CHARACTERIZING MITOCHONDRIAL CA+2 HOMEOSTASIS AND THE EFFECTS OF OXIDATIVE DAMAGE

TBI is the result of a blow to the head, leading to neuronal death, short- and long-term brain impairment, and even the death of the patient. In Western countries, such as the United States, millions of cases of TBI happen every year. Troops fighting in Operation Iraqi Freedom and Operation Enduring Freedom in Afghanistan have been particularly prone to TBI. These TBIs can occur from either blast waves or direct blows from objects, especially given our advances in protective body army, which leave the head more exposed. Although TBI is devastating for the individual, there is a lack of effective therapies, either neuroprotective or neurorestorative.After the initial diffuse or direct blow to the head, there is a cascade of secondary events that result in increased neuronal cell death. These secondary events, which can take place on the order of minutes to days, are potential therapeutic targets. The immediate secondary events are often due to membrane shearing caused by the impact, including disruption of neuronal membranes. Once this happens, neural cells become leaky, altering their cellular homeostasis. One very important downstream event is mitochondrial dysfunction, which primarily results from disrupted calcium regulation and generation of excess reactive oxygen species (ROS). These types of mitochondrial misregulation can lead to cell death.Although there is ample evidence of mitochondrial dysfunction post-TBI, there are few therapies that ameliorate the effects of TBI on mitochondria. It is imperative we identify additional drugs and treatments. Research using TBI rodent models have examined the global effects of injury on mitochondrial function, using biochemistry to look at changes in mitochondrial function by typically grinding up various regions of the central nervous system (for examples see. While these studies have underscored the immediate role mitochondrial dysfunction plays after TBI, there is a gap in our knowledge at the cellular and sub-cellular level concerning how mitochondria in the effected tissue react to injury. Mitochondria are a heterogeneous population with respect to their biochemistry, and are also very dynamic, changing shape, numbers and location frequently. A better understanding of how mitochondria react to TBI at the organelle and sub-cellular level will allow us not only to better understand the immediate changes in mitochondria, but also allow us to potentially develop better strategies for coping with the enormous loss of mitochondrial function post-TBI.The purpose of this proposal is to develop a new model for TBI using the Drosophila melanogaster larval brain. Using a less complex model system allows us to better image mitochondria, and has a superior genetic advantage to study the genes and molecular mechanisms that control mitochondrial function in the brain. In addition, much is known about the cell types in the larval brain, as well as the neural stem cells present. We are using live-imaging to detect changes in mitochondrial calcium levels and ROS production pre- and post-TBI in order to better understand this process. Our goal is to create a robust Drosophila system to study mitochondrial changes in response to TBI. The advantage this model will have over existing rodent TBI models is we can visualize mitochondria at single organelle resolution and use live-imaging to identify the sequence of changes taking place after brain injury. This information will direct future development of therapies targeting mitochondrial function.