Abstract

Mesencephalic trigeminal neurons (MTNs) are the sole primary afferent neurons with cell bodies located within the central nervous system. MTNs convey proprioceptive inputs from masticatory muscles and periodontal ligaments, thereby contributing to the precise regulation of jaw–oral motor functions. Through ionic mechanisms such as currents generated by the voltage-dependent sodium (Nav) channel isoform Nav1.6, hyperpolarization-activated currents, and persistent inward currents, MTNs generate sustained and burst firing that regulate masticatory rhythm and jaw-jerk reflex timing. Their activity is further modulated by neurotransmitters, including serotonin and norepinephrine, which provide flexibility in sensorimotor integration. Pathological conditions such as chronic stress and sodium channel dysfunction induce MTN hyperexcitability or irregular firing, contributing to bruxism, temporomandibular disorders, and feeding impairment in amyotrophic lateral sclerosis models. In addition, aging and tooth loss lead to Piezo2 downregulation and neuronal death, potentially resulting in masticatory dysfunction and cognitive decline. Recent findings suggest that interventions targeting vesicular glutamate transporter 1 projections, melanocortin 4 receptor signaling, and nitric oxide pathways represent novel therapeutic approaches. Taken together, MTNs have emerged as promising targets for treating conditions ranging from masticatory motor disorders to neurodegenerative diseases.

Graphical abstract

1 Introduction

Orofacial movements such as speaking, eating, and expressing emotions are executed unconsciously under remarkably precise control. At the foundation of this fine motor regulation lies proprioception, the sensory system that provides continuous feedback regarding the position, movement, and forces acting on the body. In mastication, during which millisecond-level precision in force adjustment is required, proprioceptive input is indispensable. Mesencephalic trigeminal neurons (MTNs) represent the central nodes of orofacial proprioception (Lazarov, 2002). MTNs are distinguished by their exceptional anatomical configuration, which underscores their role as “guardians” in maintaining orofacial function.

The most striking feature of MTNs is their highly unusual localization: unlike most first-order sensory neurons, whose cell bodies reside in peripheral sensory ganglia (e.g., the trigeminal ganglion), MTNs are located within the midbrain of the central nervous system (CNS; Ramón y Cajal, 1909). This exceptional arrangement suggests that MTNs serve as both conduits of sensory information and active participants in higher-order information processing and rapid reflex control. Their peripheral processes terminate in mechanoreceptors within the muscle spindles of jaw-closing muscles and in the periodontal ligament, which anchors the teeth to alveolar bone (Shigenaga et al., 1990; Linden et al., 1994). Whereas muscle spindles monitor muscle stretch, periodontal ligament receptors detect pressure and vibration applied to the teeth. MTNs integrate these two distinct sensory streams in real time, providing precise information on the state of the masticatory apparatus and occlusal forces (Kato et al., 1999).

This unique capacity for high-fidelity information gathering underpins the concept of MTNs as “guardians” of oral function. For example, when chewing hard food, humans can crush it efficiently without damaging their teeth or temporomandibular joint because MTNs rapidly feedback occlusal force signals from the periodontal ligament to the trigeminal motor nucleus, optimizing the contractile force of jaw-closing muscles (Morimoto et al., 1985, 1989). Excessive force elicits inhibitory input from MTNs to jaw-closing muscles, whereas insufficient force enhances contraction. Through this finely tuned feedback loop, teeth, periodontal tissues, and temporomandibular joints are protected from mechanical overload. MTNs also receive input from jaw-closing muscle spindles and serve as the afferent limb of the monosynaptic jaw-jerk reflex (Szentágothai, 1948). This rapid defensive reflex contracts jaw-closing muscles within milliseconds when the mandible is unexpectedly depressed, maintaining joint stability. The central role of MTNs in this reflex arc provides compelling evidence of their indispensable contribution to orofacial homeostasis.

Pathological consequences can arise when this protective function is compromised. Abnormally strong masticatory muscle activity observed in sleep bruxism has been linked to dysfunction of proprioceptive feedback mediated by MTNs (Giovanni and Giorgia, 2021). Similarly, in atypical odontalgia and certain forms of trigeminal neuralgia, aberrant processing of proprioceptive input has been hypothesized to contribute to altered pain perception, potentially underpinned by abnormal electrophysiological properties of MTNs (Sessle, 2006).

Taken together, MTNs, through their unique anatomical localization and functional connectivity, enable fine control of mastication, provide reflexive protection of orofacial structures, and contribute to the oral representation of “self-awareness.” These neurons are active guardians of orofacial integrity. Several review articles on MTNs have been published to date (Table 1). This review focuses on the electrophysiological properties of MTNs, highlighting the mechanisms underlying their precise functional roles and exploring the contributions of dysfunction to several orofacial disorders.

Author(s), yearJournalMain focusKey contributionsLimitations/gapsShigenaga et al. (1988)Brain Research ReviewsMorphology and connectivityDetailed anatomical mapping of MTN projectionsLimited to cat morphology; small sample, little functional or developmental dataSakata and Yoshimatsu (1995)Methods and Findings in Experimental and Clinical PharmacologyRole of hypothalamic and Vmes histamine neurons in energy balance and feedingFound that histamine suppresses feeding, regulates mastication (Vmes), controls meal size (VMH), and links energy deficit to astrocytic glycogenolysis; deficits explain obesity in Zucker ratsMostly animal models (rats); limited direct human evidenceLazarov (2000)Advances in Anatomy, Embryology and Cell BiologyNeuroanatomy and neurochemistry of cat MTNIdentified Glu as primary transmitter; presence of small GABAergic neurons, NOS, and peptidergic inputs; evidence of plastic changes after axotomySpecies specificity unclear; limited functional validation; many findings based on abnormal conditionsLazarov (2002)Progress in NeurobiologyComparison of neurochemical content of TG vs. MTNIdentified distinct neurotransmitter and peptide profiles in TG and MTN neurons; observed Glu, PV, and CB dominance in MTN vs. diverse neuropeptides in TG; highlighted unique perineuronal basket-like arborizationsLacks functional analysis of how these neurochemical differences influence sensory processing; based mostly on immunohistochemistry without physiological correlationSakata et al. (2003)Experimental Biology and MedicineRole of MTN histamine (HA) neurons in mastication and energy regulationFound that HA neurons in Vmes are activated at early feeding phase; the neurons regulate eating speed and link mastication to hypothalamic centers controlling intake and metabolismStudy limited to rats; mechanisms in humans remain unclear; exact molecular pathways beyond HA remain unresolvedKolta et al. (2007)Archives of Oral BiologyBurst generation in trigeminal neuronsRevealed these neurons generate rhythmic bursts independent of synaptic input; maturation coincides with onset of mastication; low extracellular Ca2+ helps initiate chewingStudy limited to rats; developmental stage only; no direct behavioral or human validationLazarov (2007)Brain Research ReviewsNeurochemistry of MTN in orofacial proprioceptionSummarizes neurotransmitters, neuropeptides, receptors in MTN; highlights role in masticatory proprioception; provides evidence for neuroplasticity depending on environmentMostly descriptive review; limited functional/physiological data; lacks direct experimental validation of proposed mechanismsXing et al. (2014)NeurosignalsElectrophysiological featuresSystematic overview of MTN firing propertiesLimited molecular/pathophysiological sequencingYoshida et al. (2017)Journal of Oral ScienceProjection and synaptic organization of single Vmes afferentsCompared spindle and periodontal afferents; revealed distinct projection and synaptic patterns in jaw-closing and supratrigeminal nucleiLack of detailed prior reports; limited to single-afferent levelTrigo et al. (2025)NeuroscienceRole of electrical coupling and Ih current in coincidence detection in MesV neuronsIh makes coupling time-dependent, sharpens coincidence detection of inhibitory inputsModel-based, limited in vivo evidence

Previous review articles on MTNs (Vmes/MTN).

2 Anatomical organization2.1 Location, connections, and projections of the mesencephalic trigeminal nucleus (Vmes)

The Vmes contains the cell bodies of unique primary sensory neurons mediating orofacial proprioception. Unlike conventional sensory neurons located in peripheral ganglia such as the trigeminal ganglion (TG), the cell bodies of MTNs reside within the brainstem, a central localization that reflects their specialized function (Lazarov, 2002). The Vmes forms a longitudinal column extending from the level of the superior colliculus, sometimes reaching the pretectal area and interstitial nucleus of Cajal, caudally surrounding the cerebral aqueduct and fourth ventricle through the inferior colliculus and locus coeruleus. The Vmes extends down to the pontine level adjacent to the principal sensory trigeminal nucleus and dorsomedial trigeminal motor nucleus (Vmo; Huff et al., 2025). This elongated organization corresponds to the developmental origin of MTNs from the rhombic lip rather than the neural crest (Shirasaki and Pfaff, 2002). Functionally, neurons innervating jaw-closing muscles (masseter, temporalis) occupy rostral and central portions, whereas those projecting to jaw-opening muscles and the periodontal ligament are located caudally (Luo and Dessem, 1996) (Figure 1).

Anatomical localization and projection targets of the mesencephalic trigeminal nucleus. Schematic representation of the anatomical localization of Vmes and its major peripheral and central connections. Neurons within the Vmes receive proprioceptive input from muscle spindles of jaw-closing muscles and mechanoreceptors in the periodontal ligament. Central projections from the Vmes form monosynaptic connections with Vmo, constituting the afferent limb of the jaw-jerk reflex and supporting precise control of masticatory movements.

Mesencephalic trigeminal neurons are pseudounipolar neurons whose single process bifurcates into peripheral and central branches. The peripheral branch joins the trigeminal motor root (mandibular nerve) and terminates in two major proprioceptor types: muscle spindles in jaw-closing muscles, sensing stretch (Kato et al., 1999), and mechanoreceptors in the periodontal ligament, detecting tooth pressure (Matsushita et al., 1982). Through these proprioceptors, the Vmes provides continuous information on jaw movement and occlusal force.

The central branches of MTNs project widely within the brainstem. The most prominent branch is the direct excitatory projection to the Vmo, forming the monosynaptic jaw-jerk reflex arc that elicits jaw-closing muscle contraction upon stretch (Appenteng et al., 1978; Chandler, 1989). Additional collaterals target orofacial motor and autonomic nuclei, including the trigeminal sensory nuclei, reticular formation, and superior salivatory and hypoglossal nuclei (Lazarov, 2002). In rats, jaw muscle spindle afferents can drive multiple orofacial motoneuron pools via common premotor neurons (Zhang et al., 2012), indicating that Vmes input coordinates complex multi-muscle movements such as jaw–tongue–facial interactions. Some projections also reach the cerebellum (especially the anterior lobe), contributing to motor coordination and learning, and possibly extend polysynaptically to the thalamus for proprioceptive awareness (Lazarov, 2000).

Through this unique central localization and extensive projection network, the Vmes serves as a crucial hub integrating proprioceptive input for reflexive and coordinated orofacial motor control.

2.2 Developmental origins and molecular markers

Although migratory neural crest cells give rise to nearly all primary sensory neurons located in peripheral ganglia, MTNs arise directly from the neuroepithelium of the CNS. This unique developmental trajectory is intrinsically linked to a distinctive combination of molecular markers that govern their fate, survival, and eventual specialization in proprioceptive function.

According to the classical paradigm of neurodevelopment, the central nervous system originates from the neural tube, whereas the peripheral nervous system primarily derives from the neural crest, reflecting distinct developmental lineages. Based on this framework, early studies often assumed MTNs, like TG neurons, were neural crest-derived (D'Amico-Martel and Noden, 1983). However, subsequent evidence from avian chimera experiments and lineage-tracing studies has clearly refuted this assumption, demonstrating that MTNs arise through a distinct ontogenetic pathway.

Current consensus indicates that the majority of MTNs originate from the dorsal neuroepithelium at the midbrain–hindbrain junction, encompassing the isthmic organizer region and the most rostral rhombomere (Hunter et al., 2001). This dorsal neuroepithelial domain includes the rhombic lip, a secondary germinal zone known to generate multiple neuronal populations, including cerebellar granule cells.

Lineage-tracing studies using Cre recombinase systems driven by promoters active within this neuroepithelial territory, such as Wnt1 and Engrailed-1, have provided compelling evidence that progenitor cells expressing these transcription factors migrate from the ventricular zone and differentiate into MTNs (Danielian et al., 1998; Zervas et al., 2004). The shared developmental origin of MTNs and certain cerebellar neurons, under the influence of potent signaling molecules such as Wnt1 and Fgf8 secreted from the isthmic organizer, suggests the presence of a common developmental program underlying neuronal populations involved in sensorimotor integration.

Notably, neurons derived from this isthmic organizer–associated neuroepithelium, including cerebellar granule cells, share electrophysiological properties optimized for high temporal precision, such as reliable high-frequency firing and subthreshold resonance. Consistent with this, MTNs exhibit specialized intrinsic membrane properties that support precise timing of proprioceptive signaling, reinforcing the functional relevance of their shared developmental origin.

The specification of progenitors into mature MTNs is tightly regulated by the spatiotemporal expression of transcription factors. This process likely begins with the expression of the paired-box transcription factors Pax3 and Pax7 in the dorsal neural tube (Baker et al., 1999). Once these cells exit their final mitosis and commit to the MTN lineage, they begin to express a distinctive set of transcription factors that promote their differentiation from other rhombic lip-derived neurons. The most critical of these factors are the paired-like homeodomain transcription factors Phox2a and Phox2b, known master regulators of visceral sensory and motor neuron development that are believed to contribute to MTN identity and survival (Pattyn et al., 2000). At the same time, MTNs express the LIM homeodomain protein Islet-1 and the POU domain transcription factor Brn3a (also known as Pou4f1), both broadly involved in sensory neuron differentiation and axon guidance (Fedtsova and Turner, 1995).

The establishment of MTNs as proprioceptive neurons is marked by the expression of Runt-related transcription factor 3. Runt-related transcription factor 3 defines proprioceptive neurons across the nervous system, including dorsal root ganglion (DRG) neurons, and its expression in MTNs highlights a molecular homology with peripheral proprioceptive neurons despite their distinct locations and origins (Inoue et al., 2002). As neurons mature, their survival and circuit integration increasingly depend on trophic support. Although many DRG proprioceptive neurons rely on neurotrophin-3 acting through TrkC receptors, MTNs express TrkB, implicating BDNF–TrkB signaling in their maintenance and function (Tang et al., 2010). This divergence in neurotrophic dependence underscores a fundamental distinction between central and peripheral proprioceptive systems.

2.3 Piezo2 as a molecular basis of mechanotransduction in MTNs

Mechanotransduction is the process by which mechanical stimuli are converted into electrical signals in sensory neurons. Piezo2, a mechanically activated cation channel, has been identified as the principal molecular mediator of mechanotransduction in mammalian proprioceptive neurons. Genetic ablation of Piezo2 in mice abolishes mechanically evoked receptor potentials and action potential firing in muscle spindle afferents, resulting in profound deficits in proprioceptive signaling and severe motor incoordination. These findings establish Piezo2 as an essential mechanotransducer underlying peripheral proprioception (Woo et al., 2015).

Mesencephalic trigeminal neurons exhibit functional and molecular characteristics closely resembling those of peripheral proprioceptive neurons, despite their unique localization within the central nervous system (Lazarov, 2002). MTNs innervate muscle spindles of jaw-closing muscles and mechanoreceptors in the periodontal ligament, thereby encoding muscle stretch and tooth load information critical for the precise control of mastication (Linden et al., 1994). Consistent with their proprioceptive identity, MTNs express Piezo2, providing a molecular substrate for the conversion of mechanical deformation into receptor potentials at their peripheral terminals (Florez-Paz et al., 2016).

Piezo2-mediated mechanotransduction is well suited to the physiological demands placed on MTNs. The fast activation kinetics and low mechanical threshold of Piezo2 channels enable rapid and faithful detection of subtle changes in muscle tension and occlusal force, analogous to the requirements of limb proprioceptive systems. Mechanical inputs transduced via Piezo2 are conveyed centrally to brainstem circuits, including direct projections to the trigeminal motor nucleus, thereby forming the afferent limb of the jaw-jerk reflex and contributing to fine-tuned regulation of bite force and masticatory rhythm (Linden et al., 1994; Lazarov, 2002; Woo et al., 2015; Florez-Paz et al., 2016).

Collectively, Piezo2 endows MTNs with high-fidelity mechanosensory capability, linking peripheral mechanical events in muscle spindles and periodontal ligament receptors to central sensorimotor integration. Through this mechanism, Piezo2 serves as a key molecular gatekeeper supporting the precise timing and force control required for normal mastication (Linden et al., 1994; Lazarov, 2002; Woo et al., 2015; Florez-Paz et al., 2016).

2.4 Morphological and neurochemical diversity within the Vmes population

MTNs are characterized by an unusually high degree of electrical coupling. Their morphology resembles, yet diverges from, that of peripheral sensory ganglia (Shigenaga et al., 1988; Ter-Avetisyan et al., 2018). Intracellular horseradish peroxidase tracing in cats revealed the distinctive “Y-shaped” bifurcation of Vmes axons, consisting of a peripheral branch projecting to cranial nerves and a central branch innervating brainstem motor and reticular nuclei (Shigenaga et al., 1990). These central branches exhibit axon collaterals terminating in the dorsal Vmo or trigeminal-adjacent structures, with collateral patterns correlating to group I (muscle spindle) or group II (periodontal) sensory phenotypes (Shigenaga et al., 1990). Developmental studies further indicated that these morphological features are established early. In mice, molecular pathways such as CNP–Npr2–cGKI signaling are essential for proper axonal bifurcation at the hindbrain level, highlighting canonical growth patterns that diversify morphology within the Vmes pool (Ter-Avetisyan et al., 2018).

Beyond morphology, the Vmes displays a rich neurochemical repertoire. In primates, ultrastructural studies combining immunoelectron microscopy demonstrated that although the somata are sparsely innervated, they receive diverse synaptic inputs, including excitatory and modulatory signals (Wang and May, 2023). In rodents, immunohistochemical surveys have revealed the presence of multiple neurotransmitters, including glutamate (via VGLUT1/2), GABA, serotonin (5-HT), and neuropeptides such as dynorphin A and orexin. MTNs also exhibit ATP-gated P2X receptor currents, implicating purinergic input in excitatory control (Khakh et al., 1997). Moreover, during early postnatal development, trigeminal neurons display differential expression of the NMDA receptor subunits NR2A and NR2B, with a developmental shift from NR2B to NR2A dominance that dictates excitatory postsynaptic current kinetics and synaptic plasticity rules (Turman et al., 1999; Turman et al., 2002; Turman et al., 2000).

Electrophysiologically, this molecular heterogeneity translates into functional diversity. Recordings in the cat Vmes uncovered differences between somatodendritic spikes and axonal action potentials, as well as variable responses to mechanical stretch (Shigenaga et al., 1990). Recent single-cell transcriptomic analyses in mice revealed at least five transcriptionally distinct Vmes subtypes, each expressing different combinations of ion channels, receptors, and mechanosensory proteins. These subtypes likely underlie group-specific responses to muscle versus ligament stretch (Lee et al., 2023).

Thus, the morphological and neurochemical diversity of the Vmes reflects its dual roles as both a primary sensor and integral component of premotor circuits. Its large bifurcating axons enable efficient transmission of proprioceptive signals to jaw motor nuclei while supporting local electrical propagation. Simultaneously, the broad range of synaptic inputs—from glutamatergic to peptidergic—provides dynamic modulation under different behavioral and stress conditions. The diversity within the Vmes neuronal population therefore establishes a neuroanatomical framework that supports both reflexive jaw control and adaptive plasticity.

3 The electrophysiological repertoire of MTNs3.1 Intrinsic properties: components of excitability

Mesencephalic trigeminal neurons are unique primary sensory neurons within the mammalian CNS, as their cell bodies are located inside the CNS and they transmit proprioceptive information from the muscle spindles of the masticatory muscles and the periodontal ligament (Yoshida et al., 2017). The electrophysiological excitability of these neurons is largely dependent on their intrinsic properties, particularly the expression patterns and distribution of Na+ and K+ channels, which play a crucial role in determining firing modes. MTNs exhibit intrinsically generated, depolarization-induced high-frequency spike discharges that are repeatedly and intermittently evoked during a depolarizing current pulse, rather than occurring spontaneously (Wu et al., 2001). This firing behavior is supported by membrane potential resonance arising from the interaction between persistent Na+ currents and 4-AP-sensitive non-inactivating (D-type; I_D) K+ currents, with additional contributions from other K+ conductances shaping excitability (Wu et al., 2001; Wu N. et al., 2005; Wu W. W. et al., 2005; Enomoto et al., 2006). Notably, glutamatergic synaptic activation (including mGluR-I–dependent mechanisms) can further enhance resonance-dependent oscillations and promote a switch from single spiking to bursting in MTNs (Chung et al., 2015). Furthermore, researchers demonstrated that these neurons generate persistent inward currents (PICs), which enable them to sustain firing over extended periods (Enomoto et al., 2006). These intrinsic properties are also known to change plastically during development or under pathological conditions, suggesting that MTN dysfunction can contribute to abnormal mastication or altered nociception (Turman et al., 1999). Collectively, these findings underscore the importance of intrinsic excitability in supporting the sensorimotor integration functions of MTNs.

Mesencephalic trigeminal neurons also exhibit different firing patterns than other sensory neurons. These neurons process proprioceptive input from muscle spindles and display both tonic firing and rhythmic bursting. According to prior research, MTNs can sustain stable repetitive action potential discharge, which might vary across developmental stages (Yoshida et al., 2017). Shigenaga et al. (1988), further demonstrated that during rhythmic mastication, MTNs burst in synchrony with jaw motor neurons, suggesting functional coupling with central pattern generators. These diverse firing patterns indicate that MTNs are endowed with adaptive response properties required for fine control of both static and dynamic muscle tension.

In addition to spontaneous activity, MTNs sometimes display subthreshold membrane potential oscillations, characterized by small, periodic fluctuations within a given voltage range that can lead to action potential generation under appropriate conditions. This phenomenon, observed in vitro by Pedroarena and Llinás (1997), was identified as an important electrical signature suggestive of pacemaker-like properties. Specifically, low-threshold Ca2+ currents (T-type Ca2+ currents) and persistent Na+ currents help stabilize these oscillations and form the basis for rhythmic activity (Enomoto et al., 2006). Moreover, the presence of hyperpolarization-activated (Ih) currents in MTNs, contributing to phase and frequency alignment of subthreshold oscillations via hyperpolarization sag and rebound depolarization, has long been recognized (Khakh and Henderson, 1998). This suggests that MTNs participate in rhythm generation, thereby contributing to the timing of cyclic movements such as mastication. Furthermore, MTNs receive synaptic inputs that evoke firing, reset the phase, and modulate the frequency and amplitude of intrinsic oscillations (Verdier et al., 2004). Thus, pacemaker-like activity in the Vmes functions as a “semi-autonomous oscillator” entrainable by network inputs and tightly linked to synaptic integration (Figure 2).

Electrophysiological properties of mesencephalic trigeminal neurons. Representative electrophysiological properties of mesencephalic trigeminal neurons recorded by slice patch clamp. Wild-type neurons exhibit spontaneous burst firing, subthreshold membrane potential oscillations, voltage sag responses, and membrane resonance revealed by ZAP stimulation. Voltage-clamp recordings demonstrate transient, persistent, and resurgent sodium currents. In Nav1.6 knockout mice, burst firing and subthreshold resonance are absent, and resurgent sodium currents are markedly reduced, indicating a critical role of Nav1.6 in rhythmic firing.

3.2 Ionic basis of excitability: major channels and currents

The excitability of MTNs is maintained through the coordinated action of multiple ion channels regulating membrane potential. The most critical are voltage-dependent sodium (Nav) channels, which generate action potentials with rapid upstrokes. Yoshida demonstrated that Vmes neurons regulate excitability through Na+ and K+ currents as well as Ca2+ currents (Yoshida et al., 2017). Furthermore, Del Negro and Chandler revealed that PICs are central to sustaining excitability in the Vmes, with Na+-PICs and Ca2+-PICs forming the basis for rhythmic firing (Enomoto et al., 2006). Inhibitory channels such as inwardly rectifying K+ channels and delayed rectifier K+ channels are also essential for setting firing thresholds and recovery dynamics, thereby refining motor control and preventing malfunction. Additionally, Yamada and colleagues, using neonatal rats, demonstrated that zinc deficiency alters MTN membrane properties and spike discharge patterns. Specifically, zinc-deficient neurons exhibited moderate membrane potential depolarization, reduced cell capacitance, increased afterhyperpolarization, shortened burst duration, and accelerated spike frequency adaptation. These effects were likely attributable to enhanced Ca2+-dependent K+ conductance, suggesting that the nutritional status contributes to the plasticity of firing modes (Yamada et al., 2021).

In MTNs, the interplay among voltage-dependent Na+, K+, and Ca2+ channels is crucial for action potential generation and firing pattern regulation. Notably, Nav1.6 channels mediate transient, persistent (INaP), and resurgent sodium (INaR) currents, making major contributions to firing frequency and membrane resonance properties, as demonstrated in knockout mouse studies (Enomoto et al., 2006; Seki et al., 2019). Deletion of Nav1.6 resulted in an 18% reduction in transient Na+ currents, 39% reduction in INaP currents, and 76% reduction in INaR currents, leading to marked changes in membrane resonance (Enomoto et al., 2006; Westenbroek et al., 1989). Enomoto and colleagues further demonstrated that developmental increases in INaP and INaR currents are critical for supporting high-frequency firing, providing a basis for age-dependent changes in MTN excitability (Enomoto et al., 2018; Venugopal et al., 2019).

Voltage-dependent calcium channels, particularly T-type channels, are also implicated in subthreshold oscillations and burst firing. K+ channels, including delayed rectifier K+ channels and inwardly rectifying K+ channels, contribute to repolarization and recovery, regulating firing timing and patterns (Tanaka et al., 2003). In neonatal rat MTNs, delayed rectifier and A-type K+ currents were found via both physiological recordings and computational models to primarily regulate the onset, frequency, and adaptation of repetitive firing (Del Negro and Chandler, 1997). In this context, Tanaka and colleagues reported functional maturation of inwardly rectifying K+ channels and Ih currents during development, with increased Ih currents providing low-frequency resonance and firing stability (Tanaka et al., 2003). Together, these channels establish the ionic basis that supports stable excitability and rhythmic activity in the Vmes.

The pacemaker function of MTN is sustained by INaP currents and Ih currents mediated by HCN channels (Khakh and Henderson, 1998). In rat MTNs, Ih currents have been physiologically identified as Cs+-sensitive currents activated by hyperpolarization, providing the ionic substrate for sag responses and rebound firing (Khakh and Henderson, 1998). Prior research indicated that INaP currents contribute to burst firing and resonance by facilitating subthreshold oscillations, thereby enabling rhythmic activity (Wu et al., 2005; Enomoto et al., 2006). Ih currents, mediated by HCN channels (particularly HCN1 and HCN2), are activated at subthreshold voltages, and they contribute to membrane stabilization and spike timing regulation (Wu et al., 2005). Kang et al. showed that Ih and Na+–K+ pump currents in MTNs interact bidirectionally via local Na+ microdomains: Na+ influx through h-channels activates pump currents, while pump activity modulates Ih. Colocalization of HCN1/2 channels and the Na+–K+ pumpalpha3 isoform supports this reciprocal regulation of neuronal excitability (Kang et al., 2004). Because this Na+ microdomain is also shared with AMPA receptors, it was concluded that I-AMPA evoked in MTN neurons is opposed by a transient decrease in standing inward Ih due to negative shifts in the reversal potential of Ih as a consequence of elevated Na+ concentrations within the Na+ microdomain in response to Na+ influx through AMPA channels (Kawasaki et al., 2018). Accordingly, suppression of HCN conductance-for example by Cs+ application or noradrenergic inputs from the locus coeruleus-enhances AMPA-evoked depolarization and can trigger a barrage of spikes in MTNs (Kawasaki et al., 2018; Toyoda et al., 2022). This mechanism is distinct from the previously postulated mechanisms for the inhibition of EPSPs by the activity of HCN channels (Magee, 1998; Carr et al., 2007). Tanaka and colleagues further reported that 5-HT suppresses Ih currents, particularly in late developmental stages, suggesting developmental stage-dependent serotonergic modulation of MTN frequency selectivity (Tanaka et al., 2019). This dynamic interaction tunes sag responses, rebound spikes, resonance frequency, and spike precision, thereby stabilizing tonic firing and enhancing the robustness of rhythmogenesis under sustained input (Tanaka et al., 2019). Pharmacological manipulation of HCN activity or Na+–K+ pump activity shifts these parameters in predictable ways, supporting the causal role of this bidirectional interaction. Tanaka and colleagues also reported that orexin A enhances INaP currents, increasing firing frequency and resonance, thereby suggesting a link among feeding behavior, energy metabolism, and orofacial motor control (Tanaka et al., 2021) (Figure 2).

3.3 Synaptic integration and neuromodulation

Mesencephalic trigeminal neurons, owing to their unique structure and function, are subject to both sensory input reception and integration and regulation by a wide range of neurotransmitters. Copray and coworkers immunohistochemically demonstrated the expression of receptors for 5-HT, norepinephrine (NE), dopamine, and GABA in MTNs (Copray et al., 1990). These neuromodulators can alter resting membrane potential, firing threshold, and postsynaptic potentials, thereby flexibly adjusting neuronal excitability. Synaptic integration and neuromodulation by extrinsic inputs modify MTN activity patterns and timing, enabling dynamic regulation of cyclic movements and reflexes. Purinergic modulation by ATP also plays a critical role. In rat MTNs, “native” P2X receptors mediate fast inward currents evoked by ATP, but their pharmacological and kinetic profiles do not correspond to any of the known P2X subtypes (P2X1–P2X7) characterized in heterologous systems (Khakh et al., 1997; Patel et al., 2001). Moreover, MTN afferents converge and diverge within the premotor layer, distributing in parallel to multiple motor pools via “common premotor neurons.” Thus, synaptic integration both regulates gain within a single motor nucleus and organizes coordination across muscle groups (Zhang et al., 2012). This “mismatch” suggests that P2X receptors functioning in MTNs possess unique subunit compositions or accessory factor dependencies, indicating the existence of MTN-specific purinergic regulatory mechanisms.

Autapses, self-synapses in which a neuron provides input to itself, have also been hypothesized in MTNs. Using computational modeling, Guo and colleagues demonstrated that autaptic self-regulation can contribute to firing stability and timing precision (Guo et al., 2016). If MTNs employ such a self-regulatory mechanism, it might function as an intrinsic corrective system for fine-tuning masticatory rhythms and sensorimotor integration.

MTNs are strongly influenced by extrinsic neuromodulators, particularly 5-HT and NE. 5-HT depolarizes MTNs and lowers their firing threshold, thereby increasing excitability (Hsiao et al., 1997; Tanaka and Chandler, 2006). Tanaka and colleagues also demonstrated that 5-HT suppresses Ih currents and alters low-frequency resonance, indicating that serotonergic modulation developmentally shifts frequency selectivity (Tanaka et al., 2019). Noradrenergic inputs from the locus coeruleus suppress Ih conductance in MTNs via activation of α2-adrenergic receptors, likely through volume transmission mechanisms (Toyoda et al., 2022). This suppression of Ih alters intrinsic excitability and can facilitate depolarization-induced spike barrages in response