The first few weeks in the lab I find are the most difficult. You’re in a new place, and you're doing complicated science that you likely have no experience with. To make it even more intimidating, most members in the lab are skilled, intelligent, and extremely knowledgeable of not only the lab’s research but a plethora of subjects. In my lab, we study mitosis and differentiation of neural stem cells. Our research roughly falls into the fields of molecular genetics, microbiology, developmental and stem cell biology, and of course, neuroscience. In other words, my lab is extremely interdisciplinary.
But as I spend more and more time here I am familiarizing myself with the techniques, tools, and basics of working in a neural stem cell lab. If I wanted to sum up my research in one statement I would tell you that I am doing an anatomical study of the basal process of neural stem cells during mammalian corticogenesis. But to someone who has no background knowledge of neuroscience and molecular biology this description has very little meaning. So I’m going to start with corticogenesis.
Corticogenesis is the formation of the cerebral cortex, or the outer lining of the cerebrum. The cerebral cortex, which is also commonly referred to as the neocortex in mammals, is made of six layers which vary in neuron concentration and function. The development of this complex structure, also known as arealization, is consequently a highly specific process whose formation is largely reliant on spatio-temporal timing.
During corticogensis, neural stem cells (NSCs) also known as radial glial cells (RGCs) go through a process known as neurogenesis in which mature “adult” neurons are generated. The following figure displays a simplified version of this process:
However, as shown in the figure, this pathway is not always direct. NSCs can directly differentiation into mature neurons, or they can first differentiate into basal progenitors (BPs), also known as intermediate progenitor cells (IPCs), and then later, following another round of mitosis, differentiate into mature neurons. If NSCs do not differentiate following mitosis, this is known as proliferation. Both daughter cells will be NSCs.
The following diagram displays the direct and indirect forms of neurogenesis. It is important to note that when NSCs differentiate following mitosis, one of the daughter cells remains a NSC while the other may be either a basal progenitor or a neuron. Never do both daughter cells differentiate, except when an IPC undergoes mitosis to form two mature neurons.
My research focuses on understanding neurogenesis and the several factors that contribute to neuron differentiation. One of these factors, regulation of the cell cycle, specifically mitosis, is critical for cortical development. After all, it is the process of mitosis which produces mature neurons. Understanding the mechanisms underlying the division of these stem cells, or radial glial cells is crucial. These NSCs are long neuroepithelial cells which stretch all the way from the ventricle zone (VZ) that borders the ventricles of mammalian brains, to the pial surface of the brain. To reach such great lengths, these cells have a long epithelial structure known as a basal process. At the end of the basal process are structures known as “basal endfeet.”
It has been found that cell polarity places a crucial role in neurogenesis. There is strong evidence that supports that daughter cell inheritance of the basal or apical process following mitosis plays a crucial role in the determining of cell fate. This summer I am trying to understand the importance of the basal process in neurogenesis. Using techniques such as immunohistochemistry, live imaging, and in utero electroporation (thanks to the assistance of my secondary mentor Louis-Jan!) on brain slices I am trying to identify some of the factors that influence cell fate and the mechanism by which they accomplish this.
We believe that RNAs localized at the basal process may play a critical role in neurogenesis. Using immunoflorescent antibodies we can label the nuclei, cell body, basal processes, and specific proteins within cells. Below is an example of some of the immunofluorescence we do here in the lab.
mCherry is a fluorophore that we use to label the cell body and basal process. HOECHST labels the nuclei, and FMRP targets mRNAs. FMR1 is the human gene that codes for the protein FMRP, which is critical for cortical development. Mutations in the FMR1 gene can lead to a variety of mental and physical disorders such as mental retardation, autism, fragile x syndrome, and parkinsons. It is just one example of the many proteins essential for cortical development. Hopefully I can contribute to our knowledge of neurogenesis so that we may better understand the mechanisms of brain development.
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