Endothelin-1 enhances capsaicin-evoked intracellular Ca2+ response via activation of endothelin a receptor in a protein kinase Cepsilon-dependent manner in dorsal root ganglion neurons.
Increasing evidence indicates that endothelin-1 has a role for peripheral nociceptive signaling in animals and humans. However, the mechanisms of the nociceptive effects of endothelin-1 have not been fully understood. The current study investigated the effects of endothelin-1 on the capsaicin-evoked intracellular Ca2+ response of cultured adult mice dorsal root ganglion neurons. Dorsal root ganglia were harvested from adult male C57B6N mice and were cultured. With a digital image analysis system, we detected the [Ca2+]i image of cultured dorsal root ganglion cells after loading with Fura-2 acetoxymethyl. In addition, co-localization of protein kinase Cepsilon with transient receptor potential V1 and the translocation of protein kinase Cepsilon were investigated using immunohistochemical methods. Endothelin-1 (10 nM) enhanced an increase in [Ca2+]i by capsaicin (10 nM) from 87.6+/-11.6 nM to 414.8+/-62.3 nM (71 of 156 neurons). The inhibition of endothelin A receptor (BQ-123) significantly suppressed the enhancing effect of endothelin-1. In addition, a nonselective protein kinase C inhibitor (bisindolylmaleimide I) significantly suppressed the enhancing effect of endothelin-1. A myristoyl-tagged membrane-permeant-protein kinase Cepsilon V1-2 inhibitory peptide also significantly suppressed the enhancing effect of endothelin-1. In the immunocytochemical study, protein kinase Cepsilon immunoreactivity was found in most of transient receptor potential V1-positive neurons. After endothelin-1 application, protein kinase Cepsilon immunoreactivity was observed to be translocated from the cytosol to the cell membrane in transient receptor potential V1-positive neurons. Our results indicate that endothelin-1 enhances the response of dorsal root ganglion neurons to capsaicin in a protein kinase Cepsilon-dependent manner. Our findings may lead to a new strategy to treat pain associated with endothelin-1.
Yamamoto
H
H
Department of Anesthesiology, Sapporo Medical University School of Medicine, South 1, West 16, Chuo-ku, Sapporo, Hokkaido 060-8543, Japan.
Kawamata
T
T
Ninomiya
T
T
Omote
K
K
Namiki
A
A
D000465:Algorithms; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002211:Capsaicin / Q000494:pharmacology*; D002478:Cells, Cultured; D003627:Data Interpretation, Statistical; D065130:Endothelin A Receptor Antagonists; D019332:Endothelin-1 / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005727:Ganglia, Spinal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009619:Nociceptors / Q000187:drug effects; D051744:Protein Kinase C-epsilon / Q000037:antagonists & inhibitors / Q000502:physiology*; D044022:Receptor, Endothelin A / Q000378:metabolism*; D012996:Solutions
D065130:Endothelin A Receptor Antagonists; D019332:Endothelin-1; D004791:Enzyme Inhibitors; D044022:Receptor, Endothelin A; D012996:Solutions; D051744:Protein Kinase C-epsilon; D002211:Capsaicin; D002118:Calcium