Ansa Cervicalis Stimulation: A New Direction in Neurostimulation for OSA.

BACKGROUND: Hypoglossal nerve stimulation (HNS) is an alternative treatment option for patients with OSA unable to tolerate positive airway pressure but implant criteria limit treatment candidacy. Previous research indicates that caudal tracheal traction plays an important role in stabilizing upper airway patency. RESEARCH QUESTION: Does contraction of the sternothyroid muscle with ansa cervicalis stimulation (ACS), which pulls the pharynx caudally via thyroid cartilage insertions, increase maximum inspiratory airflow (VImax)? STUDY DESIGN AND METHODS: Hook-wire percutaneous electrodes were used to stimulate the medial branch of the right hypoglossal nerve and right branch of the ansa cervicalis innervating the sternothyroid muscle during propofol sedation. VImax was assessed during flow-limited inspiration with a pneumotachometer. RESULTS: Eight participants with OSA were studied using ACS with and without HNS. Compared with baseline, the mean VImax increase with isolated ACS was 298%, or 473 mL/s (95% CI, 407-539). Isolated HNS increased mean VImax from baseline by 285%, or 260 mL/s (95% CI, 216-303). Adding ACS to HNS during flow-limited inspiration increased mean VImax by 151%, or 205 mL/s (95% CI, 174-236) over isolated HNS. Stimulation was significantly associated with increase in VImax in both experiments (P < .001). INTERPRETATION: ACS independently increased VImax during propofol sedation and drove further increases in VImax when combined with HNS. The branch of the ansa cervicalis innervating the sternothyroid muscle is easily accessed. Confirmation of the ansa cervicalis as a viable neurostimulation target may enable caudal pharyngeal traction as a novel respiratory neurostimulation strategy for treating OSA.

Functional involvement of acid-sensing ion channel 3 in the swallowing reflex in rats.

BACKGROUND: Difficulty swallowing represents a major health problem. Swallowing function is improved by incorporating weak acids in suspensions/food boluses, implicating acid-sensing ion channels (ASICs) in the swallowing reflex. However, the functional involvement of ASICs in the swallowing reflex has not been fully elucidated. METHODS: We localized ASIC3s in swallowing-related regions innervated by the superior laryngeal nerves (SLNs) and those in the nodose-petrosal-jugular ganglionic complex (NPJc) and examined their functional involvement in evoking the swallowing reflex in rats. KEY RESULTS: We localized ASIC3s on epithelial cells and nerve fibers in swallowing-related regions innervated by the SLNs. In the NPJc, around half of the SLN-afferent neurons expressed ASIC3. Two-thirds of ASIC3s were localized on unmyelinated neurons in the nodose and petrosal ganglia. In the jugular ganglia, they were equally distributed on unmyelinated and myelinated neurons. Topical application of a synthetic non-proton ASIC3 activator, 2-guanidine-4-methylquinazoline (GMQ), and its natural endogenous ligand agmatine (a metabolite of the amino acid arginine) in swallowing-related regions evoked a considerable number of swallowing reflexes. Increasing the concentration of GMQ and agmatine up to a certain concentration increased the number of evoked reflexes and shortened the interval between the evoked reflexes. Agmatine was less potent than GMQ in its ability to evoke swallowing reflexes. Prior topical application of an ASIC3 antagonist significantly attenuated the number of GMQ- and agmatine-evoked swallowing reflexes. CONCLUSIONS & INFERENCES: Acid-sensing ion channel 3s localized on nerves and epithelial cells in swallowing-related regions are functional in evoking the swallowing reflex and activation of these channels via a pharmacological agonist appears to improve swallowing behavior.

Upper GIT Endoscopic Evaluation and Psychological State Assessment of Patients with Globus Sensation.

Globus sensation is a subjective feeling of a lump or foreign body in the throat without interfering swallowing of food. It is a persistent and distressing sensation in throat. It affects about 6% of population. But cause of globus is still unknown. Exact aetiology of globus is considered to be multifactorial. Some other studies also show association between globus and psychological distress including anxiety and depression. As there is no established pharmacological treatment, adequate investigations with negative result could reassure patients and improve their symptoms. In this prospective study consecutive patients with globus symptoms examined by upper GIT endoscopy with attention to larynx, epiglottis, base of tongue, both pyriform fossa and hypo-pharynx using Olympus forward viewing video Gastroscope (GIF Q-150 & GIF Q-170) to exclude organic lesion and was conducted in the department of Gastroenterology, Bangabandhu Sheikh Mujib Medical University (BSMMU) and North East Medical College, Sylhet from 1st July 2014 to 31 December 2016. Their psychological status and epidemiological information including personal and family history were noted in a pre-designed data sheet. Total 104 patients were examined, among them definite anxiety was found in 36(34.95%) and borderline feature of anxiety was found in 19(18.44%) and 48(46.60%) were free of anxiety. Incidence of anxiety was significantly higher among females and was more prevalent among housewife, married people and people from rural community. In this series, 13(12.5%) patients had definite depression and 29(27.9%) patients had borderline depression, while 61(59.2%) patients had no feature of depression. Incidence of depression was significantly higher among females, housewife and married people. Organic lesion is rare in patients with globus symptoms. Globus sensation is more common among females. Psychological factors like anxiety and depression are frequently associated with globus sensation.

Lateral pharyngotomy.

Based on a review of the medical literature, the authors document the key technical points, variants, technical errors to avoid and main functional results of lateral pharyngotomy for resection of cancers originating from the lateral oro and/or hypopharynx.

Axon and Schwann Cell Degeneration in Nerves of Upper Airway Relates to Pharyngeal Dysfunction in Snorers and Patients With Sleep Apnea.

BACKGROUND: The pathophysiologic mechanism of nocturnal obstruction and swallowing dysfunction commonly occurring in patients with sleep apnea is unclear. The goal of this study was to investigate whether nerve injuries in the upper airways of snorers and patients with sleep apnea are associated with pharyngeal dysfunction and severity of sleep apnea. METHODS: Twenty-two patients undergoing palatal surgery due to snoring and sleep apnea were investigated for a swallowing dysfunction by using videoradiography. Twelve healthy nonsnoring subjects were included as control subjects. Tissue samples from the soft palate at the base of the uvula were obtained in all patients and control subjects. Nerves and muscle were analyzed with immunohistochemical and morphologic methods, and the findings were correlated with swallowing function and degree of sleep apnea. RESULTS: In the soft palate of patients, nerve fascicles exhibited a significantly lower density of axons (5.4 vs 17.9 × 10-3 axons/µm2; P = .02), a smaller percentage area occupied by Schwann cells (17.5% vs 45.2%; P = .001) and a larger number of circular shaped Schwann cells lacking central axons (43.0% vs 12.7%; P < 0.001) compared with control subjects. The low density of axons was significantly related to degree of swallowing dysfunction (r = 0.5; P = .03) and apnea-hypopnea index > 5 (P = .03). Regenerating axons were frequently observed in patients compared with control subjects (11.3 ± 4.2% vs 4.8 ± 2.4%; P = .02). CONCLUSIONS: Axon degeneration in preterminal nerves of the soft palate is associated with pharyngeal dysfunction in snorers and patients with sleep apnea. The most likely cause for the nerve injuries is traumatic snoring vibrations and tissue stretch, leading to swallowing dysfunction and increased risk for upper airway obstruction during sleep.

Pharyngolaryngeal Sensory Deficits in Patients with Middle Cerebral Artery Infarction: Lateralization and Relation to Overall Dysphagia Severity.

BACKGROUND: Dysphagia is a frequent and dangerous complication of acute stroke. Apart from a well-timed oropharyngeal muscular contraction pattern, sensory feedback is of utmost importance for safe and efficient swallowing. In the present study, we therefore analyzed the relation between pharyngolaryngeal sensory deficits and post-stroke dysphagia (PSD) severity in a cohort of acute stroke patients with middle cerebral artery (MCA) infarction. METHODS: Eighty-four first-ever MCA stroke patients (41 left, 43 right) were included in this trial. In all patients, fiberoptic endoscopic evaluation of swallowing (FEES) was performed according to a standardized protocol within 96 h after stroke onset. PSD was classified according to the 6-point fiberoptic endoscopic dysphagia severity scale. Pharyngolaryngeal sensation was semi-quantitatively evaluated by a FEES-based touch technique. RESULTS: PSD severity was closely related to the pharyngolaryngeal sensory deficit. With regards to lateralization of the sensory deficit, there was a slight but significant preponderance of sensory loss contralateral to the side of stroke. Apart from that, right hemispheric stroke patients were found to present with a more severe PSD. CONCLUSIONS: This study provides evidence that an intact sensory feedback is of utmost importance to perform nonimpaired swallowing and highlights the key role of disturbed pharyngeal and laryngeal afferents in the pathophysiology of PSD.

Characterization of NvLWamide-like neurons reveals stereotypy in Nematostella nerve net development.

The organization of cnidarian nerve nets is traditionally described as diffuse with randomly arranged neurites that show minimal reproducibility between animals. However, most observations of nerve nets are conducted using cross-reactive antibodies that broadly label neurons, which potentially masks stereotyped patterns produced by individual neuronal subtypes. Additionally, many cnidarians species have overt structures such as a nerve ring, suggesting higher levels of organization and stereotypy exist, but mechanisms that generated that stereotypy are unknown. We previously demonstrated that NvLWamide-like is expressed in a small subset of the Nematostella nerve net and speculated that observing a few neurons within the developing nerve net would provide a better indication of potential stereotypy. Here we document NvLWamide-like expression more systematically. NvLWamide-like is initially expressed in the typical neurogenic salt and pepper pattern within the ectoderm at the gastrula stage, and expression expands to include endodermal salt and pepper expression at the planula larval stage. Expression persists in both ectoderm and endoderm in adults. We characterized our NvLWamide-like::mCherry transgenic reporter line to visualize neural architecture and found that NvLWamide-like is expressed in six neural subtypes identifiable by neural morphology and location. Upon completing development the numbers of neurons in each neural subtype are minimally variable between animals and the projection patterns of each subtype are consistent. Furthermore, between the juvenile polyp and adult stages the number of neurons for each subtype increases. We conclude that development of the Nematostella nerve net is stereotyped between individuals. Our data also imply that one aspect of generating adult cnidarian nervous systems is to modify the basic structural architecture generated in the juvenile by increasing neural number proportionally with size.

[Progress of dopamine in obstructive sleep apnea hypopnea syndrome].

The etiology and pathogenesis of sleep obstructive apnea hypopnea syndrome (OSAHS) is not yet definitive, evidence shows that the dysfunction of pharyngeal nerve and the atonia of the muscle innervated by these nerve could play an important role in the progress of OSAHS. Dopamine is a neurotransmitter in the central nervous system which significantly affects the sleep-awake regulation. So far mounting evidence shows that dopamine has a potential role in the modulation of hypoglossal nucleus. The progress of dopamine in obstructive sleep apnea hypopnea syndrome is reviewed in this article.

Chemogenetic stimulation of the hypoglossal neurons improves upper airway patency.

Obstructive sleep apnea (OSA) is characterized by recurrent upper airway obstruction during sleep. OSA leads to high cardiovascular morbidity and mortality. The pathogenesis of OSA has been linked to a defect in neuromuscular control of the pharynx. There is no effective pharmacotherapy for OSA. The objective of this study was to determine whether upper airway patency can be improved using chemogenetic approach by deploying designer receptors exclusively activated by designer drug (DREADD) in the hypoglossal motorneurons. DREADD (rAAV5-hSyn-hM3(Gq)-mCherry) and control virus (rAAV5-hSyn-EGFP) were stereotactically administered to the hypoglossal nucleus of C57BL/6J mice. In 6-8 weeks genioglossus EMG and dynamic MRI of the upper airway were performed before and after administration of the DREADD ligand clozapine-N-oxide (CNO) or vehicle (saline). In DREADD-treated mice, CNO activated the genioglossus muscle and markedly dilated the pharynx, whereas saline had no effect. Control virus treated mice showed no effect of CNO. Our results suggest that chemogenetic approach can be considered as a treatment option for OSA and other motorneuron disorders.

Denervation of the Tensor Veli Palatini Muscle and Effusion in the Tympanic Cavity.

An English springer spaniel was presented for right-sided atrophy of the muscles of mastication, analgesia and paralysis of the face, and vestibular dysfunction. Neurological signs were consistent with a lesion involving the pons and rostral medulla resulting in deficits in the function of the trigeminal, facial, and vestibular nerves. MRI disclosed a right-sided extraparenchymal mass consistent with a trigeminal nerve sheath neoplasm that was compressing and invading the pons and medulla. Atrophy of the muscles of mastication, innervated by the trigeminal nerve, was also observed on MRI. Additionally, effusion was present in the ipsilateral tympanic cavity. Gross and microscopic evaluation of the right tensor veli palatini muscle (TVPM) was consistent with neurogenic atrophy. Effusion in the tympanic cavity was likely the result of an inability to open the auditory tube as a consequence of paralysis of the TVPM. Without the ability to open the auditory tube, gases present within the auditory tube and tympanic cavity may be absorbed, creating a negative pressure environment that leads to fluid transudation and effusion build up. To the authors' knowledge, this is the first report to document neurogenic atrophy of the TVPM with concurrent effusion in the ipsilateral tympanic cavity.

Epiglottic cyst as an etiological factor of globus sensation.

BACKGROUND/AIMS: Globus is a subjective complaint that describes a sensation of a lump or a foreign body in the throat. Despite being a well-known and common clinical condition, the etiological factors have not been definitely elucidated yet. The study was set up to ascertain the relationship between epiglottic cysts and globus sensation. MATERIALS AND METHODS: All patients undergoing investigation and treatments for globus sensation were included in the study. Patients with epiglottic cysts but no other possible causes of globus sensation were constituted the series of patients. Patients were asked to assess the levels of complaint before and after the carbon dioxide (CO2) laser excisions of the cysts. RESULTS: Epiglottic cysts were found in 10 (5.4%) of the 182 patients. Three of these 10 patients who had concomitant diseases or conditions that may cause globus sensation and one patient who refused the surgery were excluded from the study. All the remaining six patients reported relief of the globus sensation after the CO2 laser excisions of the cysts. CONCLUSION: Our results, obtained from this limited series, indicated that epiglottic cysts may be considered as one of the etiological factors of globus sensation.

Mechanisms of cough provocation and cough resolution in neonates with bronchopulmonary dysplasia.

BACKGROUND: Cough and deglutition are protective mechanisms that defend against aspiration. We identified mechanisms associated with cough provocation as well as those associated with cough resolution in infants with bronchopulmonary dysplasia (BPD). METHODS: Manometry signatures of cough were recognized in 16 premature infants with BPD undergoing concurrent esophageal manometry, respiratory inductance plethysmography, and nasal air flow measurements. Pretussive and post-tussive pharyngo-esophageal motility changes were analyzed. Mechanisms associated with cough and mechanisms that restored respiratory and esophageal normalcy were analyzed. RESULTS: We analyzed 312 cough events during 88 cough clusters; 97% were associated with recognizable manometric patterns. Initial mechanisms related with coughing included nonpropagating swallow (59%), upper esophageal sphincter (UES) reflex contraction (18%), and lower esophageal sphincter (LES) relaxation (14%). UES and LES dysfunction was present in 69% of nonpropagating swallow-associated cough clusters. Mechanisms restoring post-tussive normalcy included primary peristalsis (84%), secondary peristalsis (8%), and none recognized (8%). UES contraction reflex was associated with cough clusters more frequently in infants on nasal continuous positive airway pressure (NCPAP) (OR = 9.13, 95% CI = 1.88-44.24). CONCLUSION: Cough clusters in infants with BPD had identifiable etiologies associated with esophageal events; common initial mechanisms were of upper aerodigestive origin, while common clearing mechanisms were peristaltic reflexes.

A Perforating Artery Compressing the Nerve Rootlet and Causing Glossopharyngeal Neuralgia.

BACKGROUND: A surgical procedure for glossopharyngeal neuralgia (GPN) was selected from microvascular decompression, glossopharyngeal and upper vagal rhizotomy, or a combination of these procedures based on the presence of arteries compressing the glossopharyngeal and vagal rootlets. The offending artery is usually a main trunk or branch of the cerebellar arteries. A perforating artery is a known but uncommon variation of the offending artery that causes GPN. The appropriate procedure for such cases is unknown. OBJECTIVE: To analyze the clinical significance of the perforating artery in GPN, we describe 2 patients with a perforating artery compressing the rootlet, and its mobilization relieved neuralgia. We examined the validity of decompressing a perforating artery as an alternative to rhizotomy in such cases. METHODS: We independently reviewed 12 GPN patients treated with microvascular decompression. The patients' pain severity, medication doses, preoperative imaging studies, intraoperative findings, and outcomes were examined. RESULTS: Eleven patients had neurovascular compression of the glossopharyngeal nerve. In 2 of the patients, a perforating artery compressed the rootlet, thereby generating an indentation and creating a discoloration of the rootlet. Mobilizing the perforating artery with no additional rhizotomy provided complete pain relief with no significant complications and allowed the discontinuation of medications. CONCLUSION: Even a small perforating artery can cause GPN when it compresses the rootlet. In such cases, mobilization of the perforating artery with no additional rhizotomy is an effective surgical option.

[Upper airway stimulation in obstructive sleep apnea]. / Stellungnahme der Taskforce "Neurostimulation bei Schlafapnoe" zur Stimulation der oberen Atemwege.

The stimulation of the upper airway represents an effective treatment option in case of CPAP failure in patients with moderate to severe obstructive sleep apnea. The stimulation with respiratory sensing (Inspire Medical Systems) has shown a high level of evidence in larger cohorts and longer follow-up studies. Whether the results of the stimulation without respiratory sensing (ImThera Medical) can be compared with the therapy with sensing, remains open up to now. Additional data are awaited after the planned phase III study THN#2. To optimize both procedure and to provide long term results, more studies are needed. The workgroup "sleep medicine" of the German Society of Oto-Rhino-Laryngology, Head and Neck Surgery supports theses activities with the help of the newly founded task force "Neurostimulation in Sleep Apnea".

Effects of targeted activation of tongue muscles on oropharyngeal patency in the rat.

Laboratory rats were acutely implanted with an electrode array composed of eight independently controllable contacts applied to ventral and dorsal aspects of the left and right hypoglossal nerves (HGNs) and their branches. Bipolar intramuscular electromyographic (EMG) electrodes were implanted into the left and right genioglossus, hyoglossus and styloglossus muscles to identify which muscles were activated during stimulation via the contacts. Elicited movements, including changes in the position of the tongue and in the size and the shape of the airway, were documented video-graphically through a surgery microscope and an endoscope. Constant current electrical stimulation activated various combinations of electrode contacts and the stimulation patterns were correlated with corresponding oral movements, airway sizes, and EMG activities. Results demonstrate that graded responses and differential activation of the various tongue muscles are achievable by stimulation of specific contacts in the electrode array. These effects are interpreted to result from the targeted activation of regions of the nerve lying under and between the electrodes. Further testing established that the muscle responses elicited by unilateral electrical stimulation with the present approach can be smoothly graded, that the muscle responses resulted in opening of the airway and could be reliably maintained for long durations.

Laryngeal sensation and pharyngeal delay time after (chemo)radiotherapy.

The objective of the study was to evaluate the association between changes in laryngeal sensation and initiation of swallowing reflex or swallowing function before and after (chemo)radiotherapy. A prospective study was conducted in a tertiary referral university hospital. Thirteen patients who received (chemo)radiotherapy for treatment of laryngeal or hypopharyngeal cancer were included. Laryngeal sensation was evaluated at the tip of the epiglottis before and 1, 3 months, and 1 year after (chemo)radiotherapy. Videofluoroscopy was performed at the same time. Quantitative determinations included changes in laryngeal sensation, computed analysis of pharyngeal delay time, the distance and velocity of hyoid bone movement during the phase of hyoid excursion, and pharyngeal residue rate (the proportion of the bolus that was left as residue in the pharynx at the first swallow). Laryngeal sensation significantly deteriorated 1 month after (chemo)radiotherapy, but there was a tendency to return to pretreatment levels 1 year after treatment. Neither pharyngeal delay time nor displacement of the hyoid bone changed significantly before and after (chemo)radiotherapy. In addition, there was no significant difference in the mean velocity of hyoid bone movement and the amount of stasis in the pharynx at the first swallow before and after (chemo)radiotherapy. After (chemo)radiotherapy, laryngeal sensation deteriorated. But, in this study, videofluoroscopy showed that swallowing reflex and function were maintained.

In vivo recordings of GnRH neuron firing reveal heterogeneity and dependence upon GABAA receptor signaling.

The gonadotropin-releasing hormone (GnRH) neurons are the key cells regulating fertility in all mammalian species. The scattered distribution of these neurons has made investigation of their properties extremely difficult and the key goal of recording their electrical activity in vivo near impossible. The caudal-most extension of the GnRH neuron continuum brings some cells very close to the base of the brain at the level of the anterior hypothalamic area. Taking insight from this, we developed an experimental procedure in anesthetized GnRH-GFP mice that allows the electrical activity of these GnRH neurons to be recorded in vivo. On-cell recordings revealed that the majority of GnRH neurons (86%) were spontaneously active, exhibiting a range of firing patterns, although only a minority (15%) exhibited burst firing. Mean firing frequencies ranged from 0.06 to 3.65 Hz, with the most common interspike interval being ~500 ms. All GnRH neurons tested were activated by AMPA and kisspeptin. Whereas the GABAA receptor agonist muscimol evoked excitatory, inhibitory, or mixed effects on GnRH neuron firing, the GABAA receptor antagonist picrotoxin resulted in a consistent suppression of firing. These observations represent the first electrical recordings of GnRH neurons in vivo. They reveal that GnRH neurons in vivo exhibit considerable heterogeneity in their firing patterns with both similarities and differences to firing in vitro. These variable patterns of firing in vivo are found to be critically dependent upon ongoing GABAA receptor signaling.

Hes1 is required for the development of pharyngeal organs and survival of neural crest-derived mesenchymal cells in pharyngeal arches.

Hes genes are required to maintain diverse progenitor cell populations during embryonic development. Loss of Hes1 results in a spectrum of malformations of pharyngeal endoderm-derived organs, including the ultimobranchial body (progenitor of C cells), parathyroid, thymus and thyroid glands, together with highly penetrant C-cell aplasia (81%) and parathyroid aplasia (28%). The hypoplastic parathyroid and thymus are mostly located around the pharyngeal cavity, even at embryonic day (E) 15.5 to E18.5, indicating the failure of migration of the organs. To clarify the relationship between these phenotypes and neural crest cells, we examine fate mapping of neural crest cells colonized in pharyngeal arches in Hes1 null mutants by using the Wnt1-Cre/R26R reporter system. In null mutants, the number of neural crest cells labeled by X-gal staining is markedly decreased in the pharyngeal mesenchyme at E12.5 when the primordia of the thymus, parathyroid and ultimobranchial body migrate toward their destinations. Furthermore, phospho-Histone-H3-positive proliferating cells are reduced in number in the pharyngeal mesenchyme at this stage. Our data indicate that the development of pharyngeal organs and survival of neural-crest-derived mesenchyme in pharyngeal arches are critically dependent on Hes1. We propose that the defective survival of neural-crest-derived mesenchymal cells in pharyngeal arches directly or indirectly leads to deficiencies of pharyngeal organs.

AMP-activated kinase links serotonergic signaling to glutamate release for regulation of feeding behavior in C. elegans.

Serotonergic regulation of feeding behavior has been studied intensively, both for an understanding of the basic neurocircuitry of energy balance in various organisms and as a therapeutic target for human obesity. However, its underlying molecular mechanisms remain poorly understood. Here, we show that neural serotonin signaling in C. elegans modulates feeding behavior through inhibition of AMP-activated kinase (AMPK) in interneurons expressing the C. elegans counterpart of human SIM1, a transcription factor associated with obesity. In turn, glutamatergic signaling links these interneurons to pharyngeal neurons implicated in feeding behavior. We show that AMPK-mediated regulation of glutamatergic release is conserved in rat hippocampal neurons. These findings reveal cellular and molecular mediators of serotonergic signaling.

Pituitary adenylatecyclase-activating polypeptide-immunoreactive nerve fibers in the rat epiglottis and pharynx.

The distribution of pituitary adenylatecyclase-activating polypeptide-immunoreactive (PACAP-IR) nerve fibers was studied in the rat epiglottis and pharynx. PACAP-IR nerve fibers were located beneath the mucous epithelium, and occasionally penetrated the epithelium. These nerve fibers were abundant on the laryngeal side of the epiglottis and in the dorsal and lateral border region between naso-oral and laryngeal parts of the pharynx. PACAP-IR nerve fibers were also detected in taste buds within the epiglottis and pharynx. In addition, many PACAP-IR nerve fibers were found around acinar cells and blood vessels. The double immunofluorescence method demonstrated that distribution of PACAP-IR nerve fibers was similar to that in CGRP-IR nerve fibers in the epithelium and taste bud. However, distributions of PACAP-IR and CGRP-IR nerve fibers innervating mucous glands and blood vessels were different. The retrograde tracing method also demonstrated that PACAP and CGRP were co-expressed by vagal and glossopharyngeal sensory neurons innervating the pharynx. These findings suggest that PACAP-IR nerve fibers in the epithelium and taste bud of the epiglottis and pharynx which originate from the vagal and glossopharyngeal sensory ganglia include nociceptors and chemoreceptors. The origin of PACAP-IR nerve fibers which innervate mucous glands and blood vessels may be the autonomic ganglion.