Histochemical, ultrastructural, and electron microprobe analytical studies on the localization of calcium in rat...

The enamel organs of rat incisors were separated from the enamel surface and processed for rapid freezing and freeze-substitution. A histochemical stain for calcium (GBHA) of thick Epon sections revealed intense calcium reactions in the secretory ameloblasts, exclusively in the tubulovesicular structures extending throughout their distal cytoplasm. Electron microscopy revealed a thin layer of amorphous material with clusters of electron-dense granules along the distal surface of secretory ameloblasts. In young secretory ameloblasts without typical Tomes' processes, a considerable number of mitochondria were located in the distal cytoplasm and contained numerous electron-dense granules. Similar dense granules as well as fine ribbon-like electron-dense figures, all containing significant amounts of calcium, were observed in some of the tubulovesicular structures at the distal end of these cells. A putative exocytotic figure of such dense granules was also observed. The electron-dense granules were rare in more differentiated ameloblasts with elongated Tomes' processes, which occasionally displayed ribbon-like figures in some of the tubulovesicular structures in the process region. No significant calcium peak was detected in the extracellular amorphous material, secretory granules, or along the lateral plasma membranes. These observations may imply high calcium concentrations in mitochondria and tubulovesicular structures in the distal cytoplasm of secretory ameloblasts relative to that of the cytosol and support the possible contribution of these organelles in secretory ameloblasts to cellular calcium regulation at least in the early stage of amelogenesis.