Increase In Cellular Ca2 Research Articles

On the stimulation of rat thymocyte 3‐0‐methyl‐glucose transport by mitogenic stimuli

Pretreatment of rat thymus lymphocytes with N-ethyl-maleimide prevents the stimulation of 3-O-methyl-glucose transport by concanavalin A or ionophore A23187 but does not affect the ability of concanavalin A to induce a rapid increase in cellular Ca2+take. N-ethyl-maleikide added after concanavalin A amplifies rather than prevents the subsequent stimulation of 3-O-methyl-glucose transport. Incubation of thymocytes with concanavalin A produces a decrease of 43% in the apparent Ki for phloretin, a competitive inhibitor of 3-O-methyl-glucose transport, without affecting the apparent Km for the substrate. Similarly, very low concentrations of cytochalasin B inhibit concanavalin A-stimulated glucose transport preferentially, without markedly affecting the unstimulated transport rates. The similarity between concanavalin A-stimulated 3-O-methyl-glucose transport in thymocytes and insulin-stimulated glucose transport in adipose tissue, with particular emphasis on the effects of the modifying agents described in this paper, is discussed.

Serum‐stimulated changes in calcium transport and distribution in mouse 3T3 cells and their modification by dibutyryl cyclic AMP

Serum stimulation of quiescent 3T3 cells returns the cells to a proliferative state. Changes in Ca content, transport and distribution during the transition through G1 and S phase have been investigated following serum stimulation of these cells. 45 Ca exchange data indicate at least two kinetically defined cellular compartments for Ca; a rapidly exchanging component presumably representing surface Ca which is removable by EGTA and a slowly exchanging component presumably representing cytoplasmically located Ca. Previous studies (Tupper and Zorgniotti, '77) indicate that the approach to quiescence in the 3T3 cells is characterized by a large increase in the surface Ca component. The present data demonstrate that this component is rapidly lost following serum stimulation. Furthermore, the serum induces an 8-fold increase in Ca influx into the cytoplasmic compartment and a reduction in the unidirectional efflux rate coefficient for Ca. The increased Ca uptake peaks at approximately six hours (mid G1) and is accompanied by a parallel increase in cellular Ca. Prior to entrance of the cells into S phase (10-12 hours), Ca uptake declines. This is followed by a slower decline in cytoplasmic Ca levels. Simultaneous addition to fresh serum plus 0.5 mM dibutryl cAMP inhibits the entrance of the cells into S phase. Under these conditions the loss of surface Ca is not blocked. However, the presence of 0.5 mM dibutyryl cAMP inhibits the increase in Ca uptake and, in turn, diminishes the increase in cellular Ca following serum stimulation. In contrast, a low level of dibutyryl cAMP (0.1 mM) enhances progression through G1 phase but also reduces both Ca uptake and Ca content of the cells. The data suggest that the serum induced changes in Ca content and transport are linked to intracellular cyclic nucleotide levels and progression through G1 phase and that extracellular cAMP elevating agents may enhance of inhibit these interactions in a concentration dependent manner.

Role of membrane-bound Ca in ghost permeability to Na and K.

The permeability of red cell ghosts to K is determined by the amount of membrane-bound Mg which, in turn, depends on internal Mg. Contrasting with such effect, an increase in cellular Ca raises K permeability. To test whether this action is due to a competitive displacement of membrane Mg, the free Ca content of human red cell ghosts was altered by means of Ca-EGTA buffers. Net Na and K movements as well as Ca and Mg bindings were assessed after incubation in a Na-medium at 37 degrees C. Raising Ca from 3 X 10(-7) to 1 X 10(-2) M caused a large K efflux with very little Na gain. Under similar conditions, Ca binding was increased without affecting membrane-bound Mg. Both Ca binding and K loss were markedly diminished by either adding ATP to the hemolytic medium or increasing internal Mg at a fixed Ca concentration. A Scatchard analysis showed three Ca binding sites, two of them having high affinity. It is concluded that Ca action does not arise from a displacement of membrane-bound Mg but from binding to different sites in the membrane. Presumably, high affinity sites are involved in the control of K permeability.

Stimulation of 45Ca2+ efflux from rat pituitary by luteinizing hormone-releasing hormone and other pituitary stimulants.

1. Rat anterior pituitaries were incubated in medium containing 45Ca2+, superfused with non-radioactive medium and the efflux of 45Ca2+ studied. 2. When pituitaries from overiectomized rats were used the addition of LH-RH to the medium at a time when slow exponential efflux of Ca2+ was occurring produced a transient increase in 45Ca2+ efflux simultaneous with or before an increased release of LH. The stimulation of both 45Ca2+ efflux and LH release showed a similar concentration dependence on LH-RH. 3. Increasing the K+ concentration of the medium tenfold also stimulated 45Ca2+ efflux and LH release. The response to LH-RH of both parameters was reduced by superfusion with calcium-free medium. 4. LH-RH induced only a small increase in pituitary 45Ca2+ efflux when intact or thyroidectomized male rats were used. TRH increased 45Ca2+ efflux from thyroidectomized rat pituitaries but had only a small effect when pituitaries from intact or ovariectomized rats were studied. 5. Pretreatment of ovariectomized rats with oestradiol inhibited the subsequent increase in 45Ca2+ efflux induced by LH-RH. 6. It is concluded that the secretagogue induced increase in 45Ca2+ efflux results from an increase in cellular Ca2+ activity which is presumably active in stimulus-secretion coupling.