A patch-clamp study: secretagogue-induced currents in rat peritoneal mast cells

Ca2+ entry through plasma membrane has been considered to play a significant role in elevating cytosolic free Ca2+ concentrations during stimulus-secretion coupling in mast cells, but electrophysiological evidence of the Ca2+ channels is lacking. We examined the properties of secretagogue (compound 48/80)-induced currents in rat peritoneal mast cells, using the patch-clamp technique. In the whole cell recordings, the addition of compound 48/80 induced transient currents that were suppressed by Cd or reduced by ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). In Ringer solution containing 2 mM Ca2+, the current-voltage relation was fairly linear from -100 to 50 mV and the reversal potential was 14 +/- 10.1 mV (n = 9). When the external Ca2+ was approximately 1 microM, the compound 48/80-induced currents were marginal, but readmission of Ca2+ or Ba2+ to the bath solution led to an appearance of the currents. In the cell-attached patches, the stimulation enhanced the activity of inward current mediated by Ba2+. The unitary inward Ba2+ current was characterized by the unitary conductance of 10.5 +/- 2.0 pS (n = 10) with isotonic BaCl2 pipette solution, the extrapolated reversal potential of 60.7 +/- 16.0 mV (n = 10) positive to the resting membrane potentials. The percent open time of the inward Ba2+ current channel was not appreciably changed by voltage. In some whole cell recordings, an increase in openings of the cation-selective channel (20-45 pS) was identified in the stimulated cells. When the external Na+ was completely replaced by choline+, the compound 48/80-induced currents had a fairly linear current-voltage relation.(ABSTRACT TRUNCATED AT 250 WORDS)

Download Full-text

A patch-clamp study of histamine-secreting cells.

The Journal of General Physiology ◽

10.1085/jgp.88.3.349 ◽

1986 ◽

Vol 88 (3) ◽

pp. 349-368 ◽

Cited By ~ 102

Author(s):

M Lindau ◽

J M Fernandez

Keyword(s):

Mast Cells ◽

Patch Clamp ◽

Single Channel ◽

Resting Potential ◽

Dependent Manner ◽

Patch Clamp Technique ◽

Whole Cell ◽

Peritoneal Mast Cells ◽

Ionic Conductances ◽

Rat Peritoneal Mast Cells

The ionic conductances in rat basophilic leukemia cells (RBL-2H3) and rat peritoneal mast cells were investigated using the patch-clamp technique. These two cell types were found to have different electrophysiological properties in the resting state. The only significant conductance of RBL-2H3 cells was a K+-selective inward rectifier. The single channel conductance at room temperature increased from 2-3 pS at 2.8 mM external K+ to 26 pS at 130 mM K+. This conductance, which appeared to determine the resting potential, could be blocked by Na+ and Ba2+ in a voltage-dependent manner. Rat peritoneal mast cells had a whole-cell conductance of only 10-30 pS, and the resting potential was close to zero. Sometimes discrete openings of channels were observed in the whole-cell configuration. When the Ca2+ concentration on the cytoplasmic side of the membrane was elevated, two types of channels with poor ion specificity appeared. A cation channel, observed at a Ca2+ concentration of approximately 1 microM, had a unit conductance of 30 pS. The other channel, activated at several hundred micromolar Ca2+, was anion selective and had a unit conductance of approximately 380 pS in normal Ringer solution and a bell-shaped voltage dependence. Antigenic stimulation did not cause significant changes in the ionic conductances in either cell type, which suggests that these cells use a mechanism different from ionic currents in stimulus-secretion coupling.

Download Full-text

PT pretreatment inhibits 48/80-induced activation of Ca(+)-permeable channels in rat peritoneal mast cells

AJP Cell Physiology ◽

10.1152/ajpcell.1990.259.5.c715 ◽

1990 ◽

Vol 259 (5) ◽

pp. C715-C722 ◽

Cited By ~ 19

Author(s):

M. Kuno ◽

J. Kawaguchi ◽

M. Mukai ◽

F. Nakamura

Keyword(s):

Mast Cells ◽

Patch Clamp ◽

Room Temperature ◽

Plasma Membranes ◽

Patch Clamp Technique ◽

Permeable Channel ◽

Peritoneal Mast Cells ◽

Fluorescence Change ◽

Stimulus Secretion Coupling ◽

Rat Peritoneal Mast Cells

We recently reported that the secretagogue, compound 48/80, activated Ca2(+)-permeable channels of mast cells possibly via a second messenger [Kuno, Okada, and Shibata. Am. J. Physiol. 256 (Cell Physiol. 25): C560-C568, 1989]. The effects of pretreatment with pertussis toxin (PT) on compound 48/80 (48/80)-induced activation of the Ca2(+)-permeable channel have now been investigated by measuring Ca2+ signals of cell suspensions using the Ca2+ indicator fura-2 and by recording Ba2+ currents through the channel using the patch-clamp technique. In the presence of extracellular Ca2+, the fluorescence change was biphasic, with an immediate rise and a delayed peak at room temperature. The delayed peak, mainly due to Ca2+ entry through plasma membranes, was greatly reduced by pretreatment with PT. The quenching of the fluorescence by 48/80-induced Mn2+ influx was also decreased by PT, whereas the Ca2+ transients due to Ca2+ release from the intracellular stores apparently did not change. In patch-clamp recordings from cell-attached patches with pipettes containing isotonic Ba2+, the 48/80-induced Ba2+ currents were either suppressed or delayed in the PT-treated cells, under conditions where degranulation was absent. These results suggest that PT-sensitive GTP-binding protein is involved in activating the Ca2(+)-permeable channel in mast cells during stimulus-secretion coupling.

Download Full-text