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The immobilisation of Euglena gracilis Z cells in a calcium alginate matrix maintained respiratory and photosynthetic activities and ultrastructural integrity. Moreover, immobilization did not prevent Euglena cells from greening inside the gel beads. Electron microscopy demonstrated that the immobilized cells were fixed in the same cellular state as they were when the immobilization occurred. This can he explained by simultaneous reaction of both Ca2+ and the alginate with the cells. Some hypotheses about the role of C2+ are discussed. In addition, long term storage (2 years) in calcium alginate has been performed permitting applications in algal storage.

Primary cultivated rabbit articular chondrocytes were immobilized in calcium alginate beads. Both free and entrapped cells were allowed to grow under normal conditions. After long-term immobilization, the cells still exhibited metabolic activities, patterns of division, synthesis and secretion of extracellular matrix macromolecules such as type II collagen and proteoglycans. After 38 days, immobilized rabbit articular chondrocytes predominantly expressed type II but not type I collagen. Thus, they maintained their cartilage pheno-type. After bead lysis, harvested cells showed normal growth patterns when resuspended in culture medium. On the basis of these results, long-duration storage and large-scale production of extracellular matrix components are being investigated.

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Mouse neuroblastoma cells (N18) were immobilized in calcium-alginate gel beads. Under standard culture conditions (37 degrees C; 5% CO2), cell growth was observed inside the beads. The number of cells increased threefold during 7 days of culture with cell division and differentiation visualized by electron microscopy. Cell properties maintained after short-term storage (2-3 days at 4 degrees C) included: (i) properties of voltage-dependent ionic channels tested by patch-clamp electrophysiological techniques; (ii) expression of cell-adhesion membrane proteins tested by immunohistochemistry (iii) morphological differentiation obtained by depletion of foetal calf serum in culture medium. The advantages of such an immobilization technique as applied to neurone cells are discussed.

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Groups of five rats were fed for sixteen weeks a slightly deficient diet, supplemented with 0

Cell division in exponentially increasing populations of the wildtype, photosynthetic Z strain of Euglena gracilis Klebs cultured autotrophically on an aerated, magnetically stirred, minimal mineral medium (pH 7.0) in constant light (LL) or in a light-dark 1 hour:1 hour cycle (LD:1,1) at 250C could be synchronized by a 10-hour:10-hour low (2 micromolar):normal (200 micromolar Since intracellular free calcium is well known as a cell regulator, coordinating many kinds of intracellular reactions and even its own concentration (1,2,5,7), it is quite possible that it may play a significant role in the functioning of circadian clocks (10, 13). Indeed, it has been demonstrated in a variety of eukaryotic organisms that circadian rhythmicity of many different types of physiological processes are modulated by calcium levels: cell shape (23, 24) cell division (18) in Euglena gracilis, ocular rhythms in Bulla gouldiana (25), leaf movement in Trifolium repens (3) and Cassiafasciculata (29), conidiation in Neurospora crassa (27), and phototaxis in Chlamydomonas reinhardtii (17). Three models for circadian oscillators (10) that are based in part on cellular calcium (18,21,22) share in common a cycling ofintracellular calcium among compartments (endoplasmic reticulum, mitochondria, cytoplasm, extracellular medium-depending on the model). Although this intercompartmental calcium cycling in itself constitutes the circadian mechanism in the Kippert model (21), it is only one important element of a biochemical circadian loop in that of Goto et al. (18).In an attempt to elucidate the role of calcium in the timing mechanisms that are responsible for circadian periodicities, we have studied the effects of changes in extracellular calcium concentration on the rhythm of cell division in E. gracilis Klebs. Such changes affect the intracellular distribution of calcium via changes at the level of the plasma membrane as well as within another cellular compartment in this unicellular alga (35).