Tyrosine kinases enhance the function of glycine receptors in rat hippocampal neurons and human alpha(1)beta glycine receptors.
Glycine receptors (GlyRs) are transmitter-gated channels that mediate fast inhibitory neurotransmission in the spinal cord and brain. The GlyR beta subunit contains a putative tyrosine phosphorylation site whose functional role has not been determined. To examine if protein tyrosine kinases (PTKs) regulate the function of GlyRs, we analysed whole-cell currents activated by applications of glycine to CA1 hippocampal neurons and spinal neurons. The role of a putative site for tyrosine phosphorylation at position 413 of the beta subunit was examined using site-directed mutagenesis and expression of recombinant (alpha(1)beta(Y413F)) receptors in human embryonic kidney (HEK 293) cells. Lavendustin A, an inhibitor of PTKs, depressed glycine-evoked currents (I(Gly)) in CA1 neurons and spinal neurons by 31 % and 40 %, respectively. In contrast, the intracellular application of the exogenous tyrosine kinase, cSrc, enhanced I(Gly) in CA1 neurons by 56 %. cSrc also accelerated GlyR desensitization and increased the potency of glycine 2-fold (control EC(50) = 143 microM; cSrc EC(50) = 74 microM). Exogenous cSrc, applied intracellularly, upregulated heteromeric alpha(1)beta receptors but not homomeric alpha(1) receptors. Substitution mutation of the tyrosine to phenylalanine at position beta-413 prevented this enhancement. Furthermore, a selective inhibitor of the Src family kinases, PP2, down-regulated wild-type alpha(1)beta but not alpha(1)beta(Y413F) receptors. Together, these findings indicate that GlyR function is upregulated by PTKs and this modulation is dependent on the tyrosine-413 residue of the beta subunit.
Caraiscos
Valerie B
VB
Department of Physiology, University of Toronto, Canada.
Mihic
S John
SJ
MacDonald
John F
JF
Orser
Beverley A
BA
D000818:Animals; D015536:Down-Regulation / Q000187:drug effects; D004594:Electrophysiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006624:Hippocampus / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D009474:Neurons / Q000201:enzymology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D018009:Receptors, Glycine / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D013116:Spinal Cord / Q000502:physiology; D009435:Synaptic Transmission / Q000502:physiology; D014443:Tyrosine / Q000502:physiology; D015854:Up-Regulation / Q000187:drug effects
D004791:Enzyme Inhibitors; D018009:Receptors, Glycine; D011994:Recombinant Proteins; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases