Transgenic approaches to understanding astrocyte heterogeneity

The cells in our brain are generally divided into two major categories based on their function - neurons (commonly known as nerve cells) and glial cells. Neurons are very important to our body because they process and transmit information to control our actions in daily life, while glial cells are seen as playing supporting roles to neurons. We used to think that our brain was mainly run by neurons with a little help from glia. In recent years, with rapid advances in neuroscience, this view has started to change and we now recognize that brain function is the result of concerted activities of both neurons and glia. Astrocytes are an important class of glial cells defined by their star-like shape and other features. Despite the fact that they are the most abundant cells in the brain making up half of the brain volume, our knowledge about astrocytes is still rudimentary. Astrocytes are thought to be highly diverse, and our previous work found a link between their diversity and developmental origin - that is, we might be able to predict an astrocyte's function according to where it comes from in early development. Our proposed study aims to explore further how astrocytes diversify during development and after settling in their final resting site in the mature brain, where they come under the influence of micro-environmental signals from neurons and other cells in their vicinity. Based on our data so far, we hypothesize that astrocyte diversity is determined at both developmental and micro-environmental levels. We will test this hypothesis by experiments in mice. Through genetic ("transgenic") manipulation of mice we are able to label different populations of astrocytes with green or red fluorescent proteins and also to perform "genetic surgery" to remove particular astrocyte populations of interest. We aim to produce a map of the developmental origins of astrocyte sub-populations and to relate this to the adult functions of the astrocytes in, for example, supporting communication among neurons. In addition, we plan to identify new molecular markers for the different populations of astrocytes. The reason why we need to study astrocyte development and diversity is that different subtypes of astrocytes might be functionally distinct from each other and therefore differentially involved in brain disorders such as autism. Our proposed study will produce direct information about astrocyte functional diversity and provide useful tools for future astrocyte research that can be provided to the neuroscience community at large.

Astrocytes are a major glial cell type in the central nervous system but, despite their great abundance and widespread distribution, their functional roles are still poorly understood. We recently found evidence (Tsai, Li et al. Science 2012) that the local function of astrocytes, e.g. in synaptogenesis, is influenced by their developmental site of origin in the embryonic neural tube. I now want to build on this work, focussing on the mouse forebrain, to test the idea that astrocyte functional heterogeneity is determined by developmental cues together with local environmental factors. First, I will map the origins of astrocyte sub-populations using the Cre/lox system and will conditionally ablate those populations with Diphtheria toxin A chain (DTA) in transgenic mice; this will allow me to determine whether astrocytes with different developmental origins (i.e. from different regions of the ventricular zone, VZ) are functionally interchangeable with respect to their adult function. Second, I will investigate whether there is further diversification driven by the microenvironment. I recently identified a glial-specific G-protein-coupled receptor, GPR37L1, which is expressed in a subset of mature astrocytes, unrelated to developmental origin. Preliminary evidence indicates that GPR37L1 is involved in glia-synapse interactions. I will examine the role of GPR37L1 expressing glia in synaptogenesis in GPR37L1 knockout mice and after targeted ablation of GPR37L1(+) glia using DTA. Finally, I will look for new markers for astrocyte subtypes by transcriptional profiling of sorted subsets of astrocytes on Affymetrix arrays and by RNA-seq. This project will provide new insights into astrocyte biology and will produce useful new tools for astrocyte research.

Impacts on academic community. This project will significantly advance the research into astrocyte diversity - a basic topic in brain research. We will present solid evidence to prove that astrocytes, the most abundant cells in human brain, are heterogeneous and not just the brain glue supporting neurons, but actively involved in many brain functions. Our work will contribute new knowledge and research materials to the academic community and inspire other researchers to reevaluate astrocyte' roles in physiological and pathological conditions. The proposed transgenic methods will incentivize researchers to design their own projects for cell lineage studies. In the meantime, the postdoctoral research assistant and technical assistant employed on this project will benefit from excellent training in molecular biology, histology and neurobiology.

Impacts on Business/Industry community. This study will prove that astrocytes are more complex than originally thought. Our findings will help understand the mechanisms at work in neurological disorders and have the potential to attract the interest of pharmaceutical industry in developing/testing new drugs using our models.

Impact on funding bodies and charitable organizations. Astrocytes have been reported to be involved in many neurological diseases such as Multiple Sclerosis, Autism and Alzheimer's disease. Moreover, gene mutation in astrocytes may be the cause of some diseases (such as human Rett syndrome) to which primary neuronal dysfunction is generally attributed. Therefore, our findings could open the door to the possibility of astrocyte subset based cell replacement therapy for astrocyte related diseases. Our findings will help raise funding bodies' and charitable organizations' awareness of astrocyte research and attract more funding for astrocyte study.

Impacts on the public. Understanding how our brain works is of general interest to the public. Our research will show how fascinating our brain is in a new light and hopefully our work could inspire more of the younger generation to take up neuroscience research.