Chemical coding and central projections of gastric vagal afferent neurons

Abstract  Vagal afferents that innervate gastric muscle or mucosa transmit distinct sensory information from their endings to the nucleus of the tractus solitarius (NTS). While these afferent subtypes are functionally distinct, no neurochemical correlate has been described and it is unknown whether they terminate in different central locations. This study aimed to identify gastric vagal afferent subtypes in the nodose ganglion (NG) of ferrets, their terminal areas in NTS and neurochemistry for isolectin-B4 (IB4) and calcitonin gene-related peptide (CGRP). Vagal afferents were traced from gastric muscle or mucosa and IB4 and CGRP labelling assessed in NG and NTS. 7 ± 1% and 6 ± 1% of NG neurons were traced from gastric muscle or mucosa respectively; these were more likely to label for CGRP or for both CGRP and IB4 than other NG neurons ( P  <   0.01). Muscular afferents were also less likely than others to label with IB4 ( P  <   0.001). Less than 1% of NG neurons were traced from both muscle and mucosa. Central terminals of both afferent subtypes occurred in the subnucleus gelatinosus of the NTS, but did not overlap completely. This region also labelled for CGRP and IB4. We conclude that while vagal afferents from gastric muscle and mucosa differ little in their chemical coding for CGRP and IB4, they can be traced selectively from their peripheral endings to NG and to overlapping and distinct regions of NTS. Thus, there is an anatomical substrate for convergent NTS integration for both types of afferent input.     

TrkC kinase expression in distinct subsets of cutaneous trigeminal innervation and nonneuronal cells

Neurotrophin-activated receptor tyrosine kinases (Trks) regulate sensory neuron survival, differentiation, and function. To permanently mark cells that ever express TrkC-kinase, mice with lacZ and GFP reporters of Cre recombinase activity were crossed with mice having IRES-cre inserted into the kinase-containing exon of the TrkC gene. Prenatal reporter expression matched published locations of TrkC-expression. Postnatally, more trigeminal neurons and types of mystacial pad innervation expressed reporter than immunodetectable TrkC, indicating that some innervation transiently expresses TrkC-kinase. Reporter-tagged neurons include all those that immunolabel for TrkC, a majority for TrkB, and a small proportion for TrkA. TrkA neurons expressing TrkC-reporter range from small to large size and supply well-defined types of mystacial pad innervation. Virtually all small neurons and C-fiber innervation requires TrkA to develop, but TrkC-reporter is present in only a small proportion that uniquely innervates piloneural complexes of guard hairs and inner conical bodies of vibrissa follicle-sinus complexes. TrkC-reporter is expressed in nearly all presumptive Adelta innervation, which is all eliminated in TrkA knockouts and partially eliminated in TrkC knockouts. Many types of Abeta-fiber innervation express TrkC-reporter including all Merkel, spiny, and circumferentially oriented lanceolate endings, and some reticular and longitudinally oriented lanceolate endings. Only Merkel endings require TrkC to develop and survive, whereas the other endings require TrkA and/or TrkB. Thus, TrkC is required for the existence of some types of innervation that express TrkC, but may have different functions in others. Many types of nonneuronal cells affiliated with hair follicles and blood vessels also express TrkC-reporter but lack immunodetectable TrkC.     

Detection and characterization of a sensory microganglion associated with the spinal accessory nerve: a scanning laser confocal microscopic study of the neurons and their processes.

The spinal accessory nerve has been generally thought to be a cranial nerve with purely motor function, innervating the trapezius and sternocleidomastoid muscles. The present study identified clusters of sensory neurons consistently associated with this cranial nerve in adult rats. Either a single microganglion or several dispersed microganglia were found that adhered to the spinal root of the nerve, to small vessels, or were free within the subarachnoid space. The neurons of the ganglion had axons that joined the spinal root of the nerve proximal to its exit from the skull. Additional branches appeared to have an intracranial distribution within the arachnoid of the brainstem and along its vessels. Several findings suggest that the function of the ganglion is sensory and not autonomic. First, the architectural features of neurons within the ganglion (including their size, pseudounipolar morphology, and the lack of synaptic contacts) are similar to those of neurons in other sensory ganglia. Second, substance P and calcitonin gene-related peptide coexist within neurons of the microganglion, whereas markers for the major transmitters found in autonomic ganglia in rats are absent. Third, the expression of peptides in neurons of the ganglion was sensitive to neonatal capsaicin treatment. Finally, neurons within the ganglion were filled with a retrogradely transported dye after injection of the dye into the cervical spinal cord. Although the function of the ganglion is not known, its features are consistent with a role in nociception from the muscles of the spinal accessory complex, and it may be involved in headaches that have an occipital distribution.     

Distribution of substance P-responsive and nociceptive neurones in relation to substance P-immunoreactivity within the caudal trigeminal nucleus of the rat

Substance P is a peptide which is found in small diameter primary afferent fibres and may have a function in nociceptive afferent transmission. In order to study the role of substance P in sensory processes in depth, we have compared the distributions of nociceptive neurones and substance P-responsive neurones with the distribution of substance P in the caudal trigeminal nucleus of the rat. It was found that substance P-like immunoreactivity was located primarily in the superficial layers of nucleus caudalis (equivalent to laminae I and II of the dorsal horn) and in more ventromedially located areas (equivalent to laminae V and VI). The distribution was found to be in good agreement with the distribution of nociceptive neurones. Iontophoretically applied substance P had predominantly excitatory actions on both nociceptive and non-nociceptive nucleus caudalis neurones, although the peptide did appear to be slightly more likely to excite nociceptive neurones. Similarly, the peptide appeared slightly more likely to be excitatory in areas of nucleus caudalis showing substance P staining, but excitations were also predominantly seen in areas containing little or no apparent substance P staining. These results are consistent with the proposed role for substance P as a nociceptive afferent neurotransmitter. However, it is also possible that the peptide performs other functions in the processing of sensory information.     

Sensory neuron addition in juvenile rat: time course and specificity

The time course and specificity of neuron addition to lumbar dorsal root ganglia (DRGs) L(4)-L(6) of rats was investigated. By using methods validated by three-dimensional reconstructions, profile counts in paraffin sections of nucleoli within a nucleus were 36% greater in 100-day-old (P100) rats than in 1-day-old (P1) rats. Adult values were reached by P50. Added neurons fell disproportionately into the population of neurons whose size was below that of the mean size within the ganglion. The biochemical characteristics of small neurons were used to determine whether added neurons fall into particular subpopulations. In DRGs, L(3) and L(4), the number of neurons immunoreactive to substance P (SP) or calcitonin-gene-related peptide (CGRP) or that bound the lectin isolectin B4 (IB4) was determined. Between P5 and P100, the number of SP-stained neurons increased by 2,280 (40% increase), CGRP-stained neurons increased by 6,080 (70% increase), and IB4-stained neurons increased by 6,900 (90% increase). The increase in the number of neurons stained for CGRP or IB4 was more than twice the number of neurons found to be added to these ganglia, indicating that coexpression of these markers as well as neuron number may be developmentally regulated during postnatal life.     

Primary afferent plasticity following deafferentation of the trigeminal brainstem nuclei in the adult rat

Alpha-tyrosinated tubulin is a cytoskeletal protein that is involved in axonal growth and is considered a marker of neuronal plasticity in adult mammals. In adult rats, unilateral ablation of the left facial sensorimotor cortical areas induces degeneration of corticotrigeminal projections and marked denervation of the contralateral sensory trigeminal nuclei. Western blotting and real-time-PCR of homogenates of the contralateral trigeminal ganglion (TG) revealed consistent overexpression of growth proteins 15 days after left decortication in comparison with the ipsilateral side. Immunohistochemical analyses indicated marked overexpression of α-tyrosinated tubulin in the cells of the ganglion on the right side. Cytoskeletal changes were primarily observed in the small ganglionic neurons. Application of HRP-CT, WGA-HRP, and HRP to infraorbital nerves on both sides 15 days after left decortication showed a significant degree of terminal sprouting and neosynaptogenesis from right primary afferents at the level of the right caudalis and interpolaris trigeminal subnuclei. These observations suggest that the adaptive response of TG neurons to central deafferentation, leading to overcrowding and rearrangement of the trigeminal primary afferent terminals on V spinal subnuclei neurons, could represent the anatomical basis for distortion of facial modalities, perceived as allodynia and hyperalgesia, despite nerve integrity.     

N-methyl-D-aspartate receptor subunit phenotypes of vagal afferent neurons in nodose ganglia of the rat

Most vagal afferent neurons in rat nodose ganglia express mRNA coding for the NR1 subunit of the heteromeric N-methyl-D-aspartate (NMDA) receptor ion channel. NMDA receptor subunit immunoreactivity has been detected on axon terminals of vagal afferents in the dorsal hindbrain, suggesting a role for presynaptic NMDA receptors in viscerosensory function. Although NMDA receptor subunits (NR1, NR2B, NR2C, and NR2D) have been linked to distinct neuronal populations in the brain, the NMDA receptor subunit phenotype of vagal afferent neurons has not been determined. Therefore, we examined NMDA receptor subunit (NR1, NR2B, NR2C, and NR2D) immunoreactivity in vagal afferent neurons. We found that, although the left nodose contained significantly more neurons (7,603), than the right (5,978), the proportions of NMDA subunits expressed in the left and right nodose ganglia were not significantly different. Immunoreactivity for NMDA NR1 subunit was present in 92.3% of all nodose neurons. NR2B immunoreactivity was present in 56.7% of neurons; NR2C-expressing nodose neurons made up 49.4% of the total population; NR2D subunit immunoreactivity was observed in just 13.5% of all nodose neurons. Double labeling revealed that 30.2% of nodose neurons expressed immunoreactivity to both NR2B and NR2C, whereas NR2B and NR2D immunoreactivities were colocalized in 11.5% of nodose neurons. NR2C immunoreactivity colocalized with NR2D in 13.1% of nodose neurons. Our results indicate that most vagal afferent neurons express NMDA receptor ion channels composed of NR1, NR2B, and NR2C subunits and that a minority phenotype that expresses NR2D also expresses NR1, NR2B, and NR2C.     

Organization of calcitonin gene-related peptide-immunoreactive terminals in the primate dorsal horn

The present paper is concerned with the arrangement of axons and synaptic terminals immunostained for calcitonin gene-related peptide (CGRP), a primary afferent marker, in the primate (Macaca fascicularis) dorsal horn. The CGRP axons and terminals are uniformly distributed in laminae I and II outer (o) but they are concentrated laterally and distributed intermittently in the reticulated region of lamina V. A prominent bundle of labeled axons is seen in the sacral cord dorsal to the central canal. Emphasis is given to the relation of CGRP-immunoreactive terminals to other terminals, both labeled and unlabeled, in laminae I and IIo. In this regard, adjacent CGRP-immunoreactive terminals are often united by puncta adhaerentia. Of particular interest is the observation that CGRP-immunoreactive terminals can be found presynaptic to other terminals which sometimes resemble central primary afferent endings. In addition CGRP-immunoreactive terminals end on other CGRP terminals. Both findings suggest that primary afferent terminals interact synaptically with other primary afferent terminals.     

Expression of vesicular glutamate transporter 1 immunoreactivity in peripheral and central endings of trigeminal mesencephalic nucleus neurons in the rat

The major neuronal components of the trigeminal mesencephalic nucleus (Vmes) are primary afferent neurons that convey proprioceptive information from the cranioorofacial regions. In the present study, we examined expression of vesicular glutamate transporters (VGLUTs), VGLUT1 and VGLUT2, in the primary afferent neurons of the Vmes (Vmes neurons) in neonatal and adult rats. VGLUT1 immunoreactivity was detected in the cell bodies of Vmes neurons in neonatal rats younger than 11 days old, but not in older rats. However, in situ hybridization signals for VGLUT1 mRNA were detected in both neonatal and adult rats. No VGLUT2 immunoreactivity was detected in Vmes neurons of neonatal or adult rats. VGLUT1 immunoreactivity was also seen in the peripheral sensory endings on the equatorial regions of intrafusal fibers of muscle spindles in the masseter muscles in both neonatal and adult rats. In adult rats injected with cholera toxin B subunit (CTb) into the masseter nerve, central axon terminals of Vmes neurons were identified on masseter motoneurons within the trigeminal motor nucleus (Vm) by transganglionically and retrogradely transported CTb. VGLUT1-immunopositive axon terminals in close apposition to CTb-labeled Vm motoneurons were also detected by dual-immunofluorescence histochemistry for VGLUT1/CTb. Electron microscopy after dual immunolabeling for VGLUT1/CTb by the VGLUT1/immunoperoxidase and CTb/immunogold-silver methods further revealed synaptic contact of VGLUT1- and CTb-immunopositive axon terminals upon CTb-labeled neuronal profiles within the Vm. These data indicate that VGLUT1 is expressed in both the central axon terminals and the peripheral sensory endings of Vmes neurons, although no VGLUT1 immunoreactivity was detectable in the cell bodies of Vmes neurons in adult rats.     

Regulation of neuronal and glial galectin-1 expression by peripheral and central axotomy of rat primary afferent neurons

Galectin-1 (Gal1) is an endogenously-expressed protein important for the embryonic development of the full complement of primary sensory neurons and their synaptic connections in the spinal cord. Gal1 also promotes axonal regeneration following peripheral nerve injury, but the regulation of Gal1 by axotomy in primary afferent neurons has not yet been examined. Here, we show by immunohistochemistry and in situ hybridization that Gal1 expression is differentially regulated by peripheral nerve injury and by dorsal rhizotomy. Following peripheral nerve injury, the proportion of Gal1-positive DRG neurons was increased. An increase in the proportion of large-diameter DRG neurons immunopositive for Gal1 was paralleled by an increase in the depth of immunoreactivity in the dorsal horn, where Gal1-positive terminals are normally restricted to laminae I and II. Dorsal rhizotomy did not affect the proportions of neurons containing Gal1 mRNA or protein, but did deplete the ipsilateral dorsal horn of Gal1 immunoreactivity, indicating that it is transported centrally by dorsal root axons. Dorsal rhizotomy also resulted in an increase in Gal1 mRNA the nerve peripheral to the PNS-CNS interface (likely within Schwann cells and/or macrophages), and to a lesser extent within deafferented spinal cord regions undergoing Wallerian degeneration. This latter increase was notable in the dorsal columns and along the prior trajectories of myelinated afferents into the deeper dorsal horn. These results show that neuronal and glial expressions of Gal1 are tightly correlated with regenerative success. Thus, the differential expression pattern of Gal1 following peripheral axotomy and dorsal rhizotomy suggests that endogenous Gal1 may be a factor important to the regenerative response of injured axons.     

Localization of NADPH-diaphorase-containing neurons in sensory ganglia of the rat

The presence of reduced nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase activity was studied histochemically in the sensory ganglia of the rat. Supraspinally, the trigeminal ganglion possessed only a few cells positively stained for NADPH-diaphorase, while a large number of positive neurons was found in the nodose ganglion. In the dorsal root ganglia, the distribution of positive cells showed a peculiar pattern in relation to spinal levels. Very minor populations (less than 2% of the total ganglionic cells) exhibited positive reaction in ganglia at levels ranging from the first cervical (C1) to fourth thoracic (T4) and from the second lumber (L2) through the entire sacral levels. In the middle to lower thoracic levels (from T5 to L1), however, abundant diaphorase-positive cells were observed. From these positive neurons it was possible to trace intensely stained nerve fibers. In the lower thoracic level, for example, dense positive fibers were seen in the ramus communicans. Retrograde tracing studies revealed that diaphorase-containing neurons in the lower thoracic level project at least partly to the gastric wall and the celiac ganglion. These results indicate that the diaphorase-positive ganglionic neurons in the thoracicolumbar levels may carry autonomic visceral afferent information. Double staining with NADPH-diaphorase histochemistry and peptide immunohistochemistry revealed that NADPH-diaphorase colocalizes with calcitonin gene-related peptide and substance P in many of these visceral afferent neurons.     

Collateral sprouting of the central terminals of cutaneous primary afferent neurons in the rat spinal cord: pattern, morphology, and influence of targets

The capacity of the central terminals of primary afferents to sprout into denervated areas of neonatal spinal cord and the morphology of any novel terminals has been investigated. In rats which had undergone sciatic nerve section on the day of birth, 12 of 18 physiologically characterized intact saphenous hair follicle afferents (HFAs) were labelled intra-axonally with horseradish peroxidase (HRP) were shown to sprout up to 2,000 microns into the deafferented sciatic terminal field. The morphology of these sprouts depended on which area of the sciatic nerve territory was invaded by the afferent sprouts. Six HFAs sprouted into areas normally innervated by glabrous skin afferents and the morphology of the collateral sprouts in this region resembled that of rapidly adapting (RA) afferents. The other six saphenous HFAs had sprouted into sciatic "hairy" skin areas and the morphology of these sprouts, although abnormal, was flame shaped. In rats whose sural, saphenous, and superficial peroneal nerves were cut at birth, 4 of 7 single HRP labelled RA afferents had central terminals that had sprouted into regions of cord normally devoted to "hairy" input. These showed clear signs of HFA morphology despite their peripheral receptive fields remaining in the glabrous skin. The results show collateral sprouting of single cutaneous sensory afferent axons into adjacent inappropriate central target regions following neonatal deafferentation. Such plasticity may provide some compensation following neonatal injury. The morphology of the sprouted terminals is appropriate to the new target area rather than to its functional class and is also independent of the peripheral receptive field location providing an example of central rather than peripheral control over afferent growth patterns.     

Organization of HRP-labeled trigeminal mandibular primary afferent neurons in the rat

Horseradish peroxidase (HRP) applied to the transected mandibular division of the trigeminal (V) ganglion was transported anterogradely to pri-mary afferent terminal zones in the dorsal and dorsomedial trigeminal brain-stem nuclear complex (TBNC). Primary V afferents of ganglionic origin were also visible in the ipsilateral cerebellar cortex (crus I and II, paraflocculus) and the dentate, cuneate, solitary, supratrigeminal, and dorsal motor vagal nuclei, parvicellular reticular formation, area postrema and C1–C6 dorsal horn, laminae I–V. Contralateral subnucleus caudalis and C1–C2 dorsal horn were also innervated by primary afferents which crossed in the spinal gray to terminate medially, primarily in laminae I, II, and V. Almost all of these projections were also labeled in various combinations when HRP was applied to individual sensory branches of the mandibular nerve: lingual, infe-rior alveolar, mylohyoid, and auriculotemporal. Transganglionic transport of HRP in the latter four cases revealed strong evidence for mtradivisional somatotopy among the four branches in both the ganglion and TBNC. Cell bodies innervating posterior and/or lateral portions of the head and face (i.e., auriculotemporal and mylohyoid) were found with greater frequency in dor-sal mandibular ganglion regions, while somata supplying more rostral oral-perioral regions (i.e., lingual and inferior alveolar) were predominant ventrally. Components of the mandibular projection to the TBNC were organized topographically in at least some portion of all of its three dimen-sions. Subnuclear preferences were not clear-cut; all four nerves innervated at least some portion of principalis, oralis, interpolaris, and caudalis, save for mylohyoid, which did not project to caudalis. Lingual fibers were most prominent in principalis and oralis, occupied medial portions of the mandib-ular projection to the TBNC, and descended only to rostral caudalis, most notably laminae I-III. Inferior alveolar afferents were ubiquitous in the mandibular component of the TBNC and C1–C2, save for its far lateral bor-der. Mylohyoid terminals were sparse, most prominent in interpolaris, and occupied only dorsolateral TBNC regions and laminae III and IV of C1–C3. The auriculotemporal innervation of the mandibular TBNC was heaviest in interpolaris and was restricted to mostly ventrolateral regions. Its primary focus, however, was laminae III and IV of C1–C4. The clinical implications of this topographical organization are discussed, particularly with respect to the rostrocaudal intradivisional lamination in caudalis and the cervical dorsal horn.     

Expression of P2X3 receptor in the trigeminal sensory nuclei of the rat

Trigeminal primary afferents expressing P2X(3) receptor are involved in the transmission of orofacial nociceptive information. However, little is known about their central projection pattern and ultrastructural features within the trigeminal brainstem sensory nuclei (TBSN). Here we use multiple immunofluorescence and electron microscopy to characterize the P2X(3)-immunopositive (+) neurons in the trigeminal ganglion and describe the distribution and synaptic organization of their central terminals within the rat TBSN, including nuclei principalis (Vp), oralis (Vo), interpolaris (Vi), and caudalis (Vc). In the trigeminal ganglion, P2X(3) immunoreactivity was mainly in small and medium-sized somata, but also frequently in large somata. Although most P2X(3) (+) somata costained for the nonpeptidergic marker IB4, few costained for the peptidergic marker substance P. Most P2X(3) (+) fibers in the sensory root of trigeminal ganglion (92.9%) were unmyelinated, whereas the rest were small myelinated. In the TBSN, P2X(3) immunoreactivity was dispersed in the rostral TBSN but was dense in the superficial laminae of Vc, especially in the inner lamina II. The P2X(3) (+) terminals contained numerous clear, round vesicles and sparse large, dense-core vesicles. Typically, they were presynaptic to one or two dendritic shafts and also frequently postsynaptic to axonal endings, containing pleomorphic vesicles. Such P2X(3) (+) terminals, showing glomerular shape and complex synaptic relationships, and those exhibiting axoaxonic contacts, were more frequently seen in Vp than in any other TBSN. These results suggest that orofacial nociceptive information may be transmitted via P2X(3) (+) afferents to all TBSN and that it may be processed differently in different TBSN.     

Up-regulation of calcitonin gene-related peptide and receptor tyrosine kinase TrkB in rat bladder afferent neurons following TNBS colitis

Colonic inflammation has profound effects on the urinary bladder physiology and produces hypersensitivity of bladder afferent neurons and neurogenic bladder overactivity. Calcitonin gene-related peptide (CGRP) expressed in dorsal root ganglia (DRG) plays an important role in mediating sensory perception following visceral inflammation. In the present study, we determined that the expression of CGRP was increased in bladder afferent neurons in lumbosacral DRG following tri-nitrobenzene sulfonic acid (TNBS)-induced colitis in rat. After colitis, the percentage of bladder afferent neurons expressing CGRP was increased in L1 (61.2+/-2.9% in colitis vs. 37.7+/-5.1% in controls; p<0.05) and S1 DRG (26.3+/-2.3% in colitis vs. 15.5+/-1.9% in controls; p<0.01). We also demonstrated that the expression of tyrosine kinase receptor TrkB was increased in L1 (39.7+/-2.9% in colitis vs. 25.2+/-4.3% in controls; p<0.05) and S1 DRG (45.6+/-3.8% in colitis vs. 38.3+/-3.6% in controls; p<0.01) following colitis. CGRP and TrkB were co-stored in a subpopulation of DRG neurons in control and colitic animals and the number of DRG cells co-expressing CGRP and TrkB was significantly increased in L1 (2.7-fold, p<0.01) and S1 DRG (2.4-fold, p<0.01) following colitis. In cultured DRG, exogenous BDNF application significantly increased CGRP expression, which was blocked by TrkB selective inhibitor K252a. These results suggest that up-regulation of CGRP and TrkB in bladder afferent neurons may play a role in colon-to-bladder cross-sensitization following colitis.     

Effects of capsaicin on receptive fields and on inhibitions in rat spinal cord

The dorsal horn of the lumbar enlargement of the spinal cord is somatopically arranged such that the medial half contains cells with receptive fields on the distal parts of the leg and the cells in the lateral half have proximal receptive fields. After chronic sciatic nerve transection that removes input from the distal limb, a substantial number of cells in the medial dorsal horn begin to respond to proximal cutaneous stimulation. We attempted to discover the mechanisms responsible for this expansion of receptive fields following deafferentation. In these series of experiments we used capsaicin, a C-fiber neurotoxin that either destroys C fibers when administered neonatally or produces a functional blockade of C input to the cord when administered topically to a peripheral nerve. After both neonatal or topical capsaicin treatment there was disorganization of the somatopic map of the dorsal horn similar to that produced by nerve transection. Neonatal capsaicin treatment also produced a decrease in primary afferent depolarization and of A-afferent-mediated inhibitions in the dorsal horn, similar to that observed after nerve transection. Local capsaicin treatment, however, did not alter primary afferent depolarization or A-afferent inhibitions. It is unlikely, therefore, that these forms of inhibitions were responsible for controlling receptive field size. The actions of capsaicin, however, did imply some role for C fibers in determining the organization of receptive fields in the dorsal horn.     

VR1-immunoreactive primary sensory neurons in the rat trigeminal ganglion

Immunohistochemistry for VR1, a nociceptive transducer for vanilloid compounds, protons and heat (>43°C), was performed on the rat trigeminal ganglion (TG). The immunoreactivity (IR) was detected in 20% of TG cells and these neurons were mostly small- to medium-sized (mean±S.D. 427±189 μm²). Twenty-six percent of the TG neurons retrogradely labeled from the facial skin exhibited VR1-IR, while the IR was detected in only 8% of those labeled from the tooth pulp. Co-expression of VR1 was common among the calcitonin gene-related peptide-immunoreactive cutaneous neurons (63%) but not among the similar tooth pulp neurons (20%). The present study indicates that primary nociceptive neurons which respond to vanilloid compounds, protons and heat are abundant in the facial skin but not in the tooth pulp.     

Peripheral peptidergic fibers of the trigeminal nerve in the olfactory bulb of the rat

Axons immunoreactive for calcitonin gene-related peptide (CGRP) and substance P are present in the olfactory nerve, although few, if any, olfactory receptor cells contain immunocytochemically detectible levels of these peptides. The possible trigeminal origin of these fibers was tested by performing unilateral stereotaxic lesions of the ophthalmic division of the trigeminal nerve, followed 2–25 days later by immunocytochemistry for CGRP and substance P. As reported previously, free nerve endings immunoreactive for both peptides were found transversing the nasal epithelium on the unlesioned side. Also on the unlesioned side, peptidergic axons, immunoreactive for both CGRP and substance P, could be traced from the olfactory nerve into the glomerular layer throughout the olfactory bulb, but especially into its rostral third. Ipsilateral to the trigeminal ganglion lesion, such peptide-immunoreactive fibers were absent or markedly reduced in the bulb, nerve, and epithelium. These results indicate that the peripheral branches of the ophthalmic branch of the trigeminal nerve enter the olfactory bulb along with the olfactory nerve and terminate in the glomerular layer along with the olfactory axons. Ultrastructural analysis of the CGRP-immunoreactive terminals in the glomeruli reveal vesicle-filled axonal processes terminating in the absence of obvious pre- or postsynaptic specializations. Whether the trigeminal fibers in the bulb are functional, e.g., convey information to the olfactory bulb via an axon reflex, or relay information from the olfactory bulb to the brainstem trigeminal nuclei is unclear. © 1993 Wiley-Liss, Inc.     

Brain-derived neurotrophic factor-immunoreactive primary sensory neurons in the rat trigeminal ganglion and trigeminal sensory nuclei

Immunohistochemistry for brain-derived neurotrophic factor (BDNF) was performed on the rat trigeminal ganglion (TG). The immunoreactivity (IR) was detected in 46% of TG neurons. These neurons were mostly small- or medium-sized (range, 149.7-1246.3 microm2; mean +/- SD = 373.4 +/- 151.6 microm2). A double immunofluorescence method also revealed that 54% of BDNF-immunoreactive (IR) neurons were immunoreactive for calcitonin-gene-related peptide. In addition, 93% of BDNF-IR TG neurons contained vanilloid receptor subtype 1. However, the co-expression of BDNF and vanilloid receptor 1-like receptor was very rare (less than 1%). In the trigeminal sensory nuclei, laminae II of the medullary dorsal horn was abundant in presumed BDNF-IR axon terminals. Such profiles were also detected in the dorsolateral part of the subnucleus oralis. The retrograde tracing and immunohistochemical methods demonstrated that BDNF-IR was common among cutaneous TG neurons (47%) but not tooth pulp TG neurons (13%). The present study indicates that BDNF-IR TG neurons have unmyelinated axons and project to the superficial medullary dorsal horn. It is likely that BDNF-containing neurons in both the trigeminal and spinal sensory systems have similarities in morphology and function. However, the content of BDNF in TG neurons probably depends on their peripheral targets. BDNF seems to convey nociceptive cutaneous input to the trigeminal sensory nuclei.