GluN2A- and GluN2B-immunoreactive type I cells attached to vesicular glutamate transporter 2-immunoreactive afferent nerve terminals of the rat carotid body.

We previously reported the immunoreactivity for the vesicular glutamate transporter 2 (VGLUT2) in afferent nerve terminals attached to chemoreceptor type I cells of the carotid body (CB), suggesting that glutamate is released from afferent terminals to stimulate these cells. In the present study, we examined the immunoreactivity for the glutamate-binding subunits of N-methyl-D-aspartate (NMDA) receptors, GluN2A and GluN2B in the rat CB, and the immunohistochemical relationships between these subunits and VGLUT2. Immunoreactivities for GluN2A and GluN2B were predominant in a subpopulation of tyrosine hydroxylase-immunoreactive type I cells rather than those of dopamine beta-hydroxylase-immunoreactive cells. Punctate VGLUT2-immunoreactive products were attached to GluN2A- and GluN2B-immunoreactive type I cells. Bassoon-immunoreactive products were localized between VGLUT2-immunoreactive puncta and type I cells immunoreactive for GluN2A and GluN2B. These results suggest that afferent nerve terminals release glutamate by exocytosis to modulate chemosensory activity of a subpopulation of type I cells via GluN2A- and GluN2B subunits-containing NMDA receptors.

Morphological and neurochemical characterisation of anterogradely labelled spinal sensory and autonomic nerve endings in the mouse bladder.

The bladder is innervated by axons of sympathetic and parasympathetic efferent nerves, and by spinal afferent neurons. The objective was to characterise anatomically and immunohistochemically the terminal endings of sensory and autonomic motor nerve endings in wholemount preparations of the mouse bladder. We used both anterograde labelling of pelvic and hypogastric nerves ex vivo and anterograde labelling from lumbosacral dorsal root ganglia (DRG) in vivo in male and female mice. These were combined with immunohistochemistry for major markers of sensory, sympathetic and parasympathetic nerves. Selective labelling of spinal afferent endings following dextran biotin-labelling from DRGs in vivo showed no co-localisation of VAChT or TH in sensory terminals in the detrusor and suburothelial plexus. Biotinamide was applied ex vivo to nerve trunks arising in the pelvic ganglion and running towards the bladder. Among the filled axons, 38% of detrusor fibres and 47% of suburothelial axons were immunoreactive for calcitonin-gene related peptide (CGRP). Vesicular acetylcholine transporter (VAChT) immunoreactivity was present in 26% of both detrusor and suburothelial axons. For tyrosine hydroxylase (TH), the proportions were 15% and 17%, respectively. Three major morphological types of CGRP-immunoreactive nerve endings were distinguished in the bladder wall: simple, branching and complex. VAChT-immunoreactive parasympathetic axons had simple and branching endings; TH immunoreactive axons all had simple morphologies. Our findings revealed that different subtypes of sensory and autonomic nerve endings can be reliably identified by combining anterograde labelling ex vivo with specific immunohistochemical markers, although morphologically some of these types of endings were indistinguishable.

Cranial Pair 0: The Nervus Terminalis.

Originally discovered in elasmobranchs by Fritsh in 1878, the nervus terminalis has been found in virtually all species, including humans. After more than one-century debate on its nomenclature, it is nowadays recognized as cranial pair zero. The nerve mostly originates in the olfactory placode, although neural crest contribution has been also proposed. Developmentally, the nervus terminalis is clearly observed in human embryos; subsequently, during the fetal period loses some of its ganglion cells, and it is less recognizable in adults. Fibers originating in the nasal cavity passes into the cranium through the middle area of the cribiform plate of the ethmoid bone. Intracranially, fibers joint the telencephalon at several sites including the olfactory trigone and the primordium of the hippocampus to reach preoptic and precommissural regions. The nervus terminalis shows ganglion cells, that sometimes form clusters, normally one or two located at the base of the crista galli, the so-called ganglion of the nervus terminalis. Its function is uncertain. It has been described that its fibers facilitates migration of luteinizing hormone-releasing hormone cells to the hypothalamus thus participating in the development of the hypothalamic-gonadal axis, which alteration may provoke Kallmann's syndrome in humans. This review summarizes current knowledge on this structure, incorporating original illustrations of the nerve at different developmental stages, and focuses on its anatomical and clinical relevance. Anat Rec, 302:394-404, 2019. © 2018 Wiley Periodicals, Inc.

Vagal nerve endings in visceral pleura and triangular ligaments of the rat lung.

The inner thoracic cavity is lined by the parietal pleura, and the lung lobes are covered by the visceral pleura. The parietal and visceral plurae form the pleural cavity that has negative pressure within to enable normal respiration. The lung tissues are bilaterally innervated by vagal and spinal nerves, including sensory and motor components. This complicated innervation pattern has made it difficult to discern the vagal vs. spinal processes in the pulmonary visceral pleura. With and without vagotomy, we identified vagal nerve fibres and endings distributed extensively in the visceral pleura ('P'-type nerve endings) and triangular ligaments ('L'-type nerve endings) by injecting wheat germ agglutinin-horseradish peroxidase as a tracer into the nucleus of solitary tract or nodose ganglion of male Sprague-Dawley rats. We found the hilar and non-hilar vagal pulmonary pleural innervation pathways. In the hilar pathway, vagal sub-branches enter the hilum and follow the pleural sheet to give off the terminal arborizations. In the non-hilar pathway, vagal sub-branches run caudally along the oesophagus and either directly enter the ventral-middle-mediastinal left lobe or follow the triangular ligaments to enter the left and inferior lobe. Both vagi innervate: (i) the superior, middle and accessory lobes on the ventral surfaces that face the heart; (ii) the dorsal-rostral superior lobe; (iii) the dorsal-caudal left lobe; and (iv) the left triangular ligament. Innervated only by the left vagus is: (i) the ventral-rostral and dorsal-rostral left lobe via the hilar pathway; (ii) the ventral-middle-mediastinal left lobe and the dorsal accessory lobe that face the left lobe via the non-hilar pathway; and (iii) the ventral-rostral inferior lobe that faces the heart. Innervated only by the right vagus, via the non-hilar pathway, is: (i) the inferior (ventral and dorsal) and left (ventral only) lobe in the area near the triangular ligament; (ii) the dorsal-middle-mediastinal left lobe; and (iii) the right triangular ligament. Other regions innervated with unknown vagal pathways include: (i) the middle lobe that faces the superior and inferior lobe; (ii) the rostral-mediastinal inferior lobe that faces the middle lobe; and (iii) the ventral accessory lobe that faces the diaphragm. Our study demonstrated that most areas that face the dorsal thoracic cavity have no vagal innervation, whereas the interlobar and heart-facing areas are bilaterally or unilaterally innervated with a left-rostral vs. right-caudal lateralized innervation pattern. This innervation pattern may account for the fact that the respiratory regulation in rats has a lateralized right-side dominant pattern.

Morphology of P2X3-immunoreactive nerve endings in the rat laryngeal mucosa.

The morphological characteristics of P2X3-immunoreactive nerve endings in the laryngeal mucosa were herein examined using immunohistochemistry with confocal laser microscopy. Ramified intraepithelial nerve endings immunoreactive to P2X3 were distributed in the epiglottis and arytenoid region. The axon terminals of P2X3-immunoreactive ramified endings were beaded or flat in shape. These endings were also immunoreactive to P2X2 and not identical to the nerve endings immunoreactive to Na(+)-K(+)-ATPase α3-subunit, substance P (SP), and calcitonin gene-related peptide (CGRP). P2X3-immunoreactive axon terminals were also immunoreactive to vGLUT1, vGLUT2, and vGLUT3. In addition to ramified endings, P2X3-immunoreactive nerve endings were associated with α-gustducin-immunoreactive solitary chemosensory cells and/or SNAP25-immunoreactive neuroendocrine cells. Furthermore, P2X3-immunoreactive nerve endings were also observed in the taste bud-like chemosensory cell clusters of the stratified squamous epithelium covering epiglottic and arytenoid cartilage. The P2X3-immunoreactive nerve endings that associated with sensory and/or endocrine cells and chemosensory cell clusters were also immunoreactive to P2X2, vGLUT1, vGLUT2, and vGLUT3, but not to SP or CGRP. In conclusion, P2X3-immunoreactive nerve endings may be classified into two types, i.e., intraepithelial ramified nerve endings and nerve endings associated with chemosensory cells and neuroendocrine cells.

AVP modulation of the vestibular nucleus via V1b receptors potentially contributes to the development of motion sickness in rat.

BACKGROUND: Arginine vasopressin (AVP) is considered to be an etiologic hormone in motion sickness (MS). The present study was designed to investigate whether individual differences in AVP expression in the paraventricular nucleus (PVN) and in modulation on the vestibular nucleus (VN) are involved in MS. Systemic application or microinjection of AVP into rat VN and rotatory stimulus were used to induce conditioned taste aversion (CTA) to 0.15 % saccharin sodium solution as a model of MS. RESULTS: Intra-VN use of SSR149415, an antagonist of V1b receptors (V1bRs), blunted CTA. AVP inhibited Ca(2+) influxes through L-type Ca(2+) channels and NMDA receptor channels in cultured VN neurones, but antagonised by SSR149415. More AVP and V1bRs were expressed respectively in the PVN and VN after rotatory stimulus, especially in rats susceptible to MS. In the VN, AVP content was low, the AVP mRNA was less expressed, a few AVP-positive fibres were sparsely distributed, and fewer AVP/synaptophysin-positive terminals were identified. Almost no fluoro-ruby-labelled AVP-positive neurones in the PVN were found with retrograde tracing from the VN. SNP analysis of the reported 9 sites of the AVP gene showed significant difference between the groups susceptible and insusceptible to MS at the site rs105235842 in the allele frequencies and genotypes. However, there was not any difference between these two groups in the SNP of the reported 38 sites of V1bR gene. CONCLUSIONS: AVP, through its modulatory, possibly humoral action on the VN neurones via the mediation of V1bR, may contribute to the development of motion sickness in rats; AVP gene polymorphisms may contribute to the individual difference in the responsive expression of AVP in the PVN; and higher expressions of AVP in the PVN and V1bRs in the VN may contribute to the development of motion sickness in rats after vestibular stimulation.

The innervation of the human acetabular labrum and hip joint: an anatomic study.

BACKGROUND: The aim of the current study was to evaluate the innervation of the acetabular labrum in the various zones and to understand its potential role in nociception and proprioception in hips with labral pathology. METHODS: A total of twenty hip labrums were tagged and excised intraoperatively from patients undergoing a total hip replacement. After preparation, the specimens were cut to a thickness of 10 µm and divided into four quadrants (zones) using a clock face pattern. Neurosensory structure distribution was then evaluated using Hematoxylin and Eosin (H and E), and immunoreactivity to S-100. RESULTS: All specimens had abundant free nerve endings (FNEs). These were seen predominantly superficially and on the chondral side of the labrum. In addition, predominantly three different types of nerve end organs (NEOs) were identified in all twenty specimens. FNEs and NEOs were more frequently seen in the antero-superior and postero-superior zones. Four specimens had abundant vascularity and disorganised architecture of FNEs in the deeper zones of the antero-superior quadrant suggestive of a healed tear. Myofibroblasts were present in abundance in all the labral specimens and were distributed uniformly throughout all labral zones and depth. CONCLUSIONS: The current study shows that the human acetabular labrum has abundant FNEs and NEOs. These are more abundant in the antero-superior and postero-superior zones. The labrum, by virtue of its neural innervation, can potentially mediate pain as well as proprioception of the hip joint, and be involved in neurosecretion that can influence connective tissue repair.

Epidermal expression of Lgr6 is dependent on nerve endings and Schwann cells.

Lgr5/6 proteins are stem cell markers in various tissues. However, what determines their restricted expression pattern in these tissues remains unknown. We found that in skin, Lgr6 is not only expressed in the central isthmus, directly above the hair follicle bulge cells as reported previously, but also in the interfollicular epidermis. Lgr6 expression in skin is highly correlated with the innervation sites of cutaneous nerves. In the hair follicle, Lgr6 closely localizes with the surrounding nerve endings and their corresponding Schwann cells throughout the entire hair cycle. Furthermore, ablation of cutaneous nerves leads to degeneration of Schwann cells and diminished expression of Lgr6. Our results demonstrate that the nerve endings/Schwann cells control Lgr6 expression in skin, implying that they play a role in regulation of skin epithelial cells.

Vinpocetine and α-tocopherol prevent the increase in DA and oxidative stress induced by 3-NPA in striatum isolated nerve endings.

Vinpocetine is a neuroprotective drug that exerts beneficial effects on neurological symptoms and cerebrovascular disease. 3-nitropropionic acid (3-NPA) is a toxin that irreversibly inhibits succinate dehydrogenase, the mitochondrial enzyme that acts in the electron transport chain at complex II. In previous studies in striatum-isolated nerve endings (synaptosomes), we found that vinpocetine decreased dopamine (DA) at expense of its main metabolite 3,4-dihydroxyphenylacetic acid (DOPAC), and that 3-NPA increased DA, reactive oxygen species (ROS), DA-quinone products formation, and decreased DOPAC. Therefore, in this study, the possible effect of vinpocetine on 3-NPA-induced increase in DA, ROS, lipid peroxidation, and DA-quinone products formation in striatum synaptosomes were investigated, and compared with the effects of the antioxidant α-tocopherol. Results show that the increase in DA induced by 3-NPA was inhibited by both 25 µM vinpocetine and 50 µM α-tocopherol. Vinpocetine, as α-tocopherol, also inhibited 3-NPA-induced increase in ROS (as judged by DCF fluorescence), lipid peroxidation (as judged by TBA-RS formation), and DA-quinone products formation (as judged by the nitroblue tetrazolium reduction method). As in addition to the inhibition of complex II exerted by 3-NPA, 3-NPA increases DA-oxidation products that in turn can inhibit other sites of the respiratory chain, the drop in DA produced by vinpocetine and α-tocopherol may importantly contribute to their protective action from oxidative damage, particularly in DA-rich structures.

Glutamatergic neurons in the lateral periaqueductal gray innervate neurokinin-1 receptor-expressing neurons in the ventrolateral medulla of the rat.

The neural pathways underlying the respiratory responses elicited by electrical or chemical stimulation of the lateral part of the periaqueductal gray (lPAG) remain unsettled. In the present study, we examined the lPAG projection to neurokinin-1 receptor (NK1R)-immunoreactive (ir) neurons in the ventrolateral medulla (VLM) which have been implicated in the control of respiration. After biotinylated dextranamine (BDA) injection into the lPAG, NK1R-ir neurons in the rostral VLM were embedded in the plexus of BDA-labeled fibers. At the electron microscopic level, the BDA-labeled terminals made asymmetrical synaptic contacts predominantly with dendrites and additionally with somata of the NK1R-ir neurons. Using retrograde tracing combined with in situ hybridization, we demonstrated that the vast majority of the lPAG neurons projecting to the rostral VLM were positive for vesicular glutamate transporter 2 (VGLUT2) mRNA, but not for glutamic acid decarboxylase 67 mRNA. Using a combination of anterograde tracing and immunohistochemistry, we further demonstrated that the lPAG axon terminals with VGLUT2 immunoreactivity made close apposition with the NK1R-ir neuronal profiles in the rostral VLM. These data suggest that lPAG neurons exert an excitatory influence on NK1R-expressing neurons in the rostral VLM for the control of respiration.

Dynamic longitudinal investigation of individual nerve endings in the skin of anesthetized mice using in vivo two-photon microscopy.

Visualization of individual cutaneous nerve endings has previously relied on laborious procedures of tissue excision, fixation, sectioning and staining for light or electron microscopy. We present a method for non-invasive, longitudinal two-photon microscopy of single nerve endings within the skin of anesthetized transgenic mice. Besides excellent signal-to-background ratio and nanometer-scale spatial resolution, this method offers time-lapse "movies" of pathophysiological changes in nerve fine structure over minutes, hours, days or weeks. Structure of keratinocytes and dermal matrix is visualized simultaneously with nerve endings, providing clear landmarks for longitudinal analysis. We further demonstrate feasibility of dissecting individual nerve fibers with infra-red laser and monitoring their degradation and regeneration. In summary, our excision-free optical biopsy technique is ideal for longitudinal microscopic analysis of animal skin and skin innervations in vivo and can be applied widely in preclinical models of chronic pain, allergies, skin cancers and a variety of dermatological disorders.

Abdominal vagal afferent pathways and their distributions of intraganglionic laminar endings in the rat duodenum.

We examined abdominal vagal afferents (n = 33) and the distributions of their intraganglionic laminar endings (IGLEs) in the duodenum. Rats (male, Sprague-Dawley) received a partial subdiaphragmatic vagotomy that spared a single branch. Wheat germ agglutinin-horseradish peroxidase (0.5-1.0 µl) was injected into the nodose ganglion ipsilateral to the vagotomized side. We observed that the hepatic branch does not project to the stomach, that the accessory celiac and celiac branches course along the celiac artery and innervate the intestines, and that the left nodose afferents innervate predominantly the duodenum. The hepatic branch innervates the duodenum via the "hepatoduodenal" subbranch and has the densest IGLE distribution in both the dorsoventral and the rostrocaudal extensions of the first 4-cm segment. Both gastric branches have two subbranches that innervate the duodenum; the "lesser curvature" subbranches follow the lesser curvature artery and may join the "hepatoduodenal" subbranch, whereas the "pyloric" subbranches run through the antrum and pylorus to reach the proximal duodenum. Moreover, the subbranches of ventral and dorsal gastric branches innervate more in the ventral and dorsal parts of the duodenum, respectively, and have more IGLEs in the rostral region than in the caudal. A posteriori comparisons indicate that, in the first-centimeter segment, the ventral gastric branch has significantly more IGLEs, whereas, in the third- and fourth-centimeter segments, the hepatic branch has more IGLEs. The finding that three different vagal branches innervate the duodenum with different densities of afferent endings might indicate a viscerotopic receptive field that coordinates digestive functions in feeding.

Mass transport at rotating disk electrodes: effects of synthetic particles and nerve endings.

An unstirred layer (USL) exists at the interface of solids with solutions. Thus, the particles in brain tissue preparations possess a USL as well as at the surface of a rotating disk electrode (RDE) used to measure chemical fluxes. Time constraints for observing biological kinetics based on estimated thicknesses of USLs at the membrane surface in real samples of nerve endings were estimated. Liposomes, silica, and Sephadex were used separately to model the tissue preparation particles. Within a solution stirred by the RDE, both diffusion and hydrodynamic boundary layers are formed. It was observed that the number and size of particles decreased the following: the apparent diffusion coefficient excluding Sephadex, boundary layer thicknesses excluding silica, sensitivity excluding diluted liposomes (in agreement with results from other laboratories), limiting current potentially due to an increase in the path distance, and mixing time. They have no effect on the detection limit (6 ± 2 nM). The RDE kinetically resolves transmembrane transport with a timing of approximately 30 ms.

Live imaging of bulk endocytosis in frog motor nerve terminals using FM dyes.

We observed endocytosis in real time in stimulated frog motor nerve terminals by imaging the growth of large membrane infoldings labeled with a low concentration of FM dye. The spatial and temporal information made available by these experiments allowed us to image several new aspects of this synaptic vesicle recycling pathway. Membrane infoldings appeared near synaptic vesicle clusters and grew rapidly during long-duration, high-frequency stimulation. In some cases, we observed large, elongated infoldings growing laterally into the terminal. We used these observations to calculate infolding growth rates. A decrease in stimulation frequency caused a decrease in growth rates, but the overall length of these structures was unaffected by frequency changes. Attempts to wash the dye from these infoldings after stimulation were unsuccessful, demonstrating that the fluorescent structures had been endocytosed. We also used this technique to trigger and image infoldings during repeated, short trains. We found that membrane uptake occurred repeatedly at individual endocytosis sites, but only during a portion of the total number of trains delivered to the terminal. Finally, we showed that phosphatidylinositol 3-kinase, but not actin, was involved in this endocytosis pathway. The ability to monitor many individual bulk endocytosis sites in real time should allow for new types of endocytosis measurements and could reveal novel and unexpected mechanisms for coordinating membrane recovery during synaptic activity.

Sodium-calcium exchanger and multiple sodium channel isoforms in intra-epidermal nerve terminals.

BACKGROUND: Nociception requires transduction and impulse electrogenesis in nerve fibers which innervate the body surface, including the skin. However, the molecular substrates for transduction and action potential initiation in nociceptors are incompletely understood. In this study, we examined the expression and distribution of Na+/Ca2+ exchanger (NCX) and voltage-gated sodium channel isoforms in intra-epidermal free nerve terminals. RESULTS: Small diameter DRG neurons exhibited robust NCX2, but not NCX1 or NCX3 immunolabeling, and virtually all PGP 9.5-positive intra-epidermal free nerve terminals displayed NCX2 immunoreactivity. Sodium channel NaV1.1 was not detectable in free nerve endings. In contrast, the majority of nerve terminals displayed detectable levels of expression of NaV1.6, NaV1.7, NaV1.8 and NaV1.9. Sodium channel immunoreactivity in the free nerve endings extended from the dermal boundary to the terminal tip. A similar pattern of NCX and sodium channel immunolabeling was observed in DRG neurons in vitro. CONCLUSIONS: NCX2, as well as NaV1.6, NaV1.7, NaV1.8 and NaV1.9, are present in most intra-epidermal free nerve endings. The presence of NCX2, together with multiple sodium channel isoforms, in free nerve endings may have important functional implications.

Origins of endomorphin-2 immunopositive fibers and terminals in the spinal dorsal horn of the rat.

Endomorphin 2 (EM2) plays essential roles in regulating nociceptive transmission within the spinal dorsal horn, where EM2-immunopositive (EM2-IP) fibers and terminals are densely encountered. However, the origins of these EM2-IP structures are still obscure. Unilateral primary sensory afferents disruption (lumbar 3-6 dorsal roots rhizotomy) significantly decreased the density of EM2-IP fibers and terminals in the superficial laminae (laminae I and II) on the ipsilateral but not contralateral lumbar dorsal horn (LDH). Spinal hemisection at the 7th thoracic (T7) segment down-regulated bilateral EM2 expression, with a higher influence on the ipsilateral side of the LDH. Unilateral L3-6 dorsal roots rhizotomy combined with spinal transection but not with hemisection at T7 level completely obliterated EM2-IP fibers and terminals on the rhizotomized-side of the LDH. Disruption of bilateral (exposure to the primary afferent neurotoxin, capsaicin) primary sensory afferents combined with spinal hemisection at T7 decreased the EM2-IP density bilaterally but could obliterate it on neither side of the LDH. While in capsaicin plus transection rats, EM2 was depleted symmetrically and completely. In the colchicine treated rats, no EM2-IP neuronal cell bodies could be detected in the spinal gray matter. After injecting tetramethyl rhodamine dextran-amine (TMR) into the LDH, some of the TMR retrogradely labeled neurons in the nucleus tractus solitarii (NTS) showed EM2-immunoreactivities. The present results indicate that EM2-IP fibers and terminals in the spinal dorsal horn originate from the ipsilateral primary afferents and bilateral descending fibers from NTS.

Immunohistochemical demonstration of nerve endings in iliolumbar ligament.

STUDY DESIGN: Immunohistochemical study on fresh cadaver specimens. OBJECTIVE: Assessment of mechanoreceptor and nociceptor levels and distribution in iliolumbar ligament. SUMMARY AND BACKGROUND DATA: The function of iliolumbar ligament and its role in low back pain has not been yet fully clarified. Understanding the innervation of this ligament should provide a ground which enables formation of stronger hypotheses. METHODS: Bilateral 30 iliolumbar ligaments of 15 fresh cadavers were included in the study. Morphologic properties were recorded and the ligaments were examined by focusing on 3 main parts: ligament, bone insertions, and tendon body. Assessment of mechanoreceptor and nociceptor levels and their distribution in iliolumbar ligament were performed on the basis of immunohistochemistry using the S-100 antibody specific for nerve tissue. RESULTS: Iliac wing insertion was found to be the richest region of the ligament in terms of mechanoreceptors and nociceptors. Pacinian (type II) mechanoreceptor was determined to be the most common (66.67%) receptor followed by Ruffini (type I) (19.67%) mechanoreceptor, whereas free nerve endings (type IV) and Golgi tendon organs (type III) were found to be less common, 10.83% and 2.83%, respectively. CONCLUSION: Immunohistochemical staining has shown that iliolumbar ligament had a rich nerve tissue. Those results indicate that ILL plays an important role in proprioceptive coordination of lumbosacral region alongside its known biomechanic support function. Moreover, the presence of type IV nerve endings suggest that the injury of this ligament might contribute to the low back pain.

Distribution of TRPV1- and TRPV2-immunoreactive afferent nerve endings in rat trachea.

Nociception in the trachea is important for respiratory modulation. We investigated the distribution, neurochemical characteristics, and origin of nerve endings with immunoreactivity for candidate sensor channels, TRPV1 and TRPV2, in rat trachea. In the epithelial layer, the intraepithelial nerve endings and dense subepithelial network of nerve fibers were immunoreactive for TRPV1. In contrast, TRPV2 immunoreactivity was observed mainly in nerve fibers of the tracheal submucosal layer and in several intrinsic ganglion cells in the peritracheal plexus. Double immunostaining revealed that some TRPV1-immunoreactive nerve fibers were also immunoreactive for substance P or calcitonin gene-related peptide, but neither neuropeptide colocalized with TRPV2. Injection of the retrograde tracer, fast blue, into the tracheal wall near the thoracic inlet demonstrated labeled neurons in the jugular, nodose, and dorsal root ganglia at segmental levels of C2-C8. In the jugular and nodose ganglia, 59.3% (70/118) and 10.7% (17/159), respectively, of fast blue-labeled neurons were immunoreactive for TRPV1, compared to 8.8% (8/91) and 2.6% (5/191) for TRPV2-immunoreactive. Our results indicate that TRPV1-immunoreactive nerve endings are important for tracheal nociception, and the different expression patterns of TRPV1 and TRPV2 with neuropeptides may reflect different subpopulations of sensory neurons.

Innervation of the triangular fibrocartilage complex of the human wrist: quantitative immunohistochemical study.

The distribution of neural elements in the triangular fibrocartilage complex (TFCC) of the human wrists was studied via immunohistochemical staining of protein gene product (PGP) 9.5 and calcitonin gene-related peptide (CGRP). Articular branches projecting to the TFCC arose from the dorsal branch of the ulnar nerve in all wrists examined. The TFCC is subdivided into the following six regions: the articular disc proper (ADP), meniscus homolog (MH), radio-ulnar ligament (RUL), loose part of ulnar collateral ligament (lUCL), dense part of ulnar collateral ligament (dUCL), and internal portion (IP). The IP consists of a mixture of dense and loose connective tissues enclosed by the ADP, MH, RUL, and UCL, and resides deep in the prestyloid recess, which is a pit in the MH. The densities of PGP 9.5-positive neural elements, including free nerve endings, single nerve fibers, nerve fascicles, and perivascular neural nets, were significantly higher in the IP than in other regions. Some of the neural elements except for the perivascular neural nets were positive for CGRP. The high density of neural elements in the IP suggests that sensory nerves projecting to the TFCC enter into the IP and from there distribute to adjacent regions such as the MH and RUL. Free nerve endings are responsible for pain transmission. The high density of free nerve endings in the IP suggests that the IP is a source of ulnar side wrist pain.

Neurogenic factors in the impaired healing of diabetic foot ulcers.

BACKGROUND: We hypothesize that the reduced innervation of skin can be observed both in clinically neuropathic and non-neuropathic diabetic foot ulcers and can contribute to low inflammatory cell infiltration. MATERIALS AND METHODS: Twenty patients with type 2 diabetes and active foot ulcers, without clinical evidence of peripheral sensory neuropathy (n = 12) and with sensory neuropathy (n = 8) were involved in this study. Biopsies from ulcer margin were examined immunohistochemically. RESULTS: Studies revealed presence of protein gene product 9.5 (PGP9.5)+ nerve endings only in reticular dermis in 3 of 12 non-neuropathic subjects, however, regenerating GAP-43+ endings were seen in dermis of almost all specimens. Lack of substance P+ nerve endings was characteristic for both groups. The reduced distribution of calcitonin gene-related peptide+ nerves in epidermis and dermis was seen mainly in neuropathic group. In neo-epidermis lack of nerve growth factor expression was observed in both groups, whereas neurotrophin 3 immunostaining was characteristic for neuropathic specimens (P < 0.03). Expression of trkA and trkC receptors did not differ significantly between groups. Low inflammatory cell infiltration and moderate presence of fibroblasts was characteristic for all studied specimens. CONCLUSIONS: The observed reduction of foot skin innervation and neurogenic factors expression can be correlated with low inflammatory cell accumulation and subsequently leads to the observed chronicity of diabetic foot ulcer healing process in both neuropathic and non-neuropathic patients.