Mu-opioid receptor is an important member of G-protein-coupled receptor family, which is widely distributed in the nervous system. It is the core target of regulating pain signal transmission, and also the key molecule involved in reward loop, emotional regulation and neural plasticity. Traditional opioid drugs play a powerful analgesic role by activating mu-opioid receptor, but long-term use is easy to cause substance abuse, resulting in tolerance, dependence and addiction, and its mechanism is closely related to the imbalance of signal pathway mediated by mu-opioid receptor, neuroinflammatory response and synaptic function remodeling. In recent years, with the in-depth study on the conformational dynamics of mu-opioid receptor, ligand functional selectivity and downstream signal network, biased agonists or allosteric modulators of mu-opioid receptor may reduce the risk of addiction while retaining analgesic activity. In this paper, the distribution and function of mu-opioid receptor in nervous system are systematically reviewed, the molecular mechanism of addiction development is expounded, and the current drug research and development strategies and research progress based on mu-opioid receptor are summarized, so as to provide new ideas for clinical optimization of opioid drug use scheme and development of low addictive analgesic drugs.

• Mu-Opioid Receptor; Pain Management; Addiction Mechanisms; Neuroinflammation; Low-Addiction Analgesics

Long Y, Bai J (2026) Molecular Mechanism of Mu-Opioid Receptor and Development Strategies of Analgesic Drugs. J Subst Abuse Alcohol 13(2): 1109.

The International Association for the Study of Pain (IASP) defines pain as an unpleasant sensory and emotional experience associated with, or resembling that of, actual or potential tissue damage [1,2]. Chronic pain is categorized into primary pain (independent of other diseases and unrelated to other pain sources) and secondary pain (resulting from underlying conditions, including cancer pain, chronic postoperative pain, neuropathic pain, musculoskeletal pain, and visceral pain). It can further be classified as nociceptive pain (originating from tissue damage), neuropathic pain (originating from nerve damage), and neuropathic pain (originating from a hypersensitive nervous system) [3]. Chronic pain imposes a significant economic and quality-of-life burden on over 30% of the global population, leading to substantial healthcare utilization and associated costs [4,5].

Cancer is a major public health issue. According to a 2020 report by the Chinese Center for Disease Control and Prevention, China recorded 2.3978 million cancer-related deaths [6]. Pain is one of the symptoms experienced by cancer patients and also one of the most common symptoms, prevalent in 30%-50% of patients receiving targeted cancer therapy and over 70% of advanced cancer patients [7,8]. It may trigger inflammation and stimulate the sympathetic nervous system, potentially altering the tumor microenvironment, stimulating dormant tumor growth, and promoting micrometastasis and metastatic disease progression [9]. Most cancer patients require analgesia during and after surgery, making pain management a critical component of cancer treatment [8]. Furthermore, pain symptoms are subjectively described and lack specific clinical manifestations or prognostic biomarkers [10].

Opioids are the most effective treatment for pain patients and are widely used worldwide [11]. Approximately 44.5% of cancer patients experience pain, with 30.6% reporting moderate to severe pain [12]. According to a recent report by the Centers for Disease Control and Prevention (CDC), deaths involving opioid overdose increased by 45.2% from 2016 to 2017[13]. An integral part of pain management involves the use of analgesics. These compounds interfere with the generation or transmission of impulses following harmful stimuli (pain sensations) within the nervous system. This interference can occur at the peripheral or central levels of the neural axis. The therapeutic goal is to reduce the perception of pain. Analgesics aim to modulate the formation of harmful chemicals (e.g., prostaglandins) or the activation of neuronal receptors and ion channels (e.g., peptides, kinins, monoamine receptors, Na? channels) involved in transducing or transmitting harmful stimuli.

Although opioids play a significant role in pain management, they also carry considerable adverse consequences. Opioid use may lead to side effects including hyperalgesia, analgesic tolerance, withdrawal syndrome, neuroinflammation, and respiratory depression [14]. Current medications for chronic pain include opioids, Nonsteroidal Anti-Inflammatory Drugs (NSAIDs), serotonergic compounds, antiepileptic drugs, and antidepressants. NSAIDs, acetaminophen, Selective Serotonin Reuptake Inhibitors (SSRIs), Tricyclic Antidepressants (TCAs), and Gamma-Aminobutyric Acid (GABA) analogues are commonly used for pain treatment. However, NSAIDs carry risks of cardiovascular side effects, gastrointestinal bleeding, and renal disease; acetaminophen has hepatotoxicity; and SSRIs, TCAs, and GABA analogues offer limited analgesic efficacy. Consequently, opioids remain an effective pharmacological treatment and are among the most widely prescribed medications for acute and chronic pain conditions [15].

The Mu-Opioid Receptor (MOR) is the primary receptor for opioids, widely distributed throughout the Central Nervous System (CNS). It is highly expressed in reward circuits such as the Ventral Tegmental Area (VTA)-Nucleus Accumbens (NAc) pathway, pain regulation circuits like the Periaqueductal Gray (PAG)-dorsal horn of the spinal cord (DH) pathway, and emotional regulation regions including the Amygdala (AMY), and Prefrontal Cortex (PFC). Its functional abnormalities are closely associated with neurological disorders such as addiction, chronic pain, anxiety, and depression [16]. Recent studies have revealed that MOR mediates analgesia through G protein dependent signaling pathways, while the β-arrestin pathway is closely associated with adverse effects such as tolerance, addiction, and respiratory depression [17-19]. Furthermore, heterodimers formed between MOR and other receptors (e.g., δ-opioid receptor, neuropeptide FF receptor) and neuroinflammatory responses mediated by glial cells (microglia, astrocytes) play crucial regulatory roles in opioid addiction development [20-22]. This review will examine MOR’s distribution and function within the nervous system, its involvement in addiction-related mechanisms, and recent advances in drug development.

Distribution of MOR

Opioid receptors belong to the class of G protein-coupled receptors. The hypothesis of opioid receptor existence emerged in the 1950s [23], but it was not until 1973 that three independent research teams first demonstrated the presence of membrane receptors for opioid drugs in the brain, confirming the existence of opioid receptors [24,25]. Binding experiments using different radioligands revealed that opioid receptors exhibit stereoselectivity: [³H] naloxone [24], [³H] dihydromorphin, and [³H]etorphine [25]. They are widely expressed in the central nervous system and peripheral tissues [26]. MOR, encoded by the Oprm1 gene, is a major subtype of opioid receptors, similarly expressed in both the central nervous system and peripheral organs. As a key brain region in the endogenous analgesic pathway, the PAG integrates pain information from the forebrain and transmits neuromodulatory signals to the DH via ascending nociceptive and descending antinociceptive pathways. MOR activation inhibits nociceptive signal transmission in the dorsal spinal horn through a descending inhibitory system [27]. In the superficial DH, MOR is primarily distributed in the terminals of primary afferent neurons and interneurons, directly regulating the transmission of peripheral nociceptive signals to the central nervous system [28]. In reward and addiction-related brain regions, MORs are present on the membranes of dopaminergic neurons in the VTA. Their activation inhibits GABAergic neurons’ suppression of dopaminergic neurons, leading to increased dopamine release in the NAc. This represents the core mechanism by which opioids produce euphoria and reward effects [29,30]. MOR expression in the PFC and AMY also participates in opioid-related emotional regulation and compulsive drug-seeking behavior control [31]. Within the peripheral nervous system, MOR is expressed in DRG sensory neurons, sympathetic ganglia, and gastrointestinal plexuses. Activation of peripheral MOR exerts local analgesia by inhibiting the release of nociceptive neuropeptides (e.g., substance P, calcitonin gene-related peptide), while avoiding central adverse effects [26]. Notably, MOR expression levels in peripheral tissues are significantly upregulated during inflammation or tissue injury, and acidic microenvironments enhance MOR ligand binding affinity. This provides a theoretical basis for developing peripherally restricted MOR agonists (e.g., loperamide) [32,33].

MOR-Mediated Physiological Functions of the Nervous System

Regulation of Pain Signals: Heterotrimeric G proteins constitute the largest family of G Protein-Coupled Receptor (GPCR) signaling proteins, composed of three subunits: Gα, Gβ, and Gγ [34]. In its inactive state, the heterotrimer binds GDP. Catalyzed by guanine nucleotide exchange factors (GEFs), it stimulates GDP dissociation and promotes GTP binding. The GTP-bound Gα subunit dissociates from Gβγ, activating downstream second messengers. MOR mediated signaling activates five Gαi/o subtypes within the same Gα subfamily, each playing distinct roles in analgesia [35-37]. MOR transduces diverse chemical and physical extracellular signals—including neurotransmitters, chemokines, and hormones—by activating Gαi/o proteins, generating multiple biological effects such as cell proliferation, differentiation, and development. It can independently interact with effector enzymes or ion channels to trigger appropriate cellular responses. For example, cyclic Adenosine Monophosphate (cAMP), inositol trisphosphate (IP3), and Diacylglycerol (DAG). The second messenger cAMP activates the downstream core gene protein kinase A (PKA). PKA can translocate to the cell nucleus, cAMP response element-binding protein (CREB) and other important nuclear proteins to regulate gene expression. Most PKA activates downstream pathways by directly interacting with various membrane ion channels, such as Na? channels. This interaction modulates presynaptic and postsynaptic Ca²? currents, thereby dampening neuronal excitability and reducing presynaptic sensing and inflammatory neuropeptide release [38-40]. Additionally, MOR activation triggers the opening of G protein-coupled inwardly rectifying K? (GIRK) channels, which blocks neuronal excitation and action potential propagation. G protein-dependent signaling results in decreased cAMP levels, reduced Ca²? responses, and activation of GIRK channels [41-43] 

Rewards and Emotional Regulation: MOR is pivotal for reward signaling in the VTA-NAc pathway. As the primary source of dopaminergic neurons, the VTA’s MOR activation on its membrane surface inhibits GABAergic neurons, thereby releasing their inhibition on dopaminergic neurons. This promotes dopamine release and its projection to the NAc [29-44]. Animal studies demonstrate that blocking MOR (via knockout or antagonist use) reduces dopamine release in the VTA, leading to insufficient dopamine in the NAc. This diminishes both the motivation for natural rewards and opioid-induced addictive behaviors (such as conditioned place preference) in mice, confirming its role in establishing and maintaining addictive behaviors [45,46]. As a key brain region in the reward pathway, the NAc’s MOR expression is closely linked to the dopaminergic system. Morphine exerts potent analgesic effects by activating NAc MORs. However, prolonged use can overactivate dopaminergic reward signaling, leading to morphine dependence and subsequent addiction [47-49]. Furthermore, injecting the MOR antagonist naloxone into the NAc not only limits morphine’s analgesic effects but may also suppress morphine-induced reward responses. This finding suggests that modulating MOR activity could potentially intervene in addiction pathways, offering a promising research direction and practical approach for addiction treatment [50]. MOR is widely expressed in the AMY, particularly in the basolateral AMY and the dendritic spines, axon terminals, and Golgi apparatus of interneurons [51]. It plays a central role in pain and emotional behavior regulation. MOR inhibits excitatory signals from the thalamus and suppresses excitatory transmission from the basolateral to the central AMY. It also modulates GABAergic inhibitory synaptic activity along the basolateral-to-central AMY projection pathway. MOR mediates anti-nociception and anxiety regulation. In neuropathic pain models, AMY MOR signaling undergoes significant alterations, including reduced binding capacity and downregulated expression. Concurrently, MOR activation induces neuroplastic changes, such as upregulation of FosB/ΔFosB transcription factors and associated gene expression alterations [52-54]. Adolescent morphine exposure impairs adult spatial memory and hippocampal synaptic plasticity. Chronic stress modulates learning processes in the female hippocampal opioid system but has limited effects in males, impairing its capacity to support opioid-mediated learning. MOR also regulates morphine-dependent memory in rats by interacting with CREB [55-59].

Neuroendocrine Regulation: As a member of the GPCR family, MOR activation inhibits adenylate cyclase activity and reduces cyclic AMP production, thereby affecting downstream signaling pathways involved in endocrine hormone synthesis and release. For example, during stress responses, Corticotropin-Releasing Factor (CRF) serves as a key regulator of the pituitary adrenal axis. The endogenous opioid system, including MOR, modulates CRF release from the hypothalamic paraventricular nucleus, thereby influencing the secretion of endocrine hormones such as glucocorticoids [60-62]. Concurrently, glucocorticoids can directly upregulate the transcription of enkephalin precursor mRNA by binding to glucocorticoid response elements on DNA, forming a bidirectional regulatory loop between MOR-related signaling and the endocrine system [63,64].

Adaptive Changes in MOR Signaling Pathways

MOR belongs to the GPCR family, and its signaling pathways involved in addiction mechanisms include G protein-coupled signaling pathways and β-arrestin-mediated signaling pathways. Long-term opioid use induces adaptive changes in MOR signaling pathways, which represents one of the core molecular mechanisms by which MOR contributes to addiction.

G Protein-Coupled Receptor Kinase (GRK) phosphorylation of opioid receptors leads to β-arrestin recruitment [65, 66]. GRKs are a class of serine/threonine kinases that phosphorylate agonist-bound or activated GPCRs as their primary substrates. Based on sequence homology, GRKs are subdivided into three subfamilies: the visual subfamily (GRK1 and GRK7) [67], the GRK4 subfamily (GRK4, GRK5, and GRK6) [66], and the β-AR kinase (β-ARK) subfamily (GRK2 and GRK3) [68]. GRK1 and GRK7 are primarily expressed in retinal photoreceptor cells, while GRK4 shows the highest expression in testes. GRK2, GRK3, and GRK5 are widely expressed, though their expression levels vary across tissues. In cardiac tissue, GRK2 and GRK5 are most prominent, while GRK3 and GRK6 exhibit lower expression levels. Functionally, all GRKs mediate the decoupling and internalization of activated GPCRs [69]. All GRKs are primarily localized to the cytoplasm and plasma membrane. GRK1, GRK4, GRK5, GRK6, and GRK7 reside at the membrane base. Conversely, GRK2 and GRK3 are mainly cytoplasmic and translocate to the membrane following receptor stimulation. Different GRK subtypes exert distinct effects on tolerance, reward, addiction, and motor activity [70,71]. MOR phosphorylation requires GRK2/3 subtypes, followed by binding to β-arrestin in lectin-coated pits—a recognized downstream step in agonist-selective endocytosis mechanisms. The C-terminal PH domain of MOR recruits GRK2 from the cytoplasm to the plasma membrane, a step dependent on agonists but insufficient to explain the apparent agonist selectivity of GRK2-MOR interactions [72,73]. The findings indicate that the N-Terminal Domain (NTD) of GRK2 is the key determinant of the highly agonist-selective interaction between GRK2 and MOR. The NTD binds to the activated GPCR core, stabilizing the closed conformation of the kinase domain and thereby enhancing catalytic activity [74,75].

Arrestins constitute a family of four multifunctional adapter proteins exhibiting high sequence and structural homology. They comprise an N-terminal domain, a central polar core, a C-terminal domain, and an intrinsically disordered tail. Among these, β-arrestin1 and β-arrestin2 are ubiquitous and participate in receptor desensitization and downregulation [76,77]. β-arrestins perform three key functions: Receptor desensitization, internalization, and signal transduction. Upon receptor activation, GRKs phosphorylate the intracellular domain of GPCRs, typically at the C-terminus, thereby promoting the recruitment and tight binding of β-arrestin. This binding induces a conformational change in β-arrestin. Consequently, when opioid receptors undergo internalization and degradation, β-arrestin-dependent signaling terminates G protein signaling [78]. Long-term opioid use alters GRK and β-arrestin expression, enhancing MOR desensitization and internalization. This reduces MOR sensitivity to opioids, requiring increased drug doses to achieve original analgesic effects—a phenomenon termed tolerance. Tolerance development is closely linked to the addiction process and represents a critical step in addiction progression. Furthermore, β-arrestin interacts with multifunctional proteins to activate downstream signaling pathways such as mitogen-activated protein kinase (MAPK) and p38 [79]. The interaction between Gαi subunits and β-arrestin can activate extracellular signal-regulated kinase (ERK), illustrating the intricate signaling dynamics of opioid receptors and their therapeutic potential through biased and partial agonism [80-82]. Other pathways involved include JNK signaling and the phosphoinositide 3-kinase/ protein kinase B (PI3K-AKT) pathway.

Fentanyl mainly acts by activating MOR, and has low affinity for δ Opioid Receptor (DOR) and κ Opioid Receptor (KOR), so it is a high-performance MOR agonist. Its binding affinity to recombinant human MOR is similar to that of morphine, but its analgesic effect is much higher than that of morphine, which is related to its high lipophilicity [83,84]. After the activation of MOR, fentanyl not only mediates analgesia and euphoria through inhibitory G protein, but also produces non-G-protein-dependent signals through β-arrestin complex, and the activation of β-arrestin signal is stronger, which may be an important reason for its serious side effects such as respiratory depression. The tolerance mechanism of fentanyl is also related to MOR, which mediates short-term tolerance through GRK3, while morphine depends on c-Jun amino terminal kinase. β-arrestin2 knockout mice have no effect on fentanyl analgesia tolerance, but can reduce morphine tolerance [85,86]. Fentanyl induces respiratory inhibition and other cardiopulmonary inhibition effects by acting on MOR, and both peripheral and central MOR are involved. Peripheral MOR plays a key role in fentanyl induced respiratory depression. The use of peripheral restrictive MOR antagonist naloxone methiodide (NLXM) can effectively prevent and reverse the decrease of oxygen saturation, heart rate and respiratory rate caused by fentanyl, and the effect is equivalent to that of naloxone (NLX) which can penetrate the blood-brain barrier, and NLXM will not cause aversion caused by NLX [87].

Fentanyl substances (carfentanil, sufentanil, alfentanil, etc.) are all MOR agonists, but there are differences in signal transduction. Caffeine is a β-arrestin-biased agonist of MOR, which can induce β-arrestin2 recruitment and receptor cell surface loss more strongly. The degree of GIRK current desensitization in the nucleus locus coeruleus neurons of rats is significantly higher than that of fentanyl, while fentanyl, sufentanil and alfentanil are unbiased [88,89].

Regulatory Role of Neuroinflammation

Morphine-induced neuroinflammation is typically accompanied by microglial activation and the production of inflammatory mediators such as cytokines and chemokines [21]. Microglia are CNS-resident macrophages that express MOR in the brain and spinal cord [90], playing roles in neuronal survival and death, synaptogenesis, and infection prevention. Microglia are implicated in chronic pain and play a crucial role in emotional disorders associated with the mesocorticolimbic system (MCLS) [91]. Activated microglia and astrocytes exert significant functions in opioid analgesia by promoting neuroinflammation through the release of inflammatory mediators. Microglial polarization exhibits two distinct phenotypes: the M1 phenotype releases various proinflammatory cytokines such as tumor necrosis factor α (TNF-α), Interleukin (IL-1β), IL-6, IL-12, IL-17, and IL-23 while generating substantial nitric oxide (NO); conversely, the M2 phenotype attenuates inflammation and promotes brain repair and regeneration [92-95]. IL-1β serves as a key mediator in the interaction between neuropathic pain and neuroinflammation. Following CNS injury, activated glial cells release the inflammatory mediator IL-1β within the CNS, thereby driving the onset and exacerbation of neuropathic pain. These proinflammatory cytokines inhibit morphine analgesia and induce opioid tolerance.

Research has found that morphine activates TLR4 signaling, leading to upregulation of TLR4 expression. This subsequently activates downstream cGAS-STING pathways and NF-κB, triggering the release of pro-inflammatory cytokines such as IL-6 and TNF-α. Concurrently, it causes mitochondrial dysfunction, resulting in increased mtROS and mtDNA leakage. This further exacerbates inflammation, causing cells to polarize toward a pro-inflammatory M1 phenotype. These inflammatory responses counteract morphine’s analgesic effects and ultimately induce tolerance [96]. TAK-242, as a TLR4 antagonist, significantly reduces the expression of TLR4, cGAS, STING, and phosphorylated NF-κB, decreases the release of IL-6 and TNF-α, improves mitochondrial function, reduces mtROS production and mtDNA leakage, and inhibits cell polarization toward the M1 phenotype. Given its ability to mitigate morphine-induced immune cell inflammation, TAK-242 is considered a potential intervention to improve morphine tolerance [97,98]. Furthermore, depletion of microglia using the colony-stimulating factor 1 receptor (CSF1R) inhibitor PLX3397 suppressed proinflammatory mediator release and reduced activation of proinflammatory A1-pheno-type astrocytes, thereby alleviating morphine tolerance. This indicates that microglia activation is a critical step in the development of morphine tolerance, with its abnormal interaction with astrocytes and the neuroinflammation it mediates playing a significant role in the mechanism of morphine tolerance [99].

Synaptic Plasticity Remodeling

Under long-term morphine treatment, dopaminergic neurons in the VTA exhibit increased pre-synaptic membrane branching and synaptic vesicle density, along with enhanced synaptic connections to NAc neurons. In the NAc, medium spiny neurons exhibit heightened dendritic spine density and a shift toward more mature “mushroom-shaped” morphology. Postsynaptic dense areas expand with increased expression of scaffolding proteins, providing structural support for alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and n-methyl aspartic acid (NMDA) receptor clustering [100,101]. Concurrently, elevated expression of the NMDA receptor GluN2B subunit enhances calcium influx, activates kinases to phosphorylate AMPA receptors, prolongs their synaptic membrane residence time, increases channel opening probability, and stabilizes long-term potentiation. This enhances reward circuit signaling efficiency, sensitizes the organism to reward effects, and further reinforces drug seeking behavior [102].

Beyond the reward circuitry, opioids also contribute to addiction-related anxiety and drug craving by modulating synaptic plasticity between the AMY and PFC. Under prolonged opioid exposure, plasticity of GABAergic inhibitory synapses in the Basolateral Amygdala (BLA) decreases, with reduced GABA receptor expression and release, leading to heightened neuronal excitability. This excitability propagates to the PFC, enhancing postsynaptic NMDA receptor function in pyramidal cells to induce long-term potentiation. This weakens regulation of drug craving, exacerbates withdrawal anxiety, and collectively sustains addiction and relapse [103,104]. The use of AMPAR antagonists, such as CNQX, is closely associated with synaptic plasticity. Following CNQX injection, which blocks AMPA receptor function, morphine-induced increases in dendritic spine density were significantly inhibited, and drug-seeking behavior was reduced. This confirms the central role of synaptic plasticity remodeling in opioid addiction [105].

The signaling pathways of morphine and MOR involved in addiction are organized in Figure 1.

Figure 1 This figure depicts the signaling pathways mediated by different receptors in microglia and their associated physiological effects. The left pathway shows morphine activating downstream inflammatory signals via TLR4, including the release of TNF-α and IL-1β, and promoting inflammatory responses through pathways such as MAPK/p38 and NF-κB. It also involves signal desensitization and tolerance formation mediated by molecules including β-arrestin and GRK. The right pathway demonstrates morphine activating G protein-coupled receptors via MOR, inhibiting AC to reduce cAMP levels, thereby affecting PKA and transcription factor CREB activity. Together, these pathways regulate microglial function, participating in morphine analgesia, tolerance, and neuroimmunomodulation processes.TLR4, Toll-Like Receptor 4; TNF-α, Tumor Necrosis Factor α; interleukin -1beta (IL-1β); MAPK/p38, p38 mitogen-activated protein kinase; NF-κB, Nuclear Factor kappa-B; GRK, G protein coupled receptor kinase; MOR, Mu Opioid Receptor; AC, Adenylyl Cyclase; cAMP, cyclic Adenosine Monophosphate; PKA, Protein Kinase A; CREB, cyclic-AMP response binding protein.

Development of Multifunctional Opioid Drug Ligands

Developing multifunctional opioid ligands represents a promising strategy [106,107]. Compared to single-target drugs, multifunctional drugs offer greater advantages in addressing multifactorial diseases such as pain, diabetes, cardiovascular disease, neurodegenerative disorders, and cancer [108,109]. Pain-related receptors co-express within neuronal populations or physically proximate regions, forming heterodimers or oligomers [110]. However, rationally designing drugs and understanding their interactions with relevant targets remain significant challenges.

MCRT is a multifunctional agonist that activates receptors such as MOR, μ-δ heterodimer (MDOR), and neuropeptide FF receptor 2 (NPFFR2), exhibiting distinct nociceptive effects across acute and chronic inflammatory pain models [20-111]. Activation of NPFFR2 reduces MOR-mediated anti-nociception, resulting in bell-shaped response curves in acute pain models. However, in chronic inflammatory pain models, MDOR activation produces more potent anti-nociception. MCRT demonstrated limited tolerance and opioid-induced hyperalgesia in both acute and chronic pain models, without inducing cross-tolerance to morphine. Furthermore, MCRT exhibited no addictive properties, gastrointestinal suppression, or effects on motor coordination. As a multifunctional compound, MCRT holds promise for reducing chronic inflammatory pain while minimizing opioid-associated adverse effects [112]. 

Co-administration of novel combinations simultaneously targeting MOR and DOR in peripheral and central nervous systems produces synergistic analgesia. The analgesic effects of morphine or oxymorphindole (a partial DOR agonist) alone were compared with those of the combination of loperamide (a peripherally restricted MOR agonist) and either N-benzyl-oxymorphindole or loperamide. Compared to morphine sulfate, both combinations demonstrated higher analgesic efficacy and effectively reduced post-injury hypersensitivity without side effects [113,114].

Table 1: Summary of Mechanisms of Action and Development Strategies for Analgesics Targeting the Mu-Opioid Receptor

MOR agonists cause respiratory depression, whereas DOR agonists do not. DOR represents a potential target meeting the requirements for non-addictive analgesics. DOR agonists offer more controllable receptor activation,but their development has been hindered by uncertainties surrounding the molecular mechanisms of partial agonism. C6-Quino, a selective partial DOR agonist, is a structurally engineered bis-ligand exhibiting distinct activities in G-protein and arrestin pathways. It interacts with the sodium-binding pocket, demonstrating analgesic activity without inducing DOR-associated seizures or MOR-related adverse effects [115,116].

Designing Novel Opioid Drugs

New opioid medications offer potent analgesic effects with enhanced safety profiles. Tapentadol is an opioid analgesic that acts on the central nervous system by agonizing MOR receptors and inhibiting Norepinephrine (NE) reuptake [117]. It was first approved by the FDA in 2008 as an Immediate-Release (IR) oral tablet for treating moderate to severe pain. Its Extended-Release (ER) formulation was later approved for neuropathic pain associated with diabetic peripheral neuropathy. Tzschentke et al. also evaluated tramadol’s analgesic effects across several rat and mouse pain behavior models, including the hot plate test, tail-flick test, mustard oil induced visceral pain, Spinal Nerve Ligation (SNL), and writhing response. Results indicated that compared to morphine, tapentadol exhibited lower binding affinity to MOR receptors but demonstrated analgesic potency only 2-3 times lower than morphine, with delayed tolerance development [118]. Freynhagen et al., found tapentadol caused less nausea and constipation than other opioids [119].

Claudius E. Degro et al. designed a pH-sensitive drug, a fluorinated analog of fentanyl, (±)-N-(3-fluoro-1-phenylethylpiperidin-4-yl)-N-phenylpropionamide (NFEPP). This compound selectively targets MOR receptors within acidic microenvironments found at sites of tissue injury, inflammation, and cancer. Preclinical studies in rodents for inflammatory and cancer pain demonstrated that NFEPP produces potent analgesia without activity in non-inflamed tissues, and without inducing the common side effects or addiction potential associated with traditional opioids [32-120]. Repeated administration of NFEPP maximizes its analgesic effect when inflammation is most severe, and it loses activity once inflammation resolves [33]. In summary, compared to the traditional opioid fentanyl, the novel pH-sensitive analgesic NFEPP exhibits minimal tolerance development and maintains its analgesic efficacy in inflammatory pain models. Consequently, the effective analgesia demonstrated by the pH-sensitive analgesic NFEPP appears less susceptible to tolerance development, potentially offering a clinical alternative for treating inflammatory conditions without the risk of severe side effects associated with dose escalation seen with traditional opioids [121].

Refractory pain is defined as pain that cannot be relieved due to inadequate therapeutic response or intolerable adverse drug reactions. In such cases, rotating opioids—that is, switching from one opioid to another or altering the route of administration—can improve pain relief while reducing opioid side effects [122,123]. The most frequently used medication is methadone, which offers rapid onset, prolonged duration of action, high oral bioavailability, and incomplete cross-tolerance with other opioids [124]. A study involving 92 patients with pain demonstrated that low-dose methadone combined with another opioid is both safe and effective. Methadone can provide benefits for pain control in refractory pain, even among patients with advanced disease who have long term opioid dependence [125].

A recent study describes an innovative morphine-paracetamol adduct, termed Metamorphine, successfully synthesized via Mannich condensation by introducing a phenazone moiety at the C2 position of a morphine polysaccharide skeleton. The discovery of this adduct originated clinically: It was identified as a byproduct of morphine-phenacetin drug interactions within Patient-Controlled Analgesia (PCA) pumps used for severe pain management. Given morphine’s status as an effective selective agonist of human MOR, we evaluated Metamorphine’s binding and activation activity against MOR in vitro alongside two other MOR agonist: [Met5] enkephalin, [D-Ala,2N-Me-Phe,4Gly5-ol]enkephalin (DAMGO), and TRV-130 (oliceridine). The discovery of Metamorphine lays a foundation for a novel class of opioids with broader activity [126,127]. Multidrug or multi-target approaches represent an advantageous drug design strategy aimed at developing compounds capable of targeting multiple receptors. Compared to traditional polypharmacy regimens, strategies achieving multi-receptor regulation via a single molecule can significantly optimize clinical drug use. This includes enhancing patient compliance by reducing the number of medications administered, particularly in patients with complex diseases or multiple conditions.

Designing novel allosteric ligands can modulate MOR. Positive Allosteric Modulators (PAMs) of MOR bind to its allosteric site, stabilizing its active conformation through allosteric regulation. This enhances the efficacy of endogenous opioids (e.g., endorphins) or low-dose opioid drugs, thereby providing opioid-sparing effects. This allows for the use of lower doses of opioid agonists and may reduce associated side effects. Experiments demonstrated that halogen substitution enhances the ability of parent compound 1 (BMS-986122) to potentiate standard DAMGO activity. One of the most potent compounds, lead thiazolidine derivative 14b, exhibited in vivo activity in enhancing morphine-induced anti-nociception in mice without significant side effects [128,129]. A new negative allosteric modulator of MOR (NAM) 368 can cooperate with naloxone to regulate the interaction between fentanyl and MOR. In molecular mechanism, 368 directly forms hydrogen bonds with naloxone by binding to the extracellular vestibular region of MOR, which enhances the affinity of naloxone to it, prolongs its residence time on the receptor and reduces dissociation. At the same time, 368 can stabilize the unique inactivated conformation of the extracellular regions of MOR (such as TM2 and TM7), hinder the conformational change of receptor activation induced by fentanyl, and then inhibit the activation of Gi protein mediated by fentanyl. In vivo experiments, it is difficult to reverse the analgesic effect induced by fentanyl when low-dose naloxone is used alone, but when 368 is used in combination with low-dose naloxone, it can significantly enhance the antagonistic effect of naloxone on fentanyl-induced analgesia, respiratory inhibition and other effects, and will not aggravate the withdrawal reactions caused by naloxone alone, such as diarrhea and jumping behavior [130,131]. Additionally, dual-site ligands C6 guano and RO76 reduce adverse effects like respiratory depression and addiction by decreasing G protein efficacy and reducing β-arrestin recruitment. The former achieves this through direct binding to the sodium ion site, while the latter does so via water molecule bridging [132,133].

Other Medications for Adjuvant Analgesia

Multimodal pain management offers effective pain control while significantly reducing adverse reactions associated with opioid use [122,123]. Multimodal analgesic techniques have been extensively studied, including serotonin-norepinephrine reuptake inhibitors (SNRIs) [134]. Duloxetine, an SNRI initially approved for major depressive disorder, demonstrates analgesic effects in chronic pain conditions including diabetic peripheral neuropathic pain, fibromyalgia, and chronic pain associated with major depressive disorder [135]. While duloxetine provides analgesia for Total Knee Arthroplasty (TKA) patients, its dosing regimen remains unstandardized. Studies indicate that low-dose duloxetine reduces early postoperative morphine requirements and improves Knee Injury and Osteoarthritis Outcome Score (KOOS) symptoms at 6 and 12 weeks. However, it does not significantly alleviate pain at rest or during walking. Tolerability of low-dose duloxetine was comparable to controls. These findings suggest low-dose duloxetine may serve as an adjunct to contemporary multimodal pain management strategies in TKA patients [136]. Tramadol-Celecoxib (CTC) co-crystals provide effective analgesia in acute pain models. Co-crystallization alters the pharmacokinetics of individual components, potentially enhancing tolerability. A pooled safety analysis of phase 3 randomized controlled trials in adults with acute moderate-to-severe pain following oral surgery, bunionectomy, and elective abdominal hysterectomy demonstrated that CTC 200 mg BID exhibited superior safety and tolerability compared to tramadol 100 mg QID in acute moderate-to severe postoperative pain. The tolerability advantage and analgesic efficacy of CTC contribute to a more favorable benefit-risk profile and highlight its potential for treating acute moderate to severe pain [137-141].

Recent studies have found that antidepressants can serve as adjuncts to analgesics, and their combination with opioids may help reduce the risk of addiction [63,64]. In a chronic osteoarthritis pain model, treatment with the combination of mirtazapine and morphine was evaluated over a 25-day period. Assessments included pain symptoms, tolerance development, withdrawal symptoms, and MOR protein expression levels. Results indicated that this combination reduced analgesic tolerance and prevented the progression of withdrawal symptoms [142]. The mirtazapine-morphine combination may represent a viable option for mitigating opioid-related side effects.

Non-Opioid Analgesic Routes

Acetaminophen is a non-opioid analgesic that exerts its analgesic and antipyretic effects by metabolizing into para-aminophenol. Para-aminophenol crosses the blood-brain barrier to activate nociceptive receptors, with pain regulation mechanisms directly influencing the brain [143]. When used short-term and at appropriate doses, acetaminophen is generally well-tolerated; overdose may cause hepatotoxicity [143]. Other side effects may include rash, nephrotoxicity, and electrolyte abnormalities.

Increasing evidence suggests that opioid analgesics are involved in the upregulation or downregulation of angiogenesis [144]. Recently, the relationship between vascular endothelial growth factor-A (VEGF-A) and pain management has drawn attention. To date, no anti-angiogenic drugs have been used for pain management. N-Palmitoylethanolamine (PEA) delays the onset of morphine tolerance, enhances morphine analgesia, and reduces angiogenesis in animal models. Compared to morphine alone, PEA resulted in lower expression of VEGF-A and soluble VEGF receptor 1 (sFLT-1) in rat Dorsal Root Ganglia (DRG) and spinal cord. It is speculated that PEA may exert its effects on morphine analgesia and tolerance by mediating downregulation of VEGF-A and sFLT-1. This study provides new insights into the mechanism by which ultrafine PEA delays morphine tolerance and enhances morphine analgesia [145-147].

Research by Antoine Jouvenel et al., indicates that activation of the fms-like tyrosine kinase 3 (FLT3) receptor expressed in primary afferent sensory neurons modulates MOR-dependent classical cAMP-PKA pathways, leading to neuronal hyperexcitability that drives the initiation of adverse reactions to opioid analgesia and promotes the development of chronic Opioid-Induced Tolerance (OIT) and Hyperalgesia (OIH). Consequently, peripheral FLT3 blockade prevents OIT and OIH while substantially enhancing morphine’s analgesic efficacy in both acute and chronic pain models [148,149].

Many compounds still share the same scaffold as morphine or possess similar synthetic scaffolds, which in turn limits the diversity of active compounds. Furthermore, this scaffold has been reported to induce adverse reactions, including respiratory depression and constipation. Advances in X-ray crystallography and Cryo-EM in recent years have facilitated the resolution of more opioid receptor structures, offering unprecedented opportunities for computer-aided drug design.

As a key molecule in reward circuits, pain pathways, and emotional regulation, MOR exhibits complex and diverse functions, playing a vital role in numerous signaling pathways. Consequently, it holds a pivotal position in the mechanisms of opioid analgesia and addiction. MOR exhibits extensive distribution with specific expression in numerous brain regions, forming a multidimensional regulatory network linking “pain perception-reward feedback-emotional regulation.” This distribution pattern not only explains the biological basis for opioids’ potent analgesic effects alongside their propensity to induce euphoria and emotional dependence but also provides anatomical targets for precise intervention in MOR function. At the addiction mechanism level, MOR signaling primarily involves G protein-dependent pathways and β-arrestin pathways: prolonged morphine treatment enhances GRK-mediated MOR phosphorylation, promoting β-arrestin recruitment and receptor internalization. This simultaneously weakens G protein-mediated analgesia and activates other pathways to exacerbate tolerance. Conversely, abrupt drug withdrawal triggers compensatory increases in cAMP levels, leading to withdrawal symptoms. Concurrently, activation of the TLR4-cGAS-STING pathway in microglia triggers release of the proinflammatory cytokine IL-1β, inducing neuroinflammation. This not only directly counteracts morphine analgesia but also modulates synaptic plasticity in the VTA-NAc pathway by altering the inflammatory microenvironment within the synaptic cleft.

Current MOR-based drug development fundamentally aims to modulate its signaling bias or synergistic effects to achieve the goal of “preserving analgesia while reducing addiction.” Bias agonists like TRV-130 demonstrate lower addiction potential in clinical settings by preferentially activating G protein pathways and inhibiting β-arrestin recruitment. while multifunctional ligands like MCRT achieve a balance between analgesic efficacy and tolerance by simultaneously targeting MOR, MDOR, and NPFFR2. However, these strategies still face challenges. Future research should further explore MOR expression differences across genders and age groups, as well as the regulatory effects of environmental factors like chronic stress on MOR signaling, to provide a basis for developing personalized analgesic regimens.

In summary, the function of MOR in the nervous system is not a simple “analgesia-addiction” binary regulation, but rather involves a complex network of multiple pathways and cell types. Deeply deciphering the dynamic equilibrium mechanisms of MOR signaling, combined with structural biology and computer-aided drug design techniques, will enable the development of MOR modulators with enhanced selectivity and tissue targeting. Concurrently, exploring synergistic intervention strategies targeting related pathways such as neuroinflammation and synaptic remodeling will be the core direction for advancing the development of low-addiction analgesics and optimizing clinical pain management.

Funding

This study was supported by the National Natural Science Foundation of China (No. U2002220, 82371271), the innovation team of stress and defense of Yunnan Province (202305AS350011).

Contributions

Yang Long designed the review framework, conducted the literature search, analyzed the relevant studies, and drafted the manuscript. Jie Bai provided expert guidance, critically revised the intellectual content, and approved the final version for publication. All authors read and approved of the final manuscript.

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