Functional coupling of rat metabotropic glutamate 1a receptors to phospholipase D in CHO cells: involvement of extracellular Ca2+, protein kinase C, tyrosine kinase and Rho-A.
We report here that metabotropic glutamate 1a (mGlu1a) receptors, stably expressed in CHO cells, stimulate phospholipase D (PLD) activity. Several mGlu receptor agonists were found to exert this effect, with a rank order of potency of: L-quisqualate>L-glutamate>(1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid [(1S,3R)-ACPD]=(S)-3,5-dihydroxyphenylglycine [(S)-DHPG]. Both L-glutamate- and (1S,3R)-ACPD-stimulated PLD activity were attenuated by the selective mGlu receptor antagonist (S)-alpha-methyl-4-carboxyphenylglycine. mGlu1a receptor-stimulated PLD was inhibited either by the selective protein kinase C (PKC) inhibitor, GF109203X, or via PKC downregulation. MGlu1a receptor-PLD coupling required extracellular Ca2+ and was sensitive to La3+ and Zn2+, inhibitors of intracellular Ca2+ store-operated Ca2+ influx. mGlu1a receptor-PLD coupling was inhibited by the selective tyrosine kinase inhibitor, genistein. In addition, mGlu1a receptor-PLD coupling was also inhibited by cell transfection with the selective Rho (small GTP-binding protein) inhibitors: C3-exoenzyme and dominant negative mutant RhoA constructs. Brefeldin A, a selective ADP-ribosylation factor (ARF) inhibitor, and a dominant negative ARF6 mutant, failed to significantly influence mGlu1a receptor-stimulated PLD activity. We conclude that mGlu1a receptors activate PLD via a mechanism that is dependent on extracellular Ca2+, PKC, tyrosine kinase and RhoA but independent of ARF.
Kanumilli
S
S
Department of Pharmacology, School of Medical Sciences, University of Bristol, BS8 1TD, Bristol, UK.
Toms
N J
NJ
Venkateswarlu
K
K
Mellor
H
H
Roberts
P J
PJ
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D006224:Cricetinae; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005110:Extracellular Space / Q000187:drug effects / Q000201:enzymology; D019204:GTP-Binding Proteins / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000378:metabolism*; D014162:Transfection; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D018691:Excitatory Amino Acid Antagonists; D018094:Receptors, Metabotropic Glutamate; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D010739:Phospholipase D; D019204:GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D002118:Calcium