title: Protein kinase C-alpha regulation of gallbladder Na+ transport becomes progressively more dysfunctional during gallstone formation.
abstract: Gallbladder Na+ absorption and biliary Ca2+ are both increased during gallstone formation and may promote cholesterol nucleation. Na+/H+ exchange (NHE) is a major pathway for gallbladder Na+ transport. Ca2+-dependent second messengers, including protein kinase C (PKC), inhibit basal gallbladder Na+ transport. Multiple PKC isoforms with species- and tissue-specific expression have been reported. In this study we sought to characterize Ca2+-dependent PKC isoforms in gallbladder and to examine their roles in Na+ transport during gallstone formation. Gallbladders were harvested from prairie dogs fed either nonlithogenic chow or 1.2% cholesterol-enriched diet for varying periods to induce various stages of gallstone formation. PKC was activated with the use of phorboldibutyrate, and we assessed gallbladder NHE regulation by measuring unidirectional Na+ flux and dimethylamiloride-inhibitable 22Na+ uptake. We measured gallbladder PKC activity with the use of histone III-S phosphorylation and used Gö 6976 to determine PKC-alpha contributions. Gallbladder PKC isoform messenger RNA and protein expression were examined with the use of Northern- and Western-blot analysis, respectively. Prairie dog and human gallbladder expresses PKC-alpha, betaII, and delta isoforms. The PKC activation significantly decreased gallbladder J(Na)(ms) and reduced baseline 22Na+ uptake by inhibiting NHE. PKC-alpha mediated roughly 42% of total PKC activity under basal conditions. PKC-alpha regulates basal gallbladder Na+ transport by way of stimulation of NHE isoform NHE-2 and inhibition of isoform NHE-3. PKC-alpha blockade reversed PKC-induced inhibition of J(Na)(ms) and 22Na+ uptake by about 45% in controls but was progressively less effective during gallstone formation. PKC-alpha contribution to total PKC activity is progressively reduced, whereas expression of PKC-alpha mRNA, and protein increases significantly during gallstone formation. We conclude that PKC-alpha regulation of gallbladder NHE becomes progressively more dysfunctional and may in part account for the increased Na+ absorption observed during gallstone formation.
authors.affiliation: Department of Surgery, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102-1192, USA.
authors.forename: Seth C
authors.initials: SC
authors.lastname: Narins
authors.forename: Ramugounder
authors.initials: R
authors.lastname: Ramakrishnan
authors.forename: Eun H
authors.initials: EH
authors.lastname: Park
authors.forename: Paul B
authors.initials: PB
authors.lastname: Bolno
authors.forename: David A
authors.initials: DA
authors.lastname: Haggerty
authors.forename: Peter R
authors.initials: PR
authors.lastname: Smith
authors.forename: William C
authors.initials: WC
authors.lastname: Meyers
authors.forename: Mohammad Z
authors.initials: MZ
authors.lastname: Abedin
mesh_terms: D000818:Animals; D004848:Epithelium / Q000502:physiology; D005704:Gallbladder / Q000201:enzymology*; D042882:Gallstones / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D008297:Male; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D012333:RNA, Messenger / Q000032:analysis; D012589:Sciuridae; D012964:Sodium / Q000378:metabolism*; D012979:Sodium Radioisotopes; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D012979:Sodium Radioisotopes; D017923:Sodium-Hydrogen Exchangers; D012964:Sodium; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha