Ricardo C. Araneda, PhD - US grants

DESCRIPTION (provided by applicant): The accessory olfactory bulb (AOB) is the first relay of chemosensory information in the vomeronasal system, yet very few studies have addressed cell physiology and synaptic connectivity of cells in the AOB. Principal neurons in the AOB, unlike cells in the main olfactory bulb (MOB), send primary dendrites to more than one glomerulus, and peripheral sensory neurons expressing a single vomeronasal receptor (VR) in the vomeronasal organ (VNO) project to multiple glomeruli. There is also a segregation of projections from sensory neurons in the VNO into the anterior and posterior aspects of the AOB, and this segregation may also result in differential patterns of activity in these regions. The overall aim of this application is to study the physiological properties of principal neurons in the AOB. We will relate physiological characteristics to anatomical parameters of the cells, particularly glomerular connectivity, and their distribution in the anterior-posterior axis of the AOB. In Specific Aim 1, we will determine if mitral cells are connected only to glomeruli that receive inputs from the same receptor, or if the same mitral cell maintains connection with a mixed population of receptors. We will use two lines of transgenic mice, the V1r and V2r-GFP mice. Recording from mitral cells connected to GFP-labeled glomeruli and including a fluorescent dye in the intracellular recording solution will allow identification of the cell's glomerular connection(s). We will compare the number of GFP-labeled glomeruli with those labeled by the intracellular dye. In Specific Aim 2, we will study the pharmacological and physiological properties of dendrodendritic inhibition in mitral cells. Because VNO neurons expressing the V1r and V2r project to the anterior and posterior AOB respectively, the use of these transgenic mice will allow us to compare these subdivisions of the AOB. It is anticipated that these studies will have an impact on our understanding of the sensory coding of pheromonal information and other olfactory information by the accessory olfactory system.

DESCRIPTION (provided by applicant): The accessory olfactory bulb (AOB) is the first relay of chemosensory information in the vomeronasal system, yet very few studies have addressed cell physiology and synaptic connectivity of cells in the AOB. Principal neurons in the AOB, unlike cells in the main olfactory bulb (MOB), send primary dendrites to more than one glomerulus, and peripheral sensory neurons expressing a single vomeronasal receptor (VR) in the vomeronasal organ (VNO) project to multiple glomeruli. There is also a segregation of projections from sensory neurons in the VNO into the anterior and posterior aspects of the AOB, and this segregation may also result in differential patterns of activity in these regions. The overall aim of this application is to study the physiological properties of principal neurons in the AOB. We will relate physiological characteristics to anatomical parameters of the cells, particularly glomerular connectivity, and their distribution in the anterior-posterior axis of the AOB. In Specific Aim 1, we will determine if mitral cells are connected only to glomeruli that receive inputs from the same receptor, or if the same mitral cell maintains connection with a mixed population of receptors. We will use two lines of transgenic mice, the V1r and V2r-GFP mice. Recording from mitral cells connected to GFP-labeled glomeruli and including a fluorescent dye in the intracellular recording solution will allow identification of the cell's glomerular connection(s). We will compare the number of GFP-labeled glomeruli with those labeled by the intracellular dye. In Specific Aim 2, we will study the pharmacological and physiological properties of dendrodendritic inhibition in mitral cells. Because VNO neurons expressing the V1r and V2r project to the anterior and posterior AOB respectively, the use of these transgenic mice will allow us to compare these subdivisions of the AOB. It is anticipated that these studies will have an impact on our understanding of the sensory coding of pheromonal information and other olfactory information by the accessory olfactory system.

DESCRIPTION (provided by applicant): Remarkably, granule cells (GCs) of the olfactory bulb are continuously born throughout life, providing an excellent neuronal model for the study of both developmental and adult neurogenesis of inhibitory neurons in the brain. As with GCs born during development, cells born in the adult olfactory bulb must establish functional connections with existing neuronal components, including the fibers of the noradrenergic system. This system plays an essential role in sensory information processing in the bulb. The aims outlined in this application seek to understand the physiological role and cellular mechanisms by which the noradrenergic system modulates GC function in the olfactory bulb. In the first two aims, we propose to investigate the cellular mechanisms and physiological role of the excitatory action of noradrenaline on GCs in the main and accessory olfactory bulb. In the last aim, we propose to determine the role of the noradrenergic system on adult neurogenesis of GCs in the context of olfactory mediated behaviors. PUBLIC HEALTH RELEVANCE: The discovery that new neurons are born in the adult brain, also known as adult neurogenesis, has opened a promising area of research because of its therapeutic potential for rebuilding new circuits in neurodegenerative diseases as well as in aging. Among the cells that exhibit adult neurogenesis are the inhibitory granule cells of the olfactory bulb. These neurons are continuously born throughout life, providing an excellent neuronal model for the study of both developmental and adult neurogenesis. Throughout life, newly born granule cells perform the remarkable task of survival and proper integration into their new environment. As with granule cells born during development, the cells born in the adult must establish functional connections with existing neuronal components of the bulb. Among these components are the fibers of the noradrenergic system, an important neuromodulatory system in the bulb. Thus, the study of granule cell function in relation to the noradrenergic system also has medical implications. This system has extensive projections through the entire brain and plays an important role in neuronal excitability in physiological states such as attention, anxiety and emotions.

? DESCRIPTION (provided by applicant): Early detection of Alzheimer's disease (AD) is a critical factor in combating this devastating disease. The discovery that pathological changes underlying brain degeneration and cognitive loss begin at least 10-20 years before dementia onset has provide an important target for the improvement of disease diagnosis and therapy. The development of biomarkers to detect neuropathology associated with early-stage AD will allow the implementation of preventive treatments much earlier in the pathological process, maximizing treatment efficacy. Notably, olfactory dysfunction precedes symptoms of dementia and memory loss, which has made olfactory tests a commonly used tool in early AD detection. As olfactory decline also occurs in normal aging, an accurate diagnosis of AD relies in the proper distinction between these processes. Interestingly, it is well established that the entorhinal-hippocampal circuit, a key pathway for learning and memory, exhibits early neuropathology in AD, and that olfactory information is relayed to the hippocampus via the entorhinal cortex. Unfortunately, the mechanisms underlying olfactory deficits in AD and natural aging remain largely unknown. Here we propose to unravel the mechanisms by which olfactory information is conveyed to the entorhinal cortex and the adaptations that precede olfactory dysfunction in naturally aging mice and in a transgenic mouse model of AD. To achieve this goal, our team of investigators will use a multidisciplinary approach that combines viral-assisted retrograde labeling, electrophysiology, optogenetics, and behavioral assessment.

Chemical detection using biological cells that live on the surface of an integrated circuit chip is a promising approach to identifying odors, disease, and pathogens. For example, the principal investigators have previously demonstrated that a chip can be used to detect electrical signals generated by odor-sensing cells taken from the nose when those cells are exposed to particular odors. Despite decades of research into olfactory sensors, we are still using animals, primarily dogs for odor detection; a cell-phone like device for odor detection would have widespread applications throughout society. This project aims to develop technology that overcomes the main practical challenge for devices based on cell-based sensing: the need to supply cells to these devices. Storage or supply of cells prior to device use has been challenging: shipping cells to the device location so that they can be loaded into it just prior to use is infeasible in many scenarios, as is keeping the cells alive for long periods of time during which power may be unavailable and environmental temperatures are varying. A dryable animal cell line that can be cultured in the lab and methods for genetically engineering these cells has recently been developed by others. The proposed research will lay the necessary groundwork for using such cells on chip for cell-based sensing, so that the cells can be stored in stasis on chip and later re-animated by the addition of water. The long term goal of the research is to develop a bionose-on-a-chip that can be used in a hand-held device for odor identification, replacing dogs in applications such as security, explosives detection, and search and rescue, and opening new possibilities for monitoring food safety and origin, controlling industrial processes, and even diagnosing disease.

The proposed work comprises three efforts to demonstrate the feasibility of using dryable cell lines on chip. First, a gel system will be established that is compatible with cell patterning and subsequent dessication. In the bionose application, cells expressing distinct olfactory receptors will be patterned onto particular sensing electrodes within a hydrogel host matrix, then dried in situ. The host hydrogel should be compatible with culture of cells, maintain mechanical integrity during and after drying, and support rapid exchange of ions and small molecules. A range of gel systems that can be patterned using a bioplotter will be studied. Second, the integrated circuit sensing chip will be packaged and integrated with microfluidics. This will facilitate the dehydration-rehydration process and promote testing of the system with odorants. Both conventional micro-molding and 3-dimensional printing will be evaluated. Third, the design of the on-chip micro-electrodes that monitor the electrical activity of the odorant cells will be optimized through modeling and simulation to maximize detection of the signals and to minimize anticipated crosstalk between different cell populations on the same chip. These developments will lay the groundwork not only for the bionose-on-a-chip application, but also other cell-based bioelectronic sensing devices. The proposed work is challenging because microfluidics and other organ on a chip approaches must be coupled with state of the art circuit technologies and cutting edge biology.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.