• The evolution of C ₄ photosynthesis in plants has allowed the maintenance of high CO ₂ assimilation rates despite lower stomatal conductances. This underpins the greater water-use efficiency in C ₄ species and their tendency to occupy drier, more seasonal environments than their C ₃ relatives. • The basis of interspecific variation in maximum stomatal conductance to water (g _max), as defined by stomatal density and size, was investigated in a common-environment screening experiment. Stomatal traits were measured in 28 species from seven grass lineages, and comparative methods were used to test for predicted effects of C ₃ and C ₄ photosynthesis, annual precipitation and habitat wetness on g _max. • Novel results were as follows: significant phylogenetic patterns exist in g _max and its determinants, stomatal size and stomatal density; C ₄ species consistently have lower g _max than their C ₃ relatives, associated with a shift towards smaller stomata at a given density. A direct relationship between g _max and precipitation was not supported. However, we confirmed associations between C ₄ photosynthesis and lower precipitation, and showed steeper stomatal size-density relationships and higher g _max in wetter habitats. • The observed relationships between stomatal patterning, photosynthetic pathway and habitat provide a clear example of the interplay between anatomical traits, physiological innovation and ecological adaptation in plants.