Diacylglycerol and phorbol ester stimulate secretion without raising cytoplasmic free calcium in human platelets

Effects of growth factors on cytosolic free calcium concentration and DNA synthesis in cultured rat aortic smooth muscle cells.

“…This idea is further supported by the evidence that activation of protein kinase C by synthetic diacylglyceride or phorbol ester elicites the biological response in human platelets without accompanying the calcium mobilization Yamanishi et al 1983 ;Nishizuka 1984). Moreover, Rink et al (1983) have directly confirmed that neither diacylglycerol nor phorbol ester evokes the increase in cytosolic free calcium concentration in human platelets. In contrast to PGDF, Hesketh et al (1985) demonstrated that EGF increases cytosolic free calcium concentration in 3T3 fibroblasts without accompanying inositol phospholipid breakdown nor activation of protein kinase C. Second, the possibility must be considered that calcium release from intracellular stores, which might have been masked due to the calcium chelating effect of quin 2 (Tsien 1980), might contribute to DNA synthesis provoked by PDGF.…”

Regulation of cytosolic calcium by collagen in single human platelets

“…This experimental set up also has the advantage of providing a more physiological presentation of collagen to the platelet, i. Laminin may therefore provide a important medium for single cell calcium studies in platelets using soluble agonists. The explanation for the lack of an observed increase (Rink et al, 1983;Watson et al, 1985;Pollock et al, 1986) in [Ca2+]i in platelet populations using concentrations of collagen of 20 tg ml-' or less is not known, but a number of factors are worthy of consideration. There is a much lower increase in [Ca2+], in single platelets adhered to collagen compared with the response to thrombin (see Figure 2b); thus, in a cell population low concentrations of collagen ( < 1I0jg ml-'), which induce less than 10% of cells to adhere (Smith & Dangelmaier, 1990), the weak signal generated by collagen may not be detected.…”

“…In the presence of cyclo-oxygenase inhibitors, concentrations of collagen up to 20 g ml-' do not induce a measurable rise in cytosolic calcium in platelets loaded with the fluorescent indicators, fura-2 or quin2 (Rink et al, 1983;Watson et al, 1985;Pollock et al, 1986), despite the fact that these are sufficient to induce activation of the phosphoinositide pathway. On the other hand, if the concentration of collagen is increased to 50 lg ml-' a rise in cytosolic calcium is observed in the presence of a thromboxane receptor antagonist, ADPscavenger system and fibrinogen antagonist (Smith et al, 1992); however, despite the inclusion of these inhibitors, the possibility remains that other released substances underlie this response.…”

Reconstitution of constitutive secretion using semi-intact cells: regulation by GTP but not calcium.

“…Several considerations make the latter possibility an attractive hypothesis. (a) Previous studies have shown that the Ca 2+ sensitivity of the regulated secretory apparatus can indeed be modulated; phorbol esters, for instance, can stimulate exocytic release from storage granules without a rise in intraceHular Ca 2+, most likely by increasing the affinity of the exocytic apparatus to Ca 2+ (Rink et al, 1983;Knight and Baker, 1985). (b) Cell lines that package peptide hormones into regulated granules often release a fraction of the newly synthesized peptides in an unregulated fashion.…”