The study of isolated airway myocytes has provided important information relative to specific processes that regulate contraction, proliferation, and synthetic properties of airway smooth muscle (ASM). To place this information in physiological context, however, improved methods to examine airway biology in vivo are needed. Advances in genetic, biochemical, and optical methods provide unprecedented opportunities to improve our understanding of in vivo physiology and pathophysiology. This article describes 4 important methodologic advances in the study of ASM: (1) the development of transgenic mice that could be used to investigate ASM proliferation and phenotype switching during the development of hypersensitivity, and to investigate excitation-contraction coupling; (2) the use of CD38-deficient mice to confirm the role of CD38-dependent, cyclic adenosine diphosphate-ribose-mediated calcium release in airway responsiveness; (3) investigation of the role of actin filament length and p38 mitogen-activated protein kinase activity in regulating the mechanical plasticity-elasticity balance in contracted ASM; and (d) the use of bronchial biopsies to study ASM structure and phenotype in respiratory science.