Abstract
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.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 591-598 |
| Number of pages | 8 |
| Journal | Journal of Experimental Biology |
| Volume | 201 |
| Issue number | 4 |
| State | Published - Feb 1998 |
| Externally published | Yes |
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Keywords
- B-cell cross-linking
- Carp
- Cyprinus carpio
- Digital imaging microscopy
- Fish
- Flow cytometry
- Fluo3
- Fura-2
- Intracellular Ca
- Lipopolysaccharide
- Lymphocytes
- Phytohaemagglutinin
ASJC Scopus subject areas
- Agricultural and Biological Sciences(all)
- Agricultural and Biological Sciences (miscellaneous)
Cite this
Ca2+ signals during early lymphocyte activation in carp Cyprinus carpio L. / Verburg-Van Kemenade, B. M Lidy; Saeij, Jeroen; Flik, Gert; Willems, Peter H G M.
In: Journal of Experimental Biology, Vol. 201, No. 4, 02.1998, p. 591-598.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Ca2+ signals during early lymphocyte activation in carp Cyprinus carpio L.
AU - Verburg-Van Kemenade, B. M Lidy
AU - Saeij, Jeroen
AU - Flik, Gert
AU - Willems, Peter H G M
PY - 1998/2
Y1 - 1998/2
N2 - 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.
AB - 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.
KW - B-cell cross-linking
KW - Carp
KW - Cyprinus carpio
KW - Digital imaging microscopy
KW - Fish
KW - Flow cytometry
KW - Fluo3
KW - Fura-2
KW - Intracellular Ca
KW - Lipopolysaccharide
KW - Lymphocytes
KW - Phytohaemagglutinin
UR - http://www.scopus.com/inward/record.url?scp=0031971283&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0031971283&partnerID=8YFLogxK
M3 - Article
C2 - 9438833
AN - SCOPUS:0031971283
VL - 201
SP - 591
EP - 598
JO - Journal of Experimental Biology
JF - Journal of Experimental Biology
SN - 0022-0949
IS - 4
ER -