title: PKC delta-isoform translocation and enhancement of tonic contractions of gastrointestinal smooth muscle.
abstract: PKC is involved in mediating the tonic component of gastrointestinal smooth muscle contraction in response to stimulation by agonists for G protein-coupled receptors. Here, we present pharmacological and immunohistochemical evidence indicating that a member of the novel PKC isoforms, PKC-delta, is involved in maintaining muscarinic receptor-coupled tonic contractions of the guinea pig ileum. The tonic component of carbachol-evoked contractions was enhanced by an activator of conventional and novel PKCs, phorbol 12,13-dibutyrate (PDBu; 200 nM or 1 microM), and by an activator of novel PKCs, ingenol 3,20-dibenzoate (IDB; 100 or 500 nM). Enhancement was unaffected by concentrations of bisindolylmaleimide I (BIM-I; 22 nM) that block conventional PKCs or by a PKC-epsilon-specific inhibitor peptide but was attenuated by higher doses of BIM-I (2.2 microM). Relevant proteins were localized at a cellular and subcellular level using confocal analysis. Immunohistochemical staining of the ileum showed that PKC-delta was exclusively expressed in smooth muscles distributed throughout the layers of the gut wall. PKC-epsilon immunoreactivity was prominent in enteric neurons but was largely absent from smooth muscle of the muscularis externa. Treatment with PDBu, IDB, or carbachol resulted in a time- and concentration-dependent translocation of PKC-delta from the cytoplasm to filamentous structures within smooth muscle cells. These were parallel to, but distinct from, actin filaments. The translocation of PKC-delta in response to carbachol was significantly reduced by scopolamine or calphostin C. The present study indicates that the tonic carbachol-induced contraction of the guinea pig ileum is mediated through a novel PKC, probably PKC-delta.
authors.affiliation: Department of Anatomy and Cell Biology, University of Melbourne, Parkville, Victoria 3010, Australia. d.poole@unimelb.edu.au
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Poole
authors.forename: John B
authors.initials: JB
authors.lastname: Furness
mesh_terms: D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D019852:Diacylglycerol Kinase / Q000032:analysis / Q000378:metabolism; D004224:Diterpenes / Q000494:pharmacology; D005260:Female; D006168:Guinea Pigs; D007082:Ileum / Q000187:drug effects / Q000502:physiology*; D066298:In Vitro Techniques; D007211:Indoles / Q000494:pharmacology; D007527:Isoenzymes / Q000032:analysis / Q000378:metabolism; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D009281:Naphthalenes / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051744:Protein Kinase C-epsilon / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects; D012601:Scopolamine / Q000494:pharmacology; D013347:Subcellular Fractions / Q000737:chemistry / Q000378:metabolism; D000099293:Protein Kinase C-lambda
chemical_list: D002217:Carbachol; D019852:Diacylglycerol Kinase; D004224:Diterpenes; D007211:Indoles; D007527:Isoenzymes; D008301:Maleimides; D009281:Naphthalenes; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D047428:Protein Kinase Inhibitors; D012601:Scopolamine; D000099293:Protein Kinase C-lambda; C011875:ingenol dibenzoate; C119190:protein kinase C nu; C058819:calphostin C; C070515:bisindolylmaleimide I