Analyzing anatomy over three dimensions unpacks the differences in mesophyll diffusive area between sun and shade Vitis vinifera leaves.

Leaves grown at different light intensities exhibit considerable differences in physiology, morphology and anatomy. Because plant leaves develop over three dimensions, analyses of the leaf structure should account for differences in lengths, surfaces, as well as volumes. In this manuscript, we set out to disentangle the mesophyll surface area available for diffusion per leaf area ( ) into underlying one-, two- and three-dimensional components. This allowed us to estimate the contribution of each component to , a whole-leaf trait known to link structure and function. We introduce the novel concept of a 'stomatal vaporshed,' i.e. the intercellular airspace unit most closely connected to a single stoma, and use it to describe the stomata-to-diffusive-surface pathway. To illustrate our new theoretical framework, we grew two cultivars of L. under high and low light, imaged 3D leaf anatomy using microcomputed tomography (microCT) and measured leaf gas exchange. Leaves grown under high light were less porous and thicker. Our analysis showed that these two traits and the lower per mesophyll cell volume ( ) in sun leaves could almost completely explain the difference in . Further, the studied cultivars exhibited different responses in carbon assimilation per photosynthesizing cell volume ( ). While Cabernet Sauvignon maintained constant between sun and shade leaves, it was lower in Blaufränkisch sun leaves. This difference may be related to genotype-specific strategies in building the stomata-to-diffusive-surface pathway.