ABSTRACT
Excitotoxicity mediated by glutamate receptors plays crucial roles in ischemia and other neurodegenerative diseases. Whereas overactivation of ionotropic glutamate receptors is neurotoxic, the role of metabotropic glutamate receptors (mGluRs), and especially mGluR1, remains equivocal. Here we report that activation of NMDA receptors results in calpain-mediated truncation of the C-terminal domain of mGluR1alpha at Ser(936). The truncated mGluR1alpha maintains its ability to increase cytosolic calcium while it no longer activates the neuroprotective PI(3)K-Akt signaling pathways. Full-length and truncated forms of mGluR1alpha play distinct roles in excitotoxic neuronal degeneration in cultured neurons. A fusion peptide derived from the calpain cleavage site of mGluR1alpha efficiently blocks NMDA-induced truncation of mGluR1alpha in primary neuronal cultures and exhibits neuroprotection against excitotoxicity both in vitro and in vivo. These findings shed light on the relationship between NMDA and mGluR1alpha and indicate the existence of a positive feedback regulation in excitotoxicity involving calpain and mGluR1alpha.
Subject(s)
Calpain/physiology , Excitatory Amino Acids/toxicity , Receptors, Metabotropic Glutamate/metabolism , Animals , Cell Survival/physiology , Cells, Cultured , Cerebral Cortex/cytology , Cerebral Cortex/physiology , Electrophoresis, Polyacrylamide Gel , Electrophysiology , Excitatory Amino Acid Agonists/toxicity , Immunoblotting , Immunohistochemistry , Kainic Acid/toxicity , Neurons/physiology , Rats , Receptors, N-Methyl-D-Aspartate/physiology , Signal Transduction/physiology , Silver Staining , TransfectionABSTRACT
The balance between excitatory and inhibitory synapses is a tightly regulated process that requires differential recruitment of proteins that dictate the specificity of newly formed contacts. However, factors that control this process remain unidentified. Here we show that members of the neuroligin (NLG) family, including NLG1, NLG2, and NLG3, drive the formation of both excitatory and inhibitory presynaptic contacts. The enrichment of endogenous NLG1 at excitatory contacts and NLG2 at inhibitory synapses supports an important in vivo role for these proteins in the development of both types of contacts. Immunocytochemical and electrophysiological analysis showed that the effects on excitatory and inhibitory synapses can be blocked by treatment with a fusion protein containing the extracellular domain of neurexin-1beta. We also found that overexpression of PSD-95, a postsynaptic binding partner of NLGs, resulted in a shift in the distribution of NLG2 from inhibitory to excitatory synapses. These findings reveal a critical role for NLGs and their synaptic partners in controlling the number of inhibitory and excitatory synapses. Furthermore, relative levels of PSD-95 alter the ratio of excitatory to inhibitory synaptic contacts by sequestering members of the NLG family to excitatory synapses.
Subject(s)
Membrane Proteins/physiology , Nerve Tissue Proteins/metabolism , Nerve Tissue Proteins/physiology , Synapses/metabolism , Animals , Blotting, Western , Cell Adhesion Molecules, Neuronal , Cells, Cultured , Cloning, Molecular , DNA, Complementary/metabolism , Disks Large Homolog 4 Protein , Electrophysiology , Gene Library , Green Fluorescent Proteins/metabolism , Guanylate Kinases , Hippocampus/cytology , Hippocampus/metabolism , Image Processing, Computer-Assisted , Immunohistochemistry , Intracellular Signaling Peptides and Proteins , Membrane Proteins/metabolism , Mice , Microscopy, Fluorescence , Models, Biological , Multigene Family , Nerve Tissue Proteins/chemistry , Neurons/metabolism , Protein Binding , Rats , Rats, Wistar , Recombinant Fusion Proteins/chemistry , TransfectionABSTRACT
Dopamine D1-like receptors, composed of D1 and D5 receptors, have been documented to modulate glutamate-mediated fast excitatory synaptic neurotransmission. Here, we report that dopamine D1 receptors modulate NMDA glutamate receptor-mediated functions through direct protein-protein interactions. Two regions in the D1 receptor carboxyl tail can directly and selectively couple to NMDA glutamate receptor subunits NR1-1a and NR2A. While one interaction is involved in the inhibition of NMDA receptor-gated currents, the other is implicated in the attenuation of NMDA receptor-mediated excitotoxicity through a PI-3 kinase-dependent pathway.
Subject(s)
Receptors, Dopamine D1/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Androstadienes/pharmacology , Animals , Apoptosis , Benzazepines/pharmacology , Blotting, Western , Cells, Cultured , Corpus Striatum/metabolism , Enzyme Inhibitors/pharmacology , Hippocampus/metabolism , Models, Chemical , Phosphoinositide-3 Kinase Inhibitors , Rats , WortmanninABSTRACT
Src kinase regulation of N-methyl-D-aspartate (NMDA) subtype glutamate receptors in the central nervous system (CNS) has been found to play an important role in processes related to learning and memory, ethanol sensitivity and epilepsy. However, little is known regarding the mechanisms underlying the regulation of Src family kinase activity in the control of NMDA receptors. Here we report that the distal phosphatase domain (D2) of protein tyrosine phosphatase alpha (PTPalpha) binds to the PDZ2 domain of post-synaptic density 95 (PSD95). Thus, Src kinase, its activator (PTPalpha) and substrate (NMDA receptors) are linked by the same scaffold protein, PSD95. Removal of PTPalpha does not affect the association of Src with NMDA receptors, but turns off the constitutive regulation of NMDA receptors by the kinase. Further more, we found that application of the PTPalpha catalytic domains (D1 + D2) into neurones enhances NMDA receptor-mediated synaptic responses. Conversely, the blockade of endogenous PTPalpha inhibits NMDA receptor activity and the induction of long-term potentiation in hippocampal neurones. Thus, PTPalpha is a novel up-regulator of synaptic strength in the CNS.
