Use of Ruthenium Red staining to detect mast cell degranulation in vivo.
OBJECTIVE
To establish a method of detecting mast cell degranulation in tissues during in vivo microscopy.


METHODS
Hamster tissues were prepared for intravital microscopy. Ruthenium red (RR) was superfused over the cheek pouch at concentrations of 0.0001-0.01% to determine the optimal concentration. Mast cells were stimulated with compound 48/80, as well as with vasoactive agents not known to be stimulatory to mast cells, following which, mast cell staining was observed. Mesenteries were stained with Toluidine Blue (TB) or RR and mast cell degranulation was assessed during treatment with compound 48/80, or control.


RESULTS
During superfusion with varying concentrations of RR, a dose dependence for background staining of unstimulated cells was observed. A RR concentration of 0.001% was optimal for in vivo detection of mast cell degranulation. Mast cells exposed to 0.001% RR were stained following stimulation with compound 48/80 but not after treatment with KCl or acetylcholine. The latter agents are not known to stimulate mast cells. Thus, arteriolar vasomotor responses, per se, did not appear to play a role in mast cell RR uptake. Comparable results were obtained with RR versus TB in control or 48/80-treated mesenteries.


CONCLUSIONS
This RR technique facilitates rapid detection of mast cell degranulation in vivo and provides an opportunity to assess both mast cell and microvascular function simultaneously.
Shepherd
R K
RK
Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, Charlottesville, USA.
Duling
B R
BR
D000818:Animals; D015550:Cell Degranulation / Q000502:physiology*; D002610:Cheek; D004396:Coloring Agents; D006224:Cricetinae; D008297:Male; D008407:Mast Cells / Q000187:drug effects / Q000502:physiology*; D008647:Mesocricetus; D008833:Microcirculation / Q000187:drug effects / Q000502:physiology; D012430:Ruthenium Red; D013194:Staining and Labeling / Q000379:methods*; D014048:Tolonium Chloride; D003189:p-Methoxy-N-methylphenethylamine / Q000494:pharmacology
D004396:Coloring Agents; D012430:Ruthenium Red; D014048:Tolonium Chloride; D003189:p-Methoxy-N-methylphenethylamine