Mesencephalic Trigeminal Nucleus Neurons Research Articles

The ultrastructure of macaque mesencephalic trigeminal nucleus neurons.

Primary afferents originating from the mesencephalic trigeminal nucleus provide the main source of proprioceptive information guiding mastication, and thus represent an important component of this critical function. Unlike those of other primary afferents, their cell bodies lie within the central nervous system. It is believed that this unusual central location allows them to be regulated by synaptic input. In this study, we explored the ultrastructure of macaque mesencephalic trigeminal nucleus neurons to determine the presence and nature of this synaptic input in a primate. We first confirmed the location of macaque mesencephalic trigeminal neurons by retrograde labeling from the masticatory muscles. Since the labeled neurons were by far the largest cells located at the edge of the periaqueductal gray, we could undertake sampling for electron microscopy based on soma size. Ultrastructurally, mesencephalic trigeminal neurons had very large somata with euchromatic nuclei that sometimes displayed deeply indented nuclear membranes. Terminal profiles with varied vesicle characteristics and synaptic density thicknesses were found in contact with either their somatic plasma membranes or somatic spines. However, in contradistinction to other, much smaller, somata in the region, the plasma membranes of the mesencephalic trigeminal somata had only a few synaptic contacts. They did extend numerous somatic spines of various lengths into the neuropil, but most of these also lacked synaptic contact. The observed ultrastructural organization indicates that macaque trigeminal mesencephalic neurons do receive synaptic contacts, but despite their central location, they only avail themselves of very limited input.

Mesencephalic trigeminal nucleus neurons with collaterals to both eyelid and masseter muscles shown by fluorescent double-labeling, revealing a potential mechanism for Marcus Gunn Syndrome.

Poking palpebral conjunctiva evoked upper-eyelid retraction during ophthalmic surgery. Iatrogenic eyelid ptosis occurred if eyelid branch of lachrymal nerve was sectioned. Mesencephalic trigeminal nucleus (Vme) neurons were labeled when tracer injected into lachrymal nerve innervating eyelid Mueller's muscle. Masseter afferent Vme neurons projecting to oculomotor nucleus (III) was observed in toad and rat, which helps amphibians to stare prey when they open mouth widely to prey. We hypothesized single Vme neurons may have peripheral collaterals to both eyelid and masseter muscles. WGA-594 was injected into upper eyelid, and WGA-488 was simultaneously delivered into ipsilateral masseter muscle in the same rat. Then, double labeled Vme neurons were found under both conventional and confocal microscope. Meanwhile, contact of WGA-594 positive eyelid afferent Vme neurons with WGA-488 labeled masseter afferent ones were observed sometimes. Combined with our previous observation of oculomotor projection Vme neurons, we thought WGA-594/488 double labeled Vme cells, at least some of them, are oculomotor projecting ones. Contact between eyelid and masseter afferent Vme neurons are supposed to be electrotonically coupled, based on a line of previous studies. If exogenous or genetic factors make these Vme neurons misinterpret masseter input as eyelid afferent signals, these Vme neurons might feedforward massages to eyelid retractor motoneurons in the III. Besides, oculomotor projecting Vme neurons might be co-fired by adjacent masseter afferent Vme neurons through electrotonic coupling once the masseter muscle is activated. In these cases, Marcus Gunn Syndrome might occur. This finding leads to a new hypothesis for the Syndrome.

Transcriptional profiling of dental sensory and proprioceptive trigeminal neurons using single-cell RNA sequencing

Dental primary afferent (DPA) neurons and proprioceptive mesencephalic trigeminal nucleus (MTN) neurons, located in the trigeminal ganglion and the brainstem, respectively, are essential for controlling masticatory functions. Despite extensive transcriptomic studies on various somatosensory neurons, there is still a lack of knowledge about the molecular identities of these populations due to technical challenges in their circuit-validated isolation. Here, we employed high-depth single-cell RNA sequencing (scRNA-seq) in combination with retrograde tracing in mice to identify intrinsic transcriptional features of DPA and MTN neurons. Our transcriptome analysis revealed five major types of DPA neurons with cell type-specific gene enrichment, some of which exhibit unique mechano-nociceptive properties capable of transmitting nociception in response to innocuous mechanical stimuli in the teeth. Furthermore, we discovered cellular heterogeneity within MTN neurons that potentially contribute to their responsiveness to mechanical stretch in the masseter muscle spindles. Additionally, DPA and MTN neurons represented sensory compartments with distinct molecular profiles characterized by various ion channels, receptors, neuropeptides, and mechanoreceptors. Together, our study provides new biological insights regarding the highly specialized mechanosensory functions of DPA and MTN neurons in pain and proprioception.

The Nature of Noradrenergic Volume Transmission From Locus Coeruleus to Brainstem Mesencephalic Trigeminal Sensory Neurons.

Noradrenergic neurons in the locus coeruleus (LC) release noradrenaline (NA) that acts via volume transmission to activate extrasynaptic G-protein coupled receptors (GPCRs) in target cells throughout the brain. As the closest projection, the dorsal LC laterally adjoins the mesencephalic trigeminal nucleus (MTN), in which proprioceptive primary sensory neurons innervating muscle spindles of jaw-closing muscles are exceptionally located. MTN neurons express α2-adrenergic receptors (α2-ARs) and display hyperpolarization-activated cyclic nucleotide-gated (HCN) currents (Ihs), which is downregulated by α2-AR activation. To quantify the activity-dependent outcome of volume transmission of NA from LC to MTN, we investigated how direct LC activation inhibits Ih in MTN neurons by performing dual whole-cell recordings from LC and MTN neurons. Repetition of 20 Hz spike-train evoked with 1-s current-pulse in LC neurons every 30 s resulted in a gradual decrease in Ih evoked every 30 s, revealing a Hill-type relationship between the number of spike-trains in LC neurons and the degree of Ih inhibition in MTN neurons. On the other hand, when microstimulation was applied in LC every 30 s, an LC neuron repeatedly displayed a transient higher-frequency firing followed by a tonic firing at 5–10 Hz for 30 s. This subsequently caused a similar Hill-type inhibition of Ih in the simultaneously recorded MTN neuron, but with a smaller Hill coefficient, suggesting a lower signal transduction efficacy. In contrast, 20 Hz activity induced by a 1-s pulse applied every 5–10 s caused only a transient facilitation of Ih inhibition followed by a forced termination of Ih inhibition. Thus, the three modes of LC activities modulated the volume transmission to activate α2-adrenergic GPCR to differentially inhibit Ih in MTN neurons.

The functional connectivity of locus coeruleus and mesencephalic trigeminal nucleus neurons as an implication for stress-induced masticatory dysfunction

The functional connectivity of locus coeruleus and mesencephalic trigeminal nucleus neurons as an implication for stress-induced masticatory dysfunction

Alpha 2 adrenoceptor agonist guanabenz directly inhibits hyperpolarization-activated, cyclic nucleotide-modulated (HCN) channels in mesencephalic trigeminal nucleus neurons

Alpha 2 (α2-) adrenoceptor agonists, such as clonidine or dexmedetomidine, have been found to inhibit hyperpolarization-activated, cyclic nucleotide-modulated (HCN) channels, not only by reducing intracellular cyclic AMP levels but also by directly blocking HCN channels. In this study, we examined the inhibitory effect of guanabenz, a centrally acting α2-adrenoceptor agonist with high specificity for α2A-subtype, on HCN channels in mesencephalic trigeminal nucleus (MTN) neurons which robustly express HCN channels and have been suggested to coexpress α2A-adrenoceptors. By performing whole-cell patch-clamp recording on MTN neurons in brainstem slices, hyperpolarization-activated inward current (Ih) was examined during guanabenz treatment. Guanabenz inhibited Ih in a dose-dependent manner, which was likely to be ZD7288-sensitive HCN current as it did not affect barium-sensitive inward rectifying potassium current. Guanabenz not only inhibited Ih but also shifted the voltage-dependent activation curve to hyperpolarizing potentials. Interestingly, Ih inhibition by guanabenz was not reversed by α2-adrenoceptor antagonist atipamezole treatment or by intracellular cyclic AMP perfusion, suggesting that the inhibition may not result from α2A-adrenoceptor signalling pathway but from direct inhibition of HCN channels. Coherent to our electrophysiological results, single-cell RT-PCR revealed that most MTN neurons lack α2A-adrenoceptor mRNA. Our study demonstrates that guanabenz can directly inhibit HCN channels in addition to its primary role of activating α2A-adrenoceptors.

Inhibition of GluR Current in Microvilli of Sensory Neurons via Na+-Microdomain Coupling Among GluR, HCN Channel, and Na+/K+ Pump.

Glutamatergic dendritic EPSPs evoked in cortical pyramidal neurons are depressed by activation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels expressed in dendritic spines. This depression has been attributed to shunting effects of HCN current (Ih) on input resistance or Ih deactivation. Primary sensory neurons in the rat mesencephalic trigeminal nucleus (MTN) have the somata covered by spine-like microvilli that express HCN channels. In rat MTN neurons, we demonstrated that Ih enhancement apparently diminished the glutamate receptor (GluR) current (IGluR) evoked by puff application of glutamate/AMPA and enhanced a transient outward current following IGluR (OT-IGluR). This suggests that some outward current opposes inward IGluR. The IGluR inhibition displayed a U-shaped voltage-dependence with a minimal inhibition around the resting membrane potential, suggesting that simple shunting effects or deactivation of Ih cannot explain the U-shaped voltage-dependence. Confocal imaging of Na+ revealed that GluR activation caused an accumulation of Na+ in the microvilli, which can cause a negative shift of the reversal potential for Ih (Eh). Taken together, it was suggested that IGluR evoked in MTN neurons is opposed by a transient decrease or increase in standing inward or outward Ih, respectively, both of which can be caused by negative shifts of Eh, as consistent with the U-shaped voltage-dependence of the IGluR inhibition and the OT-IGluR generation. An electron-microscopic immunohistochemical study revealed the colocalization of HCN channels and glutamatergic synapses in microvilli of MTN neurons, which would provide a morphological basis for the functional interaction between HCN and GluR channels. Mathematical modeling eliminated the possibilities of the involvements of Ih deactivation and/or shunting effect and supported the negative shift of Eh which causes the U-shaped voltage-dependent inhibition of IGluR.

The emergence of mesencephalic trigeminal neurons

BackgroundThe cells of the mesencephalic trigeminal nucleus (MTN) are the proprioceptive sensory neurons that innervate the jaw closing muscles. These cells differentiate close to the two key signalling centres that influence the dorsal midbrain, the isthmus, which mediates its effects via FGF and WNT signalling and the roof plate, which is a major source of BMP signalling as well as WNT signalling.MethodsIn this study, we have set out to analyse the importance of FGF, WNT and BMP signalling for the development of the MTN. We have employed pharmacological inhibitors of these pathways in explant cultures as well as utilising the electroporation of inhibitory constructs in vivo in the chick embryo.ResultsWe find that interfering with either FGF or WNT signalling has pronounced effects on MTN development whilst abrogation of BMP signalling has no effect. We show that treatment of explants with either FGF or WNT antagonists results in the generation of fewer MTN neurons and affects MTN axon extension and that inhibition of both these pathways has an additive effect. To complement these studies, we have used in vivo electroporation to inhibit BMP, FGF and WNT signalling within dorsal midbrain cells prior to, and during, their differentiation as MTN neurons. Again, we find that inhibition of BMP signalling has no effect on the development of MTN neurons. We additionally find that cells electroporated with inhibitory constructs for either FGF or WNT signalling can differentiate as MTN neurons suggesting that these pathways are not required cell intrinsically for the emergence of these neurons. Indeed, we also show that explants of dorsal mesencephalon lacking both the isthmus and roof plate can generate MTN neurons. However, we did find that inhibiting FGF or WNT signalling had consequences for MTN differentiation.ConclusionsOur results suggest that the emergence of MTN neurons is an intrinsic property of the dorsal mesencephalon of gnathostomes, and that this population undergoes expansion, and maturation, along with the rest of the dorsal midbrain under the influence of FGF and WNT signalling.

Effects of acidic pH on voltage-gated ion channels in rat trigeminal mesencephalic nucleus neurons.

The effects of acidic pH on several voltage-dependent ion channels, such as voltage-dependent K+ and Ca2+ channels, and hyperpolarization-gated and cyclic nucleotide-activated cation (HCN) channels, were examined using a whole-cell patch clamp technique on mechanically isolated rat mesencephalic trigeminal nucleus neurons. The application of a pH 6.5 solution had no effect on the peak amplitude of voltage-dependent K+ currents. A pH 6.0 solution slightly, but significantly inhibited the peak amplitude of voltage-dependent K+ currents. The pH 6.0 also shifted both the current-voltage and conductance-voltage relationships to the depolarization range. The application of a pH 6.5 solution scarcely affected the peak amplitude of membrane currents mediated by HCN channels, which were profoundly inhibited by the general HCN channel blocker Cs+ (1 mM). However, the pH 6.0 solution slightly, but significantly inhibited the peak amplitude of HCN-mediated currents. Although the pH 6.0 solution showed complex modulation of the current-voltage and conductance-voltage relationships, the midpoint voltages for the activation of HCN channels were not changed by acidic pH. On the other hand, voltage-dependent Ca2+ channels were significantly inhibited by an acidic pH. The application of an acidic pH solution significantly shifted the current-voltage and conductance-voltage relationships to the depolarization range. The modulation of several voltage-dependent ion channels by an acidic pH might affect the excitability of mesencephalic trigeminal nucleus neurons, and thus physiological functions mediated by the mesencephalic trigeminal nucleus could be affected in acidic pH conditions.

Acidic pH modulation of Na+ channels in trigeminal mesencephalic nucleus neurons.

Cell bodies of trigeminal mesencephalic nucleus (Vmes) neurons are located within the central nervous system, and therefore, peripheral as well as central acidosis can modulate the excitability of Vmes neurons. Here, we report the effect of acidic pH on voltage-gated Na channels in acutely isolated rat Vmes neurons using a conventional whole-cell patch clamp technique. Acidic pH (pH 6.0) slightly but significantly shifted both the activation and steady-state fast inactivation relationships toward depolarized potentials. However, acidic pH (pH 6.0) had a minor effect on the inactivation kinetics of voltage-gated Na channels. Less sensitivity of voltage-gated Na channels to acidic pH may allow Vmes neurons to transduce the precise proprioceptive information even under acidic pH conditions.

Generation of resonance-dependent oscillation by mGluR-I activation switches single spiking to bursting in mesencephalic trigeminal sensory neurons.

The primary sensory neurons supplying muscle spindles of jaw-closing muscles are unique in that they have their somata in the mesencephalic trigeminal nucleus (MTN) in the brainstem, thereby receiving various synaptic inputs. MTN neurons display bursting upon activation of glutamatergic synaptic inputs while they faithfully relay respective impulses arising from peripheral sensory organs. The persistent sodium current (IN aP ) is reported to be responsible for both the generation of bursts and the relay of impulses. We addressed how IN aP is controlled either to trigger bursts or to relay respective impulses as single spikes in MTN neurons. Protein kinase C (PKC) activation enhanced IN aP only at low voltages. Spike generation was facilitated by PKC activation at membrane potentials more depolarized than the resting potential. By injection of a ramp current pulse, a burst of spikes was triggered from a depolarized membrane potential whereas its instantaneous spike frequency remained almost constant despite the ramp increases in the current intensity beyond the threshold. A puff application of glutamate preceding the ramp pulse lowered the threshold for evoking bursts by ramp pulses while chelerythrine abolished such effects of glutamate. Dihydroxyphenylglycine, an agonist of mGluR1/5, also caused similar effects, and increased both the frequency and impedance of membrane resonance. Immunohistochemistry revealed that glutamatergic synapses are made onto the stem axons, and that mGluR1/5 and Nav1.6 are co-localized in the stem axon. Taken together, glutamatergic synaptic inputs onto the stem axon may be able to switch the relaying to the bursting mode.

Specification of sensory neurons occurs through diverse developmental programs functioning in the brain and spinal cord.

Vertebrates possess two populations of sensory neurons located within the central nervous system: Rohon-Beard (RB) and mesencephalic trigeminal nucleus (MTN) neurons. RB neurons are transient spinal cord neurons whilst MTN neurons are the proprioceptive cells that innervate the jaw muscles. It has been suggested that MTN and RB neurons share similarities and may have a common developmental program, but it is unclear how similar or different their development is. We have dissected RB and MTN neuronal specification in zebrafish. We find that RB and MTN neurons express a core set of genes indicative of sensory neurons, but find these are also expressed by adjacent diencephalic neurons. Unlike RB neurons, our evidence argues against a role for the neural crest during MTN development. We additionally find that neurogenin1 function is dispensable for MTN differentiation, unlike RB cells and all other sensory neurons. Finally, we demonstrate that, although Notch signalling is involved in RB development, it is not involved in the generation of MTN cells. Our work reveals fundamental differences between the development of MTN and RB neurons and suggests that these populations are non-homologous and thus have distinct developmental and, probably, evolutionary origins.

Characterization of proton-induced currents in rat trigeminal mesencephalic nucleus neurons

Acid-sensing ion channels (ASICs) are widely expressed in central as well as peripheral neurons. Here we have characterized the proton-induced currents in acutely isolated rat trigeminal mesencephalic nucleus (Vmes) neurons using a whole cell patch-clamp technique. In a voltage-clamp condition, the application of acid extracellular solution (≤pH 6.5) induced the inward currents in a pH-dependent manner. The proton-induced currents disappeared in the Na+-free external solution, and were concentration-dependently blocked by amiloride, a general ASIC blocker. The reversal potential of proton-induced currents was similar to the theoretical Na+ equilibrium potential, suggesting that the proton-induced currents are mainly mediated by the activation of ASICs, which are highly selective to Na+. The modulation of proton-induced currents by divalent cations and the expression patterns of ASIC transcripts using by the multi-cell RT-PCR assay suggest that Vmes neurons express functional ASIC2a and ASIC1b subunits. In a current-clamp condition, acidic pH directly depolarized the membrane potential and generated a burst of action potentials at Vmes neurons, which innervate the masseter muscle spindles. Considering that cell bodies of Vmes neurons are located within the central nervous system, ASICs expressed on Vmes neurons, by sensing peripheral and/or central acidosis, might play pivotal roles in the transduction of proprioceptive information from the masseter muscles and periodontal ligaments.

Onecut factors control development of the Locus Coeruleus and of the mesencephalic trigeminal nucleus

The Locus Coeruleus (LC), the main noradrenergic nucleus in the vertebrate CNS, contributes to the regulation of several processes including arousal, sleep, adaptative behaviors and stress. Regulators controlling the formation of the LC have been identified but factors involved in its maintenance remain unknown. Here, we show that members of the Onecut (OC) family of transcription factors, namely HNF-6, OC-2 and OC-3, are required for maintenance of the LC phenotype. Indeed, in embryos lacking any OC proteins, LC neurons properly differentiate but abnormally migrate and eventually lose their noradrenergic characteristics. Surprisingly, the expression of Oc genes in these neurons is restricted to the earliest differentiation stages, suggesting that OC factors may regulate maintenance of the LC in a non cell-autonomous manner. Accordingly, the OC factors are present throughout development in a population directly adjacent to the LC, the rhombencephalic portion of the mesencephalic trigeminal nucleus (MTN). In the absence of OC factors, rhombencephalic MTN neurons fail to be generated, suggesting that OC proteins cell-autonomously control their production. Hence, we propose that OC factors are required at early developmental stages for differentiation of the MTN neurons that are in turn necessary for maintenance of the LC.

Action potential-triggered somatic exocytosis in mesencephalic trigeminal nucleus neurons in rat brain slices

The neurons in the mesencephalic trigeminal nucleus (MeV) play essential roles in proprioceptive sensation of the face and oral cavity. The somata of MeV neurons are generally assumed to carry out neuronal functions but not to play a direct role in synaptic transmission. Using whole-cell recording and membrane capacitance (C(m)) measurements, we found that the somata of MeV neurons underwent robust exocytosis (C(m) jumps) upon depolarization and with the normal firing of action potentials in brain slices. Both removing [Ca(2+)](o) and buffering [Ca(2+)](i) with BAPTA blocked this exocytosis, indicating that it was completely Ca(2+) dependent. In addition, an electron microscopic study showed synaptic-like vesicles approximated to the plasma membrane in somata. There was a single Ca(2+)-dependent releasable vesicle pool with a peak release rate of 1912 fF s(-1). Importantly, following depolarization-induced somatic exocytosis, GABA-mediated postsynaptic currents were transiently reduced by 31%, suggesting that the somatic vesicular release had a retrograde effect on afferent GABAergic transmission. These results provide strong evidence that the somata of MeV neurons undergo robust somatic secretion and may play a crucial role in bidirectional communication between somata and their synaptic inputs in the central nervous system.

Action Potential-Triggered Somatic Exocytosis in Mesencephalic Trigeminal Nucleus Neurons in Rat Brain Slices

The neurons in the mesencephalic trigeminal nucleus (MeV) play essential roles in proprioceptive sensation of the face and oral cavity. The somata of MeV neurons are generally assumed to carry out neuronal functions but not to play a direct role in synaptic transmission. Using whole-cell recording and membrane capacitance (Cm) measurements, we found that the somata of MeV neurons underwent robust exocytosis (Cm jumps) upon depolarization and with the normal firing of action potentials in brain slices. Both removing [Ca2+]o and buffering [Ca2+]i with BAPTA blocked this exocytosis, indicating that it was completely Ca2+-dependent. In addition, an electron microscopic study showed synaptic-like vesicles approximated to the plasma membrane in somata. There was a single Ca2+-dependent releasable vesicle pool with a peak release rate of 1912 fF/s. Importantly, following depolarization-induced somatic exocytosis, GABA-mediated postsynaptic currents were transiently reduced by 31%, suggesting that the somatic vesicular release had a retrograde effect on afferent GABAergic transmission. These results provide strong evidence that the somata of MeV neurons undergo robust somatic secretion and may play a crucial role in bidirectional communication between somata and their synaptic inputs in the central nervous system.

Drebrin (developmentally regulated brain protein) is associated with axo-somatic synapses and neuronal gap junctions in rat mesencephalic trigeminal nucleus

Drebrin (developmentally regulated brain protein)-like immunoreactivity was investigated in the adult rat mesencephalic trigeminal nucleus (MTN) using light and electron microscope. Intense immunoreactive puncta were observed on the cytoplasmic membrane and within the cytoplasm. The cytoplasm was also faintly immunopositive for drebrin, and thus MTN somata other than multipolar cells were distinguishable from non-MTN somata. These immunoreactive cell bodies were localized from the level of the superior colliculus to the pons. Electron microscopic observation showed that the post-synaptic cytoplasmic membrane at axo-somatic synapses was immunoreactive for drebrin. Drebrin-like immunoreactivity was also observed on spine-like processes emanating from MTN somata. In addition, the post-synaptic cytoplasmic membrane at axo-somatic synapses was also immunopositive for drebrin. Within the cytoplasm of MTN cell bodies, a part of the rough endoplasmic reticulum and neighboring structures were also immunopositive. Further, both ends of the somato-somatic close appositions that contained neuronal gap junctions harbored immunoreactive structures. We can infer from the results that drebrin is an ideal marker protein for MTN cell bodies. The abundance of drebrin-like immunoreactivity in the MTN neurons suggests that the MTN has highly flexible synaptogenesis.

Rapid use‐dependent down‐regulation of γ‐aminobutyric acid type A receptors in rat mesencephalic trigeminal neurons

Rundown is ubiquitously seen in response to repetitive activation of receptor or ion channels as a use-dependent down-regulation through various mechanisms. In contrast to AMPA receptors, gamma-aminobutyric acid type A receptor (GABA(A)R) are believed to display no rapid use-dependent down-regulation. We report here a rapid use-dependent down-regulation of GABA(A)R in primary sensory neurons of rat mesencephalic trigeminal nucleus (MTN), which express synaptic GABA(A)Rs in addition to extrasynaptic ones, unlike other primary sensory neurons. When muscimol was repetitively puff-applied to an MTN neuron every 2 min before, during, and after the muscimol bath application for 5 min, both the GABA(A) responses obtained under both current- and voltage-clamp conditions were almost completely depressed during the bath application. However, the former and latter GABA(A) responses recovered to 26% +/- 7% and 36% +/- 7% of their control amplitudes, respectively, 15 min after washout of the bath-applied muscimol. By contrast, when examined in the presence of chelerythrine, a protein kinase C (PKC) inhibitor, together with a stringent chelation of intracellular Ca(2+), the puff responses were almost completely recovered, whereas those were recovered to 40-60% of the control by either chelerythrine or EGTA alone. A phosphatidylinositol 3-kinase inhibitor (PI3K), wortmannin, which blocks various signal transductions, including vesicular trafficking, significantly enhanced the rundown of the puff responses examined every 2 min. These findings indicate that the rundown of GABA(A) response in MTN neurons is mediated by the use-dependent down-regulation of GABA(A)R, which is reversed by PKC inhibition together with intracellular Ca(2+) chelation, while being facilitated by PI3K inhibition.

Developmental and spatial expression pattern of α‐Taxilin in the rat central nervous system

Alpha-taxilin has been identified as a binding partner of syntaxin family members and thus has been proposed to function in syntaxin-mediated intracellular vesicle trafficking. However, the lack of detailed information concerning the cellular and subcellular localization of alpha-taxilin impedes an understanding of the role of this protein. In the present study, we characterized alpha-taxilin-expressing cells in the rat CNS with a specific antibody. During embryonic development, alpha-taxilin was prominently expressed in nestin-positive neural stem cells in vivo and in vitro. As CNS development proceeded, the alpha-taxilin expression level was rapidly down-regulated. In the postnatal CNS, alpha-taxilin expression was almost confined to the neuronal lineage, with the highest levels of expression in motor neurons within the brainstem nuclei and spinal cord and in primary sensory neurons in mesencephalic trigeminal nucleus. At the cellular level, alpha-taxilin was preferentially located in Nissl substance-like structures with a tigroid or globular morphology within the soma and proximal to dendrites, but it was excluded from terminals. Combined staining with propidium iodide demonstrated that alpha-taxilin distribution overlapped with the cytoplasmic compartment enriched in RNA species, suggesting a close association of alpha-taxilin with actively translating ribosomes or polysomes in neurons. In agreement with this, a recent study indicated the preferential binding of alpha-taxilin to the nascent polypeptide-associated complex (alphaNAC), a dynamic component of the ribosomal exit tunnel in eukaryotic cells. Taken together, these findings suggest that alpha-taxilin plays multiple roles in the generation and maintenance of neurons through modulation of the NAC-mediated translational machinary and/or the syntaxin-mediated vesicle traffic in the soma.

Involvement of Persistent Na+ Current in Spike Initiation in Primary Sensory Neurons of the Rat Mesencephalic Trigeminal Nucleus

It was recently shown that the persistent Na(+) current (I(NaP)) is generated in the proximal axon in response to somatic depolarization in neocortical pyramidal neurons, although the involvement of I(NaP) in spike initiation is still unclear. Here we show a potential role of I(NaP) in spike initiation of primary sensory neurons in the mesencephalic trigeminal nucleus (MTN) that display a backpropagation of the spike initiated in the stem axon toward the soma in response to soma depolarization. Riluzole (10 muM) and tetrodotoxin (TTX, 10 nM) caused an activation delay or a stepwise increase in the threshold for evoking soma spikes (S-spikes) without affecting the spike itself. Simultaneous patch-clamp recordings from the soma and axon hillock (AH) revealed that bath application of 50 nM TTX increased the delay in spike activation in response to soma depolarization, leaving the spike-backpropagation time from the AH to soma unchanged. This indicates that the increase in activation delay occurred in the stem axon. Furthermore, under a decreasing intracellular concentration gradient of QX-314 from the soma to AH created by QX-314-containing and QX-314-free patch pipettes, the amplitude and maximum rate of rise (MRR) of AH-spikes decreased with an increase in the activation delay following repetition of current-pulse injections, whereas S-spikes displayed decreases of considerably lesser degree in amplitude and MRR. This suggests that compared to S-spikes, AH-spikes more accurately reflect the attenuation of axonal spike by QX-314, consistent with the nature of spike backpropagation. These observations strongly suggest that low-voltage-activated I(NaP) is involved in spike initiation in the stem axon of MTN neurons.