Calcium-induced patterns of calcium-oxalate crystals in isolated leaflets of Gleditsia triacanthos L. and Albizia julibrissin Durazz

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Abstract
For experimental induction of crystal cells (=crystal idioblasts) containing calcium-oxalate crystals, the lower epidermis was peeled from seedling leaflets of Gleditsia triacanthos L., exposing the crystal-free mesophyll and minor veins to the experimental solutions on which leaflets were floated for up to 10 d under continous light. On 0.3–2.0 mM Ca-acetate, increasing numbers of crystals, appearing 96 h after peeling, were induced. The pattern of crystal distribution changed with Ca2+-concentration ([Ca2+]): at low [Ca2+], crystals formed only in the non-green bundlesheath cells surrounding the veins, believed to have a relatively low Ca2+-extrusion capacity; at higher [Ca2+], crystals developed in up to 90% of the mesophyll cells, and at supraoptimal [Ca2+], large extracellular crystals formed on the tissue surface. By sequential treatments with solutions of different [Ca2+], the following three phases were identified in the induction of crystal cells: (1) during the initial 24-h period (adaptive aging), Ca2+ is not required and crystal induction is not possible; (2) during the following 48 h (induction period), exposure to 1–2 mM Ca-acetate induces the differentiation of mesophyll cells into crystal cells; (3) crystal growth begins 72 h after the start of induction. In intact leaflets of Albizia julibrissin Durazz., calcium-oxalate crystals are found exclusively in the bundle-sheath cells of the veins, but crystals were induced in the mesophyll of peeled leaflets floating on 1 mM Ca-acetate. Exposure to inductive [Ca2+] will thus trigger the differentiation of mature leaf cells into crystal cells; the spatial distribution of crystals is determined by the external [Ca2+] and by the structural and functional properties of the cells in the tissue.