Spacial compartmentalization of Ca2+ signaling complexes in pancreatic acini.
Imaging [Ca2+]i at high temporal resolution and measuring the properties of Ca2+ signaling in streptolysin O (SLO)-permeabilized cells were used to study the spacial organization of signaling complexes. Sequential stimulation of single cells within pancreatic acini with several Ca2+-mobilizing agonists revealed an agonist-specific pattern and propagation rate of Ca2+ waves in the same cells, with CCK8 stimulating the fastest and bombesin the slowest waves. More importantly, each agonist initiated the wave in a different region of the same cell. On the other hand, repetitive stimulation with the same agonist induced Ca2+ waves of the same pattern that were initiated from the same region of the cell. The agonist-specific Ca2+ signaling does not appear to be the result of coupling to different G proteins as infusion of an anti-Galphaq antibody into the cells through a patch pipette equally inhibited Ca2+ signaling by all agonists. Further evidence for compartmentalization of signaling complexes was developed in permeabilized cells. The time-dependent loss of Ca2+ signaling due to SLO permeabilization occurred in an agonist-specific manner in the sequence cabachol > bombesin > cholecystokinin. Signaling by all agonists could be completely restored with as low as 2 micro guanosine 5'-3-O-(thio)triphosphate (GTPgammaS). At this low concentration GTPgammaS recoupled inositol 1,4,5-trisphosphate production and Ca2+ release, rather than enhancing phospholipase C activity. Priming of Ca2+ signaling by GTPgammaS was agonist-specific. Guanosine 5'-O-(thio)diphosphate (GDPbetaS) uncoupled the ability of signaling complexes to release Ca2+ much better than stimulating inositol 1,4,5-trisphosphate production. The uncoupling of Ca2+ signaling by GDPbetaS was also agonist-specific. The combined findings of agonist-specific initiation sites of the Ca2+ wave and differential access of guanine nucleotides to signaling complexes suggest spacial compartmentalization of Ca2+ signaling complexes. Each complex must include a receptor, G protein, and phospholipase C that are coupled to a specific portion of the Ca2+ pool.
Xu
X
X
Department of Physiology, University of Texas Southwestern Medical Center, Dallas, Texas 75235, USA.
Zeng
W
W
Diaz
J
J
Muallem
S
S
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D001839:Bombesin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002451:Cell Compartmentation; D002766:Cholecystokinin / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006150:Guanine Nucleotides / Q000378:metabolism; D010179:Pancreas / Q000187:drug effects / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D015398:Signal Transduction
D006150:Guanine Nucleotides; D002766:Cholecystokinin; D019204:GTP-Binding Proteins; D000109:Acetylcholine; D001839:Bombesin; D002118:Calcium