this manner after the animal's birth.

A major hurdle in investigating these questions has been the inability to rapidly and efficiently manipulate gene expression in specific postmitotic neurons. Classical electroporation works efficiently only in mitotic cells, and infection with viral vectors takes days for the transgene to be expressed. Undeterred, Denis Jabaudon, at the University of Geneva in Switzerland, and his team set out to answer these questions and to find the right tools to do so.

Jabaudon and his group wanted to change the identity of layer 4 (L4) neurons of the mouse brain, which receive input connections from the thalamus, and to find out whether doing so altered the cells' characteristic connectivity pattern.

The team first worked to optimize the protocols for electroporation of DNA plasmids into the brain of postnatal animals. But this alone didn't do the trick. “It had always puzzled me why electroporation didn't work in postmitotic neurons in an efficient and reproducible way,” says Jabaudon. “Then I stumbled across several reports that suggested that plasmids get stuck in the cytoplasm and never enter the nucleus.” So he and his team searched the literature for compounds that could help enhance the nuclear transport of the plasmids. They settled on a compound, TCHD (trans-cyclohexane-1,2-diol), which acts as a nuclear permeabilizing agent.

Injection of a volume of TCHD-solubilized DNA plasmid into the mouse brain and delivery of a specific electroporation protocol caused a reporter gene to be expressed in the targeted cortical region in just a few hours. They named the procedure 'iontoporation'.

Iontoporation worked well only during the first postnatal week, but if gene expression at later stages was desired, the group could inject tamoxifen-inducible plasmids early on and activate gene expression in adulthood.

The team used this method to ectopically express a transcription factor specific to layer 5B neurons, Fezf2, in L4 neurons 1 day after birth, and they examined the outcome a couple of weeks later. Fezf2 was an ideal candidate to act as an identity switch, as it is both necessary and sufficient to generate L5B neurons during development.

They examined the molecular signature of the targeted L4 cells, their morphology and their firing properties, and it was clear to them that the cells had become L5B neurons. Even the short- and long-range connectivity inputs to the engineered cells now resembled those of L5B neurons. Cell-type conversion, however, worked only if the ectopic gene was expressed during the animal's first 10 days of life. After that, L4 neurons were L4 neurons for good.

It will now be interesting to study whether these neurons can be switched into other neuron varieties and whether identity switching can happen in other types of neurons. To investigate these possibilities, the group has started to test the protocol's efficacy in other brain areas.