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Defining protein kinase/phosphatase isoenzymic regulation of mGlu 5 receptor‐stimulated phospholipase C and Ca 2+ responses in astrocytes
Author(s) -
Bradley SJ,
Challiss RAJ
Publication year - 2011
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.2011.01421.x
Subject(s) - protein kinase c , phosphatase , biology , inositol phosphate , phospholipase c , receptor , kinase , signal transduction , microbiology and biotechnology , metabotropic receptor , inositol , biochemistry , phosphorylation , glutamate receptor
BACKGROUND AND PURPOSE Cyclical phosphorylation and dephosphorylation of a key residue within the C‐terminal domain of the activated type 5 metabotropic glutamate (mGlu 5 ) receptor is believed to cause the synchronous, oscillatory changes in inositol 1,4,5‐trisphosphate and Ca 2+ levels observed in a variety of cell types. Here, we have attempted to better define the kinase and phosphatase enzymes involved in this modulation. EXPERIMENTAL APPROACH Ca 2+ and [ 3 H]inositol phosphate ([ 3 H]IP x ) measurements in astrocyte preparations have been used to evaluate the effects of pharmacological inhibition of protein kinase C (PKC) and protein phosphatase activities and small interfering RNA‐mediated specific PKC isoenzymic knock‐down on mGlu 5 receptor signalling. KEY RESULTS Ca 2+ oscillation frequency or [ 3 H]IP x accumulation in astrocytes stimulated by mGlu 5 receptors, was concentration‐dependently decreased by protein phosphatase‐1/2A inhibition or by PKC activation. PKC inhibition also increased [ 3 H]IP x accumulation two‐ to threefold and changed the Ca 2+ response into a peak‐plateau response. However, selective inhibition of conventional PKC isoenzymes or preventing changes in [Ca 2+ ] i concentration by BAPTA‐AM loading was without effect on mGlu 5 receptor‐stimulated [ 3 H]IP x accumulation. Selective knock‐down of PKCδ was without effect on glutamate‐stimulated Ca 2+ responses; however, selective PKCε knock‐down in astrocytes changed Ca 2+ responses from oscillatory into peak‐plateau type. CONCLUSION AND IMPLICATIONS These data confirm the acute regulation of mGlu 5 receptor signalling by protein kinases and protein phosphatases and provide novel data pinpointing the isoenzymic dependence of this regulation in the native mGlu 5 receptor‐expressing rat cortical astrocyte. These data also highlight a potential alternative mechanism by which mGlu 5 receptor signalling might be therapeutically manipulated.