Among the various VGSC isoforms, Naᵥ1.3 (encoded by SCN3A) has gained attention due to its dynamic expression pattern and strong association with neuronal hyperexcitability following nerve injury (Smith et al., 2018[218], Liao et al., 2023[154]). Under normal physiological conditions, Naᵥ1.3 is mostly generated in the central nervous system (CNS), where it is crucial for controlling the excitability and maturation of early neurons. On the other hand, its expression is either very inadequate or absent in the adult nervous system, indicating that Naᵥ1.3 is more involved in neuronal development than in mature sensory signaling (Cummins et al., 2001[49], Liao et al., 2023[154]). According to several studies, chronic hyperglycemia and diabetes cause aberrant upregulation (re-expression) of Naᵥ1.3 in DRG neurons. It is re-expressed in adult sensory neurons after nerve damage or diabetic neuropathy. This upregulation is considered a major contributor to neuronal hyperexcitability, a fundamental mechanism underlying the abnormal pain sensations observed in neuropathic conditions (Dib-Hajj et al., 1998[52], Black et al., 2004[25], Fukuoka et al., 2008[80]). Functionally, Naᵥ1.3 channels are characterized by rapid recovery from inactivation and a low activation threshold, enabling neurons to fire at higher frequencies and sustain repetitive discharges. These properties make Naᵥ1.3 particularly effective in driving ectopic or spontaneous firing in injured sensory neurons, one of the defining electrophysiological features of DN (Waxman et al., 1994[249], Liu et al., 2000[161], Zhao et al., 2006[280]). Multiple molecular pathways regulate overexpression of Naᵥ1.3, for example, it has been demonstrated that downregulating microRNA-30b increases Naᵥ1.3 expression in damaged DRG neurons, whereas increasing miR-30b level decreases aberrant pain responses (Su et al., 2017[220]). Additionally, intracellular modulators such as SIRT1 downregulation in the spinal dorsal horn increase acetylation of the SCN3A, further upregulating Naᵥ1.3 channel expression, enhancing neuronal excitability, and contributing to pain signaling, while Fibroblast Growth Factor 14 (FGF14) interacts with Naᵥ1.3 channels and regulates channel inactivation kinetics and influences the neuronal firing patterns and overall excitability (Martinez-Espinosa et al., 2021[171], Wang et al., 2024[245]). Additionally, the tetrodotoxin-sensitive current in damaged DRG neurons and biophysical properties of Naᵥ1.3 were similar, indicating a function for Naᵥ1.3 in injury-induced neuronal hyperexcitability. The DRG and sciatic nerves of diabetic neuropathic rats exhibit suppression of miR-214-3p, a conserved miRNA that targets Naᵥ1.3. By regulating Naᵥ1.3 in the DRG, overexpression of miR-214-3p reduces streptozotocin (STZ)-induced neuropathy, nerve conduction retardation, neural lesions, inflammation, and apoptosis (Wang et al., 2024[243]). These evidences indicate that Naᵥ1.3 upregulation under diabetic conditions contributes to aberrant neuronal firing and lowered pain thresholds, thereby intensifying the persistent burning and tingling sensations characteristic of PDN.

The VGSC Naᵥ1.7, regulated by SCN9A gene, has been identified as a crucial mediator of peripheral pain perception due to it is widely distribution in sympathetic neurons and dorsal root ganglia, where it controls neuronal firing properties. (Bang et al., 2018[16], Dormer et al., 2023[58]). By increasing small, gradual depolarizations that result in what scientists refer to as ramp currents, Naᵥ1.7 regulates the firing threshold of sensory neurons in normal physiology. This amplification prevents undesired spontaneous firing while ensuring that neurons react correctly to harmful stimuli. The channel maintains its availability at resting membrane potential, where it may react to slight depolarizations, due to its slow inactivation kinetics. However, at higher depolarized voltages, the spontaneous activation channel becomes inactive (Goodwin et al., 2022[93], Deng et al., 2023[51]). It is distinctive biophysical characteristics, such as quick activation and delayed kinetics of closed-state inactivation, that allow for precise modulation of nociceptive signaling without triggering undesired neuronal firing (Shields et al., 2012[210], Fouillet et al., 2017[77], Dong et al., 2025[57]). Peripheral nerve injury, diabetic neuropathy, and other diseases are caused by significant changes in Naᵥ1.7 expression level and functional features during pathological situations such as diabetes and chronic hyperglycemia (Hameed, 2019[105]). While Naᵥ1.7 amplifies external stimuli and depolarizes membrane potentials closer to the threshold for Naᵥ1.8 activation, gain-of-function mutations in Naᵥ1.7 may cause diabetes-induced increased sensitivity of DRG neurons (Yang et al., 2016[258]). These gain-of-function mutations may be linked to the severity of pain and may cause axonal degeneration through energy stress. For example, sensory axons with the Naᵥ1.7 G856D mutation associated with small fiber neuropathy have lower ATP level and more reactive oxygen species (Rolyan et al., 2016[203]). It has been consistently observed that DRG neurons with increased Naᵥ1.7 expression exhibit lower action-potential firing thresholds, increased sodium influx, and aberrant spontaneous neuronal activity, which cause hyperalgesia and allodynia (Li et al., 2019[152]). From a physiological perspective, Naᵥ1.7 channel play a key role in the neuronal hyperexcitability that characterizes neuropathic pain states because they sustain recurrent firing and amplify weak stimuli (Chang et al., 2018[41], Mehboob et al., 2021[175]).

Naᵥ1.8 (SCN10A) is a tetrodotoxin-resistant sodium channel that plays a key role in pain signaling. It is found mainly in pain-sensing nerve cells of the dorsal root ganglia, where it generates most of the sodium current needed to trigger action potential. Its unique properties allow pain neurons to fire repeatedly at high frequencies, which is essential for transmitting pain signals to the central nervous system (Heinle et al., 2024[107]). Under physiological conditions, Naᵥ1.8 has unique electrical properties that make it work differently from other sodium channels. Unlike typical sodium channels, it activates and inactivates at higher voltages, and shuts down slowly. More importantly, Naᵥ1.8 does not fully stop working when the neuron is depolarized, it keep some channels open and allows current to flow through. Means Naᵥ1.8 keeps working when other sodium channels have stopped, allowing pain neurons to keep firing even during sustained stimulation (Xiao et al., 2019[254]). Naᵥ1.8 channel expression and location change with nerve damage or metabolic stress. While adjacent undamaged sensory axons increase Naᵥ1.8 channel density, especially in the periphery of the nerve, Naᵥ1.8 mRNA and protein levels decrease in directly injured dorsal root ganglion neurons. The development and duration of neuropathic pain are caused by this redistribution, which encourages spontaneous firing and ectopic electrical activity (Ma et al., 2019[165]). Studies on methylglyoxal has shown that Naᵥ1.8 plays a crucial role in DN, where hyperglycemia increases the production of methylglyoxal. By activating Naᵥ1.8 expressed on the primary afferent sensory neurons, endogenously elevated methylglyoxal levels may trigger PDN (Bierhaus et al., 2012[22]). Additionally, a continuously active version of Naᵥ1.8 is produced when methylglyoxal binds to arginine residues. Thus, methylglyoxal depolarizes sensory neurons and induces posttranslational modifications in Naᵥ1.8, which appears to be the cause of primary hyperalgesia in diabetic people and animals (Bierhaus et al., 2012[22]).

In DN, reactive carbonyl species can modify channel residues and shift gating toward greater availability, amplifying nociceptive signaling even without overt axonal loss (Bierhaus et al., 2012[22]). Gain-of-function SCN10A variants found in small-fiber which delay inactivation and enhance persistent sodium influx, leading to sustained firing in DRG neurons and increased sensory excitability. These changes are consistent with the clinical pain phenotypes observed in affected patients (Garrison et al., 2014[85]). Studies have demonstrated that human pain-related tissues frequently exhibit elevated Naᵥ1.8 immunoreactivity, which is consistent with the burning and tingling sensations typical of neuropathic conditions (Bird et al., 2013[24]). The threshold for action potential generation is lowered by slight increases in Naᵥ1.8 conductance or a hyperpolarization in its activation curve, according to computational modeling, which also replicates important electrophysiological characteristics of neuropathic pain, such as aberrant action potential shape, increased repetitive firing, and elevated neuronal excitability (Kan et al., 2024[133]).

Naᵥ1.9 represents a tetrodotoxin-resistant voltage-gated sodium channel encoded by SCN11A that demonstrates preferential expression within small-diameter nociceptive neurons throughout dorsal root ganglia and trigeminal ganglia regions (Vanoye et al., 2013[234], Dib-Hajj et al., 2015[53]). This channel differs from Naᵥ1.7 and Naᵥ1.8 in that it shows activation characteristics close to resting membrane potential values, about −70 mV, allowing for the generation of low-threshold, sustained sodium currents that prolong activity during slight depolarizing events (Sleeper et al., 2000[217]). Functionally, Naᵥ1.9 operates as a subthreshold amplification mechanism through its persistent current, enhancing neuronal responsiveness to diminished or gradual stimulation patterns (Zhao et al., 2023[278]). Under normal conditions, the channel facilitates the firing of action potentials by increasing the resting membrane potential toward positive values and maintains the elevated sensitivity of the neurons, allowing them to recognize signals of inflammatory pain. To transmit strong, persistent pain signals to the brain, the channel produces prolonged, repeated firing and threshold activity (Bennett et al., 2019[20]). While Naᵥ1.9 generates complex firing patterns in healthy DRG neurons, such as evoked firing and spontaneous bursts, its potential to contribute to neuropathic pain caused by nerve damage is yet unclear. In contrast, Naᵥ1.9 is known to be associated with inflammatory pain, where inflammatory mediators either upregulate it or enhance its post-translationally to increase nociceptor response (Huang et al., 2014[117]). During diabetes, sustained hyperglycemia and oxidative stress trigger several mechanisms to raise sodium channel activity: (1) increased expression of Naᵥ1.9 in DRG neurons; (2) post-translational phosphorylation of Naᵥ1.9 via PKA/PKC pathways; and (3) ROS-dependent cholesterol oxidation, which hyperpolarizes Naᵥ1.9 activation thresholds. Notably, Naᵥ1.9 is upregulated mostly in large diameter neurons, which is an ectopic expression pattern different from its typical small fiber location and may be a factor in allodynia (Wang et al., 2024[243], Yang et al., 2025[259], Bigsby et al., 2022[23]). Following peripheral nerve injury, animal model investigations consistently demonstrate reductions in Naᵥ1.9 expression in sensory neurons (Amaya et al., 2006[7]). Various studies have described that Naᵥ1.9 knockout and antisense knockdown rodents consistently reduce inflammatory pain but have little effect on neuropathic pain, after nerve injury or baseline acute pain thresholds. This selectivity reflects differential channel regulation of inflammatory mediators such as PGE2, bradykinin, and IL-1β, which strongly potentiate Naᵥ1.9 through PKA/PKC pathways, whereas nerve injury triggers upregulation of Naᵥ1.7 and Naᵥ1.3 instead (Maingret et al., 2008[167], Kakimura et al., 2010[131], Lolignier et al., 2011[162]). Thus, Naᵥ1.9 acts as a crucial regulatory factor among voltage-gated sodium channel families, managing nociceptive neuron excitability and reactivity to inflammatory and metabolic signaling cascades, making it a significant contributor of DN (Amsalem et al., 2018[8]).

N-type calcium channel (Caᵥ2.2) an important voltage-activated calcium channel subtype that control presynaptic neurotransmitter release in nociceptive pathways and is encoded by CACNA1B gene (Pearson, 2007[188]). These channels, abundantly expressed in DRG neurons and their central terminals in the spinal dorsal horn, act as molecular transducers that couple membrane depolarization to calcium influx, causing release of excitatory neurotransmitters such as glutamate, substance P, and calcitonin gene-related peptide (CGRP) from synaptic vesicles (Chi et al., 2009[44], Park and Luo, 2010[187], Catterall, 2011[38]). Under physiological conditions, these regulate calcium-dependent processes and maintain normal sensory signal transmission. In DN conditions, this homeostatic process is severely disrupted by chronic hyperglycemia and related metabolic abnormalities, altering Caᵥ2.2 from a physiological mediator into a pathological cause of neuronal damage and chronic neuropathic pain (Joksimovic et al., 2022[128]).

Persistently elevated blood glucose leads to the accumulation of ROS and AGEs in the body, which triggers MAPKs, involved in downstream signaling pathways (González et al., 2023[91]). In Metabolic dysfunction, PKC, calmodulin-dependent protein kinase II (CaMKII), and MAPK kinases directly phosphorylate Caᵥ2.2 channels, increasing their opening frequency and accelerating their activation kinetics, allowing upsurged calcium influx through existing channel proteins, while concurrent upregulation of CACNA1B transcription via CREB and NF-κB increases channel expression, producing a synergistic amplification of presynaptic calcium influx (Szymanowicz et al., 2024[221]). Chronic hyperglycemia also causes the α2δ-1 auxiliary subunit (CACNA2D1) to be upregulated, which increases Caᵥ2.2 channel trafficking to the presynaptic membrane and, thereby, enhances calcium influx during action potentials (Bauer et al., 2009[19]). In nerve terminals, excessive influx dysregulates Caᵥ2.2 channels, triggering mitochondrial calcium overload, reducing ATP production, and initiating the axonal degeneration typical of progressive DN.

The L-type channels (Caᵥ1.2 and Caᵥ1.3) located in neuronal soma and dendrites are involved in signal transduction and provide a molecular intermediate between calcium influx and intracellular calcium signaling pathways rather than vesicle release (Zhang et al., 2006[271]). In normal conditions, calcium entry via these channels results in activation of calcineurin, PKA and CaMKII, which phosphorylate the downstream transcription factors cAMP response element-binding protein (CREB) and Nuclear Factor of Activated T cells (NFAT). Gene regulation by activity-dependent calcium signaling is essential for maintaining neuronal plasticity and metabolic balance (MacDonnell et al., 2009[166], Lee and Fields, 2021[149]). However, in DN, chronic hyperglycemia and oxidative stress transform this adaptive calcium signaling into a pathological process.

ROS produced from hyperglycemia activate various PKC isoforms (PKC-β and PKC-ε) (Yang et al., 2009[257]), which can phosphorylate Caᵥ1.2 channels in some contexts, but in acute or direct glucose-mediated hyperglycaemic signaling, the primary kinase phosphorylating Caᵥ1.2 at Ser1928 is PKA. Hyperglycemia also activates PKA-mediated phosphorylation at the Ser1928 site on Caᵥ1.2, enhancing Caᵥ1.2 channel activity and elevated calcium entry into neurons under hyperglycemic conditions (Nystoriak et al., 2017[184]). The resultant increase in presynaptic and somatic calcium influx exceeds physiological values. The excessive L-type calcium load also surpasses the mitochondrial calcium retention threshold, inhibiting oxidative phosphorylation, reducing neuronal ATP production and inducing opening of the mPTP (Strubbe-Rivera et al., 2021[219], Ivanova et al., 2025[121]). Diminished caveolin-1 expression promotes demyelination through enhanced Erb B2 signaling in Schwann cells. Loss of Cav-1 impairs myelin maintenance, causing secondary axonal damage and reduced neuroprotection (McGuire et al., 2009[173]). Increased calcium also activates the CaMKII and ERK/MAPK cascades, which reduce anti-apoptotic gene expression (Bcl-2) while promoting pro-apoptotic signaling. This transcriptional alteration drives the apoptosis of sensory neurons, accounting for the progressive loss of nerve fibers in DN (Timmins et al., 2009[226], Chung et al., 2018[46]).

T-type calcium channels, particularly (Caᵥ3.2), encoded by the CACNA1H gene (Cain et al., 2018[31]), key regulators which control subthreshold excitability and burst firing in nociceptive pathways DRG neurons. especially C-fiber nociceptors, express Caᵥ3.2 in abundance at the soma and peripheral terminals. These low-voltage-activated channels open near resting membrane potentials act as molecular amplifiers, converting small membrane depolarizations into calcium influx, thereby promoting repetitive firing and improving nociceptive signaling in the spinal dorsal horn (Todorovic and Jevtovic-Todorovic, 2011[227], Voisin et al., 2016[238]). This low-threshold calcium influx stabilizes neuronal firing rates and preserves normal sensory response under physiological conditions. Relying on CACNA1H transcriptional upregulation, posttranslational changes, mainly N-linked glycosylation, are the main cause of the approximately two-fold increase in Caᵥ3.2 activity in diabetic DRG neurons , T-type calcium currents are amplified, the Caᵥ3.2 pore-forming subunit is phosphorylated, oxidative changes (glycation and S-nitrosylation) occur, and neuronal excitability exceeds physiological thresholds (Joksimovic et al., 2020[127]). Elevated low-threshold calcium influx disrupts intracellular homeostasis, facilitates mitochondrial calcium overload above the organelle retention capacity, and impairs oxidative phosphorylation, ultimately decreasing ATP production and contributing to progressive axonal degeneration characteristic of diabetic neuropathy (Gleichmann and Mattson, 2011[89]). Additionally, persistent Caᵥ3.2 hyperactivity strengthens both peripheral and central sensitization mechanisms in nociceptive pathways. Increased glutamate and neuropeptide release in the dorsal horn, ectopic spontaneous firing at the peripheral terminal, and long-term potentiation-like plasticity are all facilitated by enhanced T-type channel activity, which sustains chronic diabetic neuropathic pain (Jacus et al., 2012[123]).

Among the various subtypes of VGKC, especially Kᵥ_1.1, Kᵥ_1.2, and Kᵥ_1.4, encoded by KCNA1, KCNA2, and KCNA4, are highly expressed in DRG neurons, with the expression of Kᵥ_1.1 and Kᵥ_1.2 predominantly in large-diameter neurons, and the subunit Kᵥ_1.4 in small-diameter neurons. These are present on the soma and juxtaparanodal regions of myelinated axons. These channels mediate low threshold, slowly inactivating outward K⁺ currents that stabilize the resting membrane potential and shorten the action potential, thereby limiting repetitive firing and ectopic discharges (Rasband et al., 2001[196]). These channels act as an electrical brake, they stop repolarization, ultimately increasing depolarization in neurons and more likely to fire spontaneously.

In a streptozotocin (STZ) rat model of painful diabetic neuropathy, Cao et al. (2010[35]) showed that in hyperglycemia BDNF expression is increased in DRG neuron via tyrosine phosphorylation, causing a reduction in Kᵥ currents, especially in Type-A current (Cao et al., 2010[35]). In myelinated nerve fibers, Kᵥ₁ channels are present at juxtaparanodal regions adjacent to nodes of Ranvier, where they play essential roles in action potential repolarization and nerve excitability. Hyperglycemia causes altered distribution and reduced presence of Kᵥ1.2 subunits in juxtaparanodal regions. This mis-localization disrupts the normal spatial organization of ion channels necessary for saltatory conduction (Zenker et al., 2012[269]). According to Zhao et al. (2017[279]), hyperglycemia-induced nerve damage enhances DNMT3a in DRG neurons, which reduces Kᵥ1.2 expression and methylates the Kcna2 promoter. This decrease in Kᵥ1_.2 contributes to DN by lowering potassium currents and increasing neuronal excitability (Zhao et al., 2017[279]).

Kᵥ₁ channel dysfunction is caused by metabolic and oxidative stress processes via numerous mechanisms. Hyperglycemia activates PKC, especially the β and δ isoforms, by increasing the production of diacylglycerol (DAG), while AGE accumulation in the tissue lowers Kᵥ channel current density and significantly decreases expression at both gene and protein level, initiating downstream signaling cascades that downregulate channel expression (Liu et al., 2019[160]). PKC-mediated phosphorylation inhibits Kᵥ₁ channel activity and decreases its surface expression via inducing ubiquitination-dependent endocytosis and lysosomal degradation, among other consequences. Because PKC activation may activate NADPH oxidase 2, which produces reactive oxygen species (ROS), the PKC pathway is also associated with oxidative stress. Through both functional inhibition and decreased expression, this PKC-ROS axis contributes to decreased Kᵥ channel activity (Du et al., 2021[60]). Also pro-inflammatory cytokines, particularly IL-1β, IL-6, and TNF-α, are elevated in hyperglycemia and contribute to Kᵥ1 channel dysregulation. These cytokines promote neuroinflammation, microglial activation, and neuronal hyperexcitability. Inflammatory mediators can indirectly affect Kᵥ₁ channel expression and function through activation of intracellular signaling cascades, including NF-κB and MAPK pathways. Reduction of Kᵥ channel expression leads to several pathophysiological effects such as membrane potential depolarization, hyperpolarized action potential threshold, and increased spontaneous firing rates of sensory neurons (Andrei et al., 2025[10]).

The Kᵥ3 VGKC triggers a high-voltage-activating A-type Kᵥ current found in axons and nerve terminals. These high-voltage-activated K⁺ channels have fast activation and very fast deactivation, classically associated with fast-spiking neurons. In peripheral sensory pathways, the main Kᵥ3 subunit with clear nociceptive relevance is Kᵥ3.4, encoded by the KCNC4 gene (Zemel et al., 2018[268]). Immunohistochemical and electrophysiological studies show that Kᵥ3.4 is strongly expressed in small-diameter nociceptive DRG neurons, especially C-fibers, with channel protein located in soma, axons and peripheral terminals. Kᵥ3.4 currents in DRG neurons generate a fast-inactivating A-type outward K⁺ current that helps narrow the action potential and limit Ca²⁺ entry at nerve endings, thereby regulating nociceptive signals in a homeostatic manner (Ritter et al., 2012[200]).

Ritter et al. (2015[201]) demonstrated that small-diameter DRG neurons from spinal cord injury animals show a marked reduction in Kᵥ3.4-mediated A-type potassium current due to altered channel trafficking and changes in the inactivation properties of the Kᵥ3.4 N-terminal domain, which normally ensures rapid current inactivation. As a result of this dysregulation, action potentials become broader and allow greater calcium influx into the terminals of nociceptive neurons (Ritter et al., 2015[201]). Additionally, in the streptozotocin (STZ) model of type 1 diabetes, Cao et al. (2010[35]) reported a substantial decrease in total voltage-gated potassium currents, with the downregulation of Kᵥ3.4 channel. This decrease in 3.4 current results in membrane depolarization and increasing hyperexcitability, consequently causing DN (Cao et al., 2010[35]).

The VGKC, Kᵥ4 (Kᵥ4.1, Kᵥ4.2, Kᵥ4.3, encoded by KCND1, KCND2, and KCND3, respectively), is fundamentally different from the Kᵥ₁ and Kᵥ₃ families in their biophysical properties and physiological roles in nociceptors. These channels activate at subthreshold voltages (-40 to -20 mV), rapidly activate and inactivate in response, producing a transient outward K+ current uniquely characterized by a fast recovery from inactivation (Covarrubias et al., 2008[48], Zemel et al., 2018[268]).

Chronic hyperglycemia leads to hyperexcitability in neurons. Kᵥ₄ channels undergo pronounced dysregulation. Several studies showed that sciatic nerve injury, spinal nerve ligation, and trigeminal nerve models consistently show a selective downregulation of Kᵥ_4.2 and Kᵥ_4.3, leading to a marked reduction in A-type current density (Viatchenko-Karpinski et al., 2018[237]). Thereby extending action potential, increasing calcium influx, Kᵥ_4.3 downregulation uniquely contributes to cold allodynia, and selective pharmacological obstruction of Kᵥ_4.3 in otherwise healthy animals is sufficient to induce cold hypersensitivity, providing clear evidence that this subunit acts as a critical determinant of cold-pain processing (Kanda et al., 2021[134]). Similarly, BDNF elevations, MAPK phosphorylation via Thr602, PKC activation, and inflammatory cytokines alter the expression of Kᵥ₄ subtypes (Carrillo-Reid et al., 2019[36]). In such conditions, Kᵥ4.2 and Kᵥ_4.3 mRNA levels are reduced approximately 40-50 % in DRG neurons, with protein level similarly decreased, resulting in a reduction in total Kᵥ4 current density leads to DN.

Kᵥ₇ channels, also known as M channels (Kᵥ_7.1 to Kᵥ_7.5 encoded by KCNQ1-KCNQ5), are VGKC with interesting biophysical properties (slow activation and deactivation, no inactivation, and a threshold for activation below -60 mV) (Greene and Hoshi, 2017[95]). The K⁺ current is activated near the resting membrane potential. These are expressed in small-medium DRG neurons, acting as an excitability stabilizer. Downregulation or inhibition of Kᵥ7 channels leads to enhanced excitability in central and peripheral neurons, with rapid depolarization and spontaneous firing (Barkai et al., 2017[17]). Additional injury model studies demonstrated that peripheral nerve damage increases expression of the transcriptional repressor (REST) in DRG neurons which suppresses Kcnq2 transcription and leads to a marked reduction in Kᵥ_7.2 expression. A similar breakdown occurs in DN (Rose et al., 2011[204]), where chronic hyperglycemia reduces expression of KCNQ2, KCNQ3, and KCNQ5 in DRG neurons and decreases Kᵥ_7.5 immunoreactivity in small nociceptive cells, leading to a pronounced reduction in M-current. As a result, diabetic DRG neurons shift to a depolarized, hyperexcitable state characterized by lowered spike threshold and spontaneous activity . Like Kᵥ₁, Kᵥ₃, and Kᵥ₄ channels, MAPK phosphorylation, inflammatory cytokines, and PKC activation modulate Kᵥ₇ channel expression (Yu et al., 2018[263]).

TRPV1, originally named vanilloid receptor 1 is a noxious heat-activated channel and commonly referred as the capsaicin receptor. It was initially described as a polymodal receptor that is activated by three pain-producing stimuli: vanilloid compounds (capsaicin, resiniferatoxin), at a temperature above approximately 43 °C and low pH <5.9 in chronic hyperglycemic conditions, metabolic stress and inflammatory mediators lower this thermal activation threshold (Du and Liu, 2025[59]). As a resultant Protein kinase A and protein kinase C-dependent phosphorylation sensitizes TRPV1, allowing it to open at physiological temperatures. As a result, normally non-painful warmth is recognized as burning pain. This mechanism explains the prominent thermal hyperalgesia and heat intolerance (Uchytilova et al., 2021[231]) observed during the early stages of DN. In pathological conditions TRVP1 decrease the expression of unmyelinated C-fibers which are primarily responsible for heat pain detection, while functional TRPV1 activity increases in medium-diameter A-fibers. This redistribution alters thermal sensitivity and contributes to abnormal temperature discrimination. Previous studies showed that impaired TRPV1 desensitization prolongs heat-evoked responses, enhancing thermal pain under repeated or sustained warm stimuli (Gao et al., 2024[81]).

As the disease progresses, TRPV1 undergoes fiber-specific redistribution. Its expression decreases in small unmyelinated C-fibers, which are primarily responsible for heat pain detection, while functional TRPV1 activity increases in medium-diameter A-fibers. This redistribution alters thermal encoding and contributes to abnormal temperature discrimination (Kim et al., 2008[137]). Experimental models further show that impaired TRPV1 desensitization prolongs heat-evoked responses, enhancing thermal pain under repeated or sustained warm stimuli (Luo et al., 2019[163]).

TRPV2, was discovered as a structural homologue of TRPV1 with 50 % amino acid identity, which is a very high-threshold heat sensor, activated at temperatures above ~52 °C and swelling, primarily expressed in neuronal and non-neuronal cells Aδ Aβ nociceptors fibers of DRG, trigeminal ganglia(TG) (Fricke and Leffler, 2024[79]). Its activation corresponds to extreme thermal stimuli rather than physiological warmth. Under inflammatory conditions, TRPV2 function is enhanced by growth factors, particularly Insulin-like Growth Factor-I (IGF-I). Activates the PI3-kinase signaling pathway, which triggers the rapid translocation of TRPV2 channels from intracellular to the plasma membrane, thereby sensitizing neurons to noxious stimuli (Kojima and Nagasawa, 2007[143]).

TRPV3 is a warm-sensitive channel, activated in the range of approximately 30-39 °C, with increased responses to higher noxious thermal stimuli, and expressed predominantly in epidermal keratinocytes rather than in sensory neurons, including TRG and TG. TRPV3 is also strongly activated and sensitized by camphor, irritants extracted from thyme, oregano, savory, and cloves (Lei and Tominaga, 2025[151]). In normal physiology of skin, TRPV3 in basal keratinocytes contributes to harmless warmth detection and epidermal homeostasis by mediating Ca²⁺ influx and triggering the release of paracrine mediators such as ATP, prostaglandins, and nitric oxide, which regulate proper warm sensation (Mandadi et al., 2009[168], Miyamoto et al., 2011[177]). In hyperglycemic conditions caused by phospholipase C and PKC activations, keratinocytes are significantly reduced. This down-regulation of keratinocyte TRPV3 contributes less to early painful hyperalgesia and more to progressive sensory loss, including reduced warm sensation and thermal hypoesthesia, by weakening keratinocyte nerve cross-talk (Facer et al., 2007[68], Geraldes and King, 2010[86]).

TRPV4 is a polymodal, non-selective cation channel belonging to the vanilloid subfamily of TRP channels. It is activated by mild hypotonic stress, shear stress, cell swelling, and innocuous warmth, typically above ~27 °C. In addition to physical stimuli, it can be sensitized by chemical mediators such as 4α-phorbol esters, low pH, citrate, nitric oxide, endocannabinoids, and arachidonic acid metabolites (Heller and O'Neil, 2007[108], Rodrigues et al., 2022[202]). The TRPV4 channel is widely expressed, including in DRG, cutaneous A- and C-fiber terminals, keratinocytes, and vascular endothelium. This distribution supports its dual role in sensory transduction and tissue homeostasis. Genetic deletion or knockdown studies show that TRPV4 contributes to mechanosensation and osmotic sensitivity, with TRPV4-deficient animals showing increased mechanical pain thresholds and altered responses to warm temperatures (Liedtke, 2007[155], Boudaka et al., 2020[27]).

In pathological conditions, TRPV4 plays a prominent role in inflammatory and neuropathic pain. It is strongly engaged in mechanical and osmotic hyperalgesia following exposure to inflammatory mediators such as prostaglandin E₂ and serotonin. Sensitization of TRPV4 occurs through intracellular signaling pathways involving PKA, PKC, and Src family kinases. Protease-activated receptor-2 activation during inflammation further enhances TRPV4 function, leading to increased release of neuropeptides such as substance P and calcitonin gene-related peptide (CGRP) from primary afferents and contribute to painful DN (Rodrigues et al., 2022[202]).

TRPVA1 is a TRP subfamily member; it is a protein that is overexpressed in a liposarcoma cell line and is distinguished by the presence of many ankyrin repeat motifs on the cytosolic amino-terminal domain (TRPAnkyrin). Because of its Drosophila homologue, it functions as a sensor for mechanical stimuli and contributes to mechanical nociception (Kwan et al., 2006[144]). It is expressed in inner ear, intestine myenteric plexus neurons, motor neurons, postganglionic sympathetic neurons, lung fibroblasts, and trigeminal and DRG neurons. Physical stimuli, such as extreme cold (less than 18 °C), as well as strong substances like mustard, garlic, wintergreen, clove, ginger, and cinnamon oils, activate TRPVA (Anand et al., 2008[9]). These all cause sharp, painful burning or tingling feelings. Peripheral activation of the TRPA1 channel depolarizes the nerve ending because of the influx of sodium ions, which can cause hyperexcitability and action potentials in pain-mediating nerve fibers (Koivisto et al., 2014[140]). The TRPA1 channel may be implicated in mechanical hyperalgesia and cold allodynia, according to a number of behavioral model studies (Iannone et al., 2023[118]). Reactive substances including reactive oxygen species (ROS) and inflammatory mediators like PKC, PKA, and bradykinin, which sensitize TRPA1 via PLC-dependent pathways, are among the additional stimuli that activate the TRPA1 channel (Aubdool et al., 2016[13]). Additionally, another factor is electrophilic activation. TRPA1 reacts with an electrophilic compound like Methylglyoxal, 4-hydroxynonenal, and nitrogen species bind with TRPA1 and open the Ca²⁺ and Na⁺ influx into the nociceptor the neuron becomes depolarized and hyperexcitable, leading to Spontaneous firing (ongoing burning pain). Lower threshold to touch and cold (mechanical and cold hyperalgesia), which ultimately causes DN (Eberhardt et al., 2012[63]).

Transient Receptor Potential Melastatin 8 (TRPM8) is a non-selective, calcium-permeable cation channel, the principal detector of cold sensation. It is activated by innocuous cooling (approximately 23-28 °C) and by cooling compounds such as menthol, eucalyptol, and icilin, with voltage-dependent gating properties (Izquierdo et al., 2021[122]). First discovered in the prostate gland as an androgen-responsive channel, it is a thermally regulated channel that is activated in-vitro by neurons derived from both TG and DRG. This is consistent with the percentage of cultured sensory neurons responding to cold and menthol (McKemy, 2007[174]). Its activity is modulated by intracellular calcium, pH, phosphatidylinositol-4,5-bisphosphate (PIP₂), and protein kinase C signaling (Yudin and Rohacs, 2012[264]). In normal physiology, TRPM8 is upregulated, acting like a built-in natural painkiller that activates whenever you experience cool sensations. In chronic hyperglycemia and inflammatory cytokines bind to protein Gαq and inhibits TRPM8 At the same time, inflammatory signaling activates an enzyme (calcineurin) response to cell surface become internalized as a resultant TRPM8 expression in the DRG decline and lose their natural cooling-based pain relief (Proudfoot et al., 2006[193], Zhang, 2019[275]). Without TRPM8, cool sensations no longer trigger the release of endogenous painkillers.

P2X3 is a ligand-gated ion channel of purinergic receptor that exists as homomeric P2X3 or heteromeric P2X2/3 complexes, which are expressed primarily in nociceptive neurons of the peripheral sensory neurons and are characteristically marked by rapid activation and rapid desensitization, especially in small to medium-diameter neurons of DRG, TG (Brederson and Jarvis, 2008[28]). During tissue stress, mechanical stimulation, or mild injury, extracellular ATP is released, which activates P2X3 receptors and induces Na⁺ and Ca²⁺ influx, leading to short‑lasting depolarization and a protective acute pain signal (Giniatullin and Nistri, 2023[88]). However, this ATP-mediated signaling is normally transient and tightly regulated. In persistent hyperglycemia, nerve injury, inflammation, mitochondrial dysfunction, and chronic metabolic stress, this transient signaling becomes dysregulated, resulting in elevated extracellular ATP levels and increased expression and sensitivity of P2X3 receptors, making sensory neurons hyperexcitable and abnormal spontaneous firing, which clinically manifests as burning pain, tingling, and sensory hypersensitivity (Xiang et al., 2008[253], Shcherbatko et al., 2016[209]). Thus, the P2X3 receptor is a central molecular mediator of ATP‑mediated peripheral pain transduction and, due to its predominant localization on peripheral sensory neurons, is considered an important therapeutic target in chronic painful conditions such as diabetic neuropathy (North, 2004[182]).

The P2X7 receptor is a member of the purinergic P2X family and functions as an ATP-gated ligand-gated ion channel, which differs from other P2X receptors because it requires high extracellular ATP concentration for activation (Martínez-Cuesta et al., 2020[170]). Structurally, the P2X7 receptor is homotrimeric and has a long intracellular C-terminal tail (239 amino acids), which plays an important role in its downstream signaling. Upon short-term ATP binding, the P2X7 channel allows Na⁺ and Ca²⁺ ions to enter the cell and K⁺ to exit, but when ATP exposure becomes prolonged, the receptor forms a large non-selective pore, which markedly increases membrane permeability and disturbs cellular homeostasis (Santana et al., 2024[206]). P2X7 receptor expression occurs predominantly on non-neuronal cells such as microglia, macrophages, astrocytes, and Schwann cells, whereas its expression on sensory neurons is limited, and therefore this receptor is more involved in neuroinflammation than in direct pain initiation. In pathological conditions such as nerve injury, chronic inflammation, or metabolic stress, extracellular ATP levels persistently increase, which continuously activates P2X7 receptors (Kaczmarek-Hajek et al., 2018[129], Hu et al., 2022[115]). This activation stimulates microglial cells and the NLRP3 inflammasome, thereby activating pro-inflammatory cytokines such as IL-1β and IL-18 are released (Wang et al., 2020[241]). In diabetic neuropathy, chronic hyperglycemia induces oxidative stress, mitochondrial dysfunction, and inflammation, which further increases ATP release and P2X7 receptor activation leading to increases the excitability of dorsal horn neurons, and develops central sensitization, which clinically manifests as persistent neuropathic pain (Chen et al., 2022[43]). Thus, the P2X7 receptor is a key mediator of ATP-mediated neuroinflammatory signaling and is considered an important therapeutic target in chronic neuropathic conditions such as painful diabetic neuropathy.

P2Y12 is a Gi-protein ionotropic purinergic receptor their primary ligand is ADP and mainly found more expressed in non-neuronal cells especially satellite glial cells of the DRG and microglia of the CNS than in neurons (Kawaguchi et al., 2015[135]). Under normal physiological conditions, P2Y12 plays a limited role in glial-neuronal communication and cellular homeostasis (Sipe et al., 2016[213]), but its role becomes quite prominent in pathological states such as chronic hyperglycemic condition and nerve injury (Guo et al., 2018[99]). During hyperglycemia and nerve stress, extracellular ATP/ADP is released from neurons, which activates the P2Y12 receptor, resulting in satellite glial cell activation, evidence of which is seen as an increase in GFAP expression. The activated P2Y12 receptor stimulates the downstream p38 MAPK signaling pathway, which increases the release of pro-inflammatory cytokines such as IL-1β and TNF-α. These inflammatory mediators enhance neuronal excitability and make sensory neurons hyper-responsive (Yi et al., 2018[260]), whose clinical manifestation appears in the form of mechanical and thermal hyperalgesia. In experimental diabetic rat models, P2Y12 receptor mRNA and protein expression have been found to be significantly increased in the DRG, and after gene silencing marked reductions have been observed in p38 MAPK activation, cytokine release, glial activation, and pain behaviors (Guo et al., 2018[99]). Thus, the P2Y12 receptor is considered not a primary trigger for pain initiation but rather a central molecular mediator in the maintenance and amplification of diabetic neuropathic pain, making it an important target for therapeutic intervention (Tozaki-Saitoh et al., 2008[229]).

In diabetic neuropathy, PIEZO 1, a mechanosensitive ion channel expressed in neurons, Schwann cells, endothelial cells, and immune cells, plays a crucial role at the junction between cellular signaling and mechanical stresses. Extensive metabolic and biomechanical stress, such as changed membrane tension, cytoskeletal remodelling, oxidative damage, and the accumulation of advanced glycation end products, are caused to chronic hyperglycemia (Gupta et al., 2025[102]). Peripheral nerve degeneration is ultimately triggered by these cascades, which also promote oxidative stress, mitochondrial dysfunction, synaptic impairment, and apoptosis. Increased PIEZO1 activity in dorsal root ganglion neurons increases neuronal excitability and membrane depolarization, both of which are characteristic of neuropathic pain (Yu et al., 2025[262]). This increased excitability amplifies nociceptive signaling and contributes to mechanical allodynia and hyperalgesia observed in DN (Lee et al., 2024[150]). Schwann cell function is also significantly impacted by PIEZO1 dysregulation. Aberrant mechanotransduction via PIEZO1 disrupts myelin maintenance and cytoskeletal architecture in diabetics. Persistent peripheral nerve damage is exacerbated by impaired Schwann cell activity, which alters axonal support, impairs nerve conduction, and restricts regenerative ability (Acheta et al., 2022[2]).

Long-term PIEZO1 overactivation also causes the endoplasmic reticulum calcium homeostasis to be disturbed, which leads to ER stress and the unfolded protein response, both of which accelerate neurodegenerative processes. Vascular dysfunction is another significant effect of PIEZO1 signaling disruption (Wang et al., 2016[244]). In response to shear stress, PIEZO1 normally triggers the PI3K-Akt-eNOS pathway in endothelial cells, encouraging the production of nitric oxide and preserving vascular homeostasis (Qu et al., 2023[194]). By restricting vasodilation, decreasing nitric oxide availability, and compromising endothelial signaling, hyperglycemia hinders this protective function. These alterations worsen neuropathic damage and cause ischemia by decreasing the microvascular perfusion of peripheral nerves. Additionally, PIEZO1 contributes to maintaining the integrity of the blood-nerve barrier; its dysregulation exacerbates neurovascular coupling in diabetes. PIEZO1, which regulates mechanosensitive inflammatory responses, is expressed by T cells, macrophages, and microglia (Zhang et al., 2024[276], Tabrizi et al., 2025[222]). Excessive PIEZO1 activation in diabetes induces pro-inflammatory mediators, including increased generation of reactive oxygen species and cytokines such as TNF-α, IL-6, and IL-1β. Both peripheral and central sensitization are maintained by these inflammatory mediators, which exacerbate neuronal damage. Chronic neuroinflammation and pain persistence are further reinforced in microglia by PIEZO1-driven activation of stress-related pathways, including JNK and mTOR. In general, PIEZO1 has two functions in diabetic neuropathy (Liu et al., 2021[157], Zhang et al., 2024[276], 2025[277]). While healthy PIEZO1 activity preserves neurovascular integrity and mechanosensory function, chronic hyperglycemia results in PIEZO1 maladaptive overactivation. This creates a vicious cycle where mechanical stress and metabolic damage worsen calcium overload, oxidative damage, inflammation, and vascular dysfunction. Experimental results demonstrating that PIEZO1 inhibition decreases neuronal damage and neuropathic pain highlight the importance of PIEZO1 as a possible therapeutic target in diabetic neuropathy (Velasco‐Estevez et al., 2020[235], Shin et al., 2023[211]).

PIEZO2 is a mechanically gated, non-selective cation channel that is genetically required for normal touch sensation and mechanotransduction in mammals. It is predominantly expressed in primary sensory neurons of the dorsal root ganglia, including low-threshold mechanoreceptors, proprioceptors, and subsets of nociceptors. PIEZO2 is also detected in peripheral afferent terminals, Schwann cells, satellite glial cells, and vascular endothelial cells, indicating its presence along the peripheral sensory pathway and neurovascular unit (Ranade et al., 2014[195], Shin et al., 2021[212]).

Under physiological conditions, PIEZO2 channels transduce mechanical forces such as pressure, stretch, and vibration into rapidly adapting inward currents mediated by Na⁺ and Ca²⁺ influx (Lacroix and Wijerathne, 2025[146]). Since most PIEZO2 channels are closed at normal negative resting membrane potentials and only become amenable for mechanical activation after depolarization, channel activity is strictly controlled by membrane voltage (Sánchez-Carranza et al., 2024[205]). By preventing excessive activation by harmless stimuli, this voltage-block mechanism keeps mechanical thresholds of nociceptor high. Membrane depolarization, inflammatory signaling, and modified intracellular second-messenger pathways are among the molecular and functional alterations that dorsal root ganglion neurons experience in neuropathic pain conditions, such as diabetic neuropathy (Garcia-Mesa et al., 2023[84], Fernández-Trillo et al., 2024[73]). Increased cAMP signaling leads to activation of Epac1, a cAMP-dependent exchange protein, which potentiates PIEZO2-mediated mechanotransduction. Increased cAMP signaling leads to activation of Epac1, a cAMP-dependent exchange protein, which potentiates PIEZO2-mediated mechanotransduction. Epac1 depends on cytoskeletal integrity and preferentially increases mechanically triggered PIEZO2 currents in sensory neurons without changing electrical excitability (Garcia-Mesa et al., 2023[84]). Mechanical allodynia and increased PIEZO2 activity are correlated with upregulated Epac1 expression in dorsal root ganglia during neuropathic pain. Relieving the voltage block of PIEZO2 reduces mechanical activation thresholds and promotes continuous activity in nociceptors, according to experimental research (Eijkelkamp et al., 2013[65]). Mechanosensitive currents in Aδ- and C-fiber nociceptors are increased by gain-of-function changes in PIEZO2, leading to a marked hypersensitivity to mechanical stimuli. These results suggest that nociceptor sensitization can be triggered by membrane depolarization brought on by noxious or sensitizing circumstances, which increases the availability of PIEZO2 channels (Sánchez-Carranza et al., 2024[205]). In diabetic distal symmetric polyneuropathy, PIEZO2 expression is increased in cutaneous microvessels, particularly in patients with painful neuropathy. Structural disorganization, endothelial dysfunction, and impaired vasodilation are associated with increased vascular PIEZO2 immunoreactivity. These vascular alterations are proposed to contribute to impaired blood flow and pain severity, linking mechanotransduction dysfunction to microvascular pathology in diabetic neuropathy (Garcia-Mesa et al., 2023[84]). Furthermore, Schwann cells and peripheral glial components express PIEZO2, and several inflammatory and neuropathic pain models have been shown to exhibit PIEZO2 overexpression (Wan et al., 2024[239]). While increasing PIEZO2 activity has been correlated with sensitivity to typically harmless mechanical stimuli, loss of function or knockdown of PIEZO2 lowers mechanical allodynia (Nencini et al., 2021[181]). All of these results point to PIEZO2's involvement in aberrant mechanotransduction, nociceptor sensitization, and neurovascular dysfunction in the formation and maintenance of mechanical pain in neuropathic dysfunction.