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Keywords:

  • sensory nerve supply;
  • oro-pharynx;
  • laryngopharynx;
  • mucosa;
  • Sihler's stain;
  • glossopharyngeal nerve;
  • vagus nerve;
  • internal superior laryngeal nerve;
  • swallowing

Abstract

  1. Top of page
  2. Abstract
  3. MATERIALS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. LITERATURE CITED

To date, the details of human sensory innervation to the pharynx and upper airway have not been demonstrated. In this study, a single human oro- and laryngopharynx obtained from autopsy was processed with a whole-mount nerve staining technique, Sihler's stain, to determine its entire sensory nerve supply. The Sihler's stain rendered all mucosa and soft tissue translucent while counterstaining nerves. The stained specimen was then dissected and the nerves were traced from their origins to the terminal branches. It was found that the sensory innervation of the human pharynx is organized into discrete primary branches that innervate specific areas, although these areas are often connected by small neural anastomoses. The density of innervation varied, with some areas receiving almost no identifiable nerve supply (e.g., posterior wall of the hypopharynx) and certain areas contained much higher density of sensory nerves: the posterior tonsillar pillars; the laryngeal surface of the epiglottis; and the postcricoid and arytenoid regions. The posterior tonsillar pillar was innervated by a dense plexus formed by the pharyngeal branches of the IX and X nerves. The epiglottis was densely innervated by the internal superior laryngeal nerve (ISLN) and IX nerve. Finally, the arytenoid and postcricoid regions were innervated by the ISLN. The postcricoid region had higher density of innervation than the arytenoid area. The use of the Sihler's stain allowed the entire sensory nerve supply of the pharyngeal areas in a human to be demonstrated for the first time. The areas of dense sensory innervation are the same areas that are known to be the most sensitive for triggering reflex swallowing or glottic protection. The data would be useful for further understanding swallowing reflex and guiding sensory reinnervation of the pharynx to treat neurogenic dysphagia and aspiration disorders. Anat Rec 258:406–420, 2000. © 2000 Wiley-Liss, Inc.

An important part of understanding swallowing, speech, and respiration disorders is knowledge of the sensory innervation to the human pharynx and upper airway. At present the specifics of this innervation are not known and animal models are of limited value due to the unique anatomy and physiology of the human oro- and laryngopharynx (OLP). Physiologic experiments in animals, as well as clinical observations in humans, show that the OLP is known to have specific areas that initiate pharyngeal swallowing and a variety of protective reflexes of the upper airway. Previous studies demonstrated that swallowing is best elicited from certain areas of the palate, pharynx, and epiglottis innervated by the maxillary branch of the trigeminal (V), glossopharyngeal (IX), and internal superior laryngeal nerve (ISLN) (Miller and Sherrington, 1916; Pommerenk, 1928). In the cat, stimulation of the posterior tonsillar pillars (Sinclair, 1971) and posterior pharyngeal wall (innervated by the IX nerve) (Doyle, 1923; Lewis and Dandy, 1930; Reichert, 1934), and the epiglottis and glottis (innervated by the ISLN) (Storey, 1968a) is able to initiate swallowing. In the human, the anterior and posterior tonsillar pillars and the posterior pharyngeal wall (Pommerenk, 1928; Doty, 1968; Goyal and Cobb, 1981) are the most sensitive areas for initiation of the pharyngeal phase of swallowing. Several investigators believed that the swallowing reflex is mainly triggered from the IX nerve (Levine, 1988; Kahrilas, 1994). Others reported that water stimulation of the laryngeal surface of the epiglottis and the arytenoid area (innervated by the ISLN) is more effective to initiate swallowing (Storey, 1968a; Shinghai and Shimada, 1976). However, little is known about the sensory nerve supply patterns in these areas. We hypothesized that swallowing and protective reflexes of the upper airway are best elicited from rather specific areas in the OLP where the mucosa is densely innervated. If so, delineation of the areas which have the greatest density of sensory innervation is very useful for further studying reflex swallowing and helpful for developing novel therapies to treat dysphagia caused by neurogenic diseases.

Another important aspect on the sensory innervation of the pharynx is to clarify the controversies concerning the precise distribution of the sensory nerves. It is generally believed that the oropharynx (from the soft palate to the level of the hyoid bone) and palatine tonsils receive their sensory innervation from the IX nerve, and the laryngopharynx (from the hyoid bone to the lower border of the cricoid cartilage) from the vagus (X) nerve (Doty, 1951, 1968; Miller, 1982). Anatomical studies further demonstrated that the IX nerve and the pharyngeal branch of the X (Ph-X) are the primary sources of sensory innervation of the pharynx. The Ph-X supplies the posterior pharyngeal wall and posterior tonsillar pillars. The IX supplies the lateral pharyngeal wall and the posterior third of the tongue (Sprague, 1944). Nerve section studies in the rat suggested that the IX also supplies fibers to the lingual surface of the epiglottis (Hogg and Bryant, 1969). By using both the nerve lesioning and electrophysiological techniques, Feindel (1956) reported similar results. The author found that the epiglottis remained some degree of sensibility after bilateral section of both the ISLN and recurrent laryngeal nerves (RLN). However, it remains unknown whether the IX in humans innervates the epiglottis. Additionally, nothing is known about the branching pattern of the individual sensory nerves to the OLP.

A recent advance in the study of the upper areodigective tract is the use of the Sihler's stain to study entire nerve supply within a large tissue and/or a whole organ such as the larynx (Wu and Sanders, 1992; Sanders et al., 1993a, b; 1994; Mu et al., 1994; Wu et al., 1994; Sanders and Mu, 1998) and pharynx (Mu and Sanders, 1996, 1998a, b). Sihler's stain clears soft tissue while counterstaining all nerves, from cranial nerves down to terminal axons. Therefore, precise distribution and branching of any nerves in the tissue can be studied by tracing them from the origins to the terminals. This stain is especially useful because it can be applied to human post mortem material. To date, this technique has mostly been used to study the motor nerve supply to human upper airway muscles even though the stain is just effective in displaying sensory nerves (Sanders and Mu, 1998). However, sensory and motor nerves look exactly alike in Sihler's stained specimens. Therefore, their identification as sensory or motor nerves depends on preserving the entire nerve supply to an area in continuity so that the nerves can be traced from their cranial nerve origins to their terminations. Motor nerves are easier to identify as the structure of muscular tissue is distinctive and usually well preserved in specimens stained with Sihler's stain. Sensory nerves are harder to identify as they can innervate a wide variety of structures, however, one tissue that retains its structure well in Sihler's specimens is mucosa, and this was the subject of this study.

This study was designed to determine the entire peripheral sensory nerve supply patterns of the human OLP by utilizing Sihler's stain. Our special attention was given to: 1) demonstrate the precise distribution and branching patterns of the major sensory nerves of the OLP; 2) determine if neural connections between the major sensory nerves to the OLP exist; 3) study whether areas known to trigger reflex swallowing have specific sensory innervation patterns; and 4) clarify if the human IX contributes innervation to the epiglottis.

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. MATERIALS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. LITERATURE CITED

Preparation of the Specimen

A single whole-mount specimen of human OLP (male, 67 years) that contained the tongue, pharynx, larynx, and upper esophageal sphincter was obtained from an autopsy. The IX and X nerves were excised off at the jugular foramen where they exited from the skull base and tagged for identification. The whole-mount specimen was then processed with Sihler's stain as follows.

Sihler's Stain

The procedures of the Sihler's stain have been described elsewhere (Mu and Sanders, 1996a,b Mu and Sanders,1998a,b). Briefly, eight steps are needed as follows: 1) Fixation of the whole-mount specimen in 10% unneutral formalin for 8 weeks. 2) Maceration in 3% KOH solution for 3 weeks with several changes of solution as required. 3) Decalcification in Sihler's solution I (one part glacial acetic acid, one part glycerin, and six parts 1% aqueous chloral hydrate) for 3 weeks with several changes of solution as required. 4) Staining in Sihler's solution II (one part stock Ehrlich's hematoxylin, one part glycerin, and six parts 1% aqueous chloral hydrate) for 4 weeks. 5) Destaining in Sihler's solution I for 3 hr. 6) Darkening the nerves in 0.05% lithium carbonate solution for 1 hr. 7) Clearing in 50% glycerin for 3 days. 8) Preservation in 100% glycerin with a few thymol crystals for 4 weeks for transparency. The Sihler's stained specimen was then dissected to remove unnecessary soft tissue so that the nerve supply can be clearly seen. The Sihler's stain for this whole-mount human specimen took 22.5 weeks.

Microdissection

After processing with the Sihler's stain the specimen was grossly dissected under a dissecting microscope (OPMI 99; Zeiss, Thornwood, NY) at a magnification of ×1.6 using microsurgical instruments. The anterior two-thirds of the tongue, hyoid bone, and laryngeal cartilages and muscles were removed, and the pharynx was divided in the midline of the posterior pharyngeal wall.

Although all the nerves (motor and sensory) within the specimen were stained as the same color, the sensory nerves supplying the mucosa can be differentiated from the motor nerves supplying the muscles by following the nerves from their origins to the terminals. In order to trace the individual sensory nerves, the grossly dissected specimen was placed on a 8.5 × 11″ transparency film (C-Line Products, Inc.), transilluminated with a Xenon light source (model 610, Karl Storz, Endoscopy-America, Inc., CA), and microdissected under a dissecting microscope (TYP 355110; Wild, Heerbrugg, Switzerland) at a magnification of ×10–30. The mucosa of the posterior tongue, oropharynx, epiglottis, larynx, and laryngopharynx was separated as a single sheet from muscle layer. During dissection, the sensory branches of the Ph-X and the pharyngeal branches of the IX (Ph-IX) were transected and tagged in order to completely separate the mucosal layer from the muscular layer. The mucosal layer was then spread on a transparency film. The IX and X (Ph-X and ISLN) nerves were traced from their origins at the jugular foramen to the terminal branches within the mucosa. During dissection, great care was taken to avoid damage any noticeable nerve branches and terminal twigs within the tissue. Microdissection was time-consuming work which took about 6 weeks.

Measurements

After the Sihler's stained specimen was microdisected the distribution and branching of the IX, Ph-X and ISLN in the OLP were then carefully examined. Measurements were taken to determine the following: 1) the number of the major nerve branches derived from each of the sensory nerves to the OLP; 2) the length of the major nerve branches; 3) the diameter of a given major nerve branch (5 mm distal to its diverging point); and 4) the topographic localization and number of the neural connections between different sensory nerves to the OLP.

Photographing

The completed specimen was then transilluminated with a Xenon light source (model 610; Karl Storz, Endoscopy-America, Inc., CA) and photographed with a 35 mm camera (OM-4 Olympus, Tokyo, Japan) fitted with a 20, 38, or 50 mm macrolens. First, the whole-mount specimen was photographed to show the origin and overall arrangement of the sensory nerves in the OLP. Finally, higher power views of certain areas innervated by each of the sensory nerves were taken to clearly demonstrate the details about the neural organization.

Assessment of Innervation Density

The complete specimen and photographs from various subregions of the pharynx were examined. The regions that were assessed for innervation density included: 1) the base of the tongue; 2) lateral pharyngeal wall; 3) posterior wall of the oro-pharynx; 4) laryngeal surface of the epiglottis; 5) piriform sinus; 6) arytenoid region; 7) post crico-arytenoid region; and 8) posterior wall of the laryngopharynx. The innervation density was evaluated by estimating the number of the terminal branches within a 1.0 cm2 area in a given region. The innervation density was divided into three degrees: 1) Degree 1: <5 terminal branches per square cm, 2) Degree 2: 5–10 terminal branches per square cm, and 3) Degree 3: >10 terminal branches per square cm.

RESULTS

  1. Top of page
  2. Abstract
  3. MATERIALS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. LITERATURE CITED

In this study, entire sensory nerve supply of a human OLP was studied by utilizing Sihler's stain. The pharyngeal walls are composed of an outer muscular layer and an inner mucosal layer. Both layers had to be separated from each other to illustrate the nerve supply to the mucosa of the OLP. Therefore, some nerve branches had to be transected while the mucosal and muscular layers were being separated. Figures 1A–C show the origins of the branches of the IX and X nerves supplying the mucosa of the OLP. Figures 1D and E demonstrate the peripheral branching and distribution of the sensory branches of the IX and X nerves in the pharyngeal mucosa. The entire sensory nerve supply to the OLP is demonstrated by Sihler's stained specimen (Fig. 1D) and clearly illustrated by a schematic drawing (Fig. 1E). Figures 2 to 5 are higher power views of the mucosa in different areas of the OLP to demonstrate the branching and distribution patterns of the individual nerves and the innervation density in various subregions of the pharynx. The branching patterns and distribution of the IX, X, and internal superior laryngeal nerve (ISLN) are given in Table 1.

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Figure 1. Sensory innervation of the human pharynx demonstrated by Sihler's stain. A: A whole-mount Sihler's stained muscular layer of the pharynx, showing the origins of the sensory branches of the IX and X nerves to the oro-pharyngeal mucosa (posterior view). The sensory nerve branches derived from the pharyngeal branch of the IX (Ph-IX; small solid arrows) and the pharyngeal branch of the X (Ph-X; large solid arrows) supply the lateral and posterior pharyngeal walls. These sensory branches had to be cut in order to separate the mucosa from the muscles. The transected sensory branches from the Ph-IX are indicated by the small asterisks and those from the Ph-X by the large asterisks. The remaining branches of the Ph-IX and Ph-X contribute to the formation of the pharyngeal plexus (open arrows) supplying the pharyngeal constrictors and upper esophageal sphincter. N, nodose ganglion; SCG, superior cervical sympathetic ganglion; X, vagus nerve; ISLN, internal superior laryngeal nerve; ESLN, external superior laryngeal nerve; SPC, superior pharyngeal constrictor; MPC, middle pharyngeal constrictor; IPC, inferior pharyngeal constrictor; CP, cricopharyngeus; UE, upper esophagus. B: Schematic of A, clearly illustrating the major nerves from which sensory branches are given to supply the mucosa of the oro-pharynx. RLN, recurrent laryngeal nerve. The other abbreviations are used in this figure are the same as those used in A. C: Higher power view of the right Ph-IX and Ph-X in A (posterior view). Note that the Ph-IX subdivides into three branches: superior (S), middle (M), and inferior (I). The sensory branches to the lateral pharyngeal wall are derived mainly from the superior and less from middle branch (small asterisks) of the Ph-IX (small solid arrow). The inferior branch of the Ph-IX (large open arrow) contributes to the formation of the pharyngeal plexus (small open arrow) to supply the pharyngeal constrictor muscles and upper esophageal sphincter. The Ph-X (large solid arrow) also gives off sensory branches (large asterisk) to the lateral pharyngeal wall. The distal portions of the transected sensory branches of the Ph-IX and Ph-X are those to the lateral pharyngeal walls as showed in D and E (middle). The abbreviations are used in this figure are the same as those used in A. Magnification 3×. D: Whole-mount Sihler's stained mucosa of the oro- and laryngopharynx (posterior view), showing the topographic localization and distribution of each of the L-IX, Ph-IX, Ph-X, and ISLN. The mocosa of the posterior pharyngeal wall has been opened in the midline. Note that the L-IX mainly supplies the posterior tongue (PT) and its adjacent structures such as the tonsil (T). The Ph-IX and Ph-X supply the lateral and posterior pharyngeal walls. The ISLN supplies the epiglottis (E), inside of the larynx, and the anterior wall of the laryngopharynx. LPW, lateralpharyngeal wall; PPW; posterior pharyngeal wall; L-IX, lingual division of the IX. E: Schematic of D, showing the branching patterns of the individual nerves supplying the pharyngeal mucosa. The L-IX subdivides into four branches: tonsillar (1), epiglottic (2), tongue base (3), and papillar (4). The lingual surface of the epiglottis also receives its innervation from both the epiglottic (large arrow) and tongue base (small arrow) branches of the L-IX. There is a dense plexus formed by the Ph-IX and Ph-X in the mucosa of the lateral pharyngeal wall (LPW). The ISLN supplies the epiglottis (E), inside of the larynx (IL) and laryngopharynx. LN, lingual nerve; FP, foliate papillae; VP, vallate papillae; LG, lymphoid granules; PPW, posterior pharyngeal wall; IL, inside of the larynx; AWL, anterior wall of the laryngopharynx.

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Figure 2. Branching pattern of the human L-IX (posterior view). A: Higher power view of the left L-IX in Figure 1D. Note that the L-IX divides into four branches: tonsillar (1), epiglottic (2), tongue base (3), and papillar (4). The first two branches are smaller than the last two branches. The papillar branch that is the largest divides into two branches, medial and lateral. The medial branch (M) supplies the vallate papillae (VP), whereas the lateral branch (L) supplies both the foliate papillae (FP) and VP. LN, lingual nerve; PT, posterior tongue; E, epiglottis; T, tonsil. Magnification 3×. B: Higher power view of the nerve supply to the vallate papillae (VP) on both sides in Figure 1D (posterior view). Note that the ‘central vallate papilla’ (C) is innervated bilaterally and the remaining vallate papillae are supplied ipsilaterally. 3, tongue base branch; 4, papillar branch; T, tonsil. PT, posterior tongue; FP, foliate papillae; L, lateral branch of the papillar branch; M, medial branch of the papillar branch. Magnification 3×. C: Higher power view of the nerve supply pattern of the central vallate papilla in B. Note that the nerve terminals come into the vallate papilla from its base (white arrows) and then break up into individual axons to reach its free end (black arrow). Magnification 30×. D: Higher power view of the nerve supply of the left vallate papillae in B. Note that although the left two papillae are innervated by the twigs derived from the lateral branch (L) and the right papilla from the medial branch (M) of the papillar branch of the L-IX, these terminal twigs connect with each other. Magnification 30×.

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Figure 3. Higher power view of the nerve supply to the human lateral and posterior pharyngeal walls in Figure 1D. A: Left Ph-IX (small solid arrow) and Ph-X (large solid arrow) enter the lateral pharyngeal wall (LPW) posterior to the tonsil (T) and break up numerous terminal twigs which supply the LPW and extend to the posterior pharyngeal wall (PPW). The nerve branches from the Ph-IX (large open arrow) and L-IX (small open arrow) connect with the ISLN. E, epiglottis. Magnification 3×. B: Higher power view of the left lateral pharyngeal wall (LPW) in A. Note that an extremely dense plexus is formed in the LPW. Magnification 9×. C: Higher power view of the right lateral pharyngeal wall in Figure 1C. Note that a similar nerve plexus formed by the Ph-IX and Ph-X exists. Magnification 9×. D: Higher power view of the left posterior pharyngeal wall (PPW) in Figure 1D. Note that the terminal twigs (between arrows) from the plexus in the lateral pharyngeal wall (LPW) extend to the PPW where a network is formed. A number of lymphoid granules in the PPW are seen (asterisks). Magnification 9×.

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Figure 4. Higher power view of the nerve supply to the epiglottis and laryngopharyngeal mucosa (posterior view). A: Higher power view of the branching pattern of the human superior laryngeal nerve (ISLN) in Figure 1D. Note that the human ISLN divides into three divisions, superior (S), middle (M), and inferior (I), each of which breaking up several secondary branches. The S division supplies the epiglottis (E). The M division supplies the aryepiglottic fold (AE) and inside of the larynx. Whereas the I division innervates the arytenoid (A) and postcricoid (PC) regions and anterior laryngopharyngeal wall. Magnification 3×. B: Higher power view of the nerve supply to the laryngeal surface of the epiglottis in A. Note that the nerve branches from both sides connect with each other and form a network. Magnification 9×. C: Higher power view of the nerve supply to the bilateral postcricoid regions (posterior view) in Figure 1D. Note that a number of nerve branches from the inferior division of the ISLN supply the postcricoid region and run across the midline. This region has extremely dense nerve supply. Magnification 9×. D: Higher power view of the right arytenoid area in A. Note that this area also has rich sensory innervation. Magnification 9×.

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Figure 5. Higher power view of several areas in Figure 1D where the neural connections between the IX and lingual nerve (LN), and between the IX and ISLN were identified. A: Neural connection (arrow) between the lingual branch of the IX (L-IX) and the LN on the left aspect of the posterior tongue. FP, foliate papillae. Magnification 12×. B: Neural connection (arrow) between the L-IX and LN on the right aspect of the posterior tongue. FP, foliate papillae. Magnification 12×. C: Neural connections (arrows) between the L-IX and ISLN on the right side. Note that the epiglottic branch (open arrow) breaks up three branches, two of which anastomosing with the superior division of the ISLN (arrows). E, epiglottis; T, tonsil. Magnification 3×. D: Higher power view of the anastomosing points (arrows) in C. Double arrow heads indicate the nerve branches derived from the epiglottic branch of the L-IX. E, epiglottis; S, superior division of the ISLN. Magnification 9×. E: (page 418) Higher power view of the right lower corners of the Figure 3A, showing the connecting nerves between the left IX and the superior division (S) of the ISLN. Note that there are five connecting nerves (small solid arrows) between the Ph-IX (large solid arrow; also see Fig. 3A, large open arrow) and the superior division (S) of the ISLN. There are two connecting nerves between the L-IX (open arrows) and the superior division of the ISLN (also see Fig. 3A, small open arrow). E, epiglottis. Magnification 9×.

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Table 1. Branching and distribution of the sensory nerves to the human oro- and laryngopharynx*
NervesMajor branchesOropharynxLaryngopharynx
PTVPTLPWPPWEAEILCAAWLPPW
  • *

    PT, posterior tongue; VP, vallate papillae; T, tonsil; LPW, lateral pharyngeal wall; PPW, posterior pharyngeal wall; E, epiglottis; AE, aryepiglottic fold; IL, inside of the larynx; CA, cricoarytenoid region; AWL, anterior wall of the laryngopharynx; Ph-IX, pharyngeal branch of the IX; Ph-X, pharyngeal branch of the X; S-ISLN, superior division of the ISLN; M-ISLN, middle division of the ISLN; I-ISLN, inferior division of the ISLN; −, no nerve supply; +, nerve supply; ++, dense nerve supply.

IXL-IX++++++
Ph-IX+++++
XPh-X++++
ISLNS-ISLN+
M-ISLN++++++
I-ISLN+++++++

Distribution and Branching Patterns of the Human IX Nerve

The distribution and branching of the human IX nerve are given in Table 1. After emerging from the skull base, the IX divides into a pharyngeal and a lingual division. The pharyngeal division of the IX (Ph-IX) subdivides into three major branches: superior, middle, and inferior (Fig. 1C). The sensory branches to the oro-pharynx (Figs 1D,E, middle) are given off most from the superior and a few from the middle branches (Figs 1A–C). The middle branch mainly supplies motor innervation to the stylopharyngeal muscle. However, it also gives off sensory branches to the lateral pharyngeal wall. The inferior branch contributes to the formation of the pharyngeal plexus (Fig. 1C, large open arrow) supplying the inferior pharyngeal constrictor muscle and the upper esophageal sphincter.

The lingual division of the IX (L-IX) contributes innervation mainly to the posterior tongue and its adjacent structures (Figs. 1D,E). Posterior to the tonsil, the L-IX breaks up four secondary branches to the tonsil, epiglottis, tongue base, and papillae, respectively. The first branch of the L-IX supplies the tonsil and peritonsillar areas (tonsillar branch). Before it reaches the tonsil, the tonsillar branch gives off two to three tertiary branches surrounding the tonsil. Its major trunk passes the tonsillar substance and breaks up two to three tertiary branches at the anterior aspect of the tonsil to supply the anterior tonsillar pillar (Figs 1D,E, 2A). Shortly after the tonsillar branch is given off, the L-IX sends out a small branch to the epiglottis (epiglottic branch). The epiglottic branch subdivides into two to three tertiary branches, one supplying the lingual surface of the epiglottis and one or two anastomosing with the ISLN (Figs 1D,E). Then the L-IX gives off its third branch to supply the mucosa of the tongue base (tongue base branch). The tongue base branch breaks up two to three tertiary ones and numerous twigs which run toward midline and connect with those from the opposite side (Figs 1D,E). Finally, the largest branch of the L-IX supplies the vallate and foliate papillae (papillar branch) in the posterior tongue. The papillar branch subdivides into two branches; lateral and medial. The lateral one mainly supplies the foliate papillae that are located on the lateral border of the tongue just in front of the anterior tonsillar pillar. It also sends one to two terminal twigs to the lateral last vallate papilla. In contrast, the medial one supplies the vallate papillae. There are 12 vallate papillae in this tongue, which are arranged into a so-called V shape (Figs 1D,E, 2A). The largest one is directed to the tip of the V (central papilla). Except for the ‘central’ papilla, there are five vallate papillae on the left side and six on the right side. The foliate branch subdivides into two branches. The large one breaks up four to five twigs to supply the foliate papillae and connect with the lingual nerve, while the small one contributes two to three twigs to supplying two to three lateral vallate papillae (Figs 1E, 2A). The vallate branch also subdivides into two branches, medial and lateral. The medial branch breaks up three to five twigs to supply the ‘central’ and one or two medial vallate papillae. The lateral branch also breaks up four to five twigs to innervate two to five medial vallate papillae (Figs 1E, 2A,B).

It was found that the central vallate papilla is the largest among all the vallate papillae and innervated bilaterally (Figs 1D,E, 2B). Numerous terminal fibers from both sides enter the vallate papilla at its base (Fig. 2C, white arrows). These terminal fibers further split into individual axons that reach the top surface of the papilla (Fig. 2C, black arrow). The remaining vallate papillae are innervated ipsilaterally. However, each of these vallate papillae receives its innervation from different terminal fibers that connect with each other (Fig. 2D).

The L-IX was measured to be ≈7–8 cm long. The length of the individual major branches was measured to be ≈4.5–5 cm for the tonsillar branch, 5–6 cm for the epiglottic branch, 4–5 cm for the tongue base branch, and 5.5–6.5 cm for the papillar branch, respectively. The main trunk of the L-IX was measured to be 0.8 mm thick. The tonsillar branch accounts for ≈15%, epiglottic branch for ≈10%, tongue base branch for ≈15%, and papillar branch for ≈60% of the L-IX, respectively.

Distribution of the Sensory Branch of the Ph-X in the Oropharynx

The distribution of the sensory branches of the X nerve is shown in Table 1. The Ph-X subdivides into motor and sensory divisions (Figs 1A,B). The motor division contributes to the formation of the pharyngeal plexus to innervate the pharyngeal constrictor muscles and upper esophageal sphincter (Figs 1A–C). While the sensory division (Figs 1A–C, large asterisks) contributes innervation to the lateral and posterior pharyngeal walls (Figs 1D,E, middle). The terminal nerve branches derived from the Ph–X and the Ph-IX form an extremely dense plexus in the mucosa of this region (Figs 1D,E, 3A–C). These nerve branches on one side extend from the lateral wall to the posterior wall of the oro-pharynx (Figs 1D,E, 3A). The nerve branches from both sides supply the mucosa of the posterior pharyngeal wall over the midline. The terminals in the posterior pharyngeal wall connect with each other forming a network (Fig. 3D).

Distribution and Branching Patterns of the Human ISLN

The superior laryngeal nerve (SLN) emerges from the lower portion of the nodose ganglion of the X nerve and passes downward. About 5 mm distal to the nodose ganglion, the SLN divides into a small external (ESLN) and a large internal branch (ISLN) Figs 1A–C). The ESLN supplies the cricothyroid muscle, whereas the ISLN penetrates the thyrohyoid membrane to supply the laryngeal and laryngopharyngeal mucosa. The details about the distribution and branching patterns of the human ISLN are shown in Table 1. Shortly after penetrating the thyrohyoid membrane, the ISLN divides into three major divisions: the superior, middle, and inferior (Fig. 4A). The length between the nodose ganglion and the point where the ISLN gives off its three major branches was measured to be ≈6.5 cm. The distance of the individual divisions was measured to be ≈4 cm for the superior, ≈2.5–3.0 cm for the middle, and ≈6–7 cm for the inferior division. The main trunk of the ISLN was measured to be ≈1.5 mm thick. The superior division accounts for ≈20%, the middle division for ≈30%, and the inferior division for ≈50% of the ISLN.

The superior division gives off two to three secondary branches, one supplying the pharyngeal surface of the epiglottis and others supplying the laryngeal surface of the epiglottis. The nerve branches supplying the laryngeal surface of the epiglottis on each side split into numerous twigs which connect with those from the opposite side to form a complex network (Figs 1E, 4A,B). The middle division subdivides into four to six secondary branches, one to two supplying the aryepiglottic fold, and three to four penetrating the aryepiglottic fold to supply false and true vocal folds (Figs 1E, 4A). The largest inferior division travels toward the cricoarytenoid joint where it gives off its first secondary branch to supply the arytenoid area. Then it runs downward and gives off approximately five to six secondary and numerous tertiary branches to form a brush-like arrangement. These nerve branches travel from superolateral to the inferomedial supplying the mucosa on the anterior wall of the laryngopharynx (Figs 1D,E, 4A). The postcricoid region (Fig. 4C) and arytenoid area (Fig. 4D) supplied by the inferior division of the ISLN were found to be densely innervated.

The Neural Connections Between the Sensory Nerves to the OLP

Although the anterior wall of the oro-pharynx (posterior tongue) is mainly supplied by the L-IX, the lingual nerve (LN) also sends few branches to supply the mucosa and foliate papillae (Figs 1E, 2A, 5A,B). Neural connections between the L-IX and LN were identified on each side (Figs 1E, 5A,B).

The neural connections between the IX and ISLN were observed on each side (Figs 1E, 5C–E). On the right side, the L-IX gives off its second branch (epiglottic branch). This branch subdivides into three branches, one supplying the pharyngeal surface of the epiglottis and others anastomosing with the superior division of the ISLN (Fig. 5C). There are three neural connections between the L-IX and ISLN on the right side (Figs 5C,D). On the left side, both the Ph-IX and L-IX give off branches to anastomose with the ISLN. As Figure 3A shows, the Ph-IX sends out a large branch (large open arrow) and the epiglottic branch of the L-IX gives off a small branch (small open arrow) to connect with the branches of the superior division of the ISLN. Higher power view of these nerves demonstrated that there are multiple connecting nerves between the Ph-IX and ISLN and between the L-IX and ISLN (Figs 1E, 5E).

Considerable neural connections between the Ph-IX and Ph-X were seen in the plexus in the mucosa of the lateral pharyngeal wall including the posterior tonsillar pillar on each side (Figs 1E, 3A–C).

Nerve Supply Density in Various Subregions of the Pharynx

The results demonstrated that the nerve supply density in the pharynx was different from region to region. The regions with degree 1 innervation density were found to be the piriform sinus and posterior wall of the laryngopharynx. The regions with degree 2 innervation density were the base of the tongue and posterior wall of the oro-pharynx. The regions with degree 3 innervation density were the lateral pharyngeal wall, laryngeal surface of the epiglottis, arytenoid area, and post cricoarytenoid region.

DISCUSSION

  1. Top of page
  2. Abstract
  3. MATERIALS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. LITERATURE CITED

In the present study, a single human OLP was studied by utilizing Sihler's stain to demonstrate its entire sensory nerve supply. Several important observations are that 1) the oro-pharynx is supplied by the IX and Ph-X, and the laryngopharynx by the ISLN; 2) the areas densely innervated are the lateral pharyngeal wall including the posterior tonsillar pillar, laryngeal surface of the epiglottis, and postcricoid and arytenoid regions; 3) the lateral wall of the oro-pharynx is doubly innervated by the Ph-IX and the Ph-X; 4) the human L-IX also innervate the lingual surface of the epiglottis; 5) multiple neural connections between the L-IX and lingual nerves and between the IX and the superior division of the ISLN were observed; and 6) the precise numbers and courses of the major nerve branches derived from each sensory nerve were determined. To our knowledge, this is the first demonstration of the entire human OLP sensory nerve supply. The data obtained from this study are significantly important for understanding the mechanisms of the pharyngeal swallowing. In addition, determination of the distribution and branching of the IX , Ph-X, and ISLN is helpful for guiding sensory reinnervation of the OLP.

Physiological Significance of the Areas Which Have Dense Nerve Supply and Double Innervation in the OLP

The important role of normal afferent input in the swallowing reflex has been conformed by both the electrical nerve stimulation and elimination of sensitivity of the OLP. Electrical stimulation of IX (Doty, 1968), Ph-X (Sinclair, 1971), and ISLN (Doty, 1951; Miller, 1972) initiates pharyngeal swallowing. In contrast, the pharyngeal swallow can be impaired by decreasing sensitivity in the oro-pharynx following application of topical anesthetic to the pharyngeal mucosa (Mansson and Sandberg, 1974, 1975). Decreased sensitivity of the OLP affects feeding and swallowing, particularly in neurologically impaired patients (Aviv et al., 1996).

The density of the sensory nerve supply to the OLP is much different from region to region as demonstrated by this study. For example, the posterior tonsillar pillar and lateral pharyngeal wall innervated by the Ph-IX and Ph-X, and the laryngeal surface of the epiglottis, postcricoid, and arytenoid regions innervated by the ISLN have the greatest density of the sensory innervation. These densely innervated areas correspond to the potent reflexogenic areas triggering the pharyngeal swallowing and upper airway protective reflexes (Miller and Sherrington, 1916; Pommerenk, 1928; Storey, 1968a). The results obtained from this study indicate that there is a close relationship between the density of sensory nerve supply and the reflexogenic areas in the OLP. This information would be important in better understanding the mechanisms of the reflex swallowing. The complex motor output of swallowing depends upon the patterned sensory input prior the motor response. In normal circumstance, the bolus is propelled backward by the tongue to the oro-pharynx. The tonsillar pillars and lateral pharyngeal walls move medially. Therefore, the afferent receptors in these areas are stimulated and the pharyngeal swallowing is triggered. The most effective stimuli for initiating swallowing are touch and light pressure applied to the pharyngeal mucosa (Miller et al., 1997) and water to the laryngeal mucosa (Storey, 1968a,b; Shinghai and Shimada, 1976). The laryngeal mucosa has rich sensory innervation (Sanders and Mu, 1998) and more receptors with free nerve endings which respond more to liquid than to pressure stimulation (Miller, 1986). Some investigators reported that the receptors located in the tongue base and piriform sinuses also initiate pharyngeal swallow (Derkay and Schechter, 1998). However, in the present study no rich sensory nerve supply was found in these areas. If the areas with the greatest density of the sensory nerve supply are consistent with the reflexogenic fields, it is reasonable to believe that the lateral pharyngeal wall including the posterior tonsillar pillar (innervated by both the Ph-IX and Ph-X) is a key area from which the pharyngeal swallowing is triggered.

Swallowing not only is an alimentary behavior, but also serves as a protective reflex of the upper airway by glottic closure (Rushmer and Hendron, 1951) as well as coughing (Storey, 1976). It is known that the ISLN plays a vital role in the protection of the airway by initiating reflex glottic closure during swallowing (Storey and Johnson, 1975; Stedman et al., 1980). The reflexogenic areas in the laryngopharynx controlled by the ISLN is most likely to be the laryngeal surface of the epiglottis, arytenoid area, and postcricoid region since they have rich sensory innervation as demonstrated by this study. The three divisions of the ISLN supply different regions in the larynx and laryngopharynx. Since the superior and middle divisions of the ISLN supply the epiglittis, aryepiglottic fold, and the inside of the larynx, it is reasonable to believe that they play a crucial role in protective reflexes of upper airway such as glottic closure and coughing during swallowing. Whereas the inferior division of the ISLN supplies the postcricoid region and the anterior laryngopharyngeal wall. Therefore, this area is likely to be associated with swallowing reflex.

Double innervation of certain areas of the mucosa in the OLP may have physiological significance. As demonstrated by the present study, the epiglottis and the lateral pharyngeal wall including the posterior tonsillar pillar are the doubly innervated areas in the OLP. The epiglottis is innervated by the ISLN and L-IX nerves. The L-IX not only contributes nerve branches to supply the pharyngeal surface of the epiglittis, but also sends branches to anastomose with the superior division of the ISLN which supplies the laryngeal surface of the epiglottis. Our findings are able to explain why the epiglottis still remains sensibility after bilateral section of the ISLN as reported by Feindel (1956) and Hogg and Bryant (1969). This study showed that the mucosa of the posterior tonsillar pillar and lateral pharyngeal wall is innervated by both the Ph-IX and the Ph-X. The findings about the double innervation of the mucosa in the OLP would be important for further studying the mechanisms of the pharyngeal swallowing and for explaining the results obtained from the electrophysiological experiments.

Clinical Significance of Clarifying Branching Patterns of the Sensory Nerves in the OLP

A recent significant advance in the head and neck surgery is the laryngeal transplantation. Since 1960s, many animal experimental studies on the laryngeal transplantation have been done and great progress has been made (Strome and Strome, 1994; Strome et al., 1994; Anthony, 1995; Anthony et al., 1996). More recently, human laryngeal transplantation has been reported (Birchall, 1997, 1998). One of the important problems facing in the laryngeal transplantation is how to restore the function of the transplanted larynx (Birchall, 1997). Up to now, motor reinnervation of the larynx by a variety of techniques is successful (Tucker, 1976; Crumley, 1991). However, little information can be available regarding the sensory reinnervation of the paralyzed and/or transplanted larynx which is critical for preventing post-operative aspiration.

Aspiration is the most life-threatening medical problem followed by dysphagia (Logemann, 1986). Approximately 40% to 50% of patients in nursing homes have experienced dysphagia (Logemann, 1993). Dysphagia is caused not only by motor component, but also by sensory component of swallowing structures. It has been estimated that approximately 50,000 American people are died from aspiration pneumonia after stroke each year (Brown and Glassenberg, 1983; Schmidt et al., 1988). At present, intractable aspiration is treated mainly by tracheostomy and other surgical procedures such as laryngeal suspension (Calcaterra, 1971; Hillel and Goode, 1983), glottic closure (Montgomery, 1975; Sasaki et al., 1980), laryngotracheal diversion (Lindeman, 1975), laryngoplasty (Biller et al., 1983), laryngeal stents (Eliachar et al., 1987), partial cricoid resection (Krespi et al., 1985), and even total laryngectomy (Montgomery, 1973). Although these procedures can eliminate aspiration in some extent, some procedures have distinct disadvantages. For example, total laryngectomy can deal with aspiration, but it results in permanent functional, communicative, cosmetic and psychological disability. It also eliminates the possibility of reversal in patients who might recover some or all of their function. There exists a great need to develop novel therapies to treat neurogenic dysphagia. Rehabilitation of the sensory function of the pharynx and larynx for treating aspiration is desirable. The stroke patients with dysphagia have laryngopharyngeal sensory dysfunction (Aviv et al., 1996). Based on a recent report, sensory reinnervation of the human laryngopharynx is effective to restore its sensitivity for treating stroke patients with serious aspiration caused by dysphagia (Aviv et al., 1997). Therefore, details about the distribution and branching patterns of the individual sensory nerves to the OLP would be of great benefit for the surgeons to design neurosurgical procedures to treat neurogenic dysphagia and its resultant serious aspiration. In addition, the nerve map provided by this study is helpful for surgeons to avoid damaging the sensory nerves to the OLP during head and neck surgery.

It should be emphasized that many anatomic facts about the sensory nerve supply of the human OLP might be uncovered by this study because of the limitation of the specimens. Therefore, much more work is still needed to investigate the sensory innervation of the human upper airway.

LITERATURE CITED

  1. Top of page
  2. Abstract
  3. MATERIALS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. LITERATURE CITED