Deep Brain Genetic Manipulation

Highly light sensitive property of photoactivatable Flp recombinase will be ideal for controlling genetic manipulation in deep mouse brain regions by illumination with a noninvasive light emitting diode, in an easy to use optogenetic module that will provide a side effect free, expandable genetic manipulation tool for neuroscience research.

Spatiotemporal control of gene expression has been acclaimed as a strategy for identifying function of genes with complex neural circuits. Complex brain function studies require sophisticated and robust technologies to enable labeling and genetic modification in animals. Approaches for controlling activity of protein or expression of genes in a spatiotemporal maner using light, peptides, small molecules, and hormones have been developed for manipulation of intact circuits or functions.

Recombination employing, chemically inducible system are most often used in vivo gene modification systems. Methods such as selective or conditional Cre-activation systems within subsets of green fluorescent protein expressing cells or dual promoter driven intersectional populations of cells are less often used. However such approaches are limited by the time and effort required to establish knock in mouse lines, and by constraints on spatiotemporal control which rely on limited sets of available genetic promoters and transgenic animal resources.

Optogenetic approaches allow activity of genetically defined neurons in animal brains to be controlled by high spatiotemporal resolution, however an module capable of revealing the functions of specific target genes in mice brains has remained a challenge.

The team’s findings featured photoactivatable Flp recombinase via investigating split sites of Flp recombinase not previously identified capable of reconstitution to be active; and validated highly light sensitive, efficient performance of photoactivatable Flp recombinase precise light targeting by demonstrating transgene expression within anatomically confined mouse brain regions.

A new method which has remained out of reach and no such light inducible Flp system developed for genetic labeling has been presented in this concept for genetically identifying cell subpopulations defined by spatial and temporal characteristics of light delivery, which has remained out of reach and no such light inducible Flp system developed.

Activation via noninvasive light illumination deep within the brain is beneficial as it avoids chemical or optic fiber implantation mediated side effects; and provides expandable utilities for transgene expression system upon Flp recombinase activity in vivo by designing viral vector for minimal leaky expression influenced by viral nascent promoters. Utility was demonstrated as a noninvasive in vivo optogenetic manipulation tool which was even applicable for deep brain structures as it reaches the hippocampus or medial septum using external LED illumination.

Professor Heo explains the team set out to develop a photoactivatable Flp recombinase to take advantage of high spatiotemporal control via light stimulation; and that there will be advantage to controlling specific gene expression desired by using LEDs with a little physical and chemical stimulation to affect the physiological phenomenon living within animals.