The goal of this project was to test the relationship between the angle of dividing precursor cells and daughter cell fate. Background: If you examine mitoses at the surface of the lateral ventricle, you'll notice that most divide with a vertical orientation, but that some divide with oblique or horizontal orientations (see figure at right). It is thought that the different orientations represent different types of divisions. The model, as it relates to the mammalian neocortex, predicts that a vertical cleavage plane orientation is associated with symmetric divisions, while a horizontal orientation is associated with asymmetric, neurogenic divisions. This mechanism is thought to segregate fate determinants and thus determine daughter cell fate.
This concept has been around for a very long time. Many have addressed this topic. In his 1935 paper "Mitosis in the Neural Tube" Frederick Sauer (pictured at left) wrote that the number of horizontal divisions at the ventricle was not sufficient to account for the number of cortical neurons. Yet, the hypothesis remained attractive, and was supported by work in Drosophila showing that the cleavage plane of dividing ganglion mother cells can be correlated with daughter cell fate.
But, the idea had not been tested in mammalian neocortex in experiments that recorded cleavage plane angle during division, and then tracked daughter cells over sufficient time for daughter cells to mature and express their mature phenotype. So, I labeled precursor cells with eGFP and prepared organotypic slices at early (E13-E15) or late stages (E16-E19) of rat cortical development. I recorded angle of division in the VZ and SVZ. After several days of time-lapse recording I phenotyped daughter cells by considering the morphology, behavior, and physiological recordings of the daughter cells.
The results showed major differences between mode of division in the VZ and SVZ. This movie shows a vertical RG cell division at the ventricle - at the t=2h 51m mark - producing asymmetric daughter cells. One daughter is a self renewed RG cell that inherits the pial fiber and returns to the ventricle to divide again. The second daughter is an intermediate progenitor cell that migrates away from the ventricle and divides horizontally in the SVZ to produce 2 cortical neurons.
These experiments demonstrated that vertical divisions can produce asymmetric daughter cells, and that vertical divisions were not perfectly vertical - they averaged 82.9º with the cleavage plane always tilting away from the daughter cell that inherited the RG pial fiber. The results were based on 36 RG cell divisions and 24 IP cell divisions recorded via time-lapse in vitro, and in vivo cleavage plane angle measurements of 1536 ventricular surface divisions and 879 SVZ divisions.
In sum, angle of division did not predict daughter cell fate. Instead, daughter cell fate was correlated with the stage of neurogenesis and the location of division: Divisions at the ventricle during early stages of cortical development were mostly symmetric, while during later stages divisions at the ventricle were mostly asymmetric, regardless of orientation. After the onset of neurogenesis, divisions that occurred at the ventricle were largely asymmetric and those in the SVZ were largely symmetric. This suggests that inheritance of fate determinants is not controlled by the angle of division, but by other factors I'd be happy to discuss if you give me a call!

Vertical divisions produce asymmetric daughter cells
Vertically produced, asymmetric translocating radial glial cell. This time-lapse movie is another example showing that vertical divisions can produce asymmetric daughter cells. The RG cell divided vertically at the t=24h:30m mark, yet produced morphologically asymmetric daughter cells: one daughter cell retained the pial fiber and the second daughter cell remained close to the mother RG cell over 100+ hours of time-lapse recording. After division the RG cell detached from the ventricle and translocated toward the cortical plate. The translocating RG cell remained proliferative, dividing two additional times at the t=52h time point, and the t=83h time point. After 100+ hours of time-lapse recording I transferred the organotypic slice to an Ephys rig and obtained whole-cell patch-clamp recordings from each cell. All four cells exhibited the membrane properties of astroglial cells. This result was obtained in 3 consecutive experiments. The astroglial membrane properties recorded from the translocating RG cell, and its daughters, stands in contrast to the neuronal membrane properties I readily and consistently recorded from daughter cells produced by IP cell divisions.

Vertical divisions produce translocating radial glial cells that remain proliferative
This time-lapse movie shows another vertical RG cell division yielding morphologically distinct daughter cells. The division at t=4h was vertical, and yielded one daughter cell that retained the pial fiber (red arrowhead), and a second daughter cell (red arrow) that remained close to the mother cell over 100+ hours of time-lapse recording. The RG cell loses contact with the ventricle during recording. The ventricular contacting process appears to degrade at the t=39h and t=45h time points, and was no longer visible after that point.
Vertical divisions produce translocating radial glial cells
This time-lapse movie was recorded from a radial glial cell in the E13 rat, during early stages of cortical development. Cytokinesis begins at the t=30m timepoint, and this RG cell division produces two daughter cells that have similar morphology.
In contrast to the RG cell divisions at the ventricle that were mostly vertical - 34/36 time-lapse recordings of RG cells were vertical - I found that IP cells were more likely to divide with a horizontal orientation during neurogenic stages of cortical development. The horizontal division of IP cells produced symmetrically fated daughter cells, as classified by morphology, behavior and physiological recordings.
Symmetric IP Cell Division in the SVZ
In the time-lapse recordings I observed that the RG cell pial fiber became very thin during M-phase of division. The fiber became so thin that in some cases I had to either expose the cell to higher laser power, or scan for a longer time to visualize the fiber. Of interesting, I noticed that as the fiber became thinner I could detect multiple varicosities traveling along the fiber in both directions but most often towards the RG cell soma. In many cases one or more of the varicosities entered the RG cell soma prior to division.
Pial fiber varicosities enter RG cells before division