TrkB-like immunoreactivity in trigeminal sensory nerve fibers of the rat molar tooth pulp: development and response to nerve injury

The localization of TrkB, a signal transducing receptor for brain-derived neurotrophic factor and neurotrophin-4, was studied in the rat mandibular molar pulp during development and following nerve injury. Sections were incubated with rabbit polyclonal antiserum against the catalytic part of the TrkB receptor, thus only binding to full-length TrkB receptors, and examined by immunofluorescence microscopy and EM immunocytochemistry. At embryonic day 17, strongly TrkB-positive fibers were located in mandibular nerve trunks, in nerve fibers of deeper mesenchyme in close spatial relation to sites of future tooth development and in subepithelial nerve plexa. At postnatal days 1–12 intensely TrkB-positive nerve fibers surrounded and eventually invaded the developing dental papillae and pulps. In the normal adult pulp TrkB-immunoreactive axons were seen extending through the radicular pulp into the coronal areas. Double labelling demonstrated a considerable overlap between TrkB-like immunoreactivity and low affinity neurotrophin receptor-like immunoreactivity in the pulp, although some low affinity neurotrophin receptor-positive nerve fibers lacked TrkB-like immunoreactivity. Immunogold electron microscopy showed TrkB-like immunoreactivity in myelinated as well as in unmyelinated axons. One week following inferior alveolar nerve injury there was a dramatic decrease in the level of TrkB-like immunoreactivity labelling in the pulp, which paralleled an increase in the expression of Schwann cell low affinity neurotrophin receptor-like immunoreactivity. The first signs of regenerating TrkB-positive nerve fibers were found at 4 weeks post-operative, and at 9 weeks the distribution of pulpal TrkB-like immunoreactivity had returned to normal. These data indicate that brain-derived neurotrophic factor and/or neurotropin-4 could be target-derived factors involved in sensory trigeminal tooth pulp nerve fiber development, differentiation or regeneration.     

Chronic dexamethasone treatment and its effects on sensory neuropeptides, pulpal injury reactions and reparative dentin

Initial sensory nerve reactions to dental injuries include terminal sprouting and intensified immunoreactivity for calcitonin gene-related peptide (CGRP) and substance P (SP); those reactions are reduced at 4 days after injury when rats are treated daily with dexamethasone (DEX) [17]. Here we have analyzed long-term effects of DEX (daily, 0.2 mg/kg) on wound healing, sensory nerve sprouting, and CGRP/SP intensity at 7–14 days after cavity preparation. All DEX treated rats had loss of appetite and stopped growing during the postoperative periods while controls had normal postoperative growth. After 7–14 days, CGRP immunoreactivity OR) was decreased to one-third of normal (P < 0.05) compared to vehicle in both the intact and injured molar pulp, and SP also decreased, but the neuropeptide intensity in adjacent periodontal innervation was not changed. Pulpal injury and inflammation were reduced by DEX treatment, but reparative dentin was formed just as well in the DEX rats as in the vehicle group. When the injured teeth formed fibrous dentin, there was sprouting of nerves towards that matrix, and DEX did not inhibit that reaction. The sprouts could contain intense neuropeptide immunoreactivity in DEX rats even though the CGRP/SP intensity in uninjured pulp was reduced. We conclude that (1) chronic DEX treatment causes a generalized decrease in CGRP and SP neuropeptides in pulpal nerves but not in periodontal ligament; (2) it reduces abscess formation in injured teeth; (3) it does not block reparative dentin formation; and (4) it does not block sprouting of pulpal nerves towards fibrous dentin. The selective loss of pulpal neuropeptides CGRP and SP during dexamethasone treatment may be caused by reduced dental function since there was substantial loss of appetite and chronic weight loss during the 1–2 week treatment periods.     

Neurocalcin-immunoreactive primary sensory neurons in the trigeminal ganglion provide myelinated innervation to the tooth pulp and periodontal ligament

The distribution of neurocalcin-immunoreactive (NC-ir) primary sensory neurons was examined in the trigeminal ganglion (TG), mesencephalic trigeminal tract nucleus (Mes5) and intraoral structures. NC-ir primary sensory neurons were located in the TG but not the Mes5. The coexpression study demonstrated that virtually all NC-ir TG neurons exhibited S100-immunoreactivity (-ir). In the tooth pulp, NC-ir nerve fibers were observed in the subodontoblastic and odontoblastic layers. Immunoelectron microscopic and retrograde tracing methods revealed that myelinated pulpal axons derived from the TG mostly exhibited the ir. In the periodontal ligament, bush-like endings showed NC-ir. These endings were morphologically identical to Ruffini-like endings. The present study suggests that NC-ir trigeminal primary sensory neurons have their cell bodies in the TG. Their peripheral axons are probably myelinated. Such neurons include pulpal nociceptors and low-threshold mechanoreceptors.     

Regulation of NGF-family ligands and receptors in adulthood and senescence: correlation to degenerative and regenerative changes in cutaneous innervation

During development, a highly differential neurotrophin dependency is reported for various types of nerve endings in the whisker follicle. To what extent these dependencies extend and play a role in adulthood is largely unresolved. We show here, using in situ hybridization and immunohistochemistry that the expression of neurotrophins and trk/p75 receptors persists in adulthood. As suggested by their expression profiles, many classes of cutaneous nerve endings disclose similar ligand-receptor dependencies in adult animals as during development, while other populations appear to switch their dependency. Furthermore, our data suggest that sensory endings that have a high turnover due to mechanical wear and tear, e. g. Merkel cell-neurite complexes at the level of ring sinus show a more complex ligand-receptor expression phenotype than do endings with a less vulnerable location, e.g. the Merkel cell-neurite complexes at the rete ridge collar. Thus, neurotrophin-3 (NT3)/trkA signalling is suggested to be important for a continuous terminal plasticity of Merkel cell-neurite complexes at the level of ring sinus in adulthood. Evidence supporting a role for neurotrophin signalling in maintaining the adult cutaneous innervation also comes from the close correlation between altered ligand-receptor expression(s) and axonal/terminal aberrations in senescence. Thus, an ageing-related decrease in target neurotrophin expression, notably NT3 and NT4, results in a site-specific loss of sensory terminals concomitant with an aberrant growth of regenerating/sprouting axons into new target fields. Ageing of the cutaneous innervation, manifested in degenerative and regenerative events, seems strongly associated with changes in neurotrophic interactions between sensory neurons and target tissues.     

Innervation of footpads of normal and mutant mice lacking sweat glands

Footpads of normal adult mice are innervated by sympathetic and sensory fibers. The sympathetic fibers associated with sweat glands contain acetylcholinesterase and immunoreactivity for vasoactive intestinal peptide. Although catecholamine histofluorescence is absent, the gland innervation exhibits immunoreactivity for tyrosine hydroxylase. A distinct population of sympathetic fibers, which possess catecholamines and neuropeptide Y as well as tyrosinehydroxylase immunoreactivity, innervates blood vessels. Sensory fibers containing immunoreactivity for substance P and calcitonin gene-related peptide course beneath the epidermis and some form endings in it. Treatment of neonatal mice with the adrenergic neurotoxin, 6-hydroxydopamine, results in loss of sympathetic innervation of sweat glands and blood vessels, permits growth of sensory axons into sweat glands, but does not alter the peptidergic sensory innervation of the dermis and epidermis. Three mouse mutations, Tabby (Ta), crinkled (cr), and downless (dl), disrupt the interactions between the mesenchyme and epidermis that are required for normal development of specific epidermal derivatives, including sweat glands. The sympathetic innervation of blood vessels and sensory innervation of footpad skin of the three mutant mice that lack sweat glands is indistinguishable from normal. The sympathetic fibers that normally innervate sweat glands, however, are not present. These results indicate that in the absence of their normal target, the sympathetic fibers that innervate sweat glands are lacking. Furthermore, they suggest that, although sensory fibers may sprout into sympathetic targets in the footpad, the domains occupied by sensory fibers are not normally accessible to sympathetic axons. © 1994 Wiley-Liss, Inc.     

Growth-associated protein (GAP-43)-like immunoreactivity in primary and permanent tooth pulp nerve fibers of the cat.

GAP-43-like immunoreactivity in developing and mature incisor and canine tooth pulp nerve fibers in the cat was examined with fluorescence immunohistochemistry and pre-embedding immunogold electron microscopy. As expected, pulpal and periodontal nerve fibers in primary teeth aged 2-3 weeks showed strong immunoreactivity. Double-labeling experiments demonstrated that 50-70% of primary pulpal GAP-43-positive nerve fibers showed CGRP-like immunoreactivity. However, in adult permanent teeth the vast majority of pulpal nerve fibers also displayed intense GAP-43-like immunoreactivity both when surrounding pulpal blood vessels and in the subodontoblast/odontoblast region. There was a high degree (90-95%) of simultaneous expression of GAP-43-like immunoreactivity and CGRP-like immunoreactivity in adult permanent pulps. Immunogold GAP-43 labeling was mainly associated with the cytoplasmic side of axonal membranes. However, occasional examples of immunolabeled Schwann cells were also found. High levels of GAP-43 in normal mature permanent pulpal nerves may facilitate neural plasticity after dental wear or injury.     

Distribution of central sensory axons in transgenic mice overexpressing nerve growth factor and lacking functional p75 neurotrophin receptor expression

This study examined the roles of nerve growth factor (NGF) and the p75 neurotrophin receptor (p75NTR) in the growth of dorsal root ganglion (DRG) central processes in the dorsal horn. Two genetically modified mouse strains were used: transgenic mice that overexpress NGF in the CNS under the control of the glial fibrillary acidic protein promoter, and p75NTR exon III null mutant mice that express a hypomorphic form of this receptor. In both NGF transgenic and nontransgenic mice with hypomorphic expression of p75NTR, there is a significant loss of DRG neurons compared to mice with normal p75NTR expression. This reduction in neuron number has been shown to underlie a corresponding decrease in peripheral nociceptive sensory innervation. Within the CNS, however, nociceptive innervation of the dorsal horn appears to be unaffected by hypomorphic expression of p75NTR. Comparisons of calcitonin gene-related peptide immunoreactivity in the dorsal horn revealed that the area occupied by DRG central processes was not significantly different between p75NTR hypomorphic mice and wild-type siblings, or between NGF transgenic mice with either hypomorphic or normal expression of p75NTR. We propose that DRG central processes arborize extensively in both NGF-transgenic and nontransgenic p75NTR hypomorphic mice in order to compensate for the loss of DRG neurons and restore dorsal horn innervation to normal levels. We also present evidence suggesting that NGF plays only a minor role in the growth of DRG central processes.     

Nerve growth factor-mediated collateral sprouting of central sensory axons into deafferentated regions of the dorsal horn is enhanced in the absence of the p75 neurotrophin receptor

This study examined the growth capacity of nerve growth factor (NGF)-responsive dorsal root ganglion (DRG) central processes using mice of the following genotypes: wildtype, p75 neurotrophin receptor (p75NTR) exon III null mutant, NGF transgenic, and NGF transgenic with p75NTR exon III null mutation (NGF/p75(-/-)). In wildtype and p75NTR exon III null mutant mice calcitonin gene-related peptide (CGRP) immunoreactivity in the dorsal horn is dramatically reduced at both 3 and 28 days after rhizotomy. NGF transgenic and NGF/p75(-/-) mice also display reduced CGRP immunoreactivity 3 days after rhizotomy, but by postsurgical day 28 significant increases in the density of CGRP-positive axons are observed in the injured dorsal horns of these mice. Interestingly, NGF/p75(-/-) mice displayed significantly more new axonal growth when compared to NGF transgenic mice expressing full-length p75NTR. Immunohistochemical and ultrastructural analyses revealed that this axonal growth is not the result of regeneration but rather injury-induced sprouting by intact DRG central processes into the lesion site. This collateral growth is restricted to deafferentated areas of the dorsal horn, and we therefore propose that this is an example of compensatory sprouting by NGF-sensitive axons in the spinal cord, a response that is enhanced in the absence of NGF binding to p75NTR.     

Overexpression of nerve growth factor in skin causes preferential increases among innervation to specific sensory targets

The impact of increased levels of skin-derived nerve growth factor (NGF) neurotrophin on sensory and sympathetic innervation to the mouse mystacial pad and postero-orbital vibrissae was determined. Consistent with an approximate doubling of neuron number in trigeminal and superior cervical ganglia, many components of the sensory and sympathetic innervation were substantially enhanced. Although the increased number of neurons raised the possibility that all types of innervation were increased, immunohistochemical analysis indicated that enhanced NGF production had a differential effect upon sensory innervation, primarily increasing unmyelinated innervation. This increased innervation occurred in specific locations known to be innervated by small, unmyelinated fibers, suggesting that NGF modulated sensory innervation density, but not targeting. In contrast, sympathetic innervation was not only increased but also was distributed to some aberrant locations. In the intervibrissal fur of the mystacial pad, both the number of sensory axons and branches appeared increased, whereas in vibrissal follicle sinus complexes, only branching increased. In some areas, sensory ending density was lower than expected based upon the size of the source nerve bundles suggesting that many axons and branches were surviving but failing to form functional endings. Furthermore, the immunochemical profile of innervation was altered in some sensory populations as demonstrated by the coexistence of RT-97 neurofilament labeling in calcitonin gene-related peptide (CGRP) positive axons, by the loss of substance P colocalization in some CGRP axons, and by an absence of neuropeptide Y labeling in tyrosine hydroxylase positive sympathetic axons. Collectively, these results indicate that the NGF mediated increase in neuron number may be selective for particular sets of innervation and that increases among some populations may result from phenotypic switching. J. Comp. Neurol. 387:489–506, 1997. © 1997 Wiley-Liss, Inc.     

Patterns of innervation of sympathetic vascular neurons by peptide-containing primary sensory fibers

The purpose of this study was to determine whether there is a specific organization of the primary sensory innervation on to identified vascular neurons in the inferior mesenteric ganglion (IMG) in guinea-pig. Retrograde tracers were placed intraluminally in inferior mesenteric artery (IMA) or inferior mesenteric vein (IMV) in vitro to identify ganglionic neurons as arterial, venous or unlabeled neurons. The distribution of primary sensory nerve fibers containing calcitonin gene-related peptide (CGRP), neuronal nitric oxide synthase (NOS) and substance P immunoreactivity (SP-IR) was compared before and after treatment with capsaicin. In control animals the density of immunoreactivity varied both with the transmitter and the type of neuron innervated. The density of immunoreactivity for all the three substances was reduced by capsaicin treatment. The degree of reduction of immunoreactivity in the fibers varied with the transmitter and the type of neuron. The density of CGRP and SP immunoreactive fibers was greatest around unlabeled neurons; 78% of the CGRP fibers were of primary sensory origin and all of the SP fibers were primary sensory. Around arterial neurons 44% of the CGRP fibers were of primary sensory origin and around venous 68% were primary sensory. NOS positive innervation around venous neurons was denser than around arterial neurons and all of it was completely (97%) eliminated by capsaicin, indicating that it was solely of primary sensory origin. We conclude that the primary sensory fibers innervating the IMG are differentially distributed to arterial and venous neurons and that the pattern of distribution is characteristic for each sensory neurotransmitter.     

Substance P-, calcitonin gene-related peptide,growth-associated protein-43, and neurotrophin receptor-like immunoreactivity associated with unmyelinated axons in feline ventral roots and pia mater

The spinal pia mater receives a rich innervation of small sensory axons via the ventral roots. Other sensory axons enter the ventral roots but end blindly or turn abruptly in hairpin loop-like formations and continue in a distal direction. In the present study, the content of substance P (SP)-, calcitonin gene-related peptide (CGRP)-, growth-associated protein (GAP-43)-, and low-affinity neurotrophin receptor protein (p75NGFr)-like immunoreactivity (-LI) associated with these different types of sensory axons was assessed with light and electron microscopic immunohistochemical techniques. In addition, the binding of antibodies against synthetic peptides representing unique sequences of residues in the products of the trk and trkB protooncogenes was analyzed. These genes encode membrane spanning proteins, which have been shown to constitute specific high affinity binding sites for several members of the nerve growth factor family of neurotrophic factors. The results of the present study imply that the ventral root afferents comprise several different types of sensory axons, which all contain SP-, CGRP-, GAP-43-, and p75NGFr-like immunoreactivities. In addition, at least some of the presumed sensory fiber bundles in ventral roots and the pia mater were immunoreactive for the trkB gene product. Moreover, leptomenin-geal cells and nonneuronal cells of the ventral roots were shown to bind antibodies to both the trk and trkB gene products. The ventral root afferents seem to share their immunohistochemical pattern with pain-transducing axons at some other locations, such as the tooth pulp. The contents of SP- and CGRP-LI in sensory axons that reach the central nervous system (CNS) through the ventral root indicate that ventral root afferents may be involved in sensory mechanisms, such as the ventral root pain reaction, as well as in the control of the pial blood vessels. The demonstration of GAP-43 and neurotrophin receptor-immunoreactivities associated with unmyelinated fibers in ventral roots and the pia mater is discussed in relation to previous reports on postnatal plasticity in these axonal populations. © 1994 Wiley-Liss, Inc.     

Morphological and neurochemical differences in peptidergic nerve fibers of the mouse vagina

The vagina is innervated by a complex arrangement of sensory, sympathetic, and parasympathetic nerve fibers that contain classical transmitters plus an array of neuropeptides and enzymes known to regulate diverse processes including blood flow and nociception. The neurochemical characteristics and distributions of peptide‐containing nerves in the mouse vagina are unknown. This study used multiple labeling immunohistochemistry, confocal maging and analysis to investigate the presence and colocalization of the peptides vasoactive intestinal polypeptide (VIP), calcitonin‐gene related peptide (CGRP), substance P (SP), neuropeptide tyrosine (NPY), and the nitric oxide synthesizing enzyme neuronal nitric oxide synthase (nNOS) in nerve fibers of the murine vaginal wall. We compared cervical and vulvar areas of the vagina in young nullipara and older multipara C57Bl/6 mice, and identified differences including that small ganglia were restricted to cervical segments, epithelial fibers were mainly present in vulvar segments and most nerve fibers were found in the lamina propria of the cervical region of the vagina, where a higher number of fibers containing immunoreactivity for VIP, CGRP, SP, or nNOS were found. Two populations of VIP‐containing fibers were identified: fibers containing CGRP and fibers containing VIP but not CGRP. Differences between young and older mice were present in multiple layers of the vaginal wall, with older mice showing overall loss of innervation of epithelium of the proximal vagina and reduced proportions of VIP, CGRP, and SP containing nerve fibers in the distal epithelium. The distal vagina also showed increased vascularization and perivascular fibers containing NPY. Immunolabeling of ganglia associated with the vagina indicated the likely origin of some peptidergic fibers. Our results reveal regional differences and age‐ or parity‐related changes in innervation of the mouse vagina, effecting the distribution of neuropeptides with diverse roles in function of the female genital tract.     

TrkA-expressing trigeminal sensory neurons display both neurochemical and structural plasticity despite a loss of p75NTR function: responses to normal and elevated levels of nerve growth factor

In neural crest-derived sensory ganglia, approximately half of the neuronal population expresses the transmembrane trkA receptor that is required for neuronal binding of target-derived nerve growth factor (NGF). These same neurons also express the p75 neurotrophin receptor (NTR) that increases the affinity of trkA for NGF. Depleting p75NTR expression reduces both the survival of trkA-positive sensory neurons and their afferent innervation of peripheral targets. In this investigation, we assessed the neurochemical and structural plasticity of trigeminal sensory neurons in p75NTR-deficient mice in response to either normal or elevated levels of NGF during postnatal development and into adulthood. Although p75NTR-deficient mice have 30% fewer trigeminal neurons, levels of trkA mRNA expression are modestly elevated in these mutant mice as compared to control mice. The density of central afferent axons and local levels of NGF are, however, comparable between mutant and control animals. Thus, despite the survival of fewer trigeminal neurons, neither ganglionic levels of trkA mRNA expression nor the density of central afferent projections are depleted in p75NTR-deficient mice. In response to elevated levels of NGF protein, transgenic mice with and without p75NTR expression display both increased levels of trkA mRNA expression and a greater density of trigeminal central afferent axons as compared to control mice. These data further reveal that an absence of p75NTR function in trigeminal sensory neurons does not diminish their capacity for NGF-dependent plasticity, namely trkA mRNA expression and collateral growth of central afferent axons.     

Cat dental pulp after denervation and subsequent re-innervation: changes in blood-flow regulation and distribution of neuropeptide-, GAP-43- and low-affinity neurotrophin receptor-like immunoreactivity

The effects of unilateral extramandibular inferior alveolar nerve injury on pulpal blood-flow responses to electrical stimulation and i.v. injections of substance P (SP) in cat mandibular canine teeth with a dentinal lesion were investigated with laser Doppler flowmetry. After blood-flow recordings, the teeth were fixed and the pulps were examined with light and electron microscopy. The distribution of pulpal SP, neurokinin A (NKA), calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), growth-associated protein (GAP-43) and low-affinity neurotrophin receptor (LANR)-like immunoreactivitivity was examined immunohistochemical fluorescence microscopy. Blood-flow recordings, performed at 10 days and 1 month postoperatively, showed that vasodilation, occurring in control teeth after bipolar electrical stimulation of the tooth crown, was absent in the denervated pulps, whereas at 3 months, five of six teeth had regained responsiveness, although at a low level. There was enhanced vasodilation (by 370%) to SP injections (400 fmol i.v.) at 10 days in denervated pulps. Such supersensitivity was reduced at 1 month despite the apparent lack of nerve fibers, and the response fell further towards the level in control teeth at 3 months when pulpal axons reappeared. At 10 days and 1 month postoperatively, light and electron microscopy demonstrated that surgery had resulted in total pulpal denervation. At 3 and 6 months, a large number of regenerated pulpal axons reappeared, in accordance with previous findings. At 10 days and 1 month after nerve transection immunohistochemistry showed a complete loss of pulpal immunoreactivity to all of the neuropeptides that were studied. At 3 and 6 months, neuropeptide immunoreactivity reappeared but far fewer number of pulpal nerve fibers were SP-, NKA- and CGRP-immunoreactive than under normal conditions, as demonstrated by double-labeling experiments with GAP-43- or LANR-antiserum. The results indicate that pulpal hemoregulatory functions, which are lost after denervation, do not return to normal levels after nerve regeneration. This malfunction may be caused by inadequate target re-innervation and/or a deficiency of neuropeptides in the re-innervated pulp.     

NPY Y1 receptors are present in axonal processes of DRG neurons

Using a sensitive immunohistochemical method, the localization of the neuropeptide Y (NPY) Y1 receptor (Y1R) was studied in contralateral and ipsilateral dorsal root ganglion (DRG) neurons of rats subjected to different unilateral manipulations with focus on their axonal processes and projection areas. Y1R-like immunoreactivity (LI) was observed in the contralateral sciatic nerve and dorsal roots of lesioned rats, and double staining revealed colocalization with calcitonin gene-related peptide (CGRP). Y1R-LI was also seen in fibers close to and even within the epidermis. A fairly small number of nerve endings double-labeled for Y1R and CGRP were present in the dorsal horn. After unilateral crush of the sciatic nerve Y1R- and CGRP-LI accumulated in the same axons proximal to the lesion. After dorsal rhizotomy CGRP-LI was strongly reduced in the ipsilateral dorsal horn. No certain change was observed for Y1R- or NPY-LI, but Y1R/CGRP double-labeled nerve endings disappeared after the lesion. These results strongly suggest centrifugal transport of Y1Rs in DRG neurons, mainly to the peripheral sensory branches. To what extent these Y1Rs are functional has not been analyzed here, but a recent study on Y1R null mice provides evidence for involvement of prejunctional Y1Rs in peripheral sensory functions     

Support of trigeminal sensory neurons by nonneuronal p75 neurotrophin receptors

The p75 neurotrophin receptor (p75NTR) binds all four mammalian neurotrophins, including neurotrophin-3 (NT-3) required for the development of select sensory neurons. This study demonstrated that many gustatory and somatosensory neurons of the tongue depend upon p75NTR. Each of thousands of filiform papillae at the front of the tongue as well as each somatosensory prominence at the back of the tongue has a small cluster of p75NTR-positive epithelial cells that is targeted by somatosensory innervation. This expression of p75NTR by epithelial target cells required NT-3 but not adult innervation. NT-3-secreting cells were adjacent to the p75NTR-positive target cells of each somatosensory organ, as demonstrated in NT-3(lacZneo) transgenic mice. In NT-3 null mutant mice, there were few lingual somatosensory neurons. In p75NTR null mutant mice, the lingual somatosensory axons were likewise absent or had deficient terminal arborizations. Cell culture indicated that substrate p75NTR can influence neuronal outgrowth. Specifically, dissociated trigeminal sensory neurons more than doubled their neurite lengths when grown on a lawn of p75NTR-overexpressing fibroblasts. This enhancement of neurite outgrowth by fibroblast p75NTR raises the possibility that epithelial target cell p75NTR may help to promote axonal arborization in vivo. The co-occurrence in p75NTR null mice of a 35% reduction in geniculate ganglion taste neurons and a shortfall of taste buds is consistent with the established role of gustatory innervation in prompting mammalian taste receptor cell differentiation.     

Identification of spinal afferent nerve endings in the colonic mucosa and submucosa that communicate directly with the spinal cord: The gut–brain axis

The major sensory nerve pathway between the colon and central nervous system (spinal cord and brain) that underlies the gut–brain axis, is via spinal afferent neurons, with cell bodies in dorsal root ganglia (DRG). Our aim was to identify the sensory nerve endings in the colon that arise from single colorectal-projecting DRG neurons. C57BL/6 mice were anesthetized and lumbosacral L6-S1 DRG injected with dextran biotin. Mice recovered for 7 days. The whole colon was then removed and stained to visualize single axons and nerve endings immunoreactive to calcitonin gene-related peptide (CGRP). Single axons arising from DRG were identified in the distal colon and their morphological features and CGRP immunoreactivity characterized. After entering the colon, single axons ramified rostrally or caudally along many rows of myenteric ganglia with little circumferential displacement, giving off varicose endings in multiple ganglia. Nerve endings arising from two classes of colorectal-projecting DRG neuron were identified. One class was peptidergic neurons that had nerve endings in circular muscle, myenteric ganglia, and submucosa. Another class of nonpeptidergic neurons innervated mucosal crypts, myenteric ganglia, and submucosa. Different morphological types of nerve endings which innervate different anatomical layers of colon can arise from the same axon and sensory neuron in DRG. These findings suggest single peptidergic and nonpeptidergic sensory neurons in DRG are potentially capable of detecting sensory stimuli from different anatomical layers of the colon, via different types of nerve endings.     

Mesencephalic trigeminal sensory neurons of cat: axon pathways and structure of mechanoreceptive endings in periodontal ligament

We injected 3H-proline into cat brainstem in order to label the entire mesencephalic trigeminal nucleus (Mes-V) for autoradiographic analysis of the size and pathways of Mes-V sensory axons and for microscopic study of Mes-V receptor structure in dental tissue. Labeled sensory axons were found in the trigeminal motor and sensory tracts and roots; approximately equal numbers of axons were found in both roots. The sensory root and all three divisions of the trigeminal nerve contained larger Mes-V axons than the motor root. Labeled Mes-V axons were found at the ganglion in the dorsomedial (infratrochlear) branch of the ophthalmic nerve but not in the ventrolateral branch. The mean diameter of Mes-V axons in periodontal ligament was 4.0 +/- 1.9 micron compared to 7.3 +/- 2.1 micron in maxillary and mandibular nerve, suggesting axonal arborization prior to innervation of ligament. Mes-V receptors in dental tissue were confined to ipsilateral periodontal ligament close to the root apex, with greater innervation on the posterior side. Receptor incidence was moderate for most teeth; however, maxillary first and second incisors and maxillary and mandibular canines had focal areas with remarkably dense innervation. No labeled axons were found in pulp of any ipsilateral teeth, and none was found in any contralateral dental tissue. EM-autoradiography demonstrated that Mes-V axons form unencapsulated Ruffini-like mechanoreceptors in periodontal ligament. The preterminal axons were small and myelinated. Neighboring bundles of unmyelinated axons and rare encapsulated endings were not labeled. The labeled mechanoreceptors branched to varying degrees among the ligament fibers; they contained numerous mitochondria and glycogen particles, as well as some vesicles and rare multivesicular bodies. They were surrounded by special Schwann cells that formed one or several layers around the ending. The endings were exposed to the basal lamina at numerous sites and occasionally extended fingers beyond the lamellar Schwann cells to contact ligament collagen.     

Regulation of cardiac innervation and function via the p75 neurotrophin receptor

Homeostatic regulation of cardiac function is dependent on the balance of inputs from the sympathetic and parasympathetic nervous systems. We investigated whether the p75 neurotrophin receptor plays a developmental role in cardiac innervation by analyzing sympathetic and parasympathetic fibers in the atria of p75 knockout and wildtype mice at several stages of postnatal development, and examining the effect on control of heart rate. We found that parasympathetic innervation of the atria in p75-/- mice was similar to wildtype at all time points, but that the density of sympathetic innervation was dynamically regulated. Compared to wildtype mice, the p75-/- mice had less innervation at postnatal day 4, an increase at day 28, and decreased innervation in adult mice. These changes reflect defects in initial fiber in-growth and the timing of the normal developmental decrease in sympathetic innervation density in the atria. Thus, p75 regulates both the growth and stability of cardiac sympathetic fibers. The distribution of sympathetic fibers was also altered, so that many regions lacked innervation. Basal heart rate was depressed in adult p75-/- mice, and these mice exhibited a diminished heart rate response to restraint stress. This resulted from the lack of sympathetic innervation rather than increased parasympathetic transmission or a direct effect of p75 in cardiac cells. Norepinephrine was elevated in p75-/- atria, but stimulating norepinephrine release with tyramine produced less tachycardia in p75-/- mice than wild type mice. This suggests that altered density and distribution of sympathetic fibers in p75-/- atria impairs the control of heart rate.