Authors: J. Yan, C.P. Bengtson, B. Buchthal, A.M. Hagenston, H. Bading
Summary: Introduction: N-methyl-D-aspartate receptors (NMDARs) are glutamate-gated, calcium-permeable neurotransmitter receptors. They are fundamental to brain development, they control synaptic plasticity in the adult, and they initiate transcriptional responses needed for the consolidation of adaptive processes in the nervous system such as memory and acquired neuroprotection. However, NMDARs are also potentially harmful to neurons because they can initiate transcription shutoff pathways and can lead to mitochondrial dysfunction and even to excitotoxic cell death induced by excessive glutamate. The molecular basis of toxic NMDAR signaling is unknown, although a high intracellular calcium load has been implicated. An alternative model suggests that, depending on their location (synaptic versus extrasynaptic), NMDARs can either promote neuronal survival or cause cell death.
Rationale: We hypothesized that NMDARs acquire toxic features through physical interaction with one or more other proteins that may be present at extrasynaptic but not synaptic locations. Identification of NMDAR-associated proteins and the mapping of their respective interaction domains may enable the development of innovative means to disrupt a putative death signaling complex. In particular, the search for interaction interface inhibitors could yield novel compounds that—unlike classical NMDAR blockers—would bring about neuroprotection by stripping off the toxic component of extrasynaptic NMDAR signaling without compromising the physiological functions of synaptic NMDARs.
Results: We found that the NMDAR subunits GluN2A and GluN2B form a complex with transient receptor potential cation channel subfamily M member 4 (TRPM4). The NMDAR/TRPM4 interaction is mediated by a 57–amino acid intracellular domain of TRPM4, termed TwinF, that is positioned just beneath the plasma membrane. TwinF interacts with I4, an evolutionarily highly conserved stretch of 18 amino acids with four regularly spaced isoleucines located within the intracellular, near-membrane portion of GluN2A and GluN2B. The NMDAR/TRPM4 complex can be disrupted by (i) expression of TwinF, which acts by competing with endogenous TRPM4 for binding to GluN2A and GluN2B, or (ii) small-molecule NMDAR/TRPM4 interaction interface inhibitors that we identified with a TwinF structure-based computational compound screen. Both TwinF and the small-molecule interface inhibitors provide robust protection against excitotoxic cell death in cultured neurons and in vivo in mouse models of neurodegeneration. They also eliminate excitotoxicity-associated transcription shutoff and mitochondrial dysfunction while leaving synaptic and extrasynaptic NMDAR-mediated currents and calcium signaling unaffected.
Conclusion: Our study uncovered the requirement of an NMDAR/TRPM4 complex for excitotoxicity. According to proteomics databases for synaptic proteins from mouse and human cortex and hippocampus, TRPM4 is absent from the synapse. This indicates that the NMDAR/TRPM4 complex forms extrasynaptically, which explains why extrasynaptic but not synaptic NMDARs promote death signaling. Our findings provide a conceptually new basis for therapeutic targeting of toxic NMDAR signaling, which contributes to the pathology of many neurological conditions including stroke, traumatic brain injury, Alzheimer’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and retinal degeneration. Recombinant and small-molecule NMDAR/TRPM4 interaction interface inhibitors define a class of potent neuroprotectants with a new mode of action that renders extrasynaptic NMDARs nontoxic and eliminates their transcription shutoff signaling. Given that increased toxic signaling of extrasynaptic NMDARs is a pathomechanism shared by many neurodegenerative disorders, NMDAR/TRPM4 interaction interface inhibitors may be effective, broadly applicable therapeutics.
Source: Science, 2020