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Updated: Aug 25, 2026
Isolation of Peritoneum-derived Mast Cells and Their Functional Characterization with Ca2+-imaging and Degranulation Assays
Published on: July 4, 2018
In vitro activation of murine DRG neurons by CGRP-mediated mucosal mast cell degranulation
F De Jonge1, A De Laet, L Van Nassauw
1Laboratory of Cell Biology and Histology, Department of Biomedical Sciences, University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp, Belgium. jean-pierre.timmermans@ua.ac.be
Abstract:
Upregulation of CGRP-immunoreactive (IR) primary afferent nerve fibers accompanied by mastocytosis is characteristic for the Schistosoma mansoni-infected murine ileum. These mucosal mast cells (MMC) and CGRP-IR fibers, which originate from dorsal root (DRG) and nodose ganglia, are found in close apposition. We examined interactions between primary cultured MMC and CGRP-IR DRG neurons in vitro by confocal recording of intracellular Ca(2+) concentration ([Ca(2+)](i)). The degranulatory EC(50) for the mast cell secretagogue compound 48/80 (C48/80; 10 microg/ml) and the neuropeptides CGRP (2.10(-8) M) and substance P (SP; 3.10(-8) M) were determined by measurement of extracellular release of the granule chymase, mouse mast cell protease-1. Application of C48/80 (10 microg/ml) and CGRP and SP (both 10(-7) M) to Fluo-4-loaded MMC induced a transient rise in [Ca(2+)](i) after a lag time, indicative of mast cell degranulation and/or secretion. The CGRP response could be completely blocked by pertussis toxin (2 microg/ml), indicating involvement of G(i) proteins. Application of MMC juice, obtained by C48/80 degranulation of MMC, to Fluo-4-loaded DRG neurons induced in all neurons a rise in [Ca(2+)](i), indicative of activation. Degranulation of MMC by C48/80 in culture dishes containing Fluo-4-loaded DRG neurons also caused activation of the DRG neurons. In conclusion, these results demonstrate a bidirectional cross-talk between cultured MMC and CGRP-IR DRG neurons in vitro. This indicates that such a communication may be the functional relevance for the close apposition between MMC and CGRP-IR nerve fibers in vivo.
Insights
This study reveals bidirectional communication between mucosal mast cells and CGRP-nerve fibers in Schistosoma mansoni infection. This interaction in the gut may explain the close proximity of these cells in vivo.
Area of Science:
- Neuroscience
- Immunology
- Parasitology
Background:
- Schistosoma mansoni infection leads to increased CGRP-immunoreactive nerve fibers and mastocytosis in the murine ileum.
- Mucosal mast cells (MMC) and CGRP-IR fibers are closely apposed in the infected ileum.
Purpose of the Study:
- To investigate the in vitro interactions between cultured MMC and CGRP-IR dorsal root ganglia (DRG) neurons.
- To elucidate the role of CGRP and substance P in mast cell degranulation and neuronal activation.
Main Methods:
- Primary cultured MMC and DRG neurons were used.
- Intracellular calcium concentration ([Ca(2+)](i)) was measured using confocal recording.
- Mast cell degranulation was assessed by measuring mouse mast cell protease-1 release.
- Neurons were activated by 'MMC juice' from degranulated MMC.
Main Results:
- Compound 48/80, CGRP, and substance P induced calcium transients in MMC, indicating degranulation.
- The CGRP-induced response in MMC was blocked by pertussis toxin, suggesting G(i) protein involvement.
- Degranulation of MMC led to activation of DRG neurons, evidenced by increased [Ca(2+)](i).
Conclusions:
- Demonstrates bidirectional communication between cultured MMC and CGRP-IR DRG neurons.
- Suggests this cross-talk is functionally relevant to the in vivo proximity of MMC and CGRP-IR nerve fibers.
