Inflammation and inflammatory agents activate protein kinase C epsilon translocation and excite guinea-pig submucosal neurons.
OBJECTIVE
Properties of enteric neurons are transformed by inflammation and protein kinase C (PKC) isoforms are involved both in long-term changes in enteric neurons, and in transducing the effects of substances released during inflammation. We investigated roles of PKCepsilon in submucosal neurons by studying translocation in response to inflammatory mediators, effects on neuron excitability, and the changes in PKCepsilon distribution in a trinitrobenzene sulphonate model of ileitis.


METHODS
Immunohistochemical detection and analysis of association with membrane and cytosolic fractions, and Western blot analysis of cytosolic and particulate fractions were used to quantify translocation. Electrophysiology methods were used to measure effects on neuron excitability.


RESULTS
All submucosal neurons were immunoreactive for the novel PKC, PKCepsilon, and direct PKC activators, phorbol 12,13-dibutyrate, ingenol 3,20-dibenzoate, and the PKCepsilon-specific activator, transactivator of transduction-Psiepsilon receptor for activated C kinase, all caused PKCepsilon translocation from cytoplasm to surfaces of the neurons. Electrophysiologic studies showed that the stimulant of novel PKCs, ingenol (1 micromol/L), increased excitability of all neurons. Stimulation of protease-activated receptors caused PKCepsilon translocation selectively in vasoactive intestinal peptide secretomotor neurons, whereas a neurokinin 3 tachykinin receptor agonist caused translocation in neuropeptide Y and calretinin neurons. In all cases translocation was reduced significantly by a PKCepsilon-specific translocation inhibitor peptide. Increased PKCepsilon at the plasma membrane occurred in all neurons 6-7 days after an inflammatory stimulus.


CONCLUSIONS
Major targets for PKCepsilon include ion channels near the plasma membrane. PKCepsilon is likely to have a significant role in controlling the excitability of submucosal neurons and is probably an intermediate in causing hyperexcitability after inflammation.
Poole
Daniel P
DP
Department of Anatomy and Cell Biology, Centre for Neuroscience, University of Melbourne, Parkville, Victoria, Australia.
Matsuyama
Hayato
H
Nguyen
Trung V
TV
Eriksson
Emily M Y
EM
Fowler
Christopher J
CJ
Furness
John B
JB
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D015153:Blotting, Western; D064032:Calbindin 2; D002462:Cell Membrane / Q000201:enzymology / Q000378:metabolism; D003593:Cytoplasm / Q000201:enzymology / Q000378:metabolism; D004195:Disease Models, Animal; D004224:Diterpenes / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D006168:Guinea Pigs; D007079:Ileitis / Q000139:chemically induced / Q000201:enzymology / Q000378:metabolism*; D007082:Ileum / Q000201:enzymology / Q000294:innervation / Q000378:metabolism*; D066298:In Vitro Techniques; D018836:Inflammation Mediators / Q000378:metabolism*; D007700:Kinetics; D009478:Neuropeptide Y / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D051744:Protein Kinase C-epsilon / Q000378:metabolism*; D021381:Protein Transport; D044464:Receptor, PAR-2 / Q000819:agonists / Q000378:metabolism; D018042:Receptors, Neurokinin-3 / Q000819:agonists / Q000378:metabolism; D064030:S100 Calcium Binding Protein G / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013368:Submucous Plexus / Q000201:enzymology / Q000378:metabolism*; D013373:Substance P / Q000031:analogs & derivatives / Q000494:pharmacology; D014302:Trinitrobenzenesulfonic Acid; D014357:Trypsin / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000187:drug effects
D064032:Calbindin 2; D004224:Diterpenes; D020536:Enzyme Activators; C477271:His-Asp-Ala-Pro-Ile-Gly-Tyr-Asp; D018836:Inflammation Mediators; D009478:Neuropeptide Y; D009842:Oligopeptides; D010446:Peptide Fragments; D044464:Receptor, PAR-2; D018042:Receptors, Neurokinin-3; D064030:S100 Calcium Binding Protein G; C054724:senktide; D013373:Substance P; D014660:Vasoactive Intestinal Peptide; D015240:Phorbol 12,13-Dibutyrate; C011875:ingenol dibenzoate; D014302:Trinitrobenzenesulfonic Acid; D051744:Protein Kinase C-epsilon; D014357:Trypsin