Cell Death after Spinal Cord Injury Is Exacerbated by Rapid TNFα-Induced Trafficking of GluR2-Lacking AMPARs to the Plasma Membrane

Abstract

Glutamate, the major excitatory neurotransmitter in the CNS, is implicated in both normal neurotransmission and excitotoxicity. Numerous in vitro findings indicate that the ionotropic glutamate receptor, AMPAR, can rapidly traffic from intracellular stores to the plasma membrane, altering neuronal excitability. These receptor trafficking events are thought to be involved in CNS plasticity as well as learning and memory. AMPAR trafficking has recently been shown to be regulated by glial release of the proinflammatory cytokine tumor necrosis factor ␣ (TNF␣) in vitro. This has potential relevance to several CNS disorders, because many pathological states have a neuroinflammatory component involving TNF␣. However, TNF␣-induced trafficking of AMPARs has only been explored in primary or slice cultures and has not been demonstrated in preclinical models of CNS damage. Here, we use confocal and image analysis techniques to demonstrate that spinal cord injury (SCI) induces trafficking of AMPARs to the neuronal membrane. We then show that this effect is mimicked by nanoinjections of TNF␣, which produces specific trafficking of GluR2-lacking receptors which enhance excitotoxicity. To determine if TNF␣-induced trafficking affects neuronal cell death, we sequestered TNF␣ after SCI using a soluble TNF␣ receptor, and significantly reduced both AMPAR trafficking and neuronal excitotoxicity in the injury penumbra. The data provide the first evidence linking rapid TNF␣-induced AMPAR trafficking to early excitotoxic secondary injury after CNS trauma in vivo, and demonstrate a novel way in which pathological states hijack mechanisms involved in normal synaptic plasticity to produce cell death.

Astrocytes expressing ALS‐linked mutant FUS induce motor neuron death through release of tumor necrosis factor‐alpha

“…Because TNFα has been shown to induce surface expression changes of AMPA‐type glutamate receptors (AMPARs), leading to excitotoxicity (Ferguson et al, 2008; Leonoudakis et al, 2008; Santello and Volterra, 2012), we hypothesized that the aberrant release of TNFα by mutFUS astrocytes may also lead to AMPA receptor dysregulation in motor neurons. To test this, we treated motor neurons with ACM from WTFUS or mutFUS‐expressing astrocytes and stained them for glutamate receptor 1 (GluA1), glutamate receptor 2 (GluA2) and microtubule‐associated protein 2 (MAP2) as dendritic marker.…”

“…TNFα is a pleiotropic molecule that can be produced by a number of different cell‐types, including astrocytes (Chung and Benveniste, 1990; Santello and Volterra, 2012). In the CNS in particular, TNFα signaling is complex, and its role in regulating synaptic transmission both physiologically and pathologically is increasingly being appreciated (Ferguson et al, 2008; Leonoudakis et al, 2008; Olmos and Lladó, 2014; Santello and Volterra, 2012). Importantly, soluble recombinant TNFα has been shown to be directly toxic to human fetal tissue derived neurons and motor neuron‐like NSC‐34 in vitro and knockout of TNFα receptors prevents motor neuron cell death in an axonal injury mouse model, indicating that TNFα can damage motor neurons (D'Souza, Alinauskas, McCrea, Goodyer, & Antel, 1995; He, Wen, & Strong, 2002; Raivich et al, 2002).…”

“…Interestingly, several groups have reported a role for TNFα as a regulator of surface AMPAR expression in spinal cord motor neurons. These studies have shown that TNFα triggers a change in AMPARs which can promote a shift towards surface expression of GluA2‐lacking receptors (Ferguson et al, 2008; Ogoshi et al, 2005; Stellwagen, 2005). Although these studies focused on the rapid effects of short‐term TNFα exposure, we have demonstrated here that longer incubations with TNFα–containing mutFUS‐ACM, not only alter GluA1 & GluA2 protein levels but also trigger an increase mRNA levels for GluA1.…”

“…In addition to classical caspase‐8 activation, one mechanism proposed for TNFα‐induced neuron death is through the rapid TNFα‐induced surface expression changes of AMPA‐type glutamate receptors (AMPARs) (Ferguson et al, 2008). Dysregulation of the precise trafficking can alter neuronal calcium permeability and contribute to excitotoxic vulnerability (Ferguson et al, 2008; Leonoudakis, Zhao, & Beattie, 2008; Olmos and Lladó, 2014). …”

The role of mTOR signaling pathway in spinal cord injury

“…31 Especially in the acute phase after injury, activated macrophages and microglia produce various proinflammatory cytokines, such as IL-1β and TNFα, 32,33 and contribute to secondary tissue damage, neuronal loss and demyelination. [34][35][36] Recent studies have demonstrated that inhibition of mTOR suppresses macrophage/microglia activation and reduces neuroinflammation. 22 These findings suggest that inhibition of mTOR can suppress macrophage/microglia activation and reduce the inflammation that causes secondary damage following SCI.…”