Signal transduction pathways mediating CCK-induced gallbladder muscle contraction.
The signal transduction that mediates CCK-induced contraction of gallbladder muscle was investigated in the cat. Contraction was measured by scanning micrometry in single muscle cells isolated enzymatically with collagenase. Production of D-myo-inositol 1,4, 5-trisphosphate (IP3) and sn-1,2-diacylglycerol (DAG) was quantitated using HPLC and TLC, respectively. Protein kinase C (PKC) activity was determined by measuring the phosphorylation of a specific substrate peptide from myelin basic protein, Ac-MBP-(4-14). CCK-induced contraction was blocked by incubation in strontium medium, pertussis toxin (PTx), and antibodies against Gialpha3 or betagamma-subunits but was not blocked by Ca2+-free medium or by antibodies against Gq/11alpha, Gialpha1-2, or Goalpha. The contraction induced by CCK was inhibited by the phospholipase C (PLC) inhibitor U-73122, anti-PLC-beta3 antibody, and the IP3 receptor antagonist heparin but was not inhibited by the the phospholipase D inhibitor propranolol or antibodies against PLC-beta1 or PLC-beta2. Western blot analysis of gallbladder muscle revealed the presence of PLC-beta2 and PLC-beta3 but not PLC-beta1. CCK caused a 94% increase in IP3 generation and an 86% increase in DAG generation. A low dose of CCK caused PKC translocation, and CCK-induced contraction was blocked by the PKC inhibitor H-7. A high dose of CCK, however, caused no PKC translocation, and its contraction was blocked by the calmodulin antagonist CGS9343B. In conclusion, CCK contracts cat gallbladder muscle by stimulating PTx-sensitive Gi 3 protein coupled with PLC-beta3, producing IP3 and DAG. Low doses activate PKC, whereas high doses activate calmodulin.
Yu
P
P
Department of Medicine, Rhode Island Hospital, and Brown University School of Medicine, Providence, Rhode Island 02903, USA.
Chen
Q
Q
Xiao
Z
Z
Harnett
K
K
Biancani
P
P
Behar
J
J
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D000818:Animals; D001562:Benzimidazoles / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002147:Calmodulin / Q000037:antagonists & inhibitors; D002415:Cats; D002462:Cell Membrane / Q000378:metabolism; D002766:Cholecystokinin / Q000494:pharmacology*; D004075:Diglycerides / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005704:Gallbladder / Q000187:drug effects / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology*; D037342:Pertussis Toxin; D054799:Phospholipase C beta; D011189:Potassium Chloride / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D010738:Type C Phospholipases / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D001562:Benzimidazoles; D002147:Calmodulin; D004075:Diglycerides; D007527:Isoenzymes; D010566:Virulence Factors, Bordetella; C052087:CGS 9343B; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004533:Egtazic Acid; D011189:Potassium Chloride; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D015544:Inositol 1,4,5-Trisphosphate; D002766:Cholecystokinin; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D054799:Phospholipase C beta; D019204:GTP-Binding Proteins; D002118:Calcium