Gossypium barbadense L. (Pima) and Gossypium hirsutum L. (upland) cottons are the two major fibre producing species grown in the south-western United States, where lint yields are adversely affected by high temperatures. In these environments, heat-adapted upland cultivars show higher yields and heat resistance than advanced Pima cultivars. Recent studies with an historical series of commercial Pima cultivars have shown that increases in lint yield and heat resistance are tightly coupled to increases in stomatal conductance and photosynthetic rate, and to decreases in leaf area. In the present study, Pima S-6 and Pima S-7 (advanced Pima cultivars) and Deltapine 90 (advanced upland cultivar) were compared under field and laboratory conditions to determine whether the physiological and morphological gradients found in the Pima historical series extrapolate to upland cotton. In the field, Deltapine 90 showed 25–35% higher stomatal conductance, 35–50% higher photosynthetic rate and 45% smaller leaf area than Pima S-6. The higher photosynthetic rate and stomatal conductance of Deltapine 90 leaves were partially related to their sun-tracking ability. In gas exchange experiments that prevented sun-tracking, the two cultivars had comparable photosynthetic rate as a function of incident radiation, while stomatal conductance was higher in upland cotton. In the 25–35°C range, photosynthetic rate as a function of temperature remained nearly constant in both cultivars, and was higher in upland cotton at all temperatures. Stomatal conductance showed a strong temperature-dependence, and conductance value and the slope of the stomatal response to temperature were higher in Deltapine 90. In progeny from a cross between Deltapine 90 and Pima S-7, the segregation of stomatal conductance in F1 and F2 populations showed a clear genetic component. These results indicate that the differences in photosynthetic rate, stomatal conductance and leaf area associated with increases in lint yield and heat resistance in the Pima historical series are also evident in a comparison between advanced cultivars of upland and Pima cotton. Upland cotton could be used as a source of genetic variation for high stomatal conductance in Pima breeding programs.