Dynamic behavior of individual cells in developing organotypic brain slices revealed by the photoconvertable protein Kaede.
In recent years, advances in optical imaging methods have facilitated the visualization of events in the developing cortex. In particular, the introduction of DNA encoding fluorescent protein into cells of the embryonic brain allows the visualization of progenitor cells; slice preparations of the cortex then allow the monitoring of the behavior of transfected cells in the context of the living cerebral wall by time-lapse microscopy. Such approaches have provided substantial information about the patterns of neuronal migration. However, as these techniques label large numbers of cells in the ventricular zone (VZ), it is difficult to follow individual cell shape changes or cell behaviors within the VZ, where neuron production and initial migration take place. Here, we report a unique method using the photoconvertable fluorescent protein Kaede, which emits green fluorescence and shifts to emitting red fluorescence upon radiation with UV. Using this method, we were able to follow the behavior of a particular pair of daughter cells among neighboring Kaede-positive cells in the SVZ of mouse brain slices. The spindle shape progenitor divided into two multipolar-shaped daughter cells. The cell-cell borders of daughter cells were clearly visualized, and easily describe the position and distance between two or more cells. The photoconvertable property of Kaede offers a powerful cell marking tool to identify the precise morphology and migratory behaviors of individual cells within living cortical slices.
Mutoh
T
T
Laboratory for Cell Culture Development, Advanced Technology Development Center, Brain Science Institute, Riken Saitama, Japan. tmuto@brain.riken.jp
Miyata
T
T
Kashiwagi
S
S
Miyawaki
A
A
Ogawa
M
M
D000818:Animals; D002452:Cell Count / Q000379:methods; D002454:Cell Differentiation / Q000502:physiology; D002465:Cell Movement / Q000502:physiology; D002540:Cerebral Cortex / Q000166:cytology* / Q000196:embryology; D005260:Female; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D051379:Mice; D008813:Mice, Inbred ICR; D009474:Neurons / Q000166:cytology*; D009924:Organ Culture Techniques; D038081:Organogenesis; D011247:Pregnancy; D013234:Stem Cells / Q000166:cytology*; D013997:Time Factors; D014162:Transfection / Q000379:methods; D014466:Ultraviolet Rays
C513582:Kaede protein, Trachyphyllia geoffroyi; D008164:Luminescent Proteins