Ca2+ signals during early lymphocyte activation in carp Cyprinus carpio L.
To measure cellular responses and the involvement of increased cytosolic Ca2+ levels ([Ca2+]i), peripheral blood leukocytes (PBL) of carp were loaded with the fluorescent intracellular Ca2+ indicators Fluo-3 and Fura-2. Responses of lymphocytes to T-cell mitogen (phytohaemagglutinin, PHA), to B-cell mitogen (lipopolysaccharide, LPS) and to immunoglobulin (Ig) cross-linking with a monoclonal antibody to carp Ig were measured using flow cytometry. Both T-cell stimulation by PHA and B-cell stimulation by membrane Ig cross-linking evoked a rapid elevation of [Ca2+]i. B-cell stimulation by LPS was not linked to an increase in [Ca2+]i. As judged by the percentage of reacting cells, it was concluded that all Ig-positive lymphocytes reacted to Ig cross-linking by elevating [Ca2+]i. At the single-cell level, the reactions of Fura-2-loaded cells were followed every 6 s using digital imaging microscopy. Both cells displaying spontaneous [Ca2+]i oscillations and non-oscillating cells responded to stimulation with an increase in [Ca2+]i, sometimes, in already oscillating cells, accompanied by an increase in frequency and/or amplitude of the oscillations. These results show that intracellular Ca2+ responses of PBL upon activation resemble those in mammals and form a powerful tool for studies into cell-specific regulation.
Verburg-Van Kemenade
B M
BM
Cell Biology and Immunology Laboratory, Institute of Animal Sciences, Agricultural University, Wageningen, The Netherlands. Lidy.Verburg@celb.edc.wau.nl
Saeij
J P
JP
Flik
G
G
Willems
P H
PH
D000818:Animals; D000937:Antigen-Antibody Reactions; D002118:Calcium / Q000502:physiology*; D002347:Carps / Q000276:immunology*; D003600:Cytosol / Q000502:physiology; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D008213:Lymphocyte Activation; D010835:Phytohemagglutinins / Q000494:pharmacology
D008070:Lipopolysaccharides; D010835:Phytohemagglutinins; D002118:Calcium