The effect of endotoxemia on concanavalin A induced alterations in cytoplasmic free calcium in rat spleen cells as determined with Fluo-3.
The effects of acute (3 h) and chronic (30 h) in vivo infusions of Escherichia coli endotoxin on the Ca2+ homeostasis of rat spleen cells was investigated. Conditions were established for obtaining reliable estimates of [Ca2+]i in these cells using the newly-developed Ca2+ indicator Fluo-3. The resting [Ca2+]i of splenocytes and T lymphocyte-enriched preparations were 119 +/- 35 and 102 +/- 31 nM, respectively. Treatment of the cells with concanavalin A (Con A) resulted in a rapid increase in [Ca2+]i. The magnitude of the increase was positively correlated with the concentration of Con A, whereas the time required to reach the maximum [Ca2+]i was inversely related to the amount of Con A. The peak [Ca2+]i was attained more rapidly in splenocytes (i.e. less than or equal to 30 s) than in the T cell-enriched fraction (i.e. 1.5-2.0 min). Both the resting [Ca2+]i and the Con A-induced increase in [Ca2+]i were similar to values previously reported for other lymphocyte cell types using different Ca2+ indicators, thereby supporting the values obtained with Fluo-3. Infusions of saline or endotoxin prior to the isolation of the cells did not result in significant alterations of either resting [Ca2+]i or the cells' response to Con A. Since chronic infusions of endotoxin have previously been shown to cause a reduction in blastogenic responsiveness of splenocytes to Con A, these data suggest that the endotoxin-induced lesion occurs distal to the mobilization of intracellular Ca2+.
Hagar
A F
AF
Department of Physiology, Louisiana State University Medical Center, New Orleans.
Spitzer
J A
JA
D000814:Aniline Compounds; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003208:Concanavalin A / Q000494:pharmacology*; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism*; D004731:Endotoxins / Q000494:pharmacology*; D005456:Fluorescent Dyes; D006706:Homeostasis / Q000187:drug effects / Q000502:physiology; D007700:Kinetics; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D014966:Xanthenes
D000814:Aniline Compounds; D004731:Endotoxins; D005456:Fluorescent Dyes; D014966:Xanthenes; D003208:Concanavalin A; C059715:Fluo-3; C044101:endotoxin, Escherichia coli; D002118:Calcium