title: Signal transduction pathways mediating CCK-8S-induced gastric antral smooth muscle contraction.
abstract: OBJECTIVE
  To investigate the functional and molecular mechanisms by which sulfated cholecystokinin octapeptide (CCK-8S) regulates calcium mobilization in gastric antral smooth muscle cells (SMCs) of rats.
  
  
  METHODS
  Isotonic contraction of antral strips was recorded using a polyphysiograph. Immunoprecipitation was used to determine the regulatory effect of protein kinase C (PKC) on regulating the phosphorylation of the type III inositol 1,4,5-triphosphate receptor (InsP(3)R3) in gastric SMCs. Alterations in the intracellular calcium ([Ca(2+)](i)) concentration were assayed using fura-2/AM-loaded microspectrofluorometry, and the L-type calcium current (I(Ca-L)) was recorded by patch-clamp techniques.
  
  
  RESULTS
  CCK-8S (5 x 10(-8) mol/l) significantly increased the mean contractile amplitude of circular muscle by 61.85 +/- 12.67% and the frequency of longitudinal muscle by 57.91 +/- 15.70% in gastric antral strips, which were suppressed by dexloxiglumide or thapsigargin (TG) and BAPTA-AM (BA). Treatment with chelerythrine (5 x 10(-8) mmol/l) significantly inhibited the CCK-8S-increased phosphorylation of InsP(3)R3 in SMCs. The amplitudes of the CCK-8S-triggered [Ca(2+)](i) concentration oscillations were reduced in a dose-dependent manner when the SMCs were pretreated with increasing concentrations of PMA (from 10(-8) to 10(-5) mol/l). On removal of extracellular calcium or blocking I(Ca-L) by nifedipine, a smaller but significant rise in the [Ca(2+)](i) concentration was still elicited by CCK-8S. When [Ca(2+)](i) was depleted by the administration of 10(-5) mol/l TG and 10(-5) mol/l BA or blocked by the calcium-dependent chloride current (I(Cl-Ca)) by giving 5 x 10(-6) mol/l niflumic acid, the CCK-8S-intensified I(Ca-L) (from -56.42 +/- 6.57 to -88.54 +/- 5.71 pA) was apparently inhibited by 90.34 +/- 4.71% and 82.59 +/- 4.24%.
  
  
  CONCLUSIONS
  These results demonstrate that the CCK-8S-evoked [Ca(2+)](i) concentration increase in gastric antral SMCs depends on the release of [Ca(2+)](i) stores which are negatively regulated by PKC-mediated phosphorylation of InsP(3)R3. Released calcium in turn activates I(Ca-L) through the activation of I(Cl-Ca), ultimately resulting in the contraction of the gastric smooth muscle.
authors.affiliation: Department of Digestive Medicine, Renmin Hospital, Wuhan University, 238 Jiefang Road, Hubei 430060, China.
authors.forename: Xin-Min
authors.initials: XM
authors.lastname: Si
authors.forename: Lei
authors.initials: L
authors.lastname: Huang
authors.forename: Shelley Chireyath
authors.initials: SC
authors.lastname: Paul
authors.forename: Ping
authors.initials: P
authors.lastname: An
authors.forename: He-Sheng
authors.initials: HS
authors.lastname: Luo
mesh_terms: D000818:Animals; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000378:metabolism; D005260:Female; D005746:Gastric Emptying / Q000502:physiology*; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism / Q000503:physiopathology*; D011706:Pyloric Antrum / Q000187:drug effects / Q000378:metabolism / Q000503:physiopathology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011949:Receptors, Cholecystokinin / Q000819:agonists*; D015398:Signal Transduction / Q000187:drug effects*; D012844:Sincalide / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: C090019:8-sulfocholecystokinin octapeptide; D020746:Calcium Channels, L-Type; D011949:Receptors, Cholecystokinin; D012844:Sincalide