We have recently shown that there are substantial changes in airway innervation in patients with severe asthma. In both these patients and in several mouse models of asthma, inflammatory changes lead to significant changes in nerve structure and phenotype. We hypothesize that structural and phenotypic changes in airway innervation are central to asthma pathophysiology. In order to characterize these changes functionally, we propose to develop an optogenetic approach to stimulating or silencing two populations of airway neurons: 1) cholinergic efferents, and 2) substance P containing afferents. We propose two specific aims:
SPECIFIC AIM #1 : We will apply optogenetic methods to stimulate or silence specific populations of airway neurons. We will express the light sensitive cation channel channelrhodopsin (ChR2) in two populations of neurons, by driving expression of the ChR2 with the promoters for choline acetyltransferase (for cholinergic parasympathetic neurons), and for substance P (Tac1, the promoter for the preprotachykinin gene, for substance P containing sensory neurons). This will allow us to activate these neurons selectively, and study the effects on airway function. Conversely, we will express halorhodopsin (a light-sensitive anion channel that hyperpolarized cells) in the same populations of neurons, allowing us to silence these neurons SPECIFIC AIM #2: We will apply these optogenetic methods to defining functional changes in the sensory and parasympathetic control of airway smooth muscle in a mouse model of eosinophilic asthma, using transgenic mice expressing IL5 in airway epithelial cells (causing intense airway eosinophilia and hyperinnervation). These mice display markedly increased reflex bronchoconstriction in response to inhaled serotonin. As both afferent (sensory) and efferent (parasympathetic) nerves are abnormal in asthma and reflex bronchoconstriction is increased in these animals, we will test the effects of selectively stimulating or silencing cholinergic neurons, and of stimulating or silencing substance P containing sensory neurons. At the completion of this project, we will have established a new method for investigating the neural control of the airways, and for defining the role of subpopulations of neurons as well as their changes in models of airway disease. Going forward, we will apply this method, as well as optogenetic stimulation or silencing of other populations of neurons, to the study of other models of asthma, including viral infections, antigen challenge, and ozone inhalation.
In this project, we will develop a new technique to use optogenetics to stimulate or silence specific populations of airway neurons. We will also apply this method to defining the abnormalities in neural control in a mouse model of asthma in which the innervation of the lungs is markedly increased.
