Stabled horses are susceptible to an inflammatory disease of the lower airways, recurrent airway obstruction (RAO). Recent consensus defines RAO as a complex disease that arises in mature horses in response to chronic exposure to environmental particulate, bacterial, and fungal components. The inflammation of RAO results in bronchoconstriction, and small airway obstruction from an accumulation of mucus and neutrophils--leading to the clinical signs of cough, tachypnea, nostril flare, and increased abdominal effort on exhalation. The disease has been studied extensively in an attempt to characterize the pathogenesis and to validate treatment modalities. In order to more fully characterize the mechanisms involved in development of the RAO phenotype, a standardized model for disease exacerbation was needed. Such a model would permit comparisons between studies; however, no model thus far offered has fully characterized components. Further, recent work has indicated that both Clara cells and their major product, Clara cell secretory protein (CCSP), are reduced in horses with RAO. Ultrastructural studies showed CCSP within the cytoplasm and nucleus of neutrophils of RAO-affected horses, which suggested that CCSP might function as a cytokine or chemokine. The present research project had three goals. First, molds consistently present in moldy hay were identified using sensitive techniques. Second, a nebulizable particulate suspension of accurately quantified materials was created to cause reliable and repeatable exacerbation of RAG. Third, the localization pattern of recombinant equine CCSP (reCCSP) within stimulated equine neutrophils was examined by immunoblot analysis. Results of the studies were as follows. Three fungi consistently identified in moldy hay were: 'Lichtheimia corymbifera, Aspergillus fumigatus,' and 'Eurotium amstelodami'. Further, nebulization of a mixture of fungal spores, lipopolysaccharide, and silica microspheres induced changes consistent with RAG exacerbation in susceptible horses. And finally, reCCSP may have been detected in stimulated neutrophils. These findings suggest that RAG can be reproduced in susceptible horses through inhalation of a defined mixture of challenge materials, and additional studies are required to assess the interactions of reCCSP and neutrophils.