scholarly journals Neomycin is a potent secretagogue of mast cells that directly activates a GTP-binding protein involved in exocytosis.

1990 ◽  
Vol 111 (6) ◽  
pp. 2885-2891 ◽  
Author(s):  
M Aridor ◽  
R Sagi-Eisenberg
Keyword(s):  
Mast Cells ◽  
Phosphatidic Acid ◽  
Rat Brain ◽  
G Protein ◽  
Gtp Binding Protein ◽  
Histamine Secretion ◽  
Gtp Binding ◽  
Rat Brain Membranes ◽  
Gamma S ◽  
Dose Dependent

When loaded alongside GTP-gamma-S into ATP-permeabilized cells, neomycin, at concentrations below 1 mM, inhibits GTP-gamma-S-induced histamine secretion and phosphatidic acid formation (Cockcroft, S., and B. D. Gomperts, 1985. Nature (Lond.). 314: 534-536; Aridor, M., L. M. Traub, and R. Sagi-Eisenberg. 1990. J. Cell Biol. 111:909-917). However, at higher concentrations internally applied neomycin induces histamine secretion in a process that is: (a) dose dependent; (b) dependent on the internal application of GTP; (c) independent of phosphoinositide breakdown; and (d) inhibited by pertussis toxin (PtX) treatment. These results indicate that neomycin can stimulate histamine secretion in a mechanism that bypasses phospholipase C (PLC) activation and yet involves a PtX-sensitive GTP-binding protein (G protein). Unlike its dual effects, when internally applied, neomycin induces histamine secretion from intact mast cells in a dose-dependent manner. Half-maximal and maximal effects are obtained at 0.5 and 1 mM neomycin, respectively. This process is rapid (approximately 30 s), is independent of external Ca2+, and is associated with phosphatidic acid formation, implying that neomycin can activate histamine secretion by a mechanism similar to that utilized by other basic secretagogues of mast cells. Neomycin stimulates fourfold the GTPase activity of cholate-solubilized rat brain membranes in a PtX-inhibitable manner. In addition neomycin, as well as the basic secretagogues of mast cells, compound 48/80, and mastoparan, significantly reduce (by approximately 80%) the ADP ribosylation of PtX substrates present in rat brain membranes. Taken together these data suggest that neomycin can stimulate secretion from mast cells by directly activating G proteins that play a role in stimulus-secretion coupling. When internally applied, neomycin presumably stimulates secretion by activating a G protein that is located downstream to PLC. This G protein serves as a substrate for PtX.

scholarly journals Exocytosis in mast cells by basic secretagogues: evidence for direct activation of GTP-binding proteins.

1990 ◽  
Vol 111 (3) ◽  
pp. 909-917 ◽  
Author(s):  
M Aridor ◽  
L M Traub ◽  
R Sagi-Eisenberg
Keyword(s):  
Mast Cells ◽  
G Protein ◽  
G Proteins ◽  
Independent Manner ◽  
Histamine Secretion ◽  
Free System ◽  
Stimulatory G Protein ◽  
A Cell ◽  
Secretion Inhibition ◽  
Gamma S

Histamine release induced by the introduction of a nonhydrolyzable analogue of GTP, GTP-gamma-S, into ATP-permeabilized mast cells, is associated with phosphoinositide breakdown, as evidenced by the production of phosphatidic acid (PA) in a neomycin-sensitive process. The dependency of both PA formation and histamine secretion on GTP-gamma-S concentrations is bell shaped. Whereas concentrations of up to 0.1 mM GTP-gamma-S stimulate both processes, at higher concentrations the cells' responsiveness is inhibited. At a concentration of 1 mM, GTP-gamma-S self-inhibits both PA formation and histamine secretion. Inhibition of secretion can, however, be overcome by the basic secretagogues compound 48/80 and mastoparan that in suboptimal doses synergize with 1 mM GTP-gamma-S to potentiate secretion. Secretion under these conditions is not accompanied by PA formation and is resistant both to depletion of Ca2+ from internal stores and to pertussis toxin (PtX) treatment. In addition, 48/80, like mastoparan, is capable of directly stimulating the GTPase activity of G-proteins in a cell-free system. Together, our results are consistent with a model in which the continuous activation of a phosphoinositide-hydrolyzing phospholipase C (PLC) by a stimulatory G-protein suffices to trigger histamine secretion. Basic secretagogues of mast cells, such as compound 48/80 and mastoparan, are capable of inducing secretion in a mechanism that bypasses PLC by directly activating a G-protein that is presumably located downstream from PLC (GE). Thereby, these secretagogues induce histamine secretion in a receptor-independent manner.

Secretion from permeabilised mast cells is enhanced by addition of gelsolin: contrasting effects of endogenous gelsolin

1995 ◽  
Vol 108 (2) ◽  
pp. 657-666 ◽  
Author(s):  
Y.S. Borovikov ◽  
J.C. Norman ◽  
L.S. Price ◽  
A. Weeds ◽  
A. Koffer
Keyword(s):  
Mast Cells ◽  
Resting State ◽  
Actin Filaments ◽  
Cortical Actin ◽  
Gtp Binding Protein ◽  
Independent Manner ◽  
Calcium Dependent ◽  
Gtp Binding ◽  
Gamma S ◽  
Rat Mast Cells

Permeabilised rat mast cells were exposed to gelsolin and its N-terminal half (S1-3), proteins that sever actin filaments in a calcium-dependent and independent manner, respectively. Gelsolin and S1-3 induced a decrease in cellular F-actin content and an increase in the extent of the secretory response. The calcium sensitivities of both these effects were consistent with the differential calcium requirements of the two proteins. Segment 1 (S1), which binds G-actin and caps filaments but does not sever them, did not show these effects. Thus, secretion of mast cells is promoted as a consequence of the severing activity of exogenous gelsolin or S1-3. Most of the endogenous gelsolin remained within permeabilised, washed mast cells and its distribution in resting state was predominantly cortical. Addition of calcium in the absence of MgATP did not reduce the F-actin content; by contrast, calcium with MgATP induced F-actin loss that was unaffected by the presence of anti-gelsolin. Because this antibody inhibits the severing activity of gelsolin, these results indicate that in permeabilised mast cells the severing activity of the remaining endogenous gelsolin is not involved in cortical actin filaments disassembly. Upon exposure to GTP-gamma-S in the absence of calcium, the content of cortical gelsolin was reduced. This parallels our previous observation of a GTP-gamma-S induced reduction of cortical actin filaments followed by their relocation to the cell's interior (Norman et al. (1994) J. Cell Biol. 126, 1005–1015) and suggests that actin redistribution may be a consequence of dissociation of gelsolin caps brought about by activation of a GTP-binding protein.

scholarly journals Coupling of 5-HT1A receptor with GTP-binding protein in rat brain membranes

1988 ◽  
Vol 46 ◽  
pp. 251
Author(s):  
Yoshihisa Kitamura ◽  
Shin-ichi Imai ◽  
Yasuyuki Nomura
Keyword(s):  
Rat Brain ◽  
Binding Protein ◽  
Gtp Binding Protein ◽  
Brain Membranes ◽  
Gtp Binding ◽  
Rat Brain Membranes

scholarly journals Mechanism of acetylcholine-induced inhibition of Ca current in bullfrog atrial myocytes.

1990 ◽  
Vol 96 (4) ◽  
pp. 865-885 ◽  
Author(s):  
T Nakajima ◽  
S Wu ◽  
H Irisawa ◽  
W Giles
Keyword(s):  
Adenylate Cyclase ◽  
Binding Protein ◽  
Cell Voltage ◽  
Inhibitory Effect ◽  
Atrial Myocytes ◽  
Ca Current ◽  
Gtp Binding Protein ◽  
Gtp Binding ◽  
Subsequent Application ◽  
Gamma S

The mechanism of the anti-beta-adrenergic action of acetylcholine (ACh) on Ca current, ICa, was examined using the tight-seal, whole-cell voltage clamp technique in single atrial myocytes from the bullfrog. Both isoproterenol (ISO) and forskolin increased ICa dose dependently. After ICa had been enhanced maximally by ISO (10(-6) M), subsequent application of forskolin (50 microM) did not further increase ICa, suggesting that ISO and forskolin increase ICa via a common biochemical pathway, possibly by stimulation of adenylate cyclase. ACh (10(-5) M) completely inhibited the effect of low doses of forskolin (2 x 10(-6) M), as well as ISO, but it failed to block the effects of high doses of forskolin (greater than 5 x 10(-5) M). Intracellular application of cyclic AMP (cAMP) also increased ICa. ACh (10(-5) M) failed to inhibit this cAMP effect, indicating that the inhibitory action of ACh occurs at a site proximal to the production of cAMP. ACh (10(-5) M) also activated an inwardly rectifying K+ current IK(ACh). Intracellular application of a nonhydrolyzable GTP analogue, GTP gamma S (5 X 10(-4) M), activated IK(ACh) within several minutes; subsequent application of ACh (10(-5) M) did not increase IK(ACh) further. These results demonstrate that a GTP-binding protein coupled to these K+ channels can be activated maximally by GTP gamma S even in the absence of ACh. Intracellular application of GTP gamma S also strongly inhibited the effect of ISO on ICa in the absence of ACh. Pertussis toxin (IAP) completely prevented both the inhibitory effect of ACh on ICa and the ACh-induced activation of IK(ACh). GTP gamma S (50 microM-1 mM) alone did not increase ICa significantly; however, when ISO was applied first, GTP gamma S (5 x 10(-4) M) gradually inhibited the ISO effect on ICa. These results indicate that ACh antagonizes the effect of ISO on ICa via a GTP-binding protein (Gi and/or Go). This effect may be mediated through a direct inhibition by the alpha-subunit of Gi which is coupled to the adenylate cyclase.

Involvement of a GTP-binding protein in mediation of serotonin and acetylcholine responses in Xenopus oocytes injected with rat brain messenger RNA

1986 ◽  
Vol 1 (3) ◽  
pp. 201-209 ◽  
Author(s):  
Nathan Dascal ◽  
Catherine Ifune ◽  
Rosemary Hopkins ◽  
Terry P. Snutch ◽  
Hermann Lübbert ◽  
...  
Keyword(s):  
Rat Brain ◽  
Messenger Rna ◽  
Xenopus Oocytes ◽  
Binding Protein ◽  
Gtp Binding Protein ◽  
Gtp Binding

scholarly journals Persistent activation of transducin by bleached rhodopsin in salamander rods.

1996 ◽  
Vol 108 (6) ◽  
pp. 557-563 ◽  
Author(s):  
H R Matthews ◽  
M C Cornwall ◽  
G L Fain
Keyword(s):  
Phosphodiesterase Inhibitor ◽  
Ambystoma Tigrinum ◽  
Gtp Binding Protein ◽  
Whole Cell ◽  
Persistent Excitation ◽  
Current Decay ◽  
Circulating Current ◽  
Gtp Binding ◽  
Patch Pipette ◽  
Gamma S

The hydrolysis-resistant GTP analogue GTP-gamma-S was introduced into rods isolated from the retina of the salamander Ambystoma tigrinum to study the origin of the persistent excitation induced by intense bleaching illumination. Dialysis of a dark-adapted rod with a whole-cell patch pipette containing 2 mM GTP-gamma-S resulted in a gradual decrease in circulating current. If the rod was first bleached and its sensitivity allowed to stabilize for at least 30 min, then dialysis with GTP-gamma-S produced a much faster current decay. The circulating current could be restored by superfusion with the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine, suggesting that the decay in current originated from persistent excitation of the phosphodiesterase by transducin bound to GTP-gamma-S. We conclude that the persistent excitation which follows bleaching is likely to involve the GTP-binding protein transducin, which mediates the normal photoresponse. This observation suggests that a form of rhodopsin which persists long after bleaching can activate transducin much as does photoisomerized rhodopsin, although with considerably lower gain.

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