Nancy Rawson - US grants

The olfactory system exhibits a remarkable regenerative capacity, both in its ongoing replacement of olfactor neurons and its ability for repair in response to chemical or surgical trauma. The processes of neurogenesis and synaptogenesis are nutritionally demanding, and occur only during development in other central nervous system (CNS) tissues. While much is known about the adverse impact of inadequate nutritional status on prenatal growth and development, little is known about nutritional influences on the ongoing regenerative processes within the olfactory system. Recently, food restriction was shown to impair the regeneration of olfactory epithelium following chemically-induced degeneration (Schwob et all, 1994). The mechanisms for this type of nutritional modulation of olfactory system regeneration have not been studied directly. Two nutritionally modulated factors, insulin and insulin-like growth factor I (IGF-I) influence growth, differentiation and survival of a variety of neuronal systems. Interestingly, these neurotrophic factors are also found in dramatically higher levels in the olfactory bulb than most other brain areas. These observations suggest the integrity of the olfactory system may be more sensitive to nutritional modulation than most CNS tissues. However, little is known about the functions of insulin and IGF-I in t he olfactory system, or their synthesis or distribution within the epithelium, where the initial phases of regeneration occur. This proposal postulates that nutritional status influences the ability of the olfactory system to carry out its regenerative processes, both ongoing and in response to trauma, and that insulin and IGF-I contribute to this effect. The proposed project will use a multidisciplinary approach to examine the effect of altered nutritional status ont he structural and functional integrity of the olfactory system, both under normal condiitons and following surgical trauma to trigger rapid olfactory neuron degeneration and regeneration. This goal will be pursued through following the Specific Aims: (1) Determine whether cells within olfactory epithelium express insulin or IGF-I mRNAs or their receptor mRNAs or proteins: (2) investigate the effects of protein-restriction (PR), and streptozotocin-induced diabetes (StD) on olfactory system structure and function; (3) determine whether levels of insulin or IGF-I mRNA, or their receptor proteins or mRNAs in the olfactory bulb and epithelium change in response to olfactory nerve section; (4) Determine whether altered nutritional status (PR and StD) influences repair of the olfactory system in response to surgical trauma. The results of these studies will establish the olfactory system as a model to investigate nutritional modulation of neuronal regeneration and function and will provide new insight into the roles of insulin and IGF-I in the olfactory system. In addition, results may contribute to a better understanding of the neuropathologic effects of impaired nutritional status as seen in diabetes and malnutrition.

This proposal builds on work accomplished during the previous funding period to further our understanding of the basic mechanisms of olfactory transduction and adaptation in human olfactory receptor neurons (ORNs) and extend these studies to examine the basis for olfactory dysfunction associated with aging and neurodegenerative disease (Alzheimer's dementia). Over the past decade, we have learned a great deal about the peripheral receptor and transduction mechanisms responsible for the initial events in odor perception in a wide variety of animal species. Students we have conducted of human olfactory receptor mechanisms reveal that they exhibit features unlike those observed in other species studied in they are more selective and can exhibit an unusual type of calcium response. These funding stress the importance of research with human tissue such as that proposed, in order to fully understand human physiology. The olfactory system is unique as an outpost of the nervous system accessible to biopsy. Studies into the mechanisms underlying the decline in olfactory function associated with age and various neurodegenerative diseases such as Alzheimer's dementia (AD) are of importance both for the potential they provide to identify means of preventing or alleviating this sensory deficit as well as to reveal underlying causes of neuronal dysfunction elsewhere in other parts of the nervous system. Our preliminary studies suggest that ORNs from older subjects and AD patients exhibit functional differences from those of younger subjects and from each other in their selectivity and sensitivity to odorants and pharmacological agents. This proposal comprises electrophysiological and biophysical studies of ORNs freshly isolated from biopsies of the olfactory epithelium. These studies will: elucidate the cellular mechanisms involved in the transduction of (Aims I and II) and adaptation to (Aim III) odorant stimulation, as well as investigating changes that occur in ORN function with age and in AD (Aims III-V). Results will lead to a better understanding of the basis for age- and disease-associated olfactory impairment and provide direct information about changes that may also be occurring elsewhere in the nervous system.

A grant has been awarded to Dr. Nancy E. Rawson at the Monell Chemical Senses Center to purchase a confocal microscope with associated hardware and software. The Monell Center is a nonprofit research institute focused on acquiring a fuller understanding of the chemical senses. Recent studies have revealed that cell-cell interactions play an important role in modulating the output of a variety of sensory receptor cells. The confocal microscope permits us to directly observe intracellular processes and interactions between different cell types in preparations that more closely reflect those of the intact organism. In addition, chemosensory cells undergo regeneration throughout the life of the animal. In order to better understand this capability, studies of growing cells are carried out using methods to identify the different stages in the cells' lifespan and evaluate the effect of drugs or other treatments on the growth process.
The confocal microscope will be used in projects aimed at addressing a variety of questions ranging from understanding how individual receptor cells detect and respond to chemical stimuli; how experience influences the ways that organisms respond to chemosensory stimuli; how odor qualities are encoded in neural activity patterns in the brain; how stem cells divide and differentiate into mature sensory receptor cells; and how these sensory systems recover from damage. These projects utilize standard histology, lectin and immuno-histochemistry, in situ hybridization of fixed tissue specimens, and biophysical methods to study live tissue. These techniques provide a variety of kinds of data, ranging from protein and mRNA localization in peripheral and central components of the taste, olfactory and trigeminal systems to dynamic extra- and intracellular signalling in tissue slices, cultures and cell ensembles. The system will also be included as one educational component in our Minority Student Research Apprenticeship Training Program, which has provided summer research experiences for over 300 high school and college undergraduates since its inception.
The ability to examine cells in ensembles, such as whole taste buds, or thick tissue sections will dramatically improve our ability to understand how the sensory systems of taste, smell and chemical irritation function in the intact organism. The confocal microscope will also enhance our ability to provide state-of-the-art training experiences for students participating in our high school, college, graduate and post-doctoral training programs.

Olfactory disability represents a danger to the patient resulting from inability to detect hazards such as natural gas and spoiled food and threatens quality of life through the loss of enjoyment of foods and fragrances. Rhinosinusitis is a primary cause of olfactory loss among patients presenting to cliemosensory clinics, and is the most common chronic medical condition in the United States, affecting nearly 33 million people/year. Chronic rhinosinusitis (CR) patients with similar degrees of nasal inflammation may present with normosmia, hyposmia or anosmia, and current therapies are only partially effective. CR may impair olfaction in several ways. Constriction of the airway due to inflammation could alter or impede airflow through the nasal cavity, reducing access of volatile compounds to the sensory receptor cells. Changes in the composition and transport of the watery mucus through which the odorants must diffuse could impair diffusion to or removal from the receptor sites. Proteins secreted by the infiltrating cells of the immune system may directly injure or modulate the function of ORNs or other cells within the neuroepithelium. Fibrosis, gland hyperplasia, keratinization or edema may change the structure of the epithelium or submucosa, impeding axonal outgrowth and preventing successful regeneration of the neuroepithelium. Our limited understanding and the relative importance of these diverse mechanisms and the cellular and molecular processes involved is limiting the development of diagnostic, therapeutic and prognostic tools. This project addresses this need through studies aimed at better understanding the psychophysical (Aim 1), gross anatomical and inflammatory (Aims 1, 2 and collaboration with Project 2) and sensorineural (Aims 3, 4) effects of CR. By comparing data from CR patients with olfactory loss to normosmic CR patients before and after treatment, we will identify features that may be prognostic for olfactory loss/recovery and gain insight into the causes for olfactory loss in CR. We will also develop novel tools for studying airflow dynamics (Aim 1 and collaboration with Project 2) and the effects of CR-associated inflammatory mediators on neurogenesis (Aim 4). Results from this project will enable improved diagnosis and treatment targeting by providing a thorough characterization of the patient population and identifying specific profiles of anatomic, physiologic and sensory characteristics associated with olfactory loss and recovery prospects. Data may also suggest new targets for drug development and will establish a model in vitro system for use in identification and evaluation of novel therapeutic approaches.

Quantitative Computed Tomography and Histological Analysis of Carnivoran Turbinates

Blaire Van Valkenburgh and Nancy Rawson
University of California-Los Angeles and Monell Chemical Senses Center

The snout of a mammal houses a complex skeleton made up of three paper-thin scrolls of bone known as the turbinates. The largest and most intricate are the maxilloturbinates, which play a critical role in conditioning respired air and minimizing water loss. Above and/or behind them lie the ethmoturbinates, which function in the sense of smell. The smallest are the relatively simple nasoturbinates that lie above the maxilloturbinates and are of uncertain function. Collectively, the turbinates and their chambers make up a third to a half of the volume of the head in most mammals, and yet we know very little about their anatomy and how they vary in size and arrangement among species. Given that complex turbinates appear to be present in the earliest known mammals and are associated with two fundamental features of mammals, olfaction and endothermy (warm-bloodedness), it is remarkable that so little is known about them. This is because their hidden, internal location and delicate structure make them difficult to study. However, recent advances in high-resolution CT scanning (HRCT) opened the door to discovery of mammalian turbinate structure. Preliminary studies on dogs and cats show that turbinates can be visualized with HRCT, revealing these structures in their entirety for the first time. The proposed research will expand and improve upon this previous work, including study of a wider array of mammal species, development of sophisticated image analysis software to quantify turbinate structure, and a histological analysis of tissues covering the turbinates that will distinguish olfactory versus respiratory regions. This study brings together anatomists, histologists, and computer scientists to produce the first quantitative overview of the nose that integrates both histological and skeletal data in any group of mammals. It will establish the groundwork for much future research on turbinate function. Undoubtedly, some of the most interesting questions about turbinate function (both olfactory and respiratory) concern aspects of flow through the nose. How is air directed to or away from olfactory areas? Is it possible to direct air to maximize flow during aerobic exercise, or to minimize flow to enhance water and heat conservation? This study will document the basic anatomical framework of mammalian turbinates for the first time, and provide fundamental information necessary for understanding nasal function and disorders involving smell and respiration in mammals, including humans. Graduate and undergraduate students will be trained through this project. The CT scans will be entered into the Digimorph library, and will therefore be accessible to the public and broader scientific community. The PI has plans to develop aspects of this research for use by teachers in disadvantaged L.A. school science curricula.