Calcium signals in single T cells on activation by lectin.
Succinylation of concanavalin A (Con A) reduces its oligomer size while retaining its mitogenicity, and provides a probe of T cell activation. We have observed responses of cytosolic ionized calcium to succinyl Con A in suspensions of Jurkat and rat lymph node (LN) cells, using a fluorimeter, and in single cells settled on glass, using a dual wavelength video imaging system. In the fluorimeter a mitogenic level of succinyl Con A (30 micrograms/ml) produced only a 15-30 nM rise in average cell calcium in the suspended Jurkat or rat cells whereas the use of quantitative video imaging produced asynchronous 250-1000 nM pulses of free calcium in 35% of Jurkat cells and 300-850 nM pulses in 45% of rat LN cells. In Jurkat cells these pulses were sometimes repetitive, giving rise to apparent oscillations. In the fluorimeter 30 micrograms/ml of native Con A (a supra-mitogenic concentration) produced a 300 nM rise in average cell calcium in suspended Jurkat cells, and a 100 nM rise in rat LN cells; when major histocompatibility complex class II-bearing cells were removed the response rose. Mitogenic Con A (3 micrograms/ml) produced a much lower rise in calcium. With video imaging the response seen was greater. Levels greater than 30 micrograms/ml Con A caused 700-5000 nM pulses synchronously in 94% of Jurkat cells and 250-1000 nM pulses in 73% of rat LN cells. At 3 micrograms/ml Con A produced asynchronous 300-1100 nM pulses in 36% of rat LN cells. We conclude that the absence of a calcium signal in the fluorimeter can conceal asynchronous calcium responses in individual cells and that brief asynchronous cytosolic calcium pulses are sufficient for lectin to activate rat T cells.
Tregear
R T
RT
AFRC Institute of Animal Physiology and Genetics Research, Babraham, Cambridge, GB.
Hoyland
J
J
Mason
W T
WT
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003208:Concanavalin A / Q000031:analogs & derivatives / Q000494:pharmacology; D005470:Fluorometry; D037102:Lectins / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects*; D051381:Rats; D013601:T-Lymphocytes / Q000378:metabolism*
D037102:Lectins; D003208:Concanavalin A; C019558:succinylconcanavalin A; D002118:Calcium