Stomata and Mesophyll Characteristics of Barley Leaf as Affected by Light: Stereological Analysis

The anatomical structure of the second leaf blade of barley{Hordeum vulgare L. cv. Koral) was studied in plants exposedto a photosynthetic photon flux density (PPFD) of 200 µmolm^–2 s^–1 compared with those grown under 25µmolm^–2–11. Design-based stereological methods wereused for the estimation of various leaf anatomical characteristicssuch as mesophyll volume, proportion of intercellular spaces,number of mesophyll cells, mean mesophyll cell volume, and internalleaf surface area. The structure of the mesophyll was more affectedby different levels of PPFD than were the stomatal characteristics.Increased PPFD produced thicker leaves with a larger mesophyllvolume having a higher number of less elongated mesophyll cellsand a larger internal leaf surface area. Key words: Hordeum vulgare, light effect, mesophyll, stereology, stomata     

Recent stereological methods for measuring leaf anatomical characteristics: estimation of the number and sizes of stomata and mesophyll cells

Recent design-based stereological methods for measuring stomatalfrequency, mean epidermal cell length, number of mesophyll cellsand mean cell volume are described in an easy-to-follow manner.Two leaf types are considered: grass leaf and bifacial leaf.The presented methods are discussed and compared with alternativemethods. Key words: Disector, leaf, mesophyll, stereology, stomata     

Relationships between photosynthesis, nitrogen and leaf structure in 14 grass species and their dependence on the basis of expression

The relationships between leaf structure, nitrogen concentration and CO₂ assimilation rate ( A ) were studied for 14 grass species grown in the laboratory under non-limiting nutrient conditions. Structural features included leaf thickness and density, and the proportion of leaf volume occupied by different types of tissue (mesophyll, epidermis, vessels and sclerenchyma). Relationships were assessed for data expressed per unit leaf area and fresh mass. The latter was found to be closely related to leaf volume, which allowed us to use A per unit leaf fresh mass ( A _fm) as a surrogate of A per unit leaf volume. Assimilation rate per unit leaf area ( A _a) was positively correlated with leaf thickness and with the amount of mesophyll per unit leaf area; the relationship with leaf nitrogen content per unit area was only marginally significant. A _fm was negatively correlated with leaf thickness and positively with fresh mass-based leaf organic nitrogen concentration. A multiple regression involving these two variables explained 81% of the variance in A _fm. The value of A _fm was also significantly related to the proportion of mesophyll in the leaf volume, but surprisingly the correlation was negative. This was because thin leaves with high A _fm and nitrogen concentration had proportionally more mechanically supportive tissues than thick ones; as a consequence, they also had a lower proportion of mesophyll. These data suggest that, in addition to leaf nitrogen, leaf thickness has a strong impact on CO₂ assimilation rate for the grass species studied.     

A Method for Calculating the Volume and Surface Area in Rice Mesophyll Cells

A method was developed for determining the surface area and volume of rice mesophyll cells of elaborate configuration. The method was employed to calculate these indices in several types of rice mesophyll cells found in seventy samples (53 species) of diverse origin coming from Japan, China, Korea, India, Nepal, Australia, France, Italy, Uzbekistan, Afghanistan, and Krasnodar and Primorskii regions. The cultivars of diverse geographic origin varied in cell shape and size due to the number, size, and arrangement of chloroplasts. When the volumes and surface areas of leaf mesophyll cells were compared using the method reported herein and a simple empirical model of the cell as a single ellipsoid, the two methods produced relatively similar data for cell volume; however, the surface area calculated by the former method was about two times larger than in the latter case. The method described in this paper allows for accurate calculations of the volume and surface area of rice mesophyll cells when data are available on the cell shape and linear dimensions and the number of chloroplasts per cell.     

Temporal and Spatial Development of the Cells of the Expanding First Leaf of Arabidopsis thaliana (L.) Heynh

The three-dimensional quantitative leaf anatomy in developingyoung (9–22 d) first leaves of wild type Arabidopsis thalianacv. Landsberg erecta from mitosis through cell and leaf expansionto the cessation of lamina growth has been studied. The domainsof cell division, the relative proportion of the cell typespresent during development and the production of intercellularspace in the developing leaf have been determined by image analysisof entire leaves sectioned in three planes. Mitotic activityoccurs throughout the youngest leaves prior to unfolding andcell expansion is initiated firstly at the leaf tip with a persistentzone of mitotic cells at the leaf base resulting in a gradientof development along the leaf axis, which persists in the olderleaves. Major anatomical changes which occur during the developmentare, a rapid increase in mesophyll volume, an increase in thevein network, and expansion of the intercellular spaces. Thepattern of cell expansion results in a 10-fold variation inmesophyll cell size in mature leaves. In the youngest leavesthe plan area of mesophyll cells varies between 100 µm²and 400 µm² whereas in mature leaves mesophyll cells rangein plan area from 800 µm² to 9500 µm². The volumesof mesophyll tissue and airspace under unit leaf area increase3-fold and 35-fold, respectively, during leaf expansion. Thevolume proportions of tissue types mesophyll:airspace:epiderrnal:vascularin the mature leaf are 61:26:12:1, respectively. This studyprovides comparative information for future identification andanalysis of leaf development mutants of Arabidopsis thaliana. Key words: Arabidopsis, quantitative leaf anatomy, leaf expansion, image analysis     

Leaf structure and specific leaf mass: the alpine desert plants of the Eastern Pamirs, Tadjikistan

This study examines interrelationships between eight leaf attributes (specific leaf mass, area, dry mass, lamina thickness, mesophyll cell number per cm², mesophyll cell volume, chloroplast volume, and number of chloroplasts per mesophyll cell) in field-grown plants of 94 species from the Eastern Pamir Mountains, at elevations between 3800 and 4750 m. Unlike most other mountain areas, the Eastern Pamirs, Karakorum system, Tadjikistan provide localities where low temperatures and radiation combine with moisture stress at high altitudes. For all the attributes measured, significant differences were found between plants with different mesophyll types. Leaves with dorsiventral palisade structure (dorsal palisade, ventral spongy mesophyll cells) had thicker leaves with larger but fewer mesophyll cells, containing more and larger chloroplasts. These differences in mesophyll type are reflected in differences in the total surface of mesophyll cells per unit leaf area ( A _mes/ A ) or volume ( A _mes/ V ). Plants with isopalisade leaf structure (palisade cells under both dorsal and ventral surfaces) are more commonly xerophytes and their increased values of A _mes/ A and A _mes/ V decrease CO₂ mesophyll resistance, which is an important adaptation to drought. Path analysis shows the critical importance of mesophyll cell volume in leading to the covariance between the different leaf attributes and hence to specific leaf mass (SLM), even though mesophyll cell volume is not itself strongly correlated with SLM. This is because mesophyll cell volume increases SLM through its effects on leaf thickness and chloroplast number per cell, but decreases SLM through its negative effect on mesophyll cell density.     

Improved Methods for the Measurement of Total Cell Surface Area in Leaf Mesophyll Tissue

Two improved methods are described for the measurement of thetotal surface area of mesophyll cells in leaf tissue. The firstinvolves stereological measurements on suspensions of enzymicallyisolated mesophyll cells; the second involves measurements ofthe total perimeter of mesophyll cell profiles on transversesections of the leaf, with allowance for curvature of the cellsurfaces by means of theoretically based correction factors.Application of the two methods to the mesophyll tissue of tobaccoleaves gave results which were in very good agreement with eachother. Both methods avoid the inaccuracies and uncertaintiesof previous methods for measurement of mesophyll cell surfaceareas. Key words: Mesophyll, Surface: Area     

Estimation of Mesophyll Conductance to CO(2) Flux by Three Different Methods

The resistance to diffusion of CO₂ from the intercellular airspaces within the leaf through the mesophyll to the sites of carboxylation during photosynthesis was measured using three different techniques. The three techniques include a method based on discrimination against the heavy stable isotope of carbon, ¹³C, and two modeling methods. The methods rely upon different assumptions, but the estimates of mesophyll conductance were similar with all three methods. The mesophyll conductance of leaves from a number of species was about 1.4 times the stomatal conductance for CO₂ diffusion determined in unstressed plants at high light. The relatively low CO₂ partial pressure inside chloroplasts of plants with a low mesophyll conductance did not lead to enhanced O₂ sensitivity of photosynthesis because the low conductance caused a significant drop in the chloroplast CO₂ partial pressure upon switching to low O₂. We found no correlation between mesophyll conductance and the ratio of internal leaf area to leaf surface area and only a weak correlation between mesophyll conductance and the proportion of leaf volume occupied by air. Mesophyll conductance was independent of CO₂ and O₂ partial pressure during the measurement, indicating that a true physical parameter, independent of biochemical effects, was being measured. No evidence for CO₂-accumulating mechanisms was found. Some plants, notably Citrus aurantium and Simmondsia chinensis, had very low conductances that limit the rate of photosynthesis these plants can attain at atmospheric CO₂ level.     

Structural Adaptation of the Leaf Mesophyll to Shading

Structural characteristics of the mesophyll were studied in five boreal grass species experiencing a wide range of light and water supply conditions. Quantitative indices of the palisade and spongy mesophyll tissues (cell and chloroplast sizes, the number of chloroplasts per cell, the total cell and chloroplast surface area per unit leaf surface area) were determined in leaves of each of the species. The cell surface area and the cell volume in spongy mesophyll were determined with a novel method based on stereological analysis of cell projections. An important role of spongy parenchyma in the photosynthetic apparatus was demonstrated. In leaves of the species studied, the spongy parenchyma constituted about 50% of the total volume and 40% of the total surface area of mesophyll cells. The proportion of the palisade to spongy mesophyll tissues varied with plant species and growth conditions. In a xerophyte Genista tinctoria, the total cell volume, cell abundance, and the total surface area of cells and chloroplasts were 30–40% larger in the palisade than in the spongy mesophyll. In contrast, in a shade-loving species Veronica chamaedris, the spongy mesophyll was 1.5–2 times more developed than the palisade mesophyll. In mesophyte species grown under high light conditions, the cell abundance and the total cell surface area were 10–20% greater in the palisade mesophyll than in the spongy parenchyma. In shaded habitats, these indices were similar in the palisade and spongy mesophyll or were 10–20% lower in the palisade mesophyll. In mesophytes, CO₂ conductance of the spongy mesophyll accounted for about 50% of the total mesophyll conductance, as calculated from the structural characteristics, with the mesophyll CO₂ conductance increasing with leaf shading.     

Shading-induced changes in the leaf mesophyll of plants of different functional types

Changes in the structural characteristics of mesophyll induced by shading were investigated in ten species of wild plants of diverse functional types. In all plant types, shading reduced leaf thickness and density by 30–50% and total surface of mesophyll, by 30–70%. The extent and mechanisms of mesophyll structural rearrangement depended on the plant functional type. In the ruderal plants, integral parameters of mesophyll, such as the surface of cells and chloroplasts and mesophyll resistance, changed threefold predominantly because of changes in the dimensions of the cells and chloroplasts. In these plants, shading reduced the volume of chloroplasts by 30%, and the chloroplast numbers per cell declined. The competitor plants showed a twofold increase in mesophyll resistance due to a decrease in the number of photosynthesizing cells per leaf area unit. Moreover, these plants maintained constant dimensions of mesophyll cells, ratios mesophyll surface/mesophyll volume and chloroplast surface/cell surface. In stress-tolerant plants, diffusion resistance of mesophyll remained the same irrespective of the growing conditions, and mesophyll rearrangement was associated with inversely proportional changes in the dimensions of the cells and cell volume per chloroplast. Noteworthy of these plants were relatively constant chloroplasts number per cell, per leaf area unit and total surface area of chloroplasts. The nature of relationship between the mesophyll diffusion resistance and structural parameters of leaf mesophyll differed in plants of diverse functional types.     

Changes in Size and Number of Mesophyll Cells, Nitrogen Content and Photosynthesis with Leaf Order in Brassica spp.

Numbers of mesophyll cells per unit leaf area decreased progressivelyfrom an upper leaf with a width of 3 cm towards the lower leaves.Enlargement of mesophyll cell size with leaf order accountedfor an increase or maintenance of mesophyll cell surface areaper unit leaf area. Increase of photosynthetic rates was correlatedwith increases of mesophyll cell surface area and nitrogen contentper unit leaf area. Therefore, in spite of an increase in cellsurface area to volume ratio with increase of mesophyll cellsize, it appears that increase of mesophyll cell surface areaand nitrogen content per unit leaf area enables a high rateof photosynthesis to be maintained. Brassica, photosynthesis, mesophyll surface area, nitrogen content, cell size, mesophyll resistance, leaf age     

Basic Types of Structural Changes in the Leaf Mesophyll during Adaptation of Eastern Pamir Plants to Mountain Conditions

Quantitative characteristics of mesophyll structure were compared in leaves of eleven alpine plant species grown under natural conditions in the Eastern Pamirs at various altitudes, from 3800 to 4750 m. Basic types of changes in mesophyll structure, associated with plant adaptation to mountain conditions, were characterized. These changes manifested themselves in different numbers of cell layers and cell sizes in the palisade tissue and, as a consequence, in changed leaf thickness and cell number per unit of leaf area. Three plant groups were identified by the changes in the leaf structural characteristics depending on the type of mesophyll structure, ecological group of plant species, and altitude of plant habitat. The first group comprised alpine xerophytes with an isopalisade structure, in which the volume of palisade cells decreased and their number per unit of leaf area increased with the altitude of plant habitat. The number of mesophyll layers and leaf thickness decreased or did not change with altitude. The second group comprised subalpine plant species with a dorsoventral structure of mesophyll; these species occur below the line of continuous night frost. In these plant species, the number of mesophyll layers, leaf thickness, and cell number per unit of leaf area increased with altitude. The third group comprised mesophyte plants with a dorsoventral and homogenous mesophyll structure, which are encountered in a wide range of habitats, including the nival belt (from 4700 to 5000 m). In this group, cell volume increased and cell number per unit of leaf area decreased with altitude. We present a general scheme of leaf structural changes, which explains the changes in the quantitative characteristics of mesophyll as a function of altitude and highland environmental conditions.     

Genome-Dependent Factors in the Development of Leaf Phototrophic Tissues in Diploid and Alloploid Wheat Species

Growth and mesostructure of the photosynthetic apparatus were studied in leaves of ten Triticum L. species. Plants with the A^u genome were shown to develop larger leaf assimilation areas due to expanding areas of individual leaves and an increase in the absolute growth rate. Leaf and mesophyll thickness and mesophyll cell size decreased in the G-genome species. Leaf compactness, which depended on cell size and number per unit leaf area and leaf folding, determined the specific patterns of internal leaf organization in wheat species with diverse genotypes. These patterns did not affect cell plastid-to-cytoplasm ratio as shown by the stable indices of cell surface area/cell volume, cell surface area per chloroplast, and cell volume per chloroplast. The structural indices of leaf phototrophic tissues, mesophyll density, and mesophyll CO₂ conductance in alloploids, as compared to diploid species, depended on both ploidy and genome constitution.     

The leaf anatomy of a broad-leaved evergreen allows an increase in leaf nitrogen content in winter

In temperate regions, evergreen species are exposed to large seasonal changes in air temperature and irradiance. They change photosynthetic characteristics of leaves responding to such environmental changes. Recent studies have suggested that photosynthetic acclimation is strongly constrained by leaf anatomy such as leaf thickness, mesophyll and chloroplast surface facing the intercellular space, and the chloroplast volume. We studied how these parameters of leaf anatomy are related with photosynthetic seasonal acclimation. We evaluated differential effects of winter and summer irradiance on leaf anatomy and photosynthesis. Using a broad-leaved evergreen Aucuba japonica , we performed a transfer experiment in which irradiance regimes were changed at the beginning of autumn and of spring. We found that a vacant space on mesophyll surface in summer enabled chloroplast volume to increase in winter. The leaf nitrogen and Rubisco content were higher in winter than in summer. They were correlated significantly with chloroplast volume and with chloroplast surface area facing the intercellular space. Thus, summer leaves were thicker than needed to accommodate mesophyll surface chloroplasts at this time of year but this allowed for increases in mesophyll surface chloroplasts in the winter. It appears that summer leaf anatomical characteristics help facilitate photosynthetic acclimation to winter conditions. Photosynthetic capacity and photosynthetic nitrogen use efficiency were lower in winter than in summer but it appears that these reductions were partially compensated by higher Rubisco contents and mesophyll surface chloroplast area in winter foliage.     

Influence of Genome on Leaf Anatomy of Triticum and Aegilops

The leaf mesophyll of Triticum and Aegilops is constructed fromcells with one to ten arms. Volume of mesophyll cells per unitleaf area was larger in some monogenomic (A and B genome) plantsthan in polyploids, while leaf volume per unit leaf area wassmaller in the former than in the latter. Consequently, thecompactness of leaf blade is higher in these monogenomic plantsthan in the polyploids. D genome plants showed a much lowervolume of both mesophyll cells and leaf blade per unit leafarea, but the compactness of the leaf blade was generally higherthan in the polyploids. Mesophyll surface area per unit leaf area tended to be largerin the A and B genome than in the D genome and polyploid plants.Out of the polyploids, AB genome plants showed a larger mesophyllsurface area per unit leaf area as compared with AG and ABDgenome plants. Therefore, either the D or the G genome seemsto have the effect of decreasing the mesophyll surface areaper unit leaf area. A decrease of the compactness of leaf bladeand the mesophyll surface area per unit leaf area in the polyploidswas considered to be associated with the reduction of theirphysiological activities on the unit leaf area basis. Triticum, Aegilops, wheat, mesophyll surface area, leaf anatomy, genome, photosynthesis     

The chemical composition and anatomical structure of leaves of grass species differing in relative growth rate

To arrive at a better understanding of variation in specific leaf mass (SLM, leaf weight per unit leaf area), we investigated the chemical composition and anatomical structure of the leaves of 14 grass species varying in potential relative growth rate. Expressed on a dry weight basis, the fast-growing grass species with low SLM contained relatively more minerals and organic N-compounds, whereas slow-growing species with high SLM contained more (hemi)cellulose and lignin. However, when expressed per unit leaf area, organic N-compounds, (hemi)cellulose, total structural carbohydrates and organic acids increased with increasing SLM. For the 14 grasses, no trend with SLM was found for the leaf volume per unit leaf area. Leaf density was positively correlated with SLM. Variation in density was not caused by variation in the proportion of intercellular spaces. The proportion of the total volume occupied by mesophyll and veins did not differ either. A high SLM was caused, at least partly, by a high proportion of non-veinal sclerenchymatic cells per cross-section. The epidermal cell area was negatively correlated with SLM. We conclude that the differences in SLM and in the relative growth rate (RGR) between fast- and slow-growing grass species are based partly on variation in anatomical differentiation and partly on chemical differences within cell types.     

Determination of mesophyll diffusion resistance in Chamaerion angustifolium by the method of three-dimensional reconstruction of the leaf cell packing

A detailed quantitative analysis of the three-dimensional organization of the mesophyll was performed, and mesophyll diffusion resistance to CO₂ in the leaves of Chamaerion angustifolium formed under different irradiance was calculated using an original method of stereometric cellular packing. For each type of leaves (sun and shade), we determined structural components of gas exchange: the volume of mesophyll per unit leaf area (V _mes), the volume of the intercellular space in the mesophyll (V _is), the area of the total mesophyll surface (S), the area of the free mesophyll surface facing the intercellular spaces (S _mes), and the ratios of the total and the free mesophyll surfaces to its volume (S/V and S _mes/V). As compared with sun leaves, in the shade leaves of Ch. angustifolium, S and V _mes decreased twofold, tissue density was reduced twofold, and the share of the intercellular space in the mesophyll rose from 49 to 72%. In shade, the diffusion resistance of the mesophyll increased by 1.8 times because of changes in the leaf structure. At the same time, the ratio S _mes/V was found to increase by 1.4 times, which facilitated the diffusion of CO₂. In the shade leaves of Ch. angustifolium, the diffusion resistance of the intercellular air spaces was reduced twofold as a result of an increase in their share in the leaf mesophyll and simplification of their geometry. Thus, the method of three-dimensional reconstruction of sun and shade leaves of Ch. angustifolium showed a comprehensive rearrangement of the mesophyll spatial organization in shade and revealed the structural mechanisms of changes in the resistance to CO₂ diffusion within the leaf.     

兴安落叶松针叶解剖结构变化及其光合能力对气候变化的适应性

叶片易受环境因子影响,其形态解剖结构特征不但与叶片的生理功能密切相关,而且反映树木对环境变化的响应和适应。叶片结构的改变势必会改变树木的生理功能。同一树种长期生长在异质环境条件下,经过自然选择和适应,会在形态和生理特性等方面产生变异,形成特定的地理种群。另外,母体所经受的环境胁迫也会影响到其子代的生长、发育和生理等特征。因此,了解植物叶片形态结构对环境变化的响应与适应是探索植物对环境变化的响应适应机制的基础。兴安落叶松(Larix gmelinii Rupr.)是我国北方森林的优势树种,主要分布在我国东北地区,但日益加剧的气候变化可能会改变其现有的分布区。为了区分叶片对气候变化的可塑性和适应性,本研究采用同质园法比较测定了6个不同气候条件下的兴安落叶松种源的32年生树木的针叶解剖结构和光合生理相关因子,利用石蜡切片方法分析了针叶的解剖结构特征、光合能力（Pmax-a）、水分利用效率（WUE）之间的关系及其对气候变化的适应性。结果表明：表皮细胞厚度、叶肉细胞厚度、传输组织厚度、维管束厚度、内皮层厚度以及叶片总厚度均存在显著的种源间差异（P < 0.05）。叶肉细胞厚度与Pmax-a、气孔导度和WUE之间均存在显著的正相关关系（P < 0.05）。叶肉细胞厚度、表皮细胞厚度、叶片总厚度以及叶肉细胞厚度和表皮细胞厚度在叶片总厚度中所占比例均与种源地的干燥度指数（即年蒸发量与年降水量之比）呈正线性关系。这些结果说明：不同种源兴安落叶松针叶解剖结构因对种源原地气候条件的长期适应而产生显著的差异,从而引起其针叶光合作用、水分利用等生理功能发生相应的变化,从而有利于该树种在气候变化的情景下得以生存和繁衍。     

Comparative analyses of leaf anatomy of dicotyledonous species in Tibetan and Inner Mongolian grasslands

Knowledge of the leaf anatomy of grassland plants is crucial for understanding how these plants adapt to the environment. Tibetan alpine grasslands and Inner Mongolian temperate grasslands are two major grassland types in northern China. Tibetan alpine grasslands occur in high-altitude regions where the low temperatures limit plant growth. Inner Mongolian temperate grasslands are found in arid regions where moisture is the limiting factor. Few comparative studies concerning the leaf anatomy of grassland plants of the Tibetan Plateau and Inner Mongolian Plateau have been conducted. We examined leaf characteristics at 71 sites and among 65 species, across the alpine grasslands of the Tibetan Plateau and the temperate grasslands of the Inner Mongolian Plateau. We compared the leaf structures of plants with different life forms and taxonomies, and their adaptation to arid or cold environments. We explored relationships among leaf features and the effects of climatic factors (i.e., growing season temperature and precipitation) on leaf characteristics. Our results showed that (i) there were significant differences in leaf anatomy between Tibetan alpine and Inner Mongolian temperate grasslands. Except for mesophyll cell density, the values obtained for thickness of leaf tissue, surface area and volume of mesophyll cells were larger on the Tibetan Plateau than on the Inner Mongolian Plateau. (ii) Within the same family or genus, leaf anatomy showed significant differences between two regions, and trends were consistent with those of whole species. (iii) Leaf anatomy of woody and herbaceous plants also showed significant differences between the regions. Except for mesophyll cell density, the values obtained for the thickness of leaf tissue, and the surface area and volume of mesophyll cells were larger in herbaceous than in woody plants. (iv) Leaf anatomical traits changed accordingly. Total leaf thickness, thicknesses of lower and upper epidermal cells, and surface area and volume of mesophyll cells were positively correlated, while mesophyll cell density was negatively associated with those traits. (v) Growing season temperature had stronger effects on leaf anatomy than growing season precipitation. Although the communities in Tibetan and Inner Mongolian grasslands were similar in appearance, leaf anatomy differed; this was probably due to the combined effects of evolutionary adaptation of plants to environment and environmental stress induced by climatic factors.     

Structural assessment of the impact of environmental constraints on Arabidopsis thaliana leaf growth: a 3D approach

Light and soil water content affect leaf surface area expansion through modifications in epidermal cell numbers and area, while effects on leaf thickness and mesophyll cell volumes are far less documented. Here, three-dimensional imaging was applied in a study of Arabidopsis thaliana leaf growth to determine leaf thickness and the cellular organization of mesophyll tissues under moderate soil water deficit and two cumulative light conditions. In contrast to surface area, thickness was highly conserved in response to water deficit under both low and high cumulative light regimes. Unlike epidermal and palisade mesophyll tissues, no reductions in cell number were observed in the spongy mesophyll; cells had rather changed in volume and shape. Furthermore, leaf features of a selection of genotypes affected in leaf functioning were analysed. The low-starch mutant pgm had very thick leaves because of unusually large palisade mesophyll cells, together with high levels of photosynthesis and stomatal conductance. By means of an open stomata mutant and a 9-cis-epoxycarotenoid dioxygenase overexpressor, it was shown that stomatal conductance does not necessarily have a major impact on leaf dimensions and cellular organization, pointing to additional mechanisms for the control of CO(2) diffusion under high and low stomatal conductance, respectively.