TRP Channels in Excitotoxicity

Abstract

Glutamate excitotoxicity is a central mechanism contributing to cellular dysfunction and death in various neurological disorders and diseases, such as stroke, traumatic brain injury, epilepsy, schizophrenia, addiction, mood disorders, Huntington’s disease, Alzheimer’s disease, Parkinson’s disease, multiple sclerosis, pathologic pain, and even normal aging-related changes. This detrimental effect emerges from glutamate binding to glutamate receptors, including α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, N-methyl-d-aspartate receptors, kainate receptors, and GluD receptors. Thus, excitotoxicity could be prevented by targeting glutamate receptors and their downstream signaling pathways. However, almost all the glutamate receptor antagonists failed to attenuate excitotoxicity in human patients, mainly due to the limited understanding of the underlying mechanisms regulating excitotoxicity. Transient receptor potential (TRP) channels serve as ancient cellular sensors capable of detecting and responding to both external and internal stimuli. The study of human TRP channels has flourished in recent decades since the initial discovery of mammalian TRP in 1995. These channels have been found to play pivotal roles in numerous pathologic conditions, including excitotoxicity. In this review, our focus centers on exploring the intricate interactions between TRP channels and glutamate receptors in excitotoxicity.

Introduction

Excitotoxicity contributes to neuronal degeneration in various neurologic disorders, including acute ischemic stroke and brain injury, and chronic diseases such as Alzheimer’s disease. Since it was first discovered several decades ago (Olney 1969), excitotoxicity caused by excessive glutamate has been the center of extensive research for understanding the underlying mechanisms and for developing effective therapeutics for neurodegenerative disorders (Choi 2020). One of the hallmark features of excitotoxicity is Ca²⁺ overload mediated by glutamate receptors, particularly NMDA receptors. However, excitotoxic N-methyl-d-aspartate receptor (NMDAR) antagonists all failed in clinical trials for ischemic stroke, which promoted great interest in identifying nonexcitotoxic Ca²⁺-permeable channels as potential therapeutic targets. Transient receptor potential (TRP) channels (Clapham 2003), featured with Ca²⁺ permeation and polymodal activation by various stress stimuli such as Ca²⁺ and oxidative stress, emerged as candidate targets for nonexcitotoxic Ca²⁺-induced neurotoxicity (Tymianski 2011). After the elegant study by Tymianski and colleagues showing that TRPM7 contributes to anoxic neuronal death independent of glutamate (Aarts and Tymianski 2005), several TRP channels, including TRPM2 and TRPM4 (Leiva-Salcedo and others 2017; Wang and others 2021a), have been shown to be involved in ischemic brain damage through nonexcitotoxic mechanisms. However, research in the past decade has revealed that TRP channels not only mediated nonexcitotoxic Ca²⁺-induced neurotoxicity but also contributed significantly to glutamate-dependent excitotoxicity. More important, recent studies uncovered physical interactions of TRP channels with NMDARs as the underlying mechanisms of excitotoxicity, highlighting a future direction for developing therapeutics for neurodegenerative disorders. This review focuses on TRP channels that mediate excitotoxic neuronal death and neurodegenerative disorders.

TRP Channels as Ancient Sensors

From unicellular organisms to human beings, the survival of all the life depends on their sensory systems to promptly perceive and respond to environmental challenges: both paramecia and humans know to avoid extreme temperature or other aversive stimulus (Xiao and Xu 2021). During the long process of evolution, this primordial but remarkable survival ability has become deeply embedded into our genes, as its action is innate and instantaneous: one can react instinctively, such as withdrawing their hand from a hot pot, without conscious deliberation. In animals, vision and hearing are crucial senses for detecting and responding to potential threats, such as predators, while taste and smell play essential roles in identifying potentially toxic foods. Additionally, pain sensation serves as a crucial mechanism for providing special attention and protection to internal or external injuries. At the cellular level, the ability to sense chemical, mechanical, thermal, oxidative, and various environmental cues can confer a variety of cells with defensive, protective, or adaptive responses, safeguarding the integrity of a cell or a living organism (Fig. 1). Completing such a vast task would be impossible without the cellular sensors, the transient receptor potential (TRP) channels, which have been evolved from unicellular ancestors.

Figure 1.

Transient receptor potential (TRP) channels as cellular sensors. TRP channels can be activated by various external stimuli, including light, chemicals, and temperature changes. This activation leads to the generation of sensory signals, playing crucial roles in processes such as vision, taste, smell, and temperature sensation. Additionally, TRP channels can be activated by internal stimuli, including oxidative stress and cellular injury. The ion influx mediated by TRP channels not only triggers membrane depolarization but is also essential for many cellular activities such as cell migration and inflammasome activation. Furthermore, TRP channel-mediated ion influx can initiate intracellular signaling pathways, particularly through Ca²⁺ signaling (calmodulin [CaM]), thereby influencing transcription factors (TFs) and gene expression. It is important to note that while TRP channels play an integral role in various cellular functions, their overactivation can lead to Ca²⁺ overload and ionic imbalance, causing cellular injury or cell death.

“Transient receptor potential” (TRP) was initially used by Cosens and Manning in 1969 to describe a light-sensing defective Drosophila mutant whose retina exhibited a transient receptor potential instead of the normal sustained response to light stimuli (Montell and Rubin 1989). To date, TRP channels have been identified in various living systems, ranging from unicellular organisms to mammals. The 28 mammalian TRP channels are classified into canonical (TRPCs), vanilloid (TRPVs), melastatin (TRPMs), mucolipin (TRPMLs), polycystin (TRPPs), and ankyrin (TRPAs) subfamilies (Clapham 2003) (Box 1). Structurally, TRP channels resemble voltage-gated K⁺ channels, consisting of four subunits in each homomeric or heteromeric channel. Within each subunit, there are the intracellular N-terminal and C-terminal domains, as well as six transmembrane domains (S1–S6). While the arrangement of S1 to S4 in TRP channels notably resembles the voltage-sensing domain (VSD) found in voltage-gated ion channels, TRP channels are not voltage gated (Nilius and Owsianik 2011). The channel pore formed by the S5, pore-loop, and S6 domains is permeable to various cations, including Na⁺, K⁺, Ca²⁺, and Mg²⁺, making TRP channels the largest nonselective cation channel superfamily. Most TRP channels are Ca²⁺ permeable, except for TRPM4 and TRPM5, which are Ca²⁺ impermeable and monovalent selective (Clapham 2003). In contrast, TRPV5 and TRPV6 are relatively Ca²⁺ selective (Clapham 2003; Nilius and Owsianik 2011). Moreover, some TRP channels, such as TRPM6 and TRPM7, are highly permeable to Mg²⁺ and trace metals like Zn²⁺ and Mn²⁺ (Bouron and others 2015; Clapham 2003).

BOX 1.

Introduction to transient receptor potential (TRP) channels. The TRP channels are a superfamily of evolutionally conserved calcium permeable nonselective cation channels. The 28 mammalian TRP channels are grouped into six subfamilies, including TRPC, TRPV, TRPM, TRPML, TRPP, and TRA subfamilies. Similar to voltage-gated potassium channels, TRP channels form functional homotetrameric or heterotetrameric channels, with each subunit consisting of six transmembrane spanning domains (S1–S6), a pore-forming loop between S5 and S6, and intracellular localized N-terminus (NTD) and C-terminus (CTD). The lack of the positively charged arginines in the S4 indicates weak voltage sensitivity. The NTD and CTD exhibit significant diversity among TRP channels. The ankyrin repeats at the NTD range from 3 or 4 in TRPC and TRPV to 14 in TRPA. TRPMs have long NTD consisting of four melastatin homology region domains (MHRs). Many TRP channels have coiled-coil (CC) domains in the NTD and/or CTD. The TRP box contains positively charged lysine and arginine residues, which can interact with PIP₂ in the membrane. TRPM6 and TRPM7 have a kinase domain at CTD, whereas TRPM2 has an enzymatic NUDT9H domain similar to the NUDT9 enzyme that hydrolyzes adenosine diphosphate ribose (ADPR).

Ubiquitously expressed in various cells and tissues, TRP channels are involved in sensory functions, including sensation to temperature and pain, taste perception, olfaction, hearing, and vision (Koivisto and others 2022). Moreover, the multimodal activation/regulation features of TRP channels confer various physiologic and pathologic functions, including critical roles in both inherited and acquired diseases in humans (Koivisto and others 2022; Yue and Xu 2021). Many TRP channels, including TRPM2 and several TRPCs, which can be significantly activated or potentiated by oxidative stress conditions and are considered biosensors for oxidative stress (Sakaguchi and Mori 2020), are involved in excitotoxicity-related neurologic diseases.

Canonical TRP channel subfamily (TRPCs).

TRPC1 was the first genetically identified mammalian TRP channel (Bon and others 2022). The TRPC subfamily comprises seven members (TRPC1–7), although human TRPC2 is a pseudogene. TRPC channels can be categorized into three groups (TRPC1, TRPC3/6/7, and TRPC4/5) based on sequence similarities. Except for TRPC5, other TRPCs do not respond to temperature changes (Bon and others 2022). Within the TRPC superfamily, heterotetramer formation is common, particularly within the same subgroups, such as TRPC1/4/5 and TRPC3/6 channels (Bon and others 2022). While this tendency for heteromerization significantly increases the diversity of the TRPC family, it presents technical challenges for their unique identification during electrophysiologic recordings of each TRPC channel in primary cells and animals (Bon and others 2022). TRPCs are ubiquitously expressed across all cell types and play pivotal roles in various diseases, including neurologic disorders, kidney diseases, cancer, cardiovascular diseases, and metabolic disorders (Bon and others 2022). Another feature of TRPCs is their activation secondary to the phospholipase C (PLC)–IP₃/Ca²⁺ pathway, typically downstream of G protein–coupled receptors (GPCRs) like mGluRs (Zong and others 2023). Most of the TRPC channels have been proposed to be involved in glutamate excitotoxicity via different molecular mechanisms.

Vanilloid TRP channel subfamily (TRPV).

The nomenclature “TRPV” stems from TRPV1’s capability to react to capsaicin, a compound present in chili peppers categorized as a vanilloid (Rosenbaum and Islas 2023). TRPVs are prominently present in the peripheral nervous system, acting as receptors for mechanical, chemical, and thermal stimuli (Rosenbaum and Islas 2023). The TRPV family comprises six members (TRPV1–6). TRPV1–4 respond to heat with different temperature thresholds, while TRPV5 and TRPV6 lack thermosensitivity (Rosenbaum and Islas 2023). TRPV5 and TRPV6 display significant selectivity for Ca²⁺ among the TRP channel superfamily, while TRPV1–4 function as nonselective cation channels (Rosenbaum and Islas 2023). TRPVs can be activated by a variety of inflammatory agents, including eicosanoids and arachidonic acid derivatives like prostaglandins, leukotrienes, and thromboxanes, implying their potential involvement in inflammatory conditions such as neurodegeneration (Rosenbaum and Islas 2023).

Melastatin TRP channel subfamily (TRPM).

When initially discovered in 1998, this subfamily was classified as “long TRPC” (LTRPC) channels due to their striking structural resemblance to TRPCs and the presence of a remarkably long N-terminal domain (NTD) (Chubanov and others 2024). Subsequently, they were given the name TRP melastatin (TRPM) because their identification was linked to a screening for genes associated with melanoma, and there are four highly conserved melastatin homology regions (MHR1–4) in their NTD, a distinguishing feature not found in other TRP channels (Chubanov and others 2024). Based on sequence similarities, TRPM channels are categorized into four groups: TRPM1/TRPM3, TRPM2/TRPM8, TRPM4/TRPM5, and TRPM6/TRPM7 (Chubanov and others 2024). Another common feature of TRPMs is that their NTD lacks the ankyrin repeats that is usually found in other TRP channels (Chubanov and others 2024; Clapham 2003). Compared to other TRP subfamilies, the members of the TRPM subfamily exhibit significant variability among themselves and possess several unique features (Chubanov and others 2024). One distinctive feature is that TRPM2, TRPM6, and TRPM7 have an enzymatic domain in their C-termini. They are also called “chanzymes” (Chubanov and others 2024). Whereas the pyrophosphatase in mammalian TRPM2 does not have enzymatic activity, the kinase domain in TRPM6 and TRPM7 can auto-phosphorylate as well as phosphorylate other substrates. Moreover, the channel kinases TRPM6 and TRPM7 are also permeable to Mg²⁺, Zn²⁺, and some trace metals (Clapham 2003). Furthermore, TRPM2, TRPM3, TRPM4, and TRPM5 respond to heat, whereas TRPM8 responds to cold (Chubanov and others 2024). All TRPMs are expressed in the brain at varying levels, with TRPM2 having the most abundantly expression among them (Zong and others 2022b). To date, TRPM2 and TRPM4 have demonstrated substantial contributions to excitotoxicity in neurons (Zong and others 2023).

Excitotoxicity

The amino acid glutamate is the most abundant neurotransmitter in the brain (Watkins and Jane 2006). Yet, problems arise when neurons are overstimulated. In 1957, subcutaneous injection of glutamate was first found to damage the inner layer in the retina in mice (Lucas and Newhouse 1957). The damaging effects caused by glutamate in brain and spinal cord were detailed in Olney’s seminal work (Olney 1969), and the concept of “excitotoxicity” was defined in the late 1970s by Olney (Doble 1999). Since then, excitotoxicity-induced cell death and resulting pathologic conditions have been extensively studied (Olney 1969; Plotegher and others 2021).

Glutamate receptors (GluRs) fall into two categories: cation-permeable ionotropic receptors (iGluRs) and GPCR metabotropic receptors (mGluRs) (Fig. 2). iGluRs can be grouped into four classes based on their responses to different agonists: α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), NMDARs, kainate receptors (KARs), and GluD receptors (Watkins and Jane 2006). All iGluRs are heterotetramers composed of four subunits, forming a central ion pore with various subunit compositions (GluA1–4 for AMPARs; GluN1, GluN2A–D, and GluN3A–B for NMDARs; GluK1–5 for KARs; GluD1–2 for GluD receptors), while mGluRs belong to group C family of GPCRs and are homo- or heterodimers formed by mGluR1–8 (Hansen and others 2021) (Fig. 2). Besides sharing a similar pore-loop domain, the structure of iGluRs differs substantially from that of TRP channels and voltage-gated ion channels. The N-terminal domain is located extracellularly, responsible for glutamate and other ligand binding, and there are only four transmembrane helices (M1–M4) to form the transmembrane domain. The C-terminal domain varies significantly in length among different subunits and is important for channel regulation as well as for the binding of partner proteins (Hansen and others 2021). AMPARs, NMDARs, KARs, and mGluRs all have been demonstrated to contribute to glutamate excitotoxicity (Fig. 2), with extensive research particularly focused on NMDARs (Crupi and others 2019; Guo and Ma 2021; Hardingham and Badling 2010; Vincent and Mulle 2009). The excitotoxicity of NMDARs is contingent on their composition and location. In general, GluN2A-containing NMDARs preferentially interact with the postsynaptic scaffolding protein PSD-95. This binding often restricts their expression primarily to synaptic sites. On the other hand, GluN2B-containing NMDARs exhibit a broader expression pattern and are more abundantly found at extrasynaptic sites (Papouin and Oliet 2014). Extrasynaptic NMDARs (approximately three-fourths of all the NMDARs) and those containing the GluN2B subunit enhance the activation of neuronal death–related signaling pathways. Conversely, synaptic NMDARs and those with the GluN2A subunit promote neuronal survival (Hardingham and others 2002; Zong and others 2022a). This interesting composition-dependent cytotoxicity is also observed in AMPARs. Unlike the majority of GluA2-containing AMPARs that are Ca²⁺ impermeable, Ca²⁺-permeable AMPARs lacking GluA2 are the primary contributor to AMPAR-mediated excitotocixity (Guo and Ma 2021). As NMDAR-mediated excitotoxicity is most well studied, in this review, we mainly focus on the interaction between TRP channels and NMDARs.

Figure 2.

Glutamate receptors (GluRs) and excitotoxicity. The activation of ionotropic GluRs (iGluRs), which include α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), N-methyl-d-aspartate receptors (NMDARs), and kainate receptors (KARs), results in ionic influx and subsequent membrane depolarization. Overactivation of iGluRs can lead to neuronal hyperactivity and cellular dysfunction. On the other hand, metabotropic GluRs (mGluRs), which are GPCRs, initiate downstream signaling upon activation. This signaling can activate pathways related to cellular dysfunction or death. The excessive ionic influx mediated by iGluRs, particularly Ca2⁺ overload, and the activation of their associated partners (especially NMDAR) are implicated in cellular injury, contributing to the activation of injury or death-related pathways in neurons and endothelial cells and leading to neuronal injury and endothelial leakage. It is noteworthy that GluR-mediated Ca²⁺ signaling in glial cells can lead to their activation toward a proinflammatory phenotype. This activation of glial cells further contributes to the inflammatory responses in the brain.

Besides neurons, NMDARs are also widely expressed in other cell types in the central nervous system (CNS), including glial cells and cerebral endothelial cells. Moreover, there has been growing evidence implicating that the functional NMDARs are also present in a variety of no-neuronal cells and tissues, such as cells from the immune system, heart, kidney, and pancreas, where they potentially play a preeminent role in various pathophysiologic processes outside the CNS (Bozic and Valdivielso 2015). NMDARs are typically composed of two GluN1 subunits for coagonist glycine/D-serine binding and two glutamate-binding GluN2 subunits for either identical or different agonists. Thus, they are also referred to as diheteromeric or triheteromeric NMDARs (Hansen and others 2021). Considering that GluN3 subunits do not bind to glutamate and the existence of triheteromeric GluN1/GluN2/GluN3 NMDARs is still uncertain (Hansen and others 2021), we primarily focus on GluN1/GluN2 NMDARs.

As the “backbone” composition of NMDARs, GluN1 subunits are always present as an essential component of an NMDAR and are widely expressed in the brain, whereas the expression pattern of GluN2 subunits varies among NMDARs at different locations of a neuron and in different types of neurons and may even change under certain pathophysiologic conditions (Hansen and others 2021), leading to a diversified biophysical properties of NMDARs (Hansen and others 2021). NMDAR activation causes membrane depolarization, and NMDAR-mediated Ca²⁺ influx is required for many critical neuronal activities and brain functions, such as synaptic plasticity and memory (Hansen and others 2021). Moreover, NMDARs play a pivotal role in integrating diverse signaling networks in the brain by physically interacting with various proteins, including 1) surface receptors, encompassing dopamine receptors, histamine 3 receptor, mGluRs, opioid receptors, ephrin B2 receptor, ApoE receptor, neuroligin, and IL-1 receptors; 2) ion channels such as nicotinic receptors, purinergic receptors, BK channels, pannexin 1, TRPM2, and TRPM4; (3) numerous intracellular proteins, including PSD-95 family scaffold proteins, Ca²⁺/calmodulin-dependent protein kinase II (CaMKII), cyclin dependent kinase 5 (Cdk5), death-associated protein kinase 1 (DAPK1), Src kinase, and rabphilin 3A (Gardoni and Di Luca 2021; Hansen and others 2021; Petit-Pedrol and Groc 2021; Zong and others 2022a; Zong and others 2022b). However, overactivation of GluRs induces neuronal death through several mechanisms, including excessive depolarization, Ca²⁺ overload, oxidative stress, mitochondrial dysfunction, and activation of death-related signaling (Dong and others 2009). Subsequently, it was discovered that glutamate excitotoxicity can lead to dysfunction and cell death not only in neurons but also in these nonneuronal cell populations (Belov Kirdajova and others 2020; Parfenova and others 2006), while ischemic excitotoxicity in neurons is most extensively studied (Fig. 3).

Figure 3.

Ischemic excitotoxicity. Graphic illustration showing excitotoxic events in the penumbra (red background indicates ischemic area, while blue background indicates nonischemic area) after stroke. Ischemia induces several key pathologic changes in the ischemic area: 1) rapid death of neurons, 2) glial activation, and 3) blood-brain barrier (BBB) leakage and immune cell invasion. Dying neurons are a major source of the poststroke glutamate surge, as they release stored neurotransmitters. Additionally, activated astrocytes and microglia also release glutamate and contribute to this surge, and the released glutamate from neurons may further enhance this process. Subsequently, the glutamate surge leads to neuronal death and glial activation in nonischemic areas, creating secondary surges of glutamate. This sets off vicious cycles that worsen brain injury. Glutamate can also affect the BBB, increasing its permeability. While it has been shown that glutamate receptors (GluRs) can regulate immune cell functions in the periphery, it remains unclear whether the glutamate surge during stroke and other neurodegenerative diseases modulates the activity of infiltrated immune cells.

Excitotoxicity is intricately linked to the development and progression of a variety of neurologic disorders and diseases, including stroke (Fig. 3), traumatic brain injury, epilepsy, schizophrenia, addiction, mood disorders, and neurodegenerative diseases like Huntington’s disease, Alzheimer’s disease, Parkinson’s disease, multiple sclerosis, and pathologic pain (Pal 2018). Excitotoxicity even contributes to the normal senile changes in the brain during aging (Pal 2018). However, clinical trials aimed at targeting excitotoxicity by directly inhibiting GluRs as a treatment for these diseases have consistently led to failures or rarely limited benefits, as exemplified by the extensive studies on NMDAR antagonists in stroke (Dong and others 2009). These discrepancies may be attributed to fundamental differences between model animals and human patients, who often have multiple underlying baseline diseases. Another crucial aspect is the oversight of other “evil” partners of GluRs, which can exert a substantial influence on the intensity of excitotoxicity (Koivisto and others 2022). In this review, we summarize the intricate interaction between TRP channels and GluRs and explore its potential applications in mitigating excitotoxicity and treating associated diseases. Given the extensive research on TRP channels in stroke pathology, we will primarily focus on their involvement in ischemic excitotoxicity.

TRP Channels and Ischemic Excitotoxicity

Many TRP channels have been implicated in cellular dysfunction and/or death in neuronal cells and cerebral endothelial cells in excitotoxicity-related neurologic diseases, especially stroke (Box 2), through regulating intracellular Ca²⁺ signals (Box 3). However, in most cases, the specific mechanisms underlying their connection with glutamate excitotoxicity are unexamined. Also, although some TRP channels have been shown to functionally interact with GluRs under physiologic conditions, the pathologic significance of the interaction remains unclear. Here, we focus on TRP channels that have been reported to have 1) physical and/or functional interactions with GluRs and 2) direct influences on excitotoxicity.

BOX 2.

Introduction to stroke. Stroke occurs when blood supply to the brain is compromised. There are two main types of strokes: ischemic stroke and hemorrhagic stroke. Ischemic stroke is characterized by the occlusion of a cerebral artery, typically due to the formation of a thrombus. Hemorrhagic stroke, on the other hand, is characterized by the rupture of a cerebral artery, leading to bleeding in the brain. Both types of strokes result in cerebral ischemia and subsequent tissue injury or infarction. Neurons are highly sensitive to oxygen deprivation and can die within minutes in the absence of sufficient oxygen supply. Ischemia and the resulting massive neuronal death trigger robust inflammatory responses in the brain. This leads to the activation of glial cells and the degradation of the blood-brain barrier (BBB). The disruption of the BBB allows neurotoxic substances in the blood to leak into the brain and facilitates the infiltration of peripheral leukocytes. These processes further intensify neuroinflammation and exacerbate ischemic injury.

BOX 3.

Tight control of intracellular Ca²⁺ concentration ([Ca²⁺]_i). As a critical signaling messenger, [Ca²⁺]_i is tightly regulated. While the extracellular Ca²⁺ concentration is approximately 2 mM and can reach about 1 mM in certain organelles or intracellular vesicles, the cytosolic free Ca²⁺ concentration in unstimulated cells remains around 100 nM. This extremely low Ca²⁺ concentration is primarily maintained by Ca²⁺ binding or buffering proteins. Upon activation of a typical Ca²⁺ channel, the [Ca²⁺]_i near the channel mouth can exceed 100 μM. However, within a short distance of approximately 20 nm, this concentration drops to about 10 μM before sharply reducing to the normal level of 100 nM within a few dozen nanometers further away. Thus, the brief activation of a Ca²⁺-permeable channel may only lead to a slight local membrane depolarization and will not trigger the activation of Ca²⁺ signaling pathways. However, the persistent activation of multiple Ca²⁺ channels can overwhelm the buffering capacity of nearby Ca²⁺-binding proteins, leading to the activation of Ca²⁺ signaling pathways, thereby regulating cellular activities and transcription. Dysregulation of Ca²⁺, such as Ca²⁺ overload, caused by the unrestricted activation of numerous Ca²⁺-permeable channels can trigger cell death signaling pathways.

TRPM2 in Ischemic Excitotoxicity

Initially named LTRPC2 or TRPC7 based on its sequence homology with TRPC1, TRPM2 features a NUDT9-H domain in its C-terminal domain, closely related to NUDT9 adenosine diphosphate ribose (ADPR) pyrophosphatase (Clapham 2003). However, the NUDT9-H domain in mammalian TRPM2 lacks enzymatic function. TRPM2’s activation requires the binding of ADPR and Ca²⁺. During oxidative stress conditions, such as inflammation, ADPR production is boosted by an increase of poly-ADPR polymerase activity, and intracellular Ca²⁺ levels also significantly rise. As a result, TRPM2 is often regarded as a cellular sensor for oxidative stress (Sakaguchi and Mori 2020). Additionally, TRPM2 is heat activated, with a typical temperature sensitivity threshold of approximately 47°C (Kashio and others 2012). This threshold can decrease to 36°C under inflammatory conditions, which falls below normal body temperature. This implies that TRPM2 may exhibit constitutive activity during inflammation (Kashio and Tominaga 2017). Inflammation plays a pivotal role in excitotoxicity-related disorders, but whether TRPM2’s temperature-sensing capability contributes to this process remains poorly determined.

The detrimental role of TRPM2 in stroke has been extensively studied, and it has long been postulated that TRPM2-mediated Ca²⁺ influx is responsible for exacerbating ischemic neuronal death (Belrose and others 2018). The breakthrough came in 2013 when a study demonstrated that TRPM2 knockout significantly alleviated ischemic brain injury by differentially modulating GluN2A and GluN2B expression in a mouse model of middle cerebral artery occlusion (MCAO) (Jia and others 2011). This study revealed that compared to the wild-type group, neurons lacking TRPM2 displayed resistance to neuronal death induced by oxygen-glucose deprivation (OGD). Furthermore, brain slices from TRPM2 knockout mice exhibited reduced synaptic excitability when exposed to H₂O₂ perfusion. Most notably, the absence of TRPM2 in mice provided robust protection against ischemic stroke (Jia and others 2011). Mechanistically, the absence of TRPM2 led to an up-regulation of the prosurvival NMDAR subunit, GluN2A, while down-regulating the prodeath subunit, GluN2B (Jia and others 2011). This molecular shift was associated with enhanced downstream prosurvival signaling. Also, the protective effect of TRPM2 knockout against OGD and increased neuronal excitability could be abolished by GluN2A antagonists, underscoring the pivotal role of GluN2A in these observations (Jia and others 2011). In summary, this study not only confirmed the detrimental effect of TRPM2 on neuronal survival during ischemic brain injury by using TRPM2 knockout mice but also, for the first time, pointed to a potential connection between TRPM2 and ischemic excitotoxicity. In another finding also reported in 2013, TRPM2 activation was found to be needed for NMDA-induced burst firing in GABAergic neurons in substantia nigra pars reticulata (Lee and others 2013). However, in both studies, limited evidence was provided regarding the specific mechanisms by which TRPM2 modulates NMDAR activity.

In 2022, our group bridged the previously existing gap between TRPM2 and ischemic excitotoxicity in neurons (Zong and others 2022a) (Fig. 4). In addition to neurons, TRPM2 is abundant in endothelial cells and immune cells, both of which significantly contribute to the damage associated with ischemic brain injury. To elucidate the specific impact of TRPM2 on ischemic neuronal death, we employed a targeted approach by selectively deleting TRPM2 in neurons. Our findings revealed that neuron-specific TRPM2 knockout mirrored the protective effects against ischemic stroke observed in global TRPM2 knockout mice (Zong and others 2022a). This evidence highlights the pivotal role of neuronal TRPM2 as a key contributor to ischemic excitotoxicity. At the molecular level, we unveiled a direct interaction between the EE₃ motif within the N-terminal domain of TRPM2 and the KKR motif situated in the C-terminal domain of GluN2A and GluN2B. This physical association served to enhance the surface expression and activation of NMDAR under ischemic conditions. Most important, we demonstrated that disrupting the TRPM2-NMDAR association using an interfering peptide led to a significant reduction in ischemic stroke severity in mice (Zong and others 2022a). Recently, we further identified the mechanisms through which this physical association enhances NMDAR excitotoxicity (Pengyu Zong and others 2024). We found that PKCγ, a well-known enhancer of NMDAR, physically associated with the N-terminal domain of TRPM2, and this association was significantly increased during ischemic stroke. Mechanistically, we demonstrated that TRPM2-mediated Ca²⁺ promotes PKCγ activation, which subsequently increases NMDAR activation and enhances its activation. The uncoupling of the TRPM2-PKCγ association produced an effect similar to that of the TRPM2-NMDAR association. This data provides evidence that TRPM2 enhances NMDAR-mediated ischemic excitotoxicity by recruiting and activating PKCγ. However, it is still uncertain whether the coupling between TRPM2 and NMDAR also contributes to cellular dysfunction in other CNS cell types, such as glial and endothelial cells.

Figure 4.

TRPM2-PKCγ-NMDAR interaction. During ischemic stroke, massive generation of reactive oxygen species (ROS) happens in neurons, amplifying TRPM2 activation and facilitating PKCγ binding to the TRPM2-extrasynaptic N-methyl-d-aspartate receptor (esNMDAR) complex. This cascade leads to enhanced Ca²⁺ influx via TRPM2, thereby activating PKCγ. As a positive regulator of NMDAR, PKCγ phosphorylation enhances NMDAR channel activity and increases NMDAR’s surface expression, intensifying excitotoxicity. The physical association among TRPM2, PKCγ, and NMDAR strengthens this regulatory process. Furthermore, NMDAR-mediated Ca²⁺ influx contributes to TRPM2 activation, and TRPM2-mediated Ca²⁺ influx may modulate NMDAR through glycogen synthase kinase 3β (GSK-3β). In addition, amyloid-β treatment enhances the activity of both TRPM2 and NMDAR.

TRPM4 in Ischemic Excitotoxicity

Similar to TRPM2, TRPM4 activation requires PIP2 and Ca²⁺, but unlike TRPM2, TRPM4 is relatively Ca²⁺ impermeant. The sensitivity of TRPM4 to Ca²⁺ gating is increased by PKC phosphorylation but reduced by extracellular nucleotide binding. TRPM4 exhibits wide expression pattern across various tissues, with the highest abundance in the heart and kidney. TRPM4 mutations are well known for causing heart conduction disorders in humans. In the brain, TRPM4 is broadly expressed in different cell types, and its abnormal activity is associated with various neurologic diseases.

TRPM4 has been found to aggravate ischemic brain injury through its involvement in angiogenesis, endothelial dysfunction, and astrocyte swelling, which are largely unrelated to excitotoxicity (Chen and others 2019; Loh and others 2014). The interaction between TRPM4 and NMDAR has emerged as a significant contributor to ischemic neuronal death. An initial study in 2015 highlighted TRPM4 knockout neurons’ deficiencies in NMDAR-dependent long-term potentiation (LTP), which were restored by facilitating NMDAR activation using the β-adrenergic agonist, isoprenaline (Menigoz and others 2016). Additionally, reduced Ca²⁺-dependent cation currents and Ca²⁺ transients, along with decreased neuron excitability, were observed in TRPM4 knockout neurons (Menigoz and others 2016). These findings suggested that TRPM4 activity is crucial for the initial depolarization needed for NMDAR activation and subsequent LTP induction (Menigoz and others 2016). As LTP is typically triggered by synaptic NMDAR (sNMDAR)–mediated Ca²⁺ influx, this study indicates that TRPM4 might functionally impact sNMDAR activity (Luscher and Malenka 2012). However, more recent research in 2020 demonstrates that TRPM4 physically associates not with sNMDARs but with esNMDARs during ischemic neuronal death. The TwinF domain in the NTD of TRPM4 was shown to physically interact with the I4 domain in the CTD of GluN2A and GluN2B. Overexpression of TwinF using AAV demonstrated protective effects against ischemic brain injury (Yan and others 2020). Notably, chemical compounds that disrupt the TwinF-I4 interaction provided similar protective effects. The TRPM4-NMDAR binding seemed to occur at extrasynaptic sites, as TRPM4 expression was absent in available synaptosome databases (Yan and others 2020). Also, uncoupling TRPM4-NMDAR hindered the dephosphorylation of CREB and ERK, events typically observed upon esNMDAR activation (Yan and others 2020). However, further evidence through immunofluorescent staining demonstrating direct extrasynaptic colocalization will be necessary to confirm this speculation. The mechanisms by which TRPM4 regulates NMDAR-mediated excitotoxicity remain less elucidated compared to TRPM2. This is because a simple protein-protein interaction is hard to enhance the activation of downstream signaling without additional mediators, especially in this study. TRPM4 was shown to not influence the NMDAR current and NMDAR activation kinetics, as well as NMDAR-dependent AP bursting and the accompanying Ca²⁺ signals (Yan and others 2020).

As a monovalent selective ion channel, TRPM4 lacks permeability to signaling divalent cations like Ca²⁺ and Zn²⁺, raising the possibility that its cytotoxic effects might need a mediator to exert detrimental influence. A typical example is the modulation of the water channel aquaporin 4 (AQP4) by TRPM4 (Stokum and others 2023). TRPM4 associates with sulfonylurea receptor (SUR1), and both of their expression was up-regulated during ischemic brain injury (Stokum and others 2023). Activation of the TRPM4-SUR1 complex in astrocyte endfeet caused Na⁺ influx, which triggered the activity of a Na⁺/Ca²⁺ exchanger NCX1 (Stokum and others 2023). This Na⁺ influx, then, triggered an indirect TRPM4-“mediated” Ca²⁺ influx following its activation, recruiting AQP4 into the TRPM4-SUR1 complex via calmodulin (CaM) (Stokum and others 2023). Subsequently, enhanced AQP4 activity contributes to cellular swelling, notably observed in astrocytes (Stokum and others 2023). However, it remains unexamined whether TRPM4-mediated Na⁺ influx in neurons also leads to Na⁺/Ca²⁺ exchanger activation and subsequent Ca²⁺ influx under ischemic conditions.

TRPCs in Ischemic Excitotoxicity

TRPCs, especially TRPC1, can readily form heterotetramers with other TRPCs and regulate their functions (Bon and others 2022). TRPC1, TRPC4, and TRPC5 are abundantly expressed in the brain and regulate important cellular activities and brain functions, such as glutamate signaling, neurite growth, synaptic transmission, and fear responses. Despite TRPC1 belonging to a different subfamily from TRPC4 and TRPC5, in the brain, TRPC1 frequently associates with TRPC4 and TRPC5 to form heterotetramers (Zong and others 2023). Thus, here we discuss TRPC1 alongside with TRPC4 and TRPC5. Moreover, TRPC3, TRPC6, and TRPC7 share structural similarities and usually form heterotetramers in vivo (Bon and others 2022). As TRPC1/4/5, TRPC3/6/7 are widely expressed in different cell types in the brain, they are involved in regulating glutamate signaling, neuronal survival, neurite growth, and synaptic transmission (Rather and others 2023).

TRPC1 in ischemic excitotoxicity.

Glutamate incubation in hippocampal organotypic slice cultures resulted in a rapid increase of TRPC1 expression, accompanied with wide-spread cell death (Narayanan and others 2008). However, this deleterious effect was inhibited by TRPC1 knockdown, suggesting a detrimental role of TRPC1 in glutamate-induced excitotoxicity (Narayanan and others 2008). More work will be needed to illustrate the significance of this interaction in different neurologic diseases.

TRPC6 in ischemic excitotoxicity.

In a rat MCAO model, a progressive down-regulation in TRPC6 expression in the peri-infarct area preceded ischemic neuronal death (Du and others 2010). Overexpression of TRPC6, rather than dominant negative TRPC6, conferred protection against OGD-induced neuronal death in a CREB-dependent manner, suggesting TRPC6’s role in promoting prosurvival signaling (Du and others 2010). Further evidence demonstrated that the down-regulation of TRPC6 was mediated by calpain cleavage at Lys16, a process contingent upon NMDAR-mediated Ca²⁺ influx (Du and others 2010) (Fig. 5). Blockade of TRPC6 degradation by calpain using the specific interfering peptide TAT-C6 protected against ischemic neuronal death in vitro and ameliorated ischemic stroke in mice (Du and others 2010). This protective effect was associated with enhanced activation of CREB signaling, indicating that TRPC6 is a downstream molecule of esNMDAR-mediated excitotoxicity (Du and others 2010). Subsequent studies revealed a reciprocal inhibition between TRPC6 and NMDAR activity. In rat hippocampal neurons, TRPC6-mediated Ca²⁺ influx was found to suppress NMDAR-mediated Ca²⁺ influx in a calcineurin-dependent manner (Li and others 2012). This observation is very interesting, as in the previous study, it was shown that NMDAR promoted TRPC6 degradation also through its Ca²⁺ influx (Du and others 2010). Moreover, TRPC6 overexpression inhibited, while TRPC6 knockdown aggravated the NMDA-induced Ca²⁺ overload (Li and others 2012) (Fig. 5). At the animal level, TRPC6 transgenic mice exhibited reduced infarct size and improved neurobehavioral outcomes (Li and others 2012). These findings collectively suggest that during ischemic stroke, TRPC6 activation confers benefit by inhibiting NMDAR-mediated excitotoxicity, while NMDAR exacerbates its detrimental effects by promoting TRPC6 degradation.

Figure 5.

TRPC6–N-methyl-d-aspartate receptor (NMDAR) interaction. Reciprocal regulation occurs between TRPC6 and NMDAR in ischemic excitotoxicity. First, TRPC6-mediated Ca²⁺ influx inhibits NMDAR activation via calcineurin. Conversely, NMDAR-mediated Ca²⁺ influx leads to the down-regulation of TRPC6 through calpain-dependent degradation at lysine 16.

TRPVs in Ischemic Excitotoxicity

TRPV1, renowned for its heat-sensitive properties in the dorsal root ganglion (DRG), garnered significant attention with the Nobel Prize in 2021 (Shuba 2020). Beyond being gated by heat, TRPV1 can be modulated by a plethora of exogenous and endogenous substances, including capsaicin, related compounds, animal vanillotoxins, acidic pH, endocannabinoids, arachidonic acid derivatives, and polyamines (Shuba 2020). Besides DRG neurons, TRPV1 is also expressed in the brain and plays important roles in several neurologic disorders, including pathologic pain and stroke (Zong and others 2023). While TRPV1’s association with stroke has been extensively studied, its role seems to be primarily linked to its thermosensitivity and its ability to regulate body temperature, potentially brain temperature (Zong and others 2023). Thus, this mechanism appears to be independent of excitotoxicity.

TRPV4 in ischemic excitotoxicity.

TRPV4 is widely expressed in the brain, playing a crucial role in regulating various essential physiologic functions, including glial activation and maintaining blood-brain barrier (BBB) integrity (Zong and others 2023). Mutations in TRPV4 have strong associations with several human neuropathies (Toft-Bertelsen and MacAulay 2021). While TRPV4 is not regulated by as many ligands as TRPV1, its activity can be influenced by phosphorylation (Toft-Bertelsen and MacAulay 2021). A significant characteristic of TRPV4 is its activation by mechanical stretch induced by cellular volume expansion (Toft-Bertelsen and MacAulay 2021). This feature becomes particularly important in the context of cellular swelling observed in inflammatory neurodegeneration, suggesting a potential role of TRPV4 activity in this process (Zong and others 2023).

The TRPV4 agonist 4α-PDD or activation of TRPV4 through hypotonic stimulation has been observed to enhance NMDA-induced current in hippocampal CA1 pyramidal neurons in a CaMKII- and GluN2B-dependent manner (Li and others 2013). Notably, the administration of the TRPV4 inhibitor HC-067047 resulted in a reduction in infarct size in a mouse model of MCAO, suggesting that TRPV4 contributes to ischemic excitotoxicity by potentiating NMDAR activation (Li and others 2013). In a separate study, TRPV4 agonists were found to inhibit hippocampal CA1 pyramidal neurons, and this inhibition was associated with PKC activation (Hong and others 2016). This study implies that the TRPV4-induced inhibition of GABA receptors, combined with the potentiation of NMDAR, may work synergistically to exacerbate neuronal hyperexcitability under ischemic conditions, further contributing to neuronal death.

Beyond the direct modulation of neuronal NMDAR, TRPV4 has been found to facilitate glutamate release in glial cells. Approximately 30% of astrocytes in the brain express TRPV4 (Shibasaki and others 2014). 4αPDD and arachidonic acid have been shown to increase the frequency of miniature excitatory postsynaptic currents (mEPSCs) in neurons cocultured with wild-type astrocytes compared to those with TRPV4 knockout astrocytes (Shibasaki and others 2014), suggesting that TRPV4 activation promotes excitatory transmitter release and enhances synaptic activity. Amino acid analysis revealed that 4αPDD treatment specifically augmented glutamate release in wild-type astrocytes but not in TRPV4 knockout astrocytes (Shibasaki and others 2014). Consistent with these observations, TRPV4 has also been shown to induce Ca²⁺ influx in both astrocytes and neurons, accompanied with the accumulation of extracellular glutamate near astrocytes during peri-infarct depolarization in a mouse MCAO model (Rakers and others 2017). These findings collectively indicate that TRPV4-mediated enhancement of astrocytic glutamate release may exacerbate ischemic excitotoxicity.

TRP Channels in Other Neurodegenerative Diseases Related to Excitotoxicity

In addition to ischemic excitotoxicity, TRP channels are involved in a variety of neurodegenerative diseases caused by glutamate excitotoxicity, including epilepsy, bipolar disease, and Alzheimer’s disease (Table 1).

Table I.

GluR-Interacting TRP Channels.

	Involved Disorders/Diseases	Involved Cell Types	Interacting GluRs
TRPM2	Stroke	Neuron	NMDAR
	Alzheimer’s disease	Neuron	NMDAR
	Bipolar disorders	Neuron	NMDAR
TRPM4	Stroke	Neuron	Undetermined
	Encephalomyelitis	Neuron	Undetermined
	Epilepsy	Neuron	KAR
TRPC1	Excitotoxicity in vitro	Neuron	Undetermined
	Epilepsy	Neuron	mGluR
TRPC4	Epilepsy	Neuron	mGluR
TRPC5	Epilepsy	Neuron	KAR
TRPC3	Epilepsy	Neuron	Undetermined
TRPC6	Stroke	Neuron	NMDAR
	Epilepsy	Neuron	Undetermined
TRPC7	Epilepsy	Neuron	NMDAR
TRPV1	Glutamate-induced hyperalgesia	Neuron	NMDAR
	Inflammatory pain	Neuron	mGluR
TRPV4	Stroke	Neuron, astrocyte	NMDAR, (GABAR), glutamate release
	Epilepsy	Neuron	NMDAR

Summary of TRP-GluR interactions in excitotoxicity. Current studies of excitotoxicity predominantly focus on neurons, overlooking the role of TRP-GluR interactions in nonneuronal cells in the brain and in cells in the peripheral. Please be aware that beyond the TRP channels outlined in this table, many other TRP channels have been implicated in excitotoxicity-related disorders/diseases. However, their interactions with GluRs are yet to be determined.

GABAR, γ-aminobutyric acid (GABA) receptor; GluR, glutamate receptor; KAR, kainate receptor; mGluR, metabotropic glutamate receptor;

NMDAR, N-methyl-d-aspartate receptor.

TRP Channels in Epilepsy

Several TRP channels, including TRPC1, TRPC3, TRPC4, TRPC5, TRPC7, TRPV4, TRPM2, and TRPM4, which can cause excitotoxic neuronal death, have been suggested to play a role in epilepsy (Fig. 6).

Figure 6.

Transient receptor potential (TRP) channels and excitotoxicity in epilepsy. The role of TRP–glutamate receptor (GluR) interaction during excitotoxicity in epilepsy is not as thoroughly investigated as in ischemic excitotoxicity. While several TRP channels have been implicated in the induction of neuronal hyperactivity and injury/death by GluR agonists and have been associated with seizure activities in mice, the precise mechanisms through which TRPs influence GluR activation and whether TRPs’ activation directly increases neuronal activity and death remain unclear. Also, it is unclear whether canonical TRPCs (TRPCs) are activated secondarily to metabotropic GluR (mGluR) activation and whether this activation contributes to neuronal hyperactivity and injury/death.

TRPC1, TRPC4, and TRPC5 in epilepsy.

In experiments using mGluR1/5 agonist 1-aminocyclopentane-trans-1,3-dicarboxylic acid (ACPD) to induce epileptiform activity, TRPC inhibitor SKF96365 effectively blocked burst firings and plateau potential in lateral septal neurons (Phelan and others 2012). In mice, TRPC1 single-knockout partially attenuated mGluR-mediated epileptiform activity, while TRPC4 single-knockout or TRPC1/4 double-knockout completely abolished this response (Phelan and others 2012). Furthermore, the observed seizure-induced neurodegeneration in the dorsolateral septal nucleus and hippocampus was mitigated in TRPC1/4 double-knockout mice, indicating the contribution of heteromeric TRPC1/4 channels to mGluR-mediated excitotoxicity in epilepsy (Phelan and others 2012). TRPC5 appears to engage in seizures through a distinct molecular pathway. TRPC5 knockout attenuated pilocarpine-induced seizures and hippocampal neuronal death in mice (Phelan and others 2013). Interestingly, this protective effect seemed unrelated to mGluRs, as ACPD-induced epileptiform bursting remained unaltered in TRPC5 knockout mice (Phelan and others 2013). Instead, TRPC5’s involvement in reducing high-frequency stimuli-induced LTP suggests its potential contribution to NMDAR-mediated excitotoxicity (Phelan and others 2013). In another study, inhibiting TRPC5 using NU6027 mitigated neuronal death in the pyriform cortex, amygdala, and hippocampus in a rat model of seizures induced by kainate, suggesting that TRPC5 contributes to kainate excitotoxicity in epilepsy (Park and others 2019).

TRPC3, TRPC6, and TRPC7 in epilepsy.

TRPC3 and TRPC6 were shown to be abundantly expressed in the dendrites of pyramidal cells and the cell bodies of dentate granule cells in the hippocampus (Kim and others 2013). Interestingly, the expression of TRPC3 increased, while TRPC6 expression was reduced in pyramidal and dentate granule neurons after LiCl-induced status epilepticus (Kim and others 2013). Correspondingly, intracerebroventricular infusion of TRPC3 inhibitor pyrazole 3 and TRPC6 activator hyperforin ameliorated neuronal death after status epilepticus, implicating a detrimental role for TRPC3 and a protective effect for TRPC6 in epilepsy (Kim and others 2013). Given TRPC6’s notable contribution to NMDAR-mediated excitotoxicity, the involvement of TRPC3 and TRPC6 in epilepsy is likely linked to glutamate-related excitotoxic processes (Kim and others 2013). TRPC7 knockout selectively inhibited gamma activity escalation induced by pilocarpine and prevented status epilepticus (Phelan and others 2014). Also, TRPC7 knockout suppressed epileptiform burst firing in CA3 pyramidal neurons, accompanied with reduced LTP at CA3 and CA1 synapses induced by high-frequency stimulation (Phelan and others 2014). Meanwhile, TRPC7 knockout did not affect spontaneous burst firing triggered by mGluR agonist ACPD (Phelan and others 2014). These findings indicate that TRPC7 activation contributes to seizure activities by enhancing NMDAR-dependent LTP.

TRPV4 in epilepsy.

Recent studies have revealed the involvement of TRPV4 in soman-induced status epilepticus. Rats exposed to soman, a chemical warfare agent, exhibited a substantial increase in TRPV4 expression in the hippocampus (Wang and others 2021b). Treatment with the TRPV4 blocker GSK2193874 significantly mitigated the severity of soman-induced seizures and reduced mortality rates in rats (Wang and others 2021b). Additionally, mice with TRPV4 knockout displayed a similar protective effect against seizures (Wang and others 2021b). The underlying mechanism involves the inhibition of NMDAR-mediated glutamate excitotoxicity and concurrent suppression of NLRP3 inflammasome-mediated neuroinflammation following TRPV4 knockout or inhibition (Wang and others 2021b). While this role of TRPV4 in soman-induced status epilepticus is established, further investigation is required to assess the contribution of TRPV4 in other types of epilepsy.

TRPM4 in epilepsy.

Recent evidence suggests that TRPM4 may contribute to excitotoxicity in temporal lobe epilepsy (Mundrucz et al., 2023). TRPM4 knockout inhibited the increase of spontaneous action potential frequency in hilar mossy cells of brain slices induced by glutamate challenge. Also, TRPM4 knockout protected hilar mossy cells against kainate-induced death in vivo, companied with a decreased seizure susceptibility. This study suggests that in addition to NMDAR, TRPM4 may also modulate the function of kainate receptor and the related excitotoxicity.

Potential Role of TRP Channels in Other Excitotoxic Neurodegenerative Diseases

TRPM2 in bipolar disorder.

TRPM2’s role in NMDAR-dependent long-term depression (LTD) was revealed in brain slices obtained from TRPM2 knockout mice, where NMDA application failed to induce LTD. This deficiency was linked to reduced phosphorylation of glycogen synthase kinase-3β (GSK-3β) (Xie and others 2011). Conversely, another study revealed that TRPM2 activation led to the dephosphorylation of GSK-3α and GSK-3β, while TRPM2 knockout mice exhibited elevated levels of phosphorylated GSK-3α and GSK-3β in the brain (Jang and others 2015). Considering GSK-3’s involvement in regulating NMDAR activity and its significance in promoting bipolar disorder (Jang and others 2015), these findings hint at a potential role of TRPM2 in NMDAR-mediated excitotoxicity associated with bipolar disorder.

TRPM2 in Alzheimer’s disease.

In addition to TRPM2’s modulation of NMDAR activity, there is evidence indicating that NMDAR can also influence TRPM2 function (Fig. 4). In CA1 pyramidal neurons from hippocampal slices, NMDA perfusion triggered TRPM2 activation, likely initiated by the NMDAR-mediated influx of Ca²⁺, considering that TRPM2 activation requires Ca²⁺ (Olah and others 2009). In another study, Aβ oligomer treatment sensitizes NMDA-induced TRPM2 currents in cultured neurons (Fig. 4), and TRPM2 knockout mitigated cognitive deficits and memory impairments in an Alzheimer’s mouse model by inhibiting endoplasmic reticulum stress and abnormal microglial activation (Ostapchenko and others 2015). These findings underscore a potential role of TRPM2 in exacerbating NMDAR-mediated excitotoxicity in Alzheimer’s disease.

TRPM4 in encephalomyelitis.

NMDAR appear to potentiate TRPM4 activity, contributing to neurodegeneration associated with glutamate excitotoxicity. Active demyelinating axons in brain lesion tissue from multiple sclerosis patients and white matter lesions in experimental autoimmune encephalomyelitis (EAE) mice demonstrated a marked increase in TRPM4 expression (Schattling and others 2012). Remarkably, both TRPM4 knockout and the TRPM4 inhibitor glibenclamide offered protection against acute axonal injury in the EAE model, independent of immune cell infiltration (Schattling and others 2012). Mechanistically, observations in cultured hippocampal neurons indicated that GluR-mediated Ca²⁺ influx may stimulate TRPM4 activation. Moreover, glutamate pretreatment enhanced TRPM4 activation in cultured hippocampal neurons, and TRPM4 knockout mitigated glutamate-induced excitotoxicity in these neurons (Schattling and others 2012), suggesting that GluR-mediated Ca²⁺ influx might stimulate TRPM4 activation. Later, two more specific TRPM4 inhibitors, CBA and NBA, were also shown to inhibit glutamate-induced neurodegeneration in vitro. Collectively, these findings indicate the potential role of TRPM4 in acute/subacute neurodegenerative diseases, such as encephalomyelitis and multiple sclerosis.

TRPV1 in glutamate-induced hyperalgesia and pain.

Both TRPV1 and NMDAR are implicated in inflammatory pain and hyperalgesia. Studies using TRPV1-specific blocker AMG9810 demonstrated its ability to attenuate NMDA-induced mechanical hyperalgesia in the masseter muscle (Lee and others 2012). NMDAR and TRPV1 are coexpressed in trigeminal sensory neurons and TRPV4 physically associated with GluN1 and GluN2B. Through CaMKII and PKC, NMDA perfusion increased TRPV1 phosphorylation and enhanced the TRPV1-mediated Ca²⁺ influx trigeminal sensory neurons (Lee and others 2012). Similarly, TRPV1 was found to participate in mGluR1/5-induced hyperalgesia. Inhibition of TRPV1 alleviated mechanical hyperalgesia induced by (R,S)-3,5-dihydroxyphenylglycine (DHPG), an mGlu1/5 agonist, accompanied by increased TRPV1 phosphorylation at S800 mediated by PKC in trigeminal ganglia (Chung and others 2015). These findings suggest that glutamate triggers hyperalgesia by promoting TRPV1 activation via NMDAR and mGluR1/5. Moreover, TRPV1 has been observed to modulate NMDAR phosphorylation. Administration of the TRPV1 antagonist capsazepine attenuated carrageenan-induced inflammatory pain in mice, accompanied with inhibited phosphorylation of GluN1 and GluN2B in the spinal cord (Kang and others 2021).

Perspective

The contributions of TRP channel activity in glutamate excitotoxicity, especially NMDAR-mediated neurotoxicity, has been extensively studied. Emerging evidence suggests that TRP-GluR interaction is a promising target for developing novel therapeutic strategies for excitotoxicity-related neurologic disorders. However, the wide expression pattern of GluRs across diverse cell types in both the brain and the periphery suggests a need to broaden research perspectives. There is a particular gap in understanding the TRP-GluR coupling in nonneuronal cells in the brain, such as glial cells and cerebral endothelial cells. Moreover, the pathophysiologic role of GluRs and their “excitotoxicity” in peripheral cells has been largely overlooked. For instance, both TRPM2 and TRPM4 are significant regulators of immune cell functions, and recent findings showed that NMDARs control macrophage polarization. Thus, it is plausible that the coupling between TRPM2/4 and NMDARs may also occur in immune cells and exert crucial roles in modulating immune cell functions. The importance of the interactions between TRP channels and GluRs in various pathologic conditions within the CNS implies that investigating these interactions in the periphery could also yield valuable insights into excitotoxic neurodegenerative disorders. Therefore, exploring the interplay between TRP channels and GluRs in peripheral tissues is equally promising and should not be neglected.