Related Experiment Videos
Activation by bacterial lipopolysaccharide causes changes in the cytosolic free calcium concentration in single
O Letari1, S Nicosia, C Chiavaroli
1Department of Medicine, University of Geneva, Switzerland.
Abstract:
Variations in the cytosolic free Ca2+ concentration [( Ca2+]i) upon LPS exposure were studied in single rat peritoneal macrophages loaded with fura-2 under carefully controlled conditions. Of a total of 60 cells examined, 47% responded to LPS (1 microgram/ml) with an increase in [Ca2+]i. Macrophages were heterogeneous with regard to the LPS response, with individual cells exhibiting single rapid and transient increases in [Ca2+]i, multiple transients, or slower and more sustained variations. In 62% of the responding cells, a second exposure to LPS elicited a [Ca2+]i rise, although usually to a slightly lower peak value. Thus, rapid desensitization to LPS does not occur in the majority of these macrophages. EGTA did not abolish the response of those cells that exhibited a single rapid transient in [Ca2+]i, indicating that the source of the initial [Ca2+]i rise was the intracellular stores. There was no obvious correlation between the type of response to LPS and the initial morphologic features (rounded vs polarized) of the cells. Our present work shows unequivocally that LPS induces increases in macrophage [Ca2+]i and, thereby, lends substantial support to the hypothesis that [Ca2+]i is a second messenger in LPS-mediated activation of the macrophage.
Insights
Lipopolysaccharide (LPS) triggers calcium (Ca2+) increases in rat macrophages, supporting its role as a second messenger in immune activation. Most macrophages did not rapidly desensitize to repeated LPS exposure.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Lipopolysaccharide (LPS) is a key component of Gram-negative bacteria.
- Macrophage activation is crucial for innate immunity.
- The role of intracellular calcium (Ca2+) in LPS-mediated signaling is not fully understood.
Purpose of the Study:
- To investigate the effect of LPS on cytosolic free Ca2+ concentration ([Ca2+]i) in rat peritoneal macrophages.
- To determine the source of Ca2+ involved in the LPS response.
- To assess potential desensitization to LPS.
Main Methods:
- Single rat peritoneal macrophages were loaded with the calcium indicator fura-2.
- Cells were exposed to LPS (1 microgram/ml) under controlled conditions.
- Intracellular Ca2+ variations were measured using fluorescence microscopy.
Main Results:
- 47% of macrophages responded to LPS with an increase in [Ca2+]i.
- Responses varied, including single rapid transients, multiple transients, and sustained increases.
- 62% of responding cells showed a response to a second LPS exposure, indicating limited rapid desensitization.
- EGTA treatment suggested intracellular Ca2+ stores as the primary source for initial Ca2+ release.
Conclusions:
- LPS unequivocally induces increases in macrophage [Ca2+]i.
- Cytosolic Ca2+ acts as a second messenger in LPS-mediated macrophage activation.
- Macrophages exhibit heterogeneous responses to LPS, with limited rapid desensitization.