# Exploring the EphB2-NMDA receptor interaction in spinal cord injury-induced neuropathic pain

> **NIH NIH R01** · THOMAS JEFFERSON UNIVERSITY · 2024 · $624,487

## Abstract

Project Summary / Abstract (30-line maximum)
In this R01 Renewal project, we will examine contribution of the EphB2 receptor-NMDA receptor (NMDAR)
interaction to excitatory synaptic neurotransmission in dorsal horn (DH) pain circuitry and neuropathic pain (NP)
after spinal cord injury (SCI). NP is a highly debilitating and often chronic outcome that occurs in a major portion
of SCI cases. Importantly, this NP is particularly refractory to treatment, urgently calling for identification of targets
that both robustly regulate pathological pain and avoid the devastating effects of opioid-based interventions.
 NMDAR-mediated hyperexcitability of DH circuitry is a major substrate for pathological pain. NP has been
linked to EphB receptor tyrosine kinases via potentiation of NMDAR function, as EphB signaling regulates
multiple facets of NMDAR biology, including NMDAR-dependent Ca2+ influx and synaptic plasticity. We
discovered that EphB2 modulation of NMDARs is mediated by a direct, extracellular interaction between EphB2
and the NMDAR. Importantly, we elucidated the mechanism regulating this interaction: phosphorylation of a
single extracellular amino acid of EphB2 (Y504) interacts with a positive surface charge on the N-terminal domain
of the NMDAR GluN1 subunit. We find in vitro that Y504 phosphorylation is required in spinal cord neurons for
EphB2-NMDAR interaction, NMDAR synaptic localization, and excitatory synapse function. In our cervical
contusion SCI model that results in forepaw hyperalgesia and allodynia, and ongoing pain, this NP phenotype is
accompanied by long-lasting increases in EphB2 expression and signaling, Y504 phosphorylation, and EphB2-
NMDAR co-localization at excitatory synapses in cervical DH, in particular in the highly-important population of
DH pain projection neurons. Excitingly, inducible DH neuron-specific EphB2 knockout or inducible chemogenetic
inhibition of EphB tyrosine kinase signaling robustly reverses already-established NP-related behavior after SCI.
 We therefore hypothesize that modulating EphB2-NMDAR interaction in DH pain projection neurons after
cervical contusion SCI will impact: (1) NMDAR synaptic localization, (2) excitatory synaptic transmission in DH
pain circuitry, and (3) NP. In Aim 1, we will determine whether inducible EphB2 knockout selectively in DH
projection neurons reduces NMDAR synaptic localization and NP-related behaviors using a novel floxed-EphB2
mouse. In Aim 2, we will determine if preventing EphB2-NMDAR interaction selectively in DH projection neurons
impacts NMDAR synaptic localization, excitatory synaptic transmission and NP behaviors using our innovative
knockin mouse that allows for inducible expression of EphB2-Y504F (EphB2 with single amino acid switch that
blocks interaction with the NMDAR). In Aim 3, we will determine the role of EphB2 intracellular tyrosine kinase
activity in NP using a chemogenetic mouse that allows for specific, inducible and reversible inhibition of EphB2
kin...

## Key facts

- **NIH application ID:** 10882574
- **Project number:** 2R01NS110385-06
- **Recipient organization:** THOMAS JEFFERSON UNIVERSITY
- **Principal Investigator:** MANUEL L COVARRUBIAS
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $624,487
- **Award type:** 2
- **Project period:** 2018-09-01 → 2029-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10882574

## Citation

> US National Institutes of Health, RePORTER application 10882574, Exploring the EphB2-NMDA receptor interaction in spinal cord injury-induced neuropathic pain (2R01NS110385-06). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10882574. Licensed CC0.

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