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This Article Full Text Full Text (PDF) All Versions of this Article: 294/5/G1245    most recent 00521.2007v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Poole, D. P. Articles by Furness, J. B. PubMed PubMed Citation Articles by Poole, D. P. Articles by Furness, J. B.

NEUROREGULATION AND MOTILITY

Stimulation of the neurokinin 3 receptor activates protein kinase C and protein kinase D in enteric neurons

¹Department of Anatomy and Cell Biology and Centre for Neuroscience, University of Melbourne, Parkville, Victoria, Australia; ²Departments of Surgery and Physiology, University of California, San Francisco, San Francisco, California; and ³Center for Ulcer Research and Education, Department of Medicine, University of California, Los Angeles, California

Submitted 9 November 2007 ; accepted in final form 22 February 2008

Tachykinins, acting through NK₃ receptors (NK₃R), contribute to excitatory transmission to intrinsic primary afferent neurons (IPANs) of the small intestine. Although this transmission is dependent on protein kinase C (PKC), its maintenance could depend on protein kinase D (PKD), a downstream target of PKC. Here we show that PKD1/2-immunoreactivity occurred exclusively in IPANs of the guinea pig ileum, demonstrated by double staining with the IPAN marker NeuN. PKC was also colocalized with PKD1/2 in IPANs. PKC and PKD1/2 trafficking was studied in enteric neurons within whole mounts of the ileal wall. In untreated preparations, PKC and PKD1/2 were cytosolic and no signal for activated (phosphorylated) PKD was detected. The NK₃R agonist senktide evoked a transient translocation of PKC and PKD1/2 from the cytosol to the plasma membrane and induced PKD1/2 phosphorylation at the plasma membrane. PKC translocation was maximal at 10 s and returned to the cytosol within 2 min. Phosphorylated-PKD1/2 was detected at the plasma membrane within 15 s and translocated to the cytosol by 2 min, where it remained active up to 30 min after NK₃R stimulation. PKD1/2 activation was reduced by a PKC inhibitor and prevented by NK₃R inhibition. NK₃R-mediated PKC and PKD activation was confirmed in HEK293 cells transiently expressing NK₃R and green fluorescent protein-tagged PKC, PKD1, PKD2, or PKD3. Senktide caused membrane translocation and activation of kinases within 30 s. After 15 min, phosphorylated PKD had returned to the cytosol. PKD activation was confirmed through Western blotting. Thus stimulation of NK₃R activates PKC and PKD in sequence, and sequential activation of these kinases may account for rapid and prolonged modulation of IPAN function.

tachykinins; primary afferent neurons