With $30,000 in funding from FRAXA Research Foundation in 1999, Dr. Karel Svoboda and his team at the Cold Spring Harbor Laboratory explored the changing nature of dendritic spine motility by imaging neocortical circuits in Fragile X mice.
Karel Svoboda, PhD
Principal Investigator
Cold Spring Harbor Laboratory
FRAXA Research Grant
$30,000 in 1999
by Karel Svoboda, 6/1/1999
One of the most interesting features of the Fragile X syndrome is the apparent involvement of dendritic spines. These tiny appendages stud the surface of most neurons in the cerebral cortex, and are the sites where these neurons receive most of their inputs. As a result, they play a pivotal role in the communication between neurons. It is perhaps not surprising, therefore, that in several forms of mental retardation, these structures are somehow altered.
In Fragile X syndrome, they are unusually abundant and tend to have relatively immature forms, similar to what may be seen in the brains of infants. We do not know, however, whether these structural changes in dendritic spines are directly responsible for symptoms of Fragile X. This is an especially difficult question to address since dendritic spines can change in size and shape over the course of minutes. Since this motility might itself be a structural manifestation of the biochemical changes that take place with thought processes such as learning and memory, it is important to know how it changes during normal brain development and how it is affected in disorders such as Fragile X syndrome.
The studies we propose are designed to uncover the changing nature of dendritic spine motility as an animal grows from infancy to the equivalent of childhood, and then to discover how this motility during this crucial period is affected in the knockout mouse model of Fragile X syndrome. Finally, if a disorder is detected, we propose to reintroduce the missing gene directly into individual neurons and determine whether this single manipulation is sufficient to correct the earlier-described disorder.
In order to carry out these studies, we will label neurons with a brightly fluorescent protein by infecting them with a harmless virus carrying the gene for the marker. This marker, enhanced green fluorescent protein (EGFP), fills the entire neuron, even the spines, with a bright green label. Using a special custom-designed microscope, neurons filled with this label can be visualized at very high magnification while still in the intact, living animal. The use of this technology (two-photon laser scanning microscopy) offers numerous advantages, especially very minimal damage to the animal and the ability to visualize neurons well below the brain surface. Thus neurons involved in a given pathway (in our case, in the part of the sensory system responsible for sensation using the whiskers) can be observed repeatedly over extended periods of time in normal and Fragile X animals at different ages. Finally, the same virus used to introduce the fluorescent marker can be used to introduce, at the same time, the FMRP that the animals lack, rendering bright green only those neurons whose genetic defect has been “corrected.” The effects of reintroduction of the gene can then directly be studied with the same imaging system described above.
We hope that this approach to studying Fragile X syndrome will contribute significantly to our understanding of the way the normal brain develops, as well as to how a single genetic defect can so profoundly affect cognitive function.
