Protein kinase C-mediated Ca2+ entry in HEK 293 cells transiently expressing human TRPV4

Br J Pharmacol. 2003 Sep;140(2):413-21. doi: 10.1038/sj.bjp.0705443. Epub 2003 Aug 11.

Abstract

1. We investigated whether protein kinase C (PKC) activation stimulates Ca2+ entry in HEK 293 cells transfected with human TRPV4 cDNA and loaded with fura-2. 2. Phorbol 12-myristate 13-acetate (PMA), a PKC-activating phorbol ester, increased the intracellular Ca2+ concentration ([Ca2+]i) in a dose-dependent manner, with an EC50 value of 11.7 nm. Exposure to a hypotonic solution (HTS) after PMA further increased [Ca2+]i. Two other PKC-activating phorbol esters, phorbol 12,13-didecanoate (PDD) and phorbol 12,13-dibutyrate, also caused [Ca2+]i to increase. 3. The inactive isomer 4alpha-PMA was less effective and the peak [Ca2+]i increase was significantly smaller than that induced by PMA. In contrast, 4alpha-PDD produced a monophasic or biphasic [Ca2+]i increase with a different latency, while 4alpha-phorbol had no effect. 4. The PMA-induced [Ca2+]i increase was abolished by prior exposure to bisindolylmaleimide (BIM), a PKC-specific inhibitor, and suppressed by the nonspecific PKC inhibitor 1-(5-isoquinolinesulphonyl)-2-methylpiperazine. The [Ca2+]i increase induced by 4alpha-PMA, 4alpha-PDD or HTS was not significantly affected by BIM. 5. These results suggest that both PKC-dependent and -independent mechanisms are involved in the phorbol ester-induced activation of TRPV4, and the PKC-independent pathway is predominant in HTS-induced Ca2+ entry.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Calcium / metabolism*
  • Cation Transport Proteins / genetics
  • Cation Transport Proteins / metabolism*
  • Cell Line
  • Dose-Response Relationship, Drug
  • Enzyme Activation / drug effects
  • Enzyme Inhibitors / pharmacology
  • Gene Expression
  • Humans
  • Hypotonic Solutions / pharmacology
  • Indoles / pharmacology
  • Ion Channels / genetics
  • Ion Channels / metabolism*
  • Maleimides / pharmacology
  • Phorbol 12,13-Dibutyrate / pharmacology
  • Phorbol Esters / chemistry
  • Phorbol Esters / pharmacology
  • Protein Kinase C / antagonists & inhibitors
  • Protein Kinase C / metabolism*
  • TRPV Cation Channels
  • Tetradecanoylphorbol Acetate / chemistry
  • Tetradecanoylphorbol Acetate / pharmacology

Substances

  • Cation Transport Proteins
  • Enzyme Inhibitors
  • Hypotonic Solutions
  • Indoles
  • Ion Channels
  • Maleimides
  • Phorbol Esters
  • TRPV Cation Channels
  • TRPV4 protein, human
  • phorbol-12,13-didecanoate
  • Phorbol 12,13-Dibutyrate
  • Protein Kinase C
  • bisindolylmaleimide
  • Tetradecanoylphorbol Acetate
  • Calcium