Age-related changes in airway-reactivity-to cholinergic and other agonists are well described in both humans and animals. In animal models these changes are associated with age-related pharmacological and biochemical changes in airway smooth muscle. As the mechanisms responsible for these changes, are not known it is the aim of this proposal to study specific intracellular mechanisms by which these changes occur. Our preliminary data suggests that changes are occurring in receptor coupling to G proteins. The G proteins couple the muscarinic receptor to several intracellular biochemical events. Since the G proteins play such a key regulatory role, it is our hypothesis that alterations occurring in these proteins result in the observed reductions in functional responses to muscarinic stimulation. Experiments are described in which we explore systematically the intracellular events resulting in second messenger generation, which are initiated by muscarinic receptor occupation in tissues from animals of different ages. Initially, we will characterize the muscarinic receptors in guinea pig tracheal tissues to determine which receptor subtypes are present and if the type, density or affinity of these receptors is changed with age. To evaluate the role of the G protein in age-related changes we will first identify which types are present in these tissues, then we will determine if changes are occurring in their structure, quantity or function with age. To test the hypothesis that the primary change is occurring at the level of the G proteins, it we need first to establish which second messenger enzyme systems the G proteins are coupled to. Once the sequence of steps resulting in second messenger generation is defined, we will block specific G proteins and determine second messenger production independently. Comparison of second messenger generation with and without G protein regulation, should provide information as to where in this sequence changes have occurred. These studies will clarify the precise mechanisms by which signal transduction occurs in airway smooth muscle and how these events are related to the changes seen physiologically with age. Many disease states such as chronic obstructive pulmonary disease and asthma, involve abnormalities in airway tone and reactivity. A clear understanding of the mechanism by which airway tone is regulated is a first step towards understanding and treating these diseases.
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