Purinergic P2X7 receptor signaling in anxiety: Roles of stress-related neuroinflammatory mechanisms and therapeutic horizons

Abstract

Anxiety is a psychological state marked by constant uneasiness, fear, or stress in response to an actual or perceived danger, resulting in physical changes such as elevated cardiac activity, poor concentration, and restlessness. The excessive stimulation of these responses results in anxiety disorders that disrupt daily activities and even impact familial and social life. The purinergic P2X7 receptor (P2X7R), a ligand-gated ion channel, is predominantly expressed in microglia and serves as a vital detector of cellular stress and injury in the central nervous system (CNS). P2X7R activation requires a high concentration of extracellular adenosine triphosphate (ATP); therefore, it is closely associated with cellular stress. Research has demonstrated that chronic stress inhibits Na⁺/K⁺-ATPase (NKA1) on the cell surface of microglia, thereby increasing the P2X7R expression. The hyperactivity of this receptor contributes to neuroinflammation and, consequently, promotes anxiety-like behaviours. The role of P2X7R in psychostimulant addiction, such as methamphetamine (METH), is complex, but increasing evidence illustrates its activation with neuroinflammation, relapse, and increased dependency. The cumulative stress activates the P2X7R, resulting in amplification of interleukin-17A (IL-17A) levels in the prefrontal cortex (PFC) and amygdala, which intensifies anxiety levels during a psychostimulant withdrawal. Future investigation should concentrate on the development of highly selective P2X7R antagonists and their ability to transverse the blood–brain barrier (BBB), as well as the incorporation of genetic information to enhance personalised therapy methods. This article aims to explore the molecular pathways and neurobehavioral processes of P2X7R in anxiety, highlight therapeutic advances, and outline future directions for translational neuropharmacology.

Introduction

Anxiety is a mental health condition characterized by persistent distress, uneasiness, and nervousness, often manifested by symptoms including restlessness, increased heart rate, poor concentration, irritability, and sleep disturbances, which can significantly disrupt daily activities and quality of life. It is triggered by hyperactivity in the amygdala and limbic system, along with dysfunction in the PFC, causing exaggerated fear and threat responses, while dysregulation of key neurotransmitters, including dopamine (DA), gamma-aminobutyric acid (GABA), norepinephrine (NE), and serotonin (5-HT), causes heightened neuronal excitability [1]. Anxiety disorders affect approximately 301 million people, and have become the leading mental health condition across the globe [2]. The P2X7R is a purinergic ion channel, primarily stimulated by high extracellular ATP, and is abundantly present on microglia, astrocytes, and various peripheral immune cells. It plays a key role in promoting the release of pro-inflammatory cytokines, driving neuroinflammatory activity, and causing neuronal loss. It differs functionally from other purinergic receptors as prolonged activation by ATP enables it to create large non-selective membrane pores, support the release of inflammatory molecules, and modulate several cellular processes [3]. Preclinical studies show that stimulating P2X7R in immune cells triggers the release of interleukin-1β (IL-1β) and tumor necrosis factor-alpha (TNF-α), following chronic neuroinflammation that can disrupt neurotransmitter secretion, including 5-HT and GABA, and promote anxiety-like behavioral outcomes [4]. The association between the P2X7R signalling and the anxiety-like behaviour has recently started to be unravelled due to preclinical and clinical studies. This link is well supported by experimental evidence showing that mice lacking the P2X7R gene or treated with P2X7R antagonists exhibit fewer anxiety-like behaviors, whereas P2X7R overactivation or upregulation yields more anxiety-like behaviors [5]. The accumulating evidence indicates that neuroinflammation is strongly related to the occurrence of mood and anxiety disorders [6, 7]; thus, P2X7R has become a key participant in the neurobiological process of anxiety [8, 9]. The genetic deletion of P2X7R or pharmacological inhibition in animal models has been revealed to decrease anxiety-like behaviour, supporting its potential as a novel therapeutic target [10]. Selective P2X7R antagonists would represent a new generation of anxiolytic agents, designed to modulate neuroinflammatory pathways, offering more targeted and effective therapeutic options for patients with anxiety disorders. Nevertheless, these methods still need to be proven through further studies, and the exact mechanisms of their effect, safety, and effectiveness in humans should be verified. The growing evidence demonstrated that P2X7R dysregulation may contribute to both the onset and worsening of anxiety symptoms, potentially via neuroinflammatory mechanisms and impaired neuronal communication. The potential pharmacological regulation of this receptor is being pursued in preclinical and clinical research, which provides directions for new therapies with higher effectiveness and fewer side effects than those currently available. P2X7R activation also contributes to METH-induced rewarding and locomotor stimulant effects by modulating IL-17A, dopamine transmission, and microglial activation, correlated with anxiety symptoms [11]. Preclinical and clinical studies show that METH withdrawal induces anxiety-like behaviors, reduced locomotion, and increased inflammatory cytokines. Nevertheless, more studies are required to completely characterize the P2X7R relation with anxiety and METH withdrawal, as well as to establish safe and targeted treatment. This article aims to evaluate the current insights into the P2X7R role in anxiety, while also exploring its therapeutic potential for future research and clinical development in this disease.

Structure and functional properties of P2X7R

Structural features of P2X7R

P2XR contains a family of conserved proteins and is found in a diverse range of species, including protozoa and algae, as well as mammals. It functions as a purinergic membrane receptor that forms a cation-permeable channel following activation of extracellular ATP [12]. The P2XR receptor family comprises seven distinct subtypes, P2X1 to P2X7, each encoded by a different gene [13]. Encoded by the P2X7R gene, the P2X7R displays unique structural and functional properties and is broadly expressed in mammalian cells, particularly haematopoietic cells and immune cells, including monocytes, macrophages, and lymphocytes [14]. The P2X7R gene on chromosome 12 encodes each of its subunits, which contains 595 amino acids due to the 13 exons of the messenger ribonucleic acid (mRNA) transcript. This receptor consists of an extracellular domain positioned between two transmembrane helices (TM1 and TM2) and with intracellular N- and C-terminal tails. It contains 3 subunits assembled to form a functional trimeric channel, lined by TM2 segments that govern channel opening and ion selectivity, while three ATP-binding sites are located at subunit interfaces [15]. Transient ATP released by multiple cellular sources during stress and tissue injury triggers calcium (Ca²⁺), sodium (Na⁺) influx, and potassium (K⁺) efflux [16]. Sustained and repeated activation causes an irreversible pore dilation, enabling the passage of hydrophilic molecules with a molecular weight of up to 900 Da through the cell membrane. It is primarily inactive under typical normal conditions, as extracellular ATP is maintained at nanomolar concentrations [17]. Nevertheless, extracellular ATP increases markedly following stress or tissue injury and serves as a damage-associated molecular pattern (DAMP) [18]. This triggers P2X7R and engages nuclear factor kappa B (NFκB), promoting the synthesis of pro-interleukin-1β (pro-IL-1β), pro-interleukin-18 (pro-IL-18), and NLR family pyrin domain-containing 3 (NLRP3) proteins. Its activation induces K⁺ efflux, facilitating NRLP3 formation and caspase-1 activation, culminating in the release of IL-1β and IL-18. This signalling is especially active in microglia rather than astrocytes, resulting in the secretion of TNF-α and interleukin-6 (IL-6) and exacerbates neuroinflammation [19].

Physiological role of P2X7R

P2X7R differs from other P2X receptor subtypes structurally and functionally, requiring significantly higher concentrations of ATP, generally exceeding 100 μM for activation. Increased extracellular ATP is produced in the brain during cellular stress, injury, or inflammation and activates the P2X7R [20]. This receptor forms a non-selective cation channel, allowing entry of Ca²⁺ and Na⁺ ions and exit of K⁺ ions. It undergoes a structural rearrangement that creates large, non-selective membrane pores, allowing molecules with molecular weight up to 900 Da, such as ATP, nicotinamide adenine dinucleotide (NAD⁺), and certain metabolites to pass through. P2X7R is expressed most extensively on microglia, the brain’s immune cells, and is a mediator of neuroinflammatory responses in the CNS. ATP activates the receptor and promotes the NLRP3 signalling, which facilitates the activation and maturation of IL-1β and IL-18 [21]. These cytokines are essential for immune responses, but their excessive production causes neuronal damage, synaptic dysfunction, and chronic neuroinflammation, making P2X7R a pivotal regulator of the innate immune system during extracellular threats or stimuli [22]. It also influences neuronal communication by modulating the release of neurotransmitters and neuromodulators, and overactivation of astrocytes can stimulate the release of ATP and glutamate, excitatory brain neurotransmitters [23]. This excessive or dysregulated glutamate release can contribute to excitotoxicity, linked to neurodegenerative and mental disorders, and P2X7R also influences neuronal inhibition by modulating GABAergic signalling. Thus, P2X7R contributes to maintaining the neurotransmitter equilibrium necessary for healthy brain function [24]. The persistent activation of P2X7R can lead to the formation of large membrane pores, which affect cell viability. The transient activation of P2X7R supports normal cellular functions such as proliferation and differentiation of neural progenitors in normal conditions. However, long-term or excessive activation leads to apoptosis or necrosis due to the disruption of ionic homeostasis, decreased mitochondrial activity, and the initiation of the release of pro-apoptotic proteins [25]. This dual role highlights the fact that, in different circumstances, it can contribute to both normal tissue homeostasis and pathological cell death.

Pathophysiological roles of P2X7R in stress, neuroinflammation, and anxiety

Pathophysiological role of P2X7R in stress

The stress response refers to the coordinated physiological and behavioral response of the body to any noxious stimulus to maintain or restore homeostasis [26]. Stress is generally categorized based on its duration as acute and chronic stress. Acute or short-term stress, particularly in healthy individuals, is often well tolerated and can even promote adaptive physiological responses without causing serious damage. In contrast, long-term stress is associated with harmful health consequences. The evidence from clinical and animal studies indicates that chronic stress is a significant risk factor for numerous conditions, including cardiovascular diseases, neurodegenerative disorders, depression, endocrine dysfunction, and cancer [27]. Chronic stress enhances ATP release via pannexin-1 hemichannels and vesicular exocytosis, resulting in sustained elevation of extracellular ATP that exceeds the clearance capacity of ectonucleotidases, and serves as a DAMP signal that strongly activates microglial P2X7R [28]. The increased extracellular ATP in stress is an effective activator of the P2X7R, causing rapid influx of Ca²⁺ and Na²⁺ ions and an outflow of K⁺ ions. The prolonged activation leads to the formation of a large membrane pore, which often results in necrosis or cell apoptosis [29]. These ion alterations also activate pannexin-1 channel, promoting further ATP release into the extracellular space and thereby amplifying P2X7R signaling. This transient channel activation is associated with enhanced cell proliferation and survival, whereas pore formation strongly promotes inflammasome activation. Inflammasomes are cytoplasmic multiprotein assemblies consisting of a sensor molecule such as NLRP1, an inflammatory caspase, and an adaptor protein known as apoptosis-associated speck-like protein containing a CARD (ASC). Once assembled, the inflammasome converts pro-caspase-1 into active caspase-1, which then enables the maturation and release of pro-inflammatory cytokines IL-1β and IL-18, ultimately triggering inflammation and pyroptotic cell death [30]. This enhances P2X7R expression on hippocampal microglia, promoting neuroinflammatory responses and oxidative stress that disrupts BBB integrity and normal neuronal and glial functioning. Animal studies illustrate that stress triggers the release of ATP in the brain and activates P2X7R. This activation stimulates the production of IL-1β, which subsequently acts on neuroendocrine pathways to increase the release of corticotropin-releasing hormone (CRH). Elevated CRH stimulates adrenocorticotropic hormone (ACTH) secretion from the pituitary, thereby disrupting the hypothalamic–pituitary–adrenal axis (HPA-axis) and promoting stress-related mood disturbances [31].

Pathophysiological role of P2X7R in neuroinflammation

Neuroinflammation refers to the activation of immune reactions in the CNS, driven by the release of cytokines, chemokines, reactive oxygen species (ROS), and other signalling molecules [32]. These inflammatory mediators are produced by microglia, astrocytes, and endothelial cells within the CNS and also by peripheral immune cells. Animal studies demonstrated that the activation of microglial stimulation and release of inflammatory cytokines are closely linked to behavioral transformations that are similar to anxiety and depression [33]. Microglia contribute significantly to neuroprotection, regulation of neuronal signals, and synaptic remodelling, and are highly responsive to even minor changes in the brain’s internal environment to maintain and regulate the body’s stress responses. Chronic stress activates P2X7R by releasing extracellular ATP, leading to the inflow of Ca²⁺ and Na²⁺ ions and an outflow of K⁺ ions [29]. This leads to prolonged activation of microglial cells and increases the release of pro-inflammatory cytokines, especially in PFC and limbic areas [34]. Notably, the outflow of K⁺ ions serves as a key signal for activating the NLRP3 inflammasome, an essential component of the innate immune system in microglia [35]. The NLRP3 inflammasome triggers cleavage and activation of caspase-1, an enzyme that converts pro-inflammatory cytokines such as pro-IL-1β and pro-IL-18 into their active forms for extracellular release. These cytokines are crucial for promoting and sustaining neuroinflammation by signalling to surrounding cells and recruiting peripheral immune cells to the brain [36]. It also leads to the activation of IкB kinase (IKK) complexes that phosphorylate and degrade the inhibitory protein IкBα, allowing the release of the NFкB complex, especially the p65 subunit, to translocate into the nucleus. NFкB binds to DNA promoter regions of pro-inflammatory genes, increasing the transcription of TNF-α, IL-6, and inducible nitric oxide synthase (iNOS) along with ROS [37]. These inflammatory mediators affect neurons by disrupting their normal function and connectivity by enhancing glutamatergic excitatory signaling while impairing GABAergic inhibitory transmission, leading to increased neuronal excitability in anxiety-related brain regions such as the amygdala, hippocampus, and PFC [38]. Neuronal hyperexcitability in the amygdala amplifies threat perception, fear responses, and emotional reactivity. In the hippocampus, it impairs the differentiation between safe and threatening stimuli, promoting generalized fear and dysregulated stress responses. The overactive PFC neurons diminish their ability to regulate amygdala activity, leading to impaired control of fear circuits and heightened anxiety [39]. Figure 1 illustrates the signalling cascade initiated by P2X7R activation, including ATP binding, assembly of NLRP3 inflammasome, and subsequent release of cytokines (IL-1β, IL-18) that drive neuroinflammation.

Fig. 1.

Mechanistic pathways linking P2X7 receptor activation to neuroinflammation and synaptic dysfunction. This diagram shows key pathways, including TrkB signalling disruption, glutamate release, mitochondrial dysfunction, ROS generation and NLRP3 inflammasome activation, leading to IL-1β and IL-18 release and neuroinflammatory cascades linked to anxiety and chronic stress. Abbreviation: TLR, Toll-like receptor; BDNF, brain-derived neuroprotective factor; P2X7R, purinergic receptor P2X7; TrkB, tropomyosin receptor kinase B; RAS, rat sarcoma (GTPase); Grb, growth factor receptor-bound protein; Raf, rapidly accelerated fibrosarcoma protein; MEK, mitogen-activated protein kinase kinase; ERK1/2, extracellular signal-regulated kinases 1 and 2; RSK, ribosomal S6 kinase; CREB, cAMP response element-binding protein; AMPA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor; NMDA, N-methyl-D-aspartate receptor; PI3K, phosphoinositide 3-kinase; AKT, protein kinase B; PP1, protein phosphatase 1; CaMKKII, calcium/calmodulin-dependent protein kinase kinase II; AMPK, AMP-activated protein kinase; mTOR, mammalian target of rapamycin; PKC, protein kinase C; MAPK, mitogen-activated protein kinase; ROS, reactive oxygen species; LC3, microtubule-associated proteins 1A/1B light chain 3; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; NLRP3, NOD-, LRR- and pyrin domain-containing protein 3; ASC, apoptosis-associated speck-like protein containing a CARD; IL-1β, interleukin-1 beta; IL-18, interleukin-18; GSDMD, gasdermin D

Pathophysiological role of P2X7R in anxiety

Anxiety disorders are commonly marked by persistent and excessive concerns about potential future threats, where anticipated negative events themselves become potent triggers of anxiety. Such responses can be initiated by environmental stimuli, while fear is strongly influenced by prolonged stress exposure and prior adverse experiences [40]. Consequently, therapeutic approaches for anxiety frequently emphasize stress regulation and the management of fear-related cues. Interpersonal frameworks of comorbidity propose that anxiety disorders can impair social and related functioning, which in turn heightens vulnerability to depressive illness. This model highlights how relational dysfunction can act as a pathway linking anxiety to the later emergence of depression. The P2X7R is primarily expressed on non-neuronal cells, including microglia, astrocytes, and oligodendrocytes, and acts as a sensor for extracellular ATP. The accumulation of ATP at the site of stress or injury can activate P2X7R-driven signalling cascades that promote neuroinflammation. During chronic stress, sustained purinergic signaling leads to prolonged ATP elevation, which enhances NLRP3 inflammasome activation and increases IL-1β production. This exaggerated inflammatory response disrupts neurotransmitter homeostasis and synaptic plasticity, thereby contributing to neurobiological changes that favor the development of depression [31].

Role of glutamate dysregulation

Glutamate is the primary excitatory signaling molecule in the CNS, and plays a critical role in learning, memory formation, and synaptic plasticity [41]. It mediates its action through both ionotropic receptors, namely N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), and kainate receptors, as well as metabotropic receptors, which consist of 8 subtypes, metabotropic glutamate receptor 1 (mGluR1) to metabotropic glutamate receptor 8 (mGluR8) [42]. Numerous studies indicate that glutamatergic neurotransmission within the limbic regions is essential for the emergence of anxiety disorders. The disruption of glutamate homeostasis can cause excitotoxicity, with the synaptic glutamate concentration rising to the extent of 100 µM. These elevations are neurotoxic and lead to atrophy and death of neurons [43]. Extracellular ATP released by glial cells in chronic stress activates P2X7R, resulting in elevated intracellular Ca²⁺ level, which activates intracellular kinases, including Ca²⁺/calmodulin-dependent protein kinase II (CaMKII), protein kinase C (PKC), and mitogen-activated protein kinases (MAPKs) [44]. These kinases phosphorylate the GluA1 subunit of AMPA receptors and promote the mobilization of AMPA-containing vesicles to the post-synaptic membrane [45]. The insertion of AMPA receptors at the postsynaptic site increases the glutamate postsynaptic signal, sufficient to remove the magnesium (Mg²⁺)-dependent block from the NMDA receptor. Subsequently, it promotes the opening of the NMDA receptors that permit Ca²⁺ influx into the cell and trigger long-term excitatory postsynaptic signals, resulting in excitotoxic damage [46]. However, the high intracellular Ca²⁺ results in the activation of calpains, phospholipases, and endonucleases, which degrade essential cellular components, including cytoskeletal proteins, membranes, and DNA [47]. Mitochondrial dysfunction impairs cellular energy metabolism and produces ROS formation that drives oxidative stress, further damaging proteins, lipids, and nucleic acids, and subsidiary activation of proteases and lipases triggers the pro-apoptotic factor cytochrome C (cyt C), initiating apoptosis, leading to neuronal loss [48]. These damaged neurons activate microglia and astrocytes, which release inflammatory mediators by P2X7R activation. These inflammatory signals reduce GABA synthesis and release by suppression of glutamic acid decarboxylase (GAD), the enzyme that converts glutamate to GABA. Neurons fire excessively and synchronously, disrupting normal rhythms in cortical and limbic circuits that destabilize information processing, leading to impaired regulation of fear and emotion and resulting in anxiety-like behavior [49]. The activation of P2X7R is also associated with reduced expression of the glutamate/aspartate sodium-dependent transporter (GLAST) or excitatory amino acid transporter 1 (EAAT1), thereby compromising the clearance of extracellular glutamate. The reduced glutamate uptake promotes its extracellular accumulation, resulting in excessive activation of neuronal glutamate receptors and heightened excitatory neurotransmission [50]. Experimental studies in rat hippocampus have shown that P2X7R at the nerve terminals induces the release of glutamate, which further stimulates the release of kainate receptor-mediated GABA efflux by inhibitory interneurons. This GABA-glutamate crosstalk, contributing to increasing excitatory signals, in turn, could lead to the development of anxiety-like behaviors [51]. This response is suppressed by blocking sodium channels with tetrodotoxin (TTX) or by inhibiting kainate receptors using gadolinium or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Additionally, P2X7R antagonists, including brilliant blue G (BBG) and pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), prevent the efflux of ATP or GABA [22]. As shown in Fig. 2, P2X7R activation in astrocytes and microglia modulates glutamate release and clearance, affecting AMPAR and NMDAR function and ultimately impairing synaptic plasticity.

Fig. 2.

Glutamatergic synapse and P2X7R-mediated modulation of neurotransmission and plasticity. This schematic illustrates the roles of astrocytes, presynaptic and postsynaptic neurons, and microglia in glutamate cycling, synaptic transmission, and neuroinflammation. It highlights glutamate–glutamine metabolism, GABAergic regulation, and P2X7R-driven pathways that influence AMPAR/NMDAR function, BDNF release, and synaptic plasticity. Abbreviation: GLT1, glutamate transporter 1; mGlu2/3, metabotropic glutamate receptor 2/3; P2X7R, purinergic receptor P2X7; Panx1, pannexin-1; ROS, reactive oxygen species; BDNF, brain-derived neuroprotective factor; AMPA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor; NMDA, N-methyl-D-aspartate receptor; PP1, protein phosphatase 1; TrkB, tropomyosin receptor kinase B; Akt, protein kinase B; GSK-3, glycogen synthase kinase 3; mTORC1, mammalian target of rapamycin complex 1; CREB, cAMP response element-binding protein; eEF2K, eukaryotic elongation factor 2 kinase; eEF2, eukaryotic elongation factor 2; GABA, gamma-aminobutyric acid; GABA_AR, gamma-aminobutyric acid type A receptor; GAD, glutamate decarboxylase; NLRP3, NOD-, LRR- and pyrin domain-containing protein 3; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; IL-1β, interleukin-1 beta; IL-18, interleukin-18

Role of P2X7R in neuroplasticity

Neuroplasticity is the remarkable capacity of the brain to reorganize itself by forming, modifying, and reinforcing neural connections in response to experiences and interactions with the environment [52]. Nevertheless, chronic stress and anxiety-related conditions are closely correlated with a disruption of this adaptive mechanism, manifesting as reduced synaptic connections and neuronal atrophy, especially within the medial prefrontal cortex (mPFC) and hippocampal region. Extracellular ATP activates P2X7R, promoting NLRP3 inflammasome formation in microglial cells and initiating the triggering of pro-inflammatory responses during prolonged stress. This activation modifies phosphorylation of insulin receptor substrate-1 (IRS-1), impairing tropomyosin receptor kinase B (TrkB) dependent brain-derived neurotrophic factor (BDNF) and subsequently affecting intracellular cascades that regulate plasticity-related gene expression. Chronic inhibition of BDNF expression and signalling impairs neuronal survival and synaptic plasticity, which eventually triggers neuronal atrophy and decreased synaptic function in the mPFC and hippocampus [53]. These structural and functional impairments decrease adaptive capacity and damage learning and stress coping strategies, along with overstimulating activity in limbic circuits controlled by the PFC. The mPFC has a number of downstream areas that play critical roles in affective regulation, including the amygdala, the dorsal raphe nucleus, the lateral habenula, and the bed nucleus of the stria terminalis (BNST) [54]. Chronic stress decreases the BDNF levels in these cortical and limbic areas, especially the hippocampus and PFC, which exacerbates impairments in neuroplasticity [55]. Chronic stress activates P2X7R and suppresses the activity of mammalian target of rapamycin complex 1 (mTORC1), a signalling protein crucial for synaptogenesis and plasticity through modification of phosphoinositide 3 kinase (PI3K)/AKT and AMP-activated protein kinase (AMPK) pathways [56]. AMPK phosphorylates tuberous sclerosis complex 2 (TSC2), enhancing its GTPase-activating activity to inactivate Ras homolog enriched in brain (Rheb), required for mTORC1 activation. It also phosphorylates regulatory-associated protein of mTOR (Raptor), promoting its interaction with 14–3-3 proteins and thereby inhibiting mTORC1 function [57]. Additionally, this impaired the PI3K/AKT pathway that normally stimulates the TSC2 and promotes mTORC1 activation [58]. This blockage of mTORC1 with chronic stress suppresses the process of synapse formation in the PFC and alone is enough to induce the rodents to develop anxiety-like behavior without exposure to stress, which demonstrates the direct causality of emotional behavior and the integrity of the synapses [59]. Stress is also known to cause hypertrophy of the pyramidal neurons of the basolateral amygdala, a structural change that probably leads to the abnormal anxiety states and the disruption of emotional processing. These results demonstrated that chronic stress impairs neuroplasticity in the brain, and the specific mechanisms and effects of this impairment vary depending on the unique functions of different neuronal regions [60]. This maladaptation highlights the complicated nature of the interactions involving neuroinflammation, synaptic remodelling, and the pathogenesis of anxiety disorders.

Role of P2X7R in hypothalamic–pituitary–adrenal (HPA) axis

The HPA axis is a central neuroendocrine pathway coordinating the physiological response of the body to stress factors, and it aids in maintaining homeostasis by regulating hormone secretion and ensuring balance during physiological stress. Its primary response in the state of stress is to promote the release of glucocorticoids, which induce short-term physiological changes to prevent damage from the stressors [61]. Recent research has highlighted P2X7R as an important mediator in the neuroinflammation and synaptic impairment that dysregulates HPA axis activity, contributing to anxiety-like behavior [62]. The increase in extracellular ATP during stress or injury activates P2X7R and P2X7R-mediated inflammatory signalling cascades. P2X7R activation in microglia facilitates the NLRP3 inflammasome formation and triggers the secretion of pro-inflammatory cytokines. These cytokines stimulate the paraventricular nucleus (PVN) of the hypothalamus, enhancing the release of CRH that stimulates the anterior pituitary to secrete ACTH into circulation. Subsequently, the ACTH acts on the adrenal cortex to promote the release of glucocorticoids, predominantly cortisol, thereby sustaining a hyperactive state of the HPA axis [63]. The neuroinflammatory P2X7R signalling can disrupt the negative feedback mechanism that regulates the HPA axis function and results in a prolonged secretion of glucocorticoids, exposing an individual to anxiety-like behaviors [64]. Besides the role in the inflammatory signalling, P2X7R contributes to glutamate imbalances in the PVN hypothalamic area [65]. This causes the stimulation of ionotropic glutamate receptors, specifically NMDA, that excite CRH and arginine vasopressin (AVP)-positive neurons in the PVN. Consequently, this triggers an increase in ACTH secretion and causes an abnormal stress response. It is not only in the PVN that such glutamatergic imbalance increases HPA activity, but also in the median eminence, which is essential for excitatory neurotransmission to control stress. These pathways have been closely related to mood and anxiety disorders. Together, these findings indicate the key role of P2X7R in neuroinflammation or in the maladaptive glutamatergic dysfunction in HPA axis overactivation. The link between cellular stress reactions and neuroendocrine changes involving P2X7R results in the onset and progression of anxiety states [66]. The modulation of this receptor is thus a great treatment method to reinstate regular functions of the HPA axis and to alleviate anxiety-associated symptoms. Figure 3 provides a schematic representation of HPA axis dysregulation under chronic stress, linking glucocorticoid release, P2X7R activation, and neuroinflammation pathways contributing to anxiety disorders.

Fig. 3.

Chronic stress-induced HPA axis activation and P2X7R-mediated neuroinflammation. This schematic shows how prolonged stress elevates cortisol levels, triggering P2X7R activation, and pro-inflammatory cytokine release (IL-1β, IL-18). These events lead to neuroinflammation and anxiety-related pathology. Abbreviation: PVN, paraventricular nucleus; CRH, corticotropin-releasing hormone; AVP, arginine vasopressin; GR, glucocorticoid receptor; ACTH, adrenocorticotropic hormone; HPA, hypothalamic–pituitary–adrenal; P2X7R, purinergic receptor P2X7; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; IL-1β, interleukin-1 beta; IL-18, interleukin-18; NO, nitric oxide; IL-1α, interleukin-1 alpha; TNF-α, tumor necrosis factor alpha; ROS, reactive oxygen species; Ca²⁺, calcium ion; BDNF, brain-derived neuroprotective factor

Addiction links: role of P2X7R in methamphetamine and anxiety

Methamphetamine-induced neurotoxicity and P2X7R activation

Methamphetamine (METH) is a strong CNS stimulant substance, commonly abused recreationally or as a performance-enhancing drug, and exerts its effects primarily by increasing the extracellular levels of monoamine neurotransmitters, especially DA, NE, and 5-HT [67]. Although METH itself is not a primary mechanism of anxiety, its potent neurotoxic and addictive properties robustly engage P2X7R-mediated neuroinflammation, which subsequently contributes to anxiety-like behaviors. Therefore, examining METH-induced activation of P2X7R is essential for understanding the addiction links to components of P2X7R-driven pathology. METH neurotoxicity involves a complex interplay of biological processes, causing the impairment of dopaminergic and serotonergic neurons [68]. The overproduction of DA and its accumulation at the synapse aid dopamine autoxidation, resulting in the formation of ROS. The produced ROS induce neuronal membrane disruption, oxidative stress, lipid peroxidation, protein carbonyl formation, and protein oxidation. METH induced oxidative stress disrupts mitochondrial integrity through swelling, respiratory enzyme inhibition, and upregulation of apoptosis via pro-apoptotic proteins like BCL2-associated X protein (BAX) and BH3-interacting domain death agonist (BID), while simultaneously downregulating anti-apoptotic proteins such as B-cell lymphoma 2 (Bcl-2) [69]. This pathway facilitates the release of cyt C into the cytosol, leading to the activation of caspase-3 [70]. Subsequently, these disturbances, together with endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR), increase neuronal vulnerability and lead to cell death [71]. The cumulative effect of these processes is a significant loss of neurons, particularly those in brain regions that are vital for thinking and mood regulation. Although METH does not directly stimulate the P2X7R; however, it causes cellular stress in the neurons and glia, leading to the increased extracellular release of ATP [72]. This ATP acts as a ligand for P2X7R, increasing the activation of its ion channels that activate the subsequent signalling cascades. P2X7R in itself is activated to facilitate microglial activation, migration, phagocytosis, and secretion of IL-1β, TNF-α, and IL-17A. This neuroinflammatory condition, triggered by this receptor, increases oxidative injury and aggravates neuronal damage. Chronic exposure upregulates the expression of P2X7R in the microglia, thereby amplifying inflammatory reactions [73]. It also induces the NLRP3 inflammasome formation in microglia, activates caspase-1, and facilitates the maturation and release of pro-inflammatory cytokines, thereby facilitating neuroinflammation through persistent activation of microglia and astrocytes [11]. Neuroinflammation worsens glutamatergic dysfunction by impairing glutamate uptake and increasing excessive glutamate release. Excessive extracellular glutamate overstimulates the NMDA receptors and AMPA receptors, resulting in calcium overload, mitochondrial dysfunction, and the enhancement of ROS production. These cascades lower the levels of some important synaptic proteins, including glutamate receptor 5 (GluR5), glutamic acid decarboxylase 65 (GAD65), and ATPase Na⁺/K⁺ transporting subunit α−3 (ATP1a3), therefore destabilizing synaptic plasticity [74]. The breakdown of neuronal connections in the hippocampus, PFC, and striatum causes mood deregulation, cognitive decline, and the development of anxiety-related behaviours. The oxidative damage by METH downregulates tyrosine hydroxylase expression in the striatum, highlighting the disruption of the dopaminergic system. P2X7R-mediated microglial stimulation also affects dopaminergic reward processing in the ventral tegmental area (VTA) and nucleus accumbens by enhancing the release of cytokine [75]. Simultaneously, the glutamate release mediated by P2X7R alters the excitatory input in the DA neurons, enhancing the DA release and intensifying the rewarding quality of METH, thereby facilitating the behavioural sensitisation and addiction. The malfunction of the dopamine transfer disturbs circuits related to anxiety, linking the neuroinflammation that depends on P2X7R to the predisposition to anxiety [76]. Thus, activation of P2X7R also increases the rewarding and locomotor action of METH by increasing mesolimbic DA release, therefore linking neuroinflammation and addictive behaviours. As demonstrated experimentally, repeated METH treatment increases the expression of microglial P2X7R, along with neuroinflammatory and neurotoxic indicators, highlighting the significance of the receptor in METH-induced pathology and related behavioral effects [77]. These processes eventually destroy synaptic and neuronal integrity in regions that control reward and mood, thereby facilitating the addictive properties of METH as well as anxiety-like behaviour that the drug produces. Thus, P2X7R represents a potential therapeutic target for mitigating METH-induced neurotoxicity and alleviation associated neuropsychiatric symptoms [78]. Figure 4 demonstrates that METH exposure amplifies P2X7R expression on microglia, leading to mitochondrial dysfunction, oxidative stress, and dopamine-related neurotoxicity.

Fig. 4.

Methamphetamine-induced neurotoxicity and P2X7R-mediated inflammatory pathways. The diagram highlights key mechanisms, including dopamine oxidation, mitochondrial dysfunction, ER stress, and excessive glutamatergic signalling. It also shows P2X7R activation led to NMDA receptor-mediated Ca²⁺ influx, ROS generation, cytokine release, and caspase-dependent neuronal apoptosis. Abbreviation: DAT, dopamine transporter; VMAT, vesicular monoamine transporter; MAO-B, monoamine oxidase B; COMT, catechol-O-methyltransferase; NMDA, N-methyl-D-aspartate receptor; GluR, glutamate receptor; ROS, reactive oxygen species; CHOP, C/EBP-homologous protein; CytC, cytochrome c; DAQ, dopamine quinone; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; P2X7R, purinergic receptor P2X7; Ca²⁺, calcium ion; ONOO, peroxynitrite; NOO, nitroxyl; ER, endoplasmic reticulum; Caspase-9, caspase 9 protease; Caspase-3, caspase 3 protease; BDNF, brain-derived neuroprotective factor

Neuroimmune-neurotransmitter feedback loops linking P2X7R, anxiety and METH addiction

The neuroimmune response initiated through P2X7R during METH exposure contributes to sustained inflammatory signaling that leads to neurotransmitter dysregulation [79]. METH exposure causes significant alterations in DA release across the mesocorticolimbic reward circuitry, including the VTA, nucleus accumbens, and the PFC, contributing to potent euphoric and reinforcing effects [80]. Nonetheless, METH neurochemical activity extends beyond the DA pathway. It strongly activates glial cells, particularly microglia and astrocytes, largely through ATP-mediated stimulation of P2X7R. It initiates the secretion of pro-inflammatory cytokines and intensifies neuroinflammatory signaling [81]. Beyond acute neurotoxicity, chronic P2X7R activation disrupts dopaminergic and glutamatergic balance, which drives both compulsive drug use and heightened anxiety susceptibility. These signals intensify inflammation-driven glutamate overflow, impair synaptic plasticity, and contribute to anxiety-like behaviors [82]. The cytokines produced through microglial and astrocytes interfere with the dopaminergic signalling, which regulates the production, secretion by synapses, and the uptake of DA by transporters and receptors [83]. These changes interfere with the neuronal systems for reward processing, motivation, and executive control. This disturbance not only augments the reinforcing effects of METH, but also promotes compulsive drug-seeking and drug-taking behaviors [84]. This vicious cycle is also aggravated by stress, which sustains the HPA axis hyperactivity and persistent glucocorticoid release. Prolonged exposure to glucocorticoids increases the production of cytokines and impairs dopamine and glutamate, increasing neuroinflammation and neurotransmitter maladaptation. This bidirectional relationship, in which neuroimmune signalling worsens the neurotransmitter functions, and the neurotransmitter dysregulation increases the effects of inflammatory pathways, ultimately resulted in impairment of neuroplasticity in the PFC, amygdala, hippocampus, and nucleus accumbens [85]. These structural and functional defects are associated with dendritic spine loss and neuronal rearrangement at the synapses and neural network rearrangement. This results in the disruption of inhibitory control, decision-making, as well as susceptibility to relapse and diminished stress resilience, which are manifest as persistent anxiety and emotional instability [86]. It is important to note that this neuroimmune-neurotransmitter feedback loop is the key to understanding the biological origin of comorbidity between METH addiction and anxiety. Therapeutic strategies focus on reducing microglial overactivation, suppression of pro-inflammatory cytokine release, regulation of glutamate receptors to avoid excitotoxicity, and selective blockage of P2X7R are the possible approaches [87]. By modulating these maladaptive pathways, there is the possibility of restoring synaptic integrity, re-equilibrating neural networks, removing anxiety symptoms, decreasing drug reinforcement, and ultimately, improving the outcome of those with both an anxiety and METH addiction [88].

Therapeutic targeting of P2X7R in anxiety and METH addiction

Rationale for targeting P2X7R

P2X7R as a therapeutic target is an effective way of treating anxiety and METH addiction because it is a key mediator of neuroinflammation and neurotransmitter dysregulation, two core processes that are similar in both disorders. P2X7R activation on microglia facilitates the release of IL-1β, IL-18, and TNF-α that disrupts neuronal circuits controlling mood and reward through DA, 5-HT, and glutamate signalling dysfunction [89]. These neurochemical adaptations increase anxiety symptoms and drug-seeking behaviour with an increase in stress response and reward processing. The interplay between neuroinflammation and neurotransmitter imbalances establishes a self-perpetuating cycle driven by P2X7R activity, as its activation by elevated extracellular ATP triggers neuroinflammation and disrupts neurotransmitter balance, leading to ongoing excitotoxicity and synaptic dysfunction [90]. Pharmacological inhibition of this receptor may disrupt this cycle, attenuating both anxiety-related symptoms and addictive behaviours simultaneously. Furthermore, genetic studies indicate that individuals with P2RX7 gene variations exhibit heightened susceptibility to anxiety and substance abuse, supporting the rationale for personalised therapeutic strategies [91]. Although a clinical study is still underway, modulating P2X7R provides an opportunity to target converging neurobiological pathways in comorbid anxiety and addiction, potentially offering more effective and comprehensive treatment than conventional approaches addressing these disorders in isolation [92].

Potential therapeutic strategies in preclinical and clinical trials

The preclinical evidence involves the investigations of the development of the P2X7R antagonism, which can be successful in reducing anxiety-like behavior and simultaneously preventing drug-seeking behavior [93]. The pharmacological inhibition by A438079 has been shown to inhibit hyperlocomotion caused by methamphetamine and conditioned place preference in rodents, which are common behavioral effects of stimulant abuse [94]. Besides behavioral control, the P2X7R inhibition also restores neuroimmune homeostasis by normalizing excessive levels of pro-inflammatory cytokines such as IL-17A within the brain, which is responsible for reward processing. It demonstrates that immune modulation is one of the key mechanisms of the formation of the therapeutic effect of P2X7R antagonists [95]. Several possible therapeutic agents have been pursued, and they include JNJ-54471300 and A-804598, which inhibit neuroinflammation in psychiatric diseases that are associated with over-activation of the P2X7R. Such agents inhibit microglial hyperactivation, reduce pro-inflammatory cytokine release, and inhibit synaptic and behavioural defects in chronic neuroinflammation. Despite the positive outcomes obtained, it has some limitations in the sense of extrapolating the preclinical findings into clinical therapies, as well as ensuring that the treatment has sufficient delivery to the CNS, more specificity of its targets to minimize off-target effects, and in the sense of identifying patients who may respond. The P2RX7 gene variants are able to stratify the human population and can be used to develop certain treatments that may be applied depending on neuroimmune susceptibility. This convergence between preclinical and first clinical research has raised a great deal of optimism that P2X7R antagonists will be a new therapeutic category capable of targeting the immune-based pathways to comorbid anxiety and substance use disorders (SUDs). JNJ-54175446 is the most promising of them. This selective P2X7R antagonist has already been through phase one and two trials, although the majority of studies have focused on patients suffering from major depressive disorder (MDD). But there are high chances that it will be successful in anxiety and addiction, as the mechanism of inflammation in the two disorders is very similar. JNJ-54175446 is a safe and well-tolerated clinical agent and has been demonstrated to have the ability to reduce peripheral cytokine production, namely IL-1β-mediated neuroinflammation, and may play a role in modulating anxiety and addiction-related behaviors [96]. This further reduced mood disorders caused by psychostimulants in experimental human models, which shows that the compound could control reward systems based on DA and reduce vulnerability to addictive behavior. An overview of the preclinical and clinical outcomes of P2X7R antagonists, including JNJ-54471300, A438079, and A-804598, and their influence on anxiety and SUDs paradigm is provided in Table 1.

Table 1.

Therapeutic agents targeting P2X7R and their preclinical and clinical status

Sr. No	Experimental Model	Animal	Drug	Dose	Route	Frequency	Study Outcomes	Reference
1	Unpredictable Chronic Mild Stress (UCMS)	Mice (BALB/c) of 8 weeks Stressor- 2 per day for 9 weeks	Brilliant Blue G (BBG)	50 mg/kg	i.p	Daily for 2 weeks after UCMS start	These results with 50 mg but anhedonic behavior and microglial activation is reduced by 25 mg and 50 mg both • ↓se anhedonic behavior • ↓se in mRNA level of P2X7R • ↓se in caspase-1 • ↓se in ASC, NFкB level • ↓se in microglial activation • ↓se in IL-1β, IL-6	[86]
2	UCMS	Male rats (Wistar-Albino) of 8–10 weeks	BBG	25 & 50 mg/kg	i.p	Every 24 h during the last 3 weeks of UCMS	• ↓se anhedonic behavior • ↓se in mRNA level of P2X7R • ↓se in caspase-1 • ↓se in microglial activation • ↓se in IL-1β, IL-6	[91]
3	Chronic Unpredictable Stress (CUS)	Male rats (Sprague–Dawley)	BBG & A-438079	BBG 1 µl (1 pM), A-438079 1 µl (1.75 nM)	Bilateral microinjection into hippocampus via an implanted canula	Once daily during CUS; injections were performed 30 min after each stress exposure	• ↓se hippocampal extracellular ATP • ↓se cleaved-caspase-1 • ↓se ASC and NLRP3 levels • ↓se IL-1β	[92]
4	CUS	Male rats (Sprague–Dawley)	A-804598	5 mg/kg	i.p	Twice daily in the last 4 weeks of CUS	• Reversed depressive/anxiety phenotype • ↓se neuroinflammation	[93]
5	Repeated Social Defeat Stress (RSD)	Male mice (C57BL/6) of 6–8 weeks and CD-1 retired breeder mice of 12 months	JNJ-54471300 (P2X7R antagonists)	10 mg/kg	i.p	30 min prior to each RSD	• ↓se microglial reactivity • ↓se IL-1β expression • ↓se gene expression of cytokine signalling • ↓se in neuroinflammatory gene expression (NFкB) • ↓se social avoidance	[94]
6	Locomotor activity, intracranial self-stimulation (ICSS) experiments, conditioned place preference (CPP)	Adult male C57Bl/6 mice	A438079	METH dose- 1 mg/kg A438079 dose- 5, 10, 50 mg/kg	i.p	Once daily for 7 days 30 min before METH for behavioral tests 15 min before METH for cytokine release	• Inhibited METH-induced hyperlocomotion and reward • Normalized the elevations of IL-17A levels in PFC	[95]

UCMS, unpredictable chronic mild stress; BBG, Brilliant Blue G; i.p., intra-peritoneal; mRNA, messenger ribonucleic acid; P2X7R, purinergic P2X7 receptor; ASC, apoptosis-associated speck-like protein containing a CARD; NFкB, nuclear factor kappa B; IL-1β, interleukin-1β; IL-6, interleukin-6; CUS, chronic unpredictable stress; ATP, adenosine triphosphate; NLRP3, NOD-like receptor family pyrin domain containing 3; RSD, repeated social defeat stress; PFC, pre-frontal cortex; METH, methamphetamine; ICSS, intracranial self-stimulation

Conclusions and future directions

P2X7R has emerged as a significant target in anxiety and METH addiction due to its role in neuroinflammation and neurotransmitter homeostasis. P2X7R activation in microglia induces the secretion of pro-inflammatory mediators, which disrupt dopaminergic, glutamatergic, and serotonergic neurotransmission, affecting mood stability, stress reactivity, and reward circuitry. This inflammatory cascade establishes self-perpetuating feedback loops that exacerbate both anxiety symptoms and addictive behaviors. As such, P2X7R emerges as a biomarker of stress-induced neuroinflammation as well as a potential pharmacological target in the treatment of comorbid anxiety and METH-induced neurotoxicity. The recent developments of precision medicine allow genetic screening of P2RX7 variants linked to increased receptor activity and neuropsychiatric risk, and offer more prominent treatment approaches. Selective antagonists, JNJ-54175446, are currently in preclinical trials and have shown positive safety profiles as well as early activity in the development of mood disorders. Future research that integrates genetic profiling and neuroimaging of P2X7R expression will enhance the therapeutic accuracy and reduce relapse risk.

Limitations and challenges

Although the evidence implicating P2X7R in the pathophysiology of anxiety is encouraging, there are several limitations and challenges to the existing knowledge and the possible clinical application of the results. To a great extent, the studies that have been conducted so far are based on animal models, which, although educational in terms of understanding the mechanisms of molecular processes, cannot reproduce the complexity and heterogeneity of human anxiety disorders. Even the neuroimmune system is very complex, with the P2X7R interacting with a large number of other purinergic and cytokine receptors, frequently leading to overlapping or compensatory signalling that is difficult to distinguish. Also, the genetic variability of the P2X7R gene has the potential to modify the functioning of the receptor and the personal susceptibility to anxiety, but the effect of these forms of variability is inconsistent across diverse populations, which requires larger and more diverse genetic studies. There are also pharmacological issues, such as ensuring that P2X7R antagonists are able to enter the BBB, the long-term safety, and preventing any unintended side effects, since the receptor is involved in the functioning of the brain in a multi-factorial way. The development of non-invasive biomarkers for measuring central P2X7R activity in humans is still in its early stages and does not allow for the effect of treatment on patients. Lastly, the interactions between P2X7R activity and environmental factors, including early-life stress, are insufficiently known, and longitudinal studies are needed to make it clear how the two combined contribute to the risk of anxiety. Additionally, METH exposure exacerbates neuroimmune dysregulation via P2X7R activation, contributing to neuroinflammation and anxiety vulnerability. These challenges highlighted the need for multidisciplinary approaches, better translational models, and well-designed clinical studies to optimize the P2X7R-targeted approach for anxiety and related disorders.

Abbreviations

P2X7R

Purinergic P2X7 receptor

CNS

Central nervous system

ATP

Adenosine triphosphate

NKAα1

Na⁺/K⁺-ATPase

METH

Methamphetamine

IL-17A

Interleukin-17A

PFC

Pre-frontal cortex

BBB

Blood-brain barrier

5-HT

Serotonin

DA

Dopamine

GABA

Gamma-aminobutyric acid

NE

Norepinephrine

IL-1β

Interleukin-1β

TNF-α

Tumor necrosis factor- α

mRNA

Messenger ribonucleic acid

TM1

Transmembrane 1

TM2

Transmembrane 2

Ca²⁺

Calcium

Na⁺

Sodium

K⁺

Potassium

DAMP

Damage-associated molecular pattern

NFкB

Nuclear factor kappa B

pro- IL-1β

Pro-interleukin-1β

pro- IL-18

Pro-interleukin-18

NLRP3

NOD-like receptor family pyrin domain containing 3

NAD

Nicotinamide adenine dinucleotide

IL-6

Interleukin-6

ASC

Adaptor protein apoptosis-associated speck-like protein containing a CARD

CRH

Corticotropin releasing hormone

ACTH

Anterior pituitary to release adrenocorticotropic hormone

HPA-axis

Hypothalamic-pituitary-adrenal axis

ROS

Reactive oxygen species

IL-1β

Interleukin-1β

IKK

IкB kinase

iNOS

Inducible nitric oxide synthase

NMDA

N-methyl-D-aspartate

AMPA

α-Amino-3-hydroxy-5-methyl-4-isoxazole propionic acid

mGluR1

Metabotropic glutamate receptor 1

mGluR8

Metabotropic glutamate receptor 8

CaMKII

Ca²⁺/calmodulin-dependent protein kinase II

PKC

Protein kinase C

MAPKs

Mitogen-activated protein kinase

Mg²⁺

Magnesium

cyt C

Cytochrome C

GAD

Glutamic acid decarboxylase

GLAST

Glutamate/aspartate sodium-dependent transporter

EAAT1

Excitatory amino acid transporter

TTX

Tetrodotoxin

CNQX

6-Cyano-7-nitroquinoxaline-2,3-dione

BBG

Brilliant blue G

PPADS

Pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid

mPFC

Medial prefrontal cortex

IRS-1

Insulin receptor substrate-1

TrkB

Tropomyosin receptor kinase B

BDNF

Brain-derived neurotrophic factor

BNST

Bed nucleus of the stria terminalis

mTORC1

Mammalian target of rapamycin complex 1

PI3K

Phosphoinositide 3 kinase

AMPK

AMP-activated protein kinase

TSC2

Tuberous sclerosis complex 2

Rheb

Ras homolog enriched in brain

Raptor

Regulatory-associated protein of mTOR

PVN

Paraventricular nucleus

AVP

Arginine vasopressin

BAX

BCL2- associated X protein

BID

BH3-interacting domain death agonist

Bcl-2

B-cell lymphoma 2

ER

Endoplasmic reticulum

UPR

Unfolded protein response

GluR5

Glutamate receptor 5

GAD65

Glutamic acid decarboxylase 665

ATP1a3

ATPase Na⁺/K⁺ transporting subunit α-3

VTA

Ventral tegmental area

SUDs

Substance use disorders

MDD

Major depressive disorder

Simranpreet Kaur

is a Pharm D research scholar with an interest in clinical practice and academic research. Involved in scientific writing, literature review, and poster presentations. Possesses working knowledge of pharmacovigilance, clinical data interpretation, and evidence-based medicine. Committed to learning and contributing to healthcare research while gaining hands-on hospital experience through an internship.

Author contributions

SK, VKG: Writing – review & editing the manuscript. KRA: Conceptualization, review, editing, visualization.

Funding

There is no funding agency in this article.

Data availability

Not applicable.

Compliance with ethical standards

Conflicts of interest

The authors declare no competing interests.

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