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EFFECT OF STYLOPHARYNGEUS MUSCLE ACTIVITY
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EXERCISING HORSES

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CAROLINE DOMINIQUE TESSIER

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EFFECT OF STYLOPHARYNGEUS MUSCLE ACTIVITY ON DORSAL
NASOPHARYNGEAL FUNCTION IN EXERCISING HORSES

By

Caroline Dominique Tessier

A THESIS

Submitted To
Michigan State University
In partial fulfillment of the requirements
For the degree of

MASTER OF SCIENCE

Department of Large Animal Clinical Sciences

2006

ABSTRACT
EFFECT OF STYLOPHARYNGEUS MUSCLE ACTIVITY ON DORSAL
NASOPHARYNGEAL FUNCTION IN EXERCISING HORSES
By

Caroline Dominique Tessier

Nasopharyngeal collapse has been clinically observed in horses as a cause of upper
respiratory obstruction and exercise intolerance, but the etiology of this condition is
unknown. The stylopharyngeus muscle, innervated by the glossopharyngeal nerve, is an
important pharyngeal dilating muscle in other species, and dysfunction of this muscle
results in nasopharyngeal collapse. Six horses were exercised at speeds corresponding to
50%, 75%, and 100% of HRmax on the treadmill while upper airway pressures and
respiratory frequencies were measured before and after induction of stylopharyngeus
muscle dysfunction by bilateral anesthesia of the glossopharyngeal nerve. Next, the
electromyographic activity of the stylopharyngeus muscle was recorded in six normal
horses performing the same exercise protocol. Bilateral glossopharyngeal nerve block
caused stylopharyngeus muscle dysfunction and dorsal nasopharyngeal collapse in all
horses. The stylopharyngeus muscle exhibited peak phasic inspiratory activity that
increased from 2.5 to 13.6 arbitary units, or 556%, as treadmill speed increased from
HRmaxso to HRmax. Therefore, the stylopharyngeus muscle is an important
nasopharyngeal dilator and stylopharyngeus dysfunction may be implicated in clinical

nasopharyngeal collapse in athletic horses.

TABLE OF CONTENTS

LIST OF FIGURES ................................................................................... iv
INTRODUCTION ................................................................................... 1
CHAPTER 1 : EFFECT OF STYLOPHARYNGEUS MUSCLE DYSFUNCTION ON
THE NASOPHARYNX IN EXERCISIN G HORSES .......................................... 12
Summary ..................................................................................... 12
Introduction ................................................................................... 1 3
Materials and Methods ..................................................................... 14
Results ........................................................................................ 18
Discussion .................................................................................... 19

CHAPTER 2 : ELECTROMYOGRAPHIC ACTIVITY OF THE

STYLOPHARYNGEUS MUSCLE IN EXERCISING HORSES ............................ 25
Summary ..................................................................................... 25
Introduction ................................................................................... 26
Materials and Methods ..................................................................... 27
Results ........................................................................................ 30
Discussion .................................................................................... 31

APPENDIX ........................................................................................... 34

REFERENCES ...................................................................................... 41

iii

LIST OF FIGURES

Figure 1. Illustration of the origin and insertion of the stylopharyngeus muscle ........... 35

Figure 2a: Endoscopic image of the guttural pouch. Notice the origin of the
stylopharyngeus muscle (arrow) on the medial aspect of the stylohyoid bone ............. 36

Figure 2b: Cadaveric specimen showing the insertion of the stylopharyngeus muscle
(white asterisk) on to the musculature of the dorsal nasopharynx ............................ 36

Figure 3: Endoscopic image of the nasopharynx of a horse running on the treadmill
following bilateral glossopharyngeal nerve block. Notice that the dorsal wall of the
nasopharynx has collapsed, obscuring the view of the corniculate processes of the
arytenoids cartilages ................................................................................. 37

Figure 4: Endoscopic view of the nasopharynx of a horse following right
glossopharyngeal nerve block, while the nares are occluded. Notice that only the right
hemipharynx has collapsed, while the left side remains in a normal position ............... 37

Figure 5: White bars: unblocked tn'als; Grey bars: with glossopharyngeal nerve
anesthesia ............................................................................................. 38

Figure 6. Recording of upper airway pressure (top tracing) and moving time averaged
EMG activity of the stylopharyngeus muscle. Notice that the EMG activity of the
stylopharyngeus muscle peaks during the early inspiration phase ........................... 39

Figure 7. Moving time averaged EMG activity of the stylopharyngeus muscle (top
tracing) and raw EMG activity (bottom tracing) recorded at HRmaxso, HRmax 75, and
HRmax. Notice that the activity increases at each speed ....................................... 39

Figure 8. Graph of the peak EMG activity of the stylopharyngeus muscle in 5 exercising

horses. Notice that the activity increases at each speed for each horse. The thick black
line represents the average of the 5 horses ....................................................... 40

iv

INTRODUCTION

Poor performance in the equine athlete is a problem with multiple facets. However, upper
airway disorders seem to have an important role to play (Martin 2000). Since the advent
of high-speed treadmill examination, we have been able to recognize a large number of
upper airway disorders present only during exercise and therefore high-speed treadmill
examination has been recommended to evaluate horses presenting for poor performance
with or without respiratory noise (Lumsden 1995). Dynamic upper airway obstructions
occur in horses affected by a certain number of disorders such as dorsal displacement of
the sofi palate, pharyngeal collapse, axial deviation of the aryepiglottic folds, epiglottic
entrapment, epiglottic retroversion (Martin 2000). Because of the dynamic and
multifactorial nature of these disorders, they are difficult to diagnose and treat effectively.
Furthermore, the pathogenesis of dynamic upper airway obstruction remains enigmatic in

some diseases such as nasopharyngeal collapse.

Nasopharyngeal collapse in horses

Nasopharyngeal collapse is a disease observed in adult horses that causes upper airway
obstruction, exercise intolerance and abnormal upper respiratory noise.

While a few horses affected by nasopharyngeal collapse will show signs of upper airway
obstruction or abnormal noise at rest (Smith 1994), the majority of them will only exhibit
clinical signs during exercise, making the condition difficult to diagnose. Poor
performance and abnormal respiratory noise are some of the owner’s common

complaints.

The features accompanying this disease are dorsal pharyngeal collapse and collapse of
the lateral walls of the nasopharynx. Dorsal pharyngeal collapse can be uni- or bilateral

(Smith 1994).

Use of high-speed treadmill endoscopy as a diagnostic tool has improved our recognition
of the disease. Reported prevalence rates vary from 3 to 29% (Kannegieter and Dore
1995; Dart 2001; Martin 2000; Durando 2002). In one study, 40 out of 256 horses
presented for poor performance had pharyngeal collapse. Thirty of 40 horses with
pharyngeal collapse had abnormal upper airway noise that was inspiratory and best heard
early or late in the exercise cycle. Collapse of the pharyngeal walls obstructed part or
most of the tracheal opening (Martin 2000). Pharyngeal collapse can also be associated
with other lesions of the upper respiratory tract. In 92 horses diagnosed with dorsal
displacement of the soft palate (DDSP) during treadmill videoendoscopy, 11/92 horses
had associated dynamic pharyngeal collapse (Parente 2002). In another retrospective
study of 54 horses diagnosed with upper respiratory tract abnormalities during treadmill
videoendoscopy, 16 had pharyngeal collapse alone. In this study, pharyngeal collapse
was the most represented abnormality related to the presence of abnormal blood gases
(decreased Pa02 and/or increased PaC02) (Durando 2002). Like many other dynamic
upper airway disorders, the cause of this disease in horses has not been elucidated and
there is no effective treatment available.

Anatomy of the nasopharynx and the pharyngeal musculature in horses

In other species, the action of several groups of muscles, including muscles of the tongue,
hyoid apparatus and nasopharynx, is responsible for stiffening and dilating the
nasopharynx (F eroah 2000; Kuna 2001; Kuna and Vanoye 1999; Kuna et al.1997). These
same muscles are also important during deglutition (Kuna and Vanoye 1999). Knowledge
of the anatomy of the nasopharynx and the relationships between different muscles is
therefore essential to understand pathophysiologic mechanisms responsible for dynamic

upper airway disorders.

The pharynx is a musculo-membranous sac that shares digestive and respiratory functions. Horses
have no communication between their 0m- and nasopharynx because their soft palate contacts the
larynx. They are therefore obligate nasal breathers. The pharynx is a funnel-shaped chamber
contained between the base of the skull and first couple of cervical vertebrae dorsally, the larynx
ventrally, and the pterygoid muscles, the mandible and the dorsal part of the hyoid apparatus
laterally. The nasopharynx is about 150m long and its long axis is directed downward and
backward. It is attached by its muscles to the palatine, pterygoid, and hyoid bones, and to the
cricoid and thyroid cartilages of the larynx. However, the nasopharynx is not directly supported
by bony structures and is therefore susceptible to large amounts of deformation by the negative

pressures applied during inhalation.

Its principal relations are: dorsally, to the base of the cranium and guttural pouches; ventrally, to
the larynx and soft palate, the latter extending caudally from the hard palate to the base of the
larynx and forming the floor of the nasopharynx, and laterally, to the medial pterygoid muscle,
the great cornu of the hyoid bone, the external carotid and external maxillary arteries, the
glossopharyngeal, anterior laryngeal, and hypoglossal nerves, the mandibular salivary gland, and

the parapharyngeal lymph glands (Sisson 1976).

Two groups of muscles will modify the shape and diameter of the pharyngeal opening:
intrinsic pharyngeal muscles that form the walls of the nasopharynx and extrinsic
pharyngeal muscles which action will also deform the nasopharynx. The intrinsic muscle
group is composed of: cricopharyngeus, hyopharyngeus, thyropharyngeus,
palatopharyngeus, pterigopharyngeus and stylopharyngeus. These muscles are
responsible for constriction and dilation of the nasopharynx (Sisson 1976). The
cricopharyngeus muscle originates on the lateral aspect of the cricoid cartilage and
attaches on the median raphe. The hyopharyngeus muscle attaches on the hyoid bone and
ends at the median raphe, and the thyropharyngeus muscle originates on the lateral
lamina of the thyroid cartilage and inserts on the median raphe. These three muscles are
pharyngeal constrictors and their shortening depresses the pharyngeal walls down and
inward. The palato-pharyngeus arises on the aponeurosis of the soft palate and inserts in
part into the upper edge of the thyroid cartilage and in part on the median raphe. The
pterygo-pharyngeus muscle lies on the anterior part of the lateral wall of the pharynx.
Both of these muscles act to shorten the pharynx and to draw the larynx and esophagus
toward the root of the tongue in swallowing. The stylopharyngeus muscle arises from the
medial surface of the great comu of the stylohyoid bone, passes ventromedially and
enters the wall of the pharynx by passing between the pterygo-pharyngeus and the palato-
pharyngeus muscles. Its fibers radiate, many bundles passing forward, others inward or
backward beneath the hyopharyngeus. It raises and dilates the pharynx to receive the
bolus in swallowing (Sisson and Grossman 1975). In people, it is known that contraction
of the stylopharyngeus muscle dilates the dorsal part of the nasopharynx (Kuna and

Vanoye 1999).

The extrinsic muscles are part of the hyoid apparatus: geniohyoid, genioglossus,
stemohyoid, sternothyroid and hyoepiglotticus muscles. The hyoid apparatus is attached
to the petrous temporal bone and supports the root of the tongue, pharynx and larynx. The
basihyoid bone is closely related to the base of the tongue via its attachments to the
genioglossus and hyoglossus muscles. It articulates with the short ceratohyoid bone
located just cranial to it. The latter joints the stylohyoid bone, which extends from the
skull cranially. Forward and downward movement of the tongue will therefore enlarge
the nasopharynx by extending the ceratohyoid-stylohyoid articulation, thus enlarging the

dorsoventral diameter of the nasopharynx.

Role of the pharyngeal dilator muscles in people and laboratory animals

Because of the increased recognition of sleep disorders in people, and sleep apneas in
particular, many studies have been conducted in man and laboratory animals in order to
understand the role of intrinsic pharyngeal muscles, in particular those responsible for
dilation of the nasopharynx. Among intrinsic upper airway muscles, the pharyngeal
dilators responsible for enlarging the nasopharynx in response to different stimuli such as
upper airway negative pressures. The stylopharyngeus muscle is part of this group of
muscles. In normal awake goats, the stylopharyngeus muscle was found to exhibit tonic
expiratory and phasic inspiratory activity, whereas the pharyngeal constrictors were
active during expiration. This duality of action allows the nasopharynx to adapt its size
and shape to the airflow and resist collapse when large negative pressures are applied to
the upper airway (Feroah 2000). Furthermore, when undergoing slow wave (SWS) and

rapid eye movement (REM) sleep, the same animals showed a decrease in

stylopharyngeus muscle activity from awake to SWS and from SWS to REM sleep,
indicating a possible involvement of stylopharyngeus muscle in sleep apnea (Feroah
2001a). In those same conditions, when negative pressure was applied, the
stylopharyngeus muscle responded by increasing its activity both in awake and SWS, but
the activity did not change in REM sleep compared to controls. These data indicate that
the stylopharyngeus muscle does participate in the compensatory mechanisms that adjust
to increasing loads of negative pressure (Feroah 2001b). In one study performed on
anesthetized rabbits, it was found that another pharyngeal dilator, the genioglossus
muscle, responds to augmented upper airway negative pressures by increasing its activity
by several folds. Section of the motor output to the genioglossus muscle abolished this
response. The pharyngeal dilator muscles have a role in compensation for added
inspiratory load and the activation of these muscles facilitates the load compensating
function of diaphragmatic muscles by decreasing airway resistance (Aleksandrova 2002).
Furthermore, in a study conducted in man, the investigators found that sudden upper
airway negative pressure applied at the end of expiration activates the genioglossus
muscle. They concluded that a reflex pathway was present that resulted in pharyngeal
dilator muscle activation in the presence of intrapharyngeal forces that otherwise may

narrow or close the pharynx (Homer 1991).

Role of the stylopharyngeus muscle
The stylopharyngeus muscle is innervated by the glossopharyngeal nerve, which is a
mixed nerve containing both motor and sensory fibers (Sisson 1976). The latter constitute

the bulk of the nerve and include those that mediate the sense of taste. The

glossopharyngeal nerve, after exiting the cranial cavity, curves ventrad and rostrad over
the guttural pouch and caudal to the thyrohyoid bone, crosses the deep face of the
external carotid artery and divides into pharyngeal and lingual branches. It also gives off
collateral branches, which are the tympanic nerve, the lesser petrosal nerve, and a branch
to the carotid sinus. The very small branch of the glossopharyngeal nerve to the
stylopharyngeus muscle arises from the dorsal border of the nerve. The pharyngeal
branch runs rostrad across the deep face of the stylohyoid bone and joins the pharyngeal
branches of the vagus and sympathetic filaments in forming the pharyngeal plexus;
several branches pass to the muscles and mucous membranes of the pharynx. The lingual
branch is the continuation of the trunk and runs along the caudal border of the stylohyoid
bone rostral to the linguofacial trunk before it dives under the hyoglossus muscle (Sisson
and Grossman 1976, Ozveren 2003). This cranial nerve has mostly sensory fibers, but
provides motor innervation to only one skeletal muscle, the stylopharyngeus. Sensory
fibers are distributed in the mucous membranes of the pharyngeal walls, soft palate,

isthmus faucium or the mucosa of the oral pharynx, and tonsils (Sisson 1976).

The role of cranial nerves, glossopharyngeal nerve in particular, has been extensively
studied in a human model for obstructive sleep apnea. Results show that electrical
stimulation of the glossopharyngeal nerve causes pharyngeal dilation in cats, by
stimulating contraction of the stylopharyngeus muscle (Iizuka 2001; Kuna 2001).
Furthermore, in adult rats, the glossopharyngeal nerve exhibits respiratory burst activity
that starts in the late expiratory phase and terminates at the end of the inspiratory phase

(Frugiere and Barillot 1994). Afferent branches of the glossopharyngeal nerve have

therefore been implicated in the pathogenesis of dynamic collapse of the upper airway in

other species.

The upper airway dilating muscles are activated in sequence

In many species, the dilating muscles of the nasopharynx are recruited in a coordinated
sequence within the respiratory pattern in order to stabilize the nasopharynx when
negative upper airway pressures are produced. In general, phasic activation of upper
airway muscles precedes inspiratory activity of the diaphragm, and augmented upper
airway pressures increase this amount of pre-activation (Van Lunteren 1984). Negative
upper airway pressure has also been shown to reflexly augment phasic inspiratory and
tonic electromyographic (EMG) activity in many upper airway dilator muscles and to
play an important role in regulating EMG activity of these muscles during breathing in
animals and humans (Van Lunteren 1984, Homer 1991, Van Der Touw 1994). In awake
goats, the stylopharyngeus muscle has been shown to display phasic inspiratory discharge
before the onset of diaphragmatic activity and its activity peaks before peak
diaphragmatic activity. A tonic discharge was also observed throughout respiration,
except at the beginning of expiration (Feroah 2000). Reflex augmentation of upper
airway muscle activity may have functional significance in the maintenance of upper
airway patency. It potentially prevents upper airway collapse in the face of negative
pressure oscillations in the upper airway or facilitates recovery when large negative
pressure changes are produced by inspiratory efforts against an obstructed airway

(Mathew 1984).

Influence of upper airway receptors on recruitment of upper airway muscles

Glossopharyngeal afferent fibers found beneath the nasopharyngeal mucosa mediate the
sniff-like aspiration reflex elicited by mechanical deformation of the nasopharyngeal
mucosa (Sant’Ambrogio et al. 1995). Some of the receptors within the nasopharynx
innervated by the glossopharyngeal nerve respond to positive and negative pressure, but
the pharynx is not an important site for reflex generation and stimulation of upper airway
patency maintaining muscles. (Homer et al. 1994) Sectioning the glossopharyngeal
nerve caused increased rather than decreased response to pressure with in the

nasopharynx (Mathew et al. 1988).

The laryngeal mucosa has an abundant supply of sensory receptors controlling a complex
pattern of respiratory reflexes that influence the patency of the upper airway and the
breathing pattern. Three types of laryngeal receptors have been identified, responding
specifically to transmural pressure, airflow and contraction of the laryngeal muscles
(Sant’Ambrogio 1985). Laryngeal pressure receptors respond to either negative or
positive pressure and a few respond to both. Reflex alterations in the breathing pattern
and upper airway muscle activity during upper airway pressure changes are mediated by
such receptors. (Mathew 1984). The laryngeal afferents follow 3 routes: the internal
branch of the superior laryngeal nerve (SLN), the external branch of the same nerve, and
the recurrent laryngeal nerve (Mathew and Sant’Ambrogio 1988). Section of the superior
laryngeal nerve has been shown to decrease the EMG activity of the pharyngeal dilator

muscles in response to upper airway negative pressure and to decrease the level of

negative pressures required to elicit upper airway collapse (Hornet 1991). Furthermore,
the reflex activation of upper airway dilators by increase of negative pressures in the
upper airway is abolished by topical anesthesia of the laryngeal mucosa (Van Lunteren
1984). Desensitization of the laryngeal mucosa has been shown to induce nasopharyngeal

collapse and upper airway obstruction in normal exercising horses (Holcombe 2001).

The laryngeal mucosa has been shown to contain multiple mechanoreceptors involved in
the reflex pathways involving recruitment of the upper airway muscles (Sant’Ambrogio
1983). The stimulus capable of eliciting such reflex is the presence of subatrnospheric
pressure in the upper airway. Laryngeal receptors are believed to be the principal
transducing elements that initiate this response (Kuna 1991). In human studies, it has
been shown that upper airway dilator muscles can be activated throughout inspiration via
ongoing upper airway mechanoreceptor reflexes. Such a feedback mechanism protects
upper airway patency within each breath in awake humans (Akahoshi 2001). These
receptors are thought to respond to negative pressures and their activity increases in face
of upper airway obstruction, therefore promoting upper airway patency (Sant’Ambrogio

1983).

We have seen that upper airway dilating muscles and the stylopharyngeus muscle in
particular play an important role in supporting the nasopharynx during inspiration in
people and laboratory animals. These muscles are recruited in sequence during the
breathing effort to maintain nasophaynrgeal patency when negative inspiratory pressures

are applied to the nasopharynx. In horses, little is known about the pathogenesis of

10

nasopharyngeal collapse and it seemed likely that the stylopharyngeus muscle would be
involved in this phenomenon. Furthermore, it is yet unknown how the upper airway
dilating muscles are recruited and activated during exercise in horses, information which

could be very useful in the understanding of dynamic upper airway disorders.

Aim of the thesis

The aim of this thesis is to investigate the role of the stylopharyngeus muscle in dorsal
nasopharyngeal function in exercising horses and attempt to explain the pathophysiology
of dorsal nasopharyngeal collapse in exercising horses.

Our first hypothesis was that bilateral anesthesia of the glossopharyngeal nerves causes
stylopharyngeus muscle dysfunction and subsequent dorsal pharyngeal collapse and
upper airway obstruction in exercising horses. I determined if bilateral local anesthesia of
the glossopharyngeal nerve resulted in stylopharyngeus muscle dysfunction and
subsequent dorsal pharyngeal collapse. The second hypothesis was that the
stylopharyngeus muscle has inspiratory activity that increases with exercise intensity. I
then aimed to correlate the electromyographic activity of the stylopharyngeus muscle to

the breathing pattern in horses exercising on the treadmill.

ll

CHAPTER 1

Effect of stylopharyngeus muscle dysfunction on the nasopharynx in exercising horses
Reasons for performing the study: To determine the function of the stylopharyngeus
muscle on nasopharyngeal function in horses and to investigate a potential cause of
dorsal nasopharyngeal collapse in horses.

Objective: To determine the effect of bilateral glossopharyngeal nerve block and
stylopharyngeus muscle dysfimction on nasopharyngeal function and airway pressures in
exercising horses.

Methods: Endoscopic examinations were performed on horses at rest and while the horses
were running on a treadmill at speeds corresponding to HRmaxso, HRmax75, and HRmax
while upper airway pressures were measured, with and without bilateral
glossopharyngeal nerve block.

Results: Bilateral glossopharyngeal nerve block caused stylopharyngeus muscle
dysfunction and dorsal nasopharyngeal collapse in all horses. Peak inspiratory upper
airway pressure was significantly (P = 0.0069) more negative at all speeds and respiratory
frequency was lower (P = 0.017) in horses with bilateral glossopharyngeal nerve block
and stylopharyngeus muscle dysfimction compared to control values.

Conclusions: Bilateral glossopharyngeal nerve anesthesia produced stylopharyngeus
muscle dysfimction, dorsal pharyngeal collapse, and airway obstruction in all horses.
Potential relevance: The stylopharyngeus muscle is likely an important nasopharyngeal
dilating muscle in horses and dysfunction of this muscle may be implicated in clinical

cases of dorsal nasopharyngeal collapse.

12

Introduction

Nasopharyngeal collapse has been clinically observed in horses as a potential cause of
exercise intolerance and upper respiratory noise (Kanniegeter and Dore 1995). Ventral
displacement of the roof of the nasopharynx and/or axial displacement of the lateral walls
of the nasopharynx are common features observed in exercising horses with this disease
(Kanniegeter and Dore 1995). Unfortunately, the cause of this obstructive disease is
unknown. No treatment is available for this condition and affected horses are often

retired from performance.

The action of several groups of muscles is responsible for stiffening and dilating the
nasopharynx (F eroah 2000; Kuna 2001; Kuna and Vanoye 1999; Kuna et al.1997). These
same muscles are also important during deglutition (Kuna and Vanoye 1999). This
dichotomy of action, contracting the pharyngeal walls during swallowing and dilating the
airway during breathing, seems contradictory. However, these muscles are uniquely
situated to perform both activities, because the pharynx is a conduit for both food and air.
The dorsal or superior pharyngeal constrictor muscles contract during swallowing,
forming a sphincter and moving the food bolus into the esophagus. These same muscles
have tonic activity during respiration such that they form a rigid pharyngeal roof during
breathing (Kuna 2001; Kuna and Vanoye 1999). Contraction of the stylopharyngeus
muscle dilates the dorsal part of the nasopharynx in people (Kuna and Vanoye 1999). In
horses, this muscle originates on the axial aspect of the distal portion of the stylohyoid
bone. It courses rostroventrally and ramifies in the wall of the dorsal nasopharynx, by

passing between the pterygopharyngeus and palatopharyngeus muscles (Fig 1). In

13

horses, contraction of the stylopharyngeus muscles raises the roof of the pharynx and
dilates the pharynx to receive the bolus during swallowing (Sisson 1976). In a similar
manner, we hypothesized that during breathing, contraction of the stylopharyngeus
muscle pulls the nasopharyngeal wall dorsally thereby supporting the dorsal wall of the

nasopharynx and preventing dynamic collapse of this area during inspiration.

The glossopharyngeal nerve provides motor innervation to the stylopharyngeus muscle.
This cranial nerve has mostly sensory fibers, but provides exclusive motor innervation to
only one skeletal muscle, the stylopharyngeus. Electrical stimulation of the
glossopharyngeal nerve causes pharyngeal dilation in cats, by stimulating contraction of
the stylopharyngeus muscle (Iizuka 2001; Kuna 2001). Therefore, the purpose of our
investigation was to determine if bilateral glossopharyngeal nerve blockade and the
resulting stylopharyngeus dysfunction would cause collapse of the dorsal nasopharynx in

exercising horses and if the induced collapse would result in airway obstruction.

Material and Methods

Horses—Six horses (4 Standardbreds and 2 mixed-breeds, 2 geldings and 4 mares, 4 to
13 years old, weighing between 400 and 427kg) were used in this experiment, which was
approved by the All-University Committee for Animal Use and Care at Michigan State
University. These horses had normal upper airway function based on physical
examinations and endoscopic examinations of the larynx and nasopharynx at rest and

during high-speed treadmill exercise.

14

Training procedure—All horses were trained to run on the treadmill prior to onset of the
study. Maximum heart rate (HRmax) was determined during an incremental exercise test
that consisted of a warm-up period of 3 minutes at a speed of 4 m/s, 2 minutes at a speed
of 6 m/s, and 1 minute at increasing speeds of 8, 10, 11, 12, and 13m/s or until the horses
became fatigued and were unable to maintain their position on the treadmill despite
humane encouragement. During this test, heart rate was recorded by use of a telemetry
system.1 The speeds corresponding to 50% of maximum heart rate (HRmaxso) and 75% of

maximum heart rate (HRmax75) were interpolated from this data.

Glossopharyngeal nerve anesthesia - An injection apparatus was made using a 12 ml
syringe attached to a 30 cm length of polyethylene tubing (I.D. 1.19 mm, 0D. 1.7 mm)
with a 22 gauge needle without its hub attached to the end of the tubing. The tubing was
inserted through a sclerotherapy needle2 apparatus after the inner tubing of this apparatus
was removed. The apparatus was then passed through the biopsy channel of the
endoscope. The apparatus was used to inject local anesthetic around the glossopharyngeal
nerve, as it coursed rostroventrally through the medial compartment of the guttural
pouch. The injection site was rostral to the external carotid artery. This placement was
chosen because the hypoglossal nerve dives deep and caudal to the external carotid
artery. In this location, the glossopharyngeal nerve is isolated such that the nerve block

would be specific for the glossopharyngeal nerve.

 

' Digital telemetry system, MI 403A. Hewlett Packard, Palo Alto, California, USA

2 Millrose Laboratories, Mentor, Ohio, USA

15

Horses were restrained in a set of stocks and a lip twitch was applied. The video-
endoscope was passed through the right naris, into the right guttural pouch. The
glossopharyngeal nerve was identified coursing across the external carotid artery within
the medial compartment of the guttural pouch. Perineural anesthesia was performed by
injecting 0.5 to 1 cc of 2% mepivicaine hydrochloride3 beneath the guttural pouch
mucosa, directly over the glossopharyngeal nerve, just rostral to the external carotid
artery. Anesthesia of the left glossopharyngeal nerve was achieved in a similar manner.

Upper airway pressure measurements - Upper airway pressures were measured in horses
running on the treadmill. Using endoscopic guidance, a ISO-cm polyethylene (inner
diameter, 2.15 mm; outer diameter, 3.25 mm) catheter with 6 holes on the side of the
catheter beginning a distance of 26 mm from the sealed tip was passed through the right
nostril and through the laryngeal opening into the trachea (N ielan et al. 1992). The tip of
this catheter was placed approximately at the junction of the proximal and middle thirds
of the cervical portion of the tracheal. The catheter was connected to a differential
pressure transducer4 and upper airway pressures were recorded by use of a chart
recorders. The differential pressure transducer was calibrated by use of a water
manometer before each experiment. Peak inspiratory and expiratory tracheal pressure
and respiratory frequency were determined from the pressure recorder tracings. Ten

consecutive breaths were averaged to determine each data point.

 

3 The Upjohn Company, Kalamazoo, Michigan, USA
4 DP-4522, Validyne Engineering Sales, Northridge, California, USA

5 Dash 11, Astromed, West Warwick, Rhode Island, USA

16

Experimental Design- Horses were fasted for a minimum of 3 hours before the
experiment to minimize gastric contents. Twenty minutes after the bilateral
glossopharyngeal nerve block was performed, the horse was restrained with a lip twitch
and the endoscope was passed through the right nostril. The horse's nares were occluded
for 60 second while observing the nasopharynx with the endoscope to assess upper
airway function. The endoscopic examination was recorded. To test if the
glossopharyngeal nerves had been anesthetized, a biopsy instrument was used to probe
the dorsal and lateral walls of the nasopharyngeal mucosa and the soft palate. Lack of
swallowing or gagging in response to the tactile stimulation indicated successful
anesthesia of the glossopharyngeal nerves. With endoscopic guidance, a catheter was
placed into the trachea. Horses were then exercised on the treadmill for 3 minutes at 4
m/s and l min at the speed corresponding to HRmaxso. Subsequently, a videoendoscope
was placed through the right naris and secured to the halter with rubber tubing. Horses
were exercised at HRmaxso, Hmens, and HRmax for 1 min at each speed. Upper airway
pressure measurements were recorded throughout the exercise trial. All endoscopic
examinations were videotaped so that upper airway function could be assessed. Horses
performed the exercise trial twice: once following bilateral glossopharyngeal nerve block
and once without the nerve block. The sequence of exercise trials was randomized so
that 3 horses performed the trial with anesthesia first, and 3 horses performed the trial
first without anesthesia.

Data analysis

17

Data were analyzed using a two-way analysis of variance, with speed and nerve block as
the main factors. Post hoc comparisons, where appropriate, were made using the Student-
Newman-Keuls test. A significance level of P < 0.05 was selected.

Results

All horses tolerated the glossopharyngeal nerve block well and bilateral glossopharyngeal
nerve anesthesia was obtained in every horse, such that none of the horses reacted to
tactile stimulation of the dorsal pharynx by gagging or swallowing. All horses completed
the exercise protocol. None of the horses showed signs of dysphagia, coughing, or

respiratory difficulty following bilateral glossopharyngeal nerve anesthesia.

All horses had normal upper airway function during upper airway endoscopy, both at rest
while the nares were occluded and during exercise, without bilateral glossopharyngeal
nerve block. In contrast, nasal occlusion at rest caused dorsal pharyngeal collapse in 4
out of 6 horses following bilateral glossopharyngeal nerve anesthesia (Fig 3).
Furthermore, the dorsal nasopharynx collapsed in all horses during treadmill exercise
(Fig 4). Specifically, during inspiration, the dorsal wall of the nasopharynx collapsed
ventrally such that the corniculate processes of the arytenoid cartilages could not be seen.
None of the horses displaced their soft palate or collapsed the lateral walls of the
nasopharynx. Two horses had endoscopic evidence of only unilateral collapse during
resting and treadmill endoscopy. In these horse the dorsal hemi-nasopharynx collapsed
ventrally during inhalation, while the contralateral side maintained a normal position (Fig
5). This unilateral collapse was likely due to ineffective glossopharyngeal nerve block on

the non—collapsing side. Bilateral glossopharyngeal anesthesia was performed

18

successfully on these two horses 10 days to 2 weeks later, and only data obtained from

successful bilateral blockade protocols was analyzed.

As treadmill speed increased from HRmaxso to HRmms and from HRmms to HRmax, peak
inspiratory upper airway pressure became significantly more negative. Also, as speed
increased from HRmaxso to HRmax, respiratory frequency increased. There was no
significant effect of treadmill speed on peak expiratory upper airway pressure. The main
effects of the bilateral glossopharyngeal nerve block were significantly (P=0.0069) more
negative peak upper airway inspiratory pressure (Fig. 6) and significantly (P = 0.017)
lower respiratory frequency. With bilateral glossopharyngeal nerve block, peak
inspiratory upper airway pressures (mean 3: SEM) at HRmaxso, HRmms, and HRmax were —
21.0 :1: 2.67, -32.1 :I: 3.41 and —37.4 :t 2.71 cm H20, respectively, compared with -15.0 i
0.94, -23.3 :t 1.57, and —30.4 d: 1.50 cm H20 without the nerve block. Also, respiratory
frequency at HRmaxso, HRmms, and HRmx was 63 :t 4.8, 79 i8.6, and 96 i: 11.8 breaths
per minute with the nerve block compared to 71 i 4.2, 83 :l: 9.2, and 99 i 10 breaths per
minute without the nerve block. We were unable to detect a significant effect of bilateral

glossopharyngeal nerve block on peak upper airway expiratory pressures.

Discussion

The results of the study reported here documented that dysfunction of the
stylopharyngeus muscle, induced by bilateral glossopharyngeal nerve block, resulted in
ventral displacement of the dorsal nasopharyngeal wall in exercising horses. Following

bilateral glossopharyngeal nerve block, peak inspiratory airway pressures were more

19

negative at all speeds, suggesting that stylopharyngeus dysfunction and the resulting
nasopharyngeal collapse caused airway obstruction. Therefore, based upon this
information, we conclude that the stylopharyngeus muscle is an important

nasopharyngeal dilating and stabilizing muscle in horses.

Bilateral glossopharyngeal nerve block resulted in dynamic collapse of the dorsal
nasopharynx, both at rest when the horses’ nares were occluded, and while the horses ran
on the treadmill. Specifically, the dorsal wall of the nasopharynx collapsed ventrally
during inspiration, obscuring the view of the corniculate processes of the arytenoid
cartilages and partially obstructing the rima glottidis. Anatomic considerations suggest
that this dynamic obstruction was caused by stylopharyngeus muscle dysfunction. The
stylopharyngeus muscles insert on the dorsal nasopharyngeal wall, perpendicular to it,
such that contraction of the stylopharyngeus muscles raises the wall of the dorsal
nasopharynx, expanding and supporting the dorsal nasopharynx, and preventing collapse

as pressures within the airway become more negative during inspiration.

Bilateral glossopharyngeal nerve anesthesia also resulted in more negative peak
inspiratory airway pressures indicating that the induced nasopharyngeal collapse caused
airway obstruction. The average decrease in peak inspiratory pressure was approximately
8 cm H2O or a 25% decrease or more negative inspiratory pressure. This change in
inspiratory pressure was small compared to the change in inspiratory pressure measured
in horses with left laryngeal hemiplegia, which approached 30 to 40 cm H2O. (Tetens et

al. 1996) Despite this observation, the obstruction resulted in an approximate 25%

20

decrease in peak inspiratory pressure, suggesting that seemingly mild nasopharyngeal

collapse in horses, clinically, may have significant functional consequences.

Interestingly, the horses with the most severe collapse of the dorsal nasopharynx, based
on endoscopic examination, did not have the largest changes in peak inspiratory airway
pressure: some horses had minor collapse, as viewed endoscopically, with large changes
in inspiratory pressures. This might be explained by the fact that while a decrease in
nasopharyngeal diameter is easily observed through the endoscope, the length of the
obstruction is harder to evaluate. The stylopharyngeus muscle has a broad insertion that
is approximately 4 cm long (based upon measurements made in 10 cadaver horse heads).
The nasopharyngeal collapse induced by bilateral glossopharyngeal nerve anesthesia
obstructed the dorsal portion of the rima glottidis in a sagittal plane but also obstructed
the nasopharynx along a length of at least 4 cm, based upon the broad insertion of the
stylopharyngeus muscle. This is important because airway resistance is indirectly
proportional to the radius but also directly proportional to the length of the affected
segment. Therefore, the airway pressure measurements may more accurately describe the

degree of airway obstruction in each horse.

The glossopharyngeal nerve is primarily a sensory nerve and provides exclusive motor
innervation to the stylopharyngeus muscle (Sisson 1976). The pharyngeal mucosa
contains a variety of sensory receptors including pressure and stretch receptors, chemical
and mechanical receptors and nocioceptive endings that when triggered, stimulate the

swallowing reflex.(Sant’Ambrogio et al. 1995) Glossopharyngeal afferent fibers found

21

beneath the nasopharyngeal mucosa mediate the sniff-like aspiration reflex elicited by
mechanical deformation of the nasopharyngeal mucosa.( Sant’Ambrogio et al. 1995)
Some of the receptors within the nasopharynx innervated by the glossopharyngeal nerve
respond to positive and negative pressure, but the pharynx is not an important site for
reflex generation and stimulation of upper airway patency maintaining muscles. (Homer
et al. 1991) Sectioning the glossopharyngeal nerve caused increased rather than
decreased response to pressure with in the nasopharynx (Mathew et al 1988). Therefore,
it is unlikely that the nasopharyngeal collapse created by bilateral glossopharyngeal nerve

block was due to decreased sensory activity within the nasopharynx.

In addition to providing motor innervation to the stylopharyngeus muscle, studies in
rhesus monkeys, dogs and rats indicate that branches of the glossopharyngeal nerve could
also provide some motor fibers to the levator veli palatini, and superior or dorsal
pharyngeal constrictor muscles, via fibers entering the pharyngeal plexus and joining
fibers from the vagus nerve (Furusawa et a1. 1996, Nishio et al. 1976, Venker-van
Haagen et al. 1986). It is unlikely that the dorsal nasopharyngeal collapse created here by
bilateral glossopharyngeal nerve block was caused by dysfunction of the levator veli
palatini or superior pharyngeal constricting muscles, because the principle motor supply
to these muscles is the vagus nerve, which was not anesthetized in this study. As well,
we did not appreciate dysphagia in any of the horses following bilateral glossopharyngeal
nerve block, which would occur with levator veli palatini and dorsal pharyngeal

constricting muscle dysfunction (DeLahunta, 1977; Mayhew 1989).

22

Glossopharyngeal nerve integrity is traditionally evaluated along with the vagus nerve by
testing swallowing function. Interestingly, none of the horses in this study showed
clinical signs of dysphagia following bilateral glossopharyngeal nerve anesthesia. The
horses never had saliva pooling at the nares or coughed during the exercise trial.
Furthermore, approximately 30 min after the end of the exercise trial the horses were
given water and hay and monitored closely for 2 hours. No signs of dysphagia were
observed in any of the horses in this study. During swallowing, constriction of the
pharyngeal lumen, in a peristaltic sequence, results from displacement of the tongue,
hyoid bone and larynx, and from contraction of the pharyngeal constrictor and dilator
muscles (Bosman et al. 1986). It is possible that, rather than causing a complete
dysphagia with lack of propulsive activity from the pharynx, bilateral glossopharyngeal
nerve block resulted in changes in the sequence or relative timing of swallowing that
could not be detected clinically. Similarly, following bilateral glossopharyngeal nerve
sectioning in dogs, the animals showed no clinical signs of dysphagia, but swallowing
dynamics and sequence was altered, based on fluoroscopic studies (V enker—van Haagen

et al. 1986).

All horses significantly decreased their respiratory frequencies after glossopharyngeal
blockade. The decrease in respiratory frequency was small, approximately 3 to 10
breaths per minute, but it occurred in 5 of 6 of horses at all speeds. We did not measure
tidal volume or airflow in this study, and therefore, attempted to explain the decreased
respiratory frequency as a technique employed by the horses to compensate for the

imposed airway obstruction. In the face of upper airway obstruction caused by the dorsal

23

nasopharyngeal collapse, the resistance to the airflow through the upper airway likely
increased. With increased resistance to breathing inspiratory time is increased, in an
attempt to maintain tidal volume, ultimately resulting in decreased respiratory frequency
(Daubenspeck 1981). Therefore, the horses modified their breathing technique by
lowering respiratory frequency in response to the imposed airway obstruction caused by

nasopharyngeal collapse.

24

CHAPTER 2

Electromyographic activity of the stylopharyngeus muscle in exercising horses.
Summary

Reasons for performing the study: To determine if the stylopharyngeus muscle has
respiratory-related activity that increases with exercise intensity in horses.

Objective: To measure the electromyographic activity of the stylopharyngeus muscle in
exercising horses and correlate it with the breathing pattern.

Methods: Electromyographic activity of the stylopharyngeus muscle and upper airway
pressures were recorded in normal horses running on a treadmill.

Results: The stylopharyngeus muscle has phasic inspiratory activity that increases with
speed.

Conclusions: The stylopharyngeus muscle is an upper airway dilating muscle that
functions principally during inspiration and its activity increases with exercise intensity.
Potential relevance: The stylopharyngeus muscle is one of a group of upper airway
dilating muscles that functions to support structures of the nasopharynx during

inspiration.

25

Introduction

Nasopharyngeal collapse has been clinically observed in horses as a cause of exercise
intolerance and upper respiratory noise (Kannegieter 1995). When this disease occurs,
collapse of the dorsal wall of the nasopharynx is one of the features observed in horses
during treadmill exercise. In horses, little is known about the activity of the pharyngeal
muscles in relation to the breathing pattern and the pathways by which they become
recruited during respiration to maintain nasopharyngeal patency. In a previous study we
documented that anesthesia of the glossopharyngeal nerves and the resulting
stylopharyngeus muscle dysfunction caused dorsal pharyngeal collapse and inspiratory
upper airway obstruction in normal horses during exercise. We concluded that the
stylopharyngeus muscle is an important dilator of the dorsal nasopharynx in horses and
that dysfunction of this muscle could be implicated in clinical cases of dorsal pharyngeal

collapse (Tessier 2004).

Before and during inspiration, the pharyngeal muscles are recruited and activated in a
coordinated sequence in order to stabilize the nasopharynx in response to negative
intraluminal pressures applied to the upper airway (Van Lunteren 1988). Upper airway
muscles show a very rapid increase in activity that precedes the onset of respiratory pump
muscle activation by as much as 200ms (Van Lunteren 1983; Kuna 1991). In other
species, the stylopharyngeus muscle has been shown to exhibit respiratory related activity
(Feroah 2000). Therefore, we hypothesized that in horses the stylopharyngeus muscle
would have inspiratory related activity that would increase with increased breathing

intensity induced by increasing levels of exercise.

26

Material and Methods

Horses—Five Standardbred horses (3 geldings and 2 mares, aged 5-12 years) were used
in the experiment, which was approved by the All-University Committee for Animal Use
and Care at Michigan State University. The horses were determined to be normal based
on physical and endoscopic examinations of the larynx and nasopharynx at rest and
during high-speed treadmill exercise. All horses were trained to run on the treadmill
prior to the experiment. Maximum heart rate (HRmax) was determined by use of a
telemetry system1 and an incremental exercise test. The speeds corresponding to 50%
(HRmaxSO) and 75% (HRmax75) of maximum heart rate were interpolated from these data.
Electrode placement- Bipolar fine wire electrodes and a ground wires were constructed
using Teflon-coated wirez. A small amount of resin cement3 was applied to the end of
the electrode. The electrode was placed in the stylopharyngeus muscle through the
nasopharyngeal opening of the guttural pouch while the horse was anesthetized. The
horse was pre-medicated with xylazine (0.04 mg/kg, IV), anesthesia was induced with
ketamine (2.2 mg/kg, IV) and diazepam (0.1 mg/kg, IV) and the horse was positioned in
stemal recumbency with its head elevated. Anesthesia was maintained by use of an
intravenous infusion of 5% guaifenesin containing 0.05 mg of xylazine/ml and 2.0 mg of
ketamine/ml, delivered at 2 ml/kg/h. Twenty gauge needles with the hubs removed were
secured to each end of the bipolar electrode. The needle was grasped with the biopsy

instrument and pulled into the biopsy channel.

 

1 Digital telemetry system, MI 403A. Hewlett Packard, Palo Alto, California, USA
2 Teflon-coated wire, No. A5637, Coonerwire, Chatsworth, PA

3 Resin cement kit, Justi Products, Oxnard, CA

27

The endoscope was then passed through the right nostril and into the right guttural pouch.
The bipolar electrode was positioned into the stylopharyngeus muscle by pushing the
needle attached to the electrode wire out of the biopsy channel and through the origin of
the stylopharyngeus muscle on the stylohyoid bone, using the biopsy instrument. This
procedure was performed twice to seat both segments of the bipolar electrode. The
electrode wires exited the guttural pouch and were passed through the nostril using a 14-
gauge needle and secured to the horse's muzzle using elastic tape. A ground wire was
placed in the subcutaneous space, just over the right sternocephalicus muscle. The
following day, muscle function measurements were made while the horses exercised on a

treadmill.

Experimental design- A 150cm long catheter4 Qsolyethylene tubing, 2.15 mm ID, 3.25
mm OD) with 6 side holes beginning a distance of 8 catheter diameters from the sealed
tip, was placed through the left naris and secured to the muzzle with tape. The catheter tip
was placed at the level of the nasopharyngeal opening of the guttural pouch in the
nasopharynx to measure upper airway pressures so that the EMG activity could be
correlated with respiration. Horses were initially exercised on the treadmill for 3 minutes
at 4 m/s and 1 min at the speed corresponding to HRmaxso. Then, the exercise trial
consisted of three continuous one-minute segments of exercise at HRmaxso, HRmms, and
HRmax respectively. Upper airway pressure measurements and EMG activity were
recorded throughout the exercise trial. Peak inspiratory and expiratory pressures and

respiratory frequency were determined from the pressure tracings.

 

4 Baxter Scientific Products, McGaw Park, IL

28

Ten consecutive breaths were averaged to determine each data point. Following the
exercise trial, the horses were rested until their heart and respiratory rates had returned to
normal.

Upper airway pressure measurements - The nasopharyngeal catheter was connected to a
differential pressure transducer5 and upper airway pressures were recorded by use of a
chart recorder6 used to record the EMG tracings. The differential pressure transducer was
calibrated by use of a water manometer to 5, 10, and 20 cm of H20 before each
experiment. The pressure tracing was used to correlate the EMG activity of the
stylopharyngeus muscle with the breathing pattern.

EMG measurements- The EMG signals were processed through a sixth-order Butterworth
filter (band pass, 50 to 5000 Hz), amplified, rectified, and moving time-averaged with a
constant of 100 milliseconds. Both raw EMG and moving time average signals were
recorded7. Quantification of the EMG was performed by digitization of the moving time-
averaged EMG signal using an image acquisition and analysis, flame-grabber softwares.
Data analysis- Respiratory timing was based on the pharyngeal pressures and determined
using the EMG signals that were recorded simultaneously with pharyngeal pressure.
Mean electrical activity was calculated by dividing the total area under each moving time

average waveform above baseline by the duration of the electrical activity. The average

 

5 Dash 11, Astromed, West Warwick, Rhode Island, USA
6 DP-4522, Validyne Engineering Sales, Northridge, California, USA
7 LabVIEW 5.0, National Instruments Software, Austin, Texas

8 Scion Image, Scion Corp, Frederick, MD

29

mean electrical activity for 10 consecutive breaths at each speed was calculated for each
horse and expressed in arbitrary units. Data was analyzed using a one-way analysis of

variance with speed as the main factor. A significance level of P < 0.05 was selected.

Results

Electrodes were placed successfully in all horses. Complications included inadvertent
puncture of the external carotid artery during electrode implantation with minor
hemorrhage (n=2) and electrode removal by the animal before the data was collected

(n=2). When this occurred, the procedure was aborted and repeated two weeks later.

Phasic stylopharyngeus muscle activity increased at the end of expiration and peaked
early during inspiration while tonic activity was present during expiration (Figure 6).

The peak EMG activity of the stylopharyngeus muscle increased significantly with
exercise intensity (Figure 7). As treadmill speed increased from HRmaxSO to HRmms and
from Hmes to HRmx, peak inspiratory EMG activity of the stylopharyngeus muscle
increased (P=0.001) in each horse: mean value :t SEM were 2.44 i 0.493 arbitrary units
(a.u.) at HRmaxso, 5.18 i 0.965 a.u. at HRmax75 and 8.36 i: 1.321 a.u. at HRmax (Figure 8).
As speed increased from HRmaxso to HRmms and from HRmax'js to HRmax, mean electrical
activity of the stylopharyngeus muscle increased significantly (P< 0.001) in each horse
from 0.468 i 0.0628 a.u., to 1.34 i 0.294 a.u. and to 2.58 i 0.471 a.u., respectively
(Figure 7). There was no significant effect of treadmill speed on tonic electromyographic
activity of the stylopharyngeus muscle, but there was a trend towards an increase in

activity (P=0.089). As speed increased from HRmaxso to HRmms and from HRmax75 to

30

HRmax, tonic activity increased from 1.38 i 0.376 a.u. to 1.78 i- 0.414 a.u. and to 1.96 i

0.42 a.u., respectively.

Discussion

The results of this study demonstrate that the stylopharyngeus muscle has a phasic
inspiratory activity that increases with speed and breathing intensity in exercising horses.
Therefore, we conclude that the stylopharyngeus muscle functions to support and dilate

the dorsal nasopharyngeal wall in exercising horses.

The electromyographic activity of the stylopharyngeus muscle was synchronous with the
breathing cycle and tonic activity was present during early expiration. Phasic activity
began at the end of expiration and increased to peak activity during early inspiration,
similar to the activity measured in goats (Feroah 2000). This pattern is a characteristic
feature of upper airway muscles in mammals, in which a very rapid increase in phasic
activity precedes the onset of respiratory pump muscle activation (Kuna 1991). Such a
sequence of activity is essential to preserve airway stability and patency prior to the onset
of negative driving pressures that generate airflow. Minute ventilation increases from 60
liters to in excess of 1700 liters as horses accelerate from a resting state to maximum
speed (Erickson et al. 1991). Driving pressures of —40 cm H2O are required to create
airflows needed to maintain minute ventilation in horses during intense exercise.
Contraction of skeletal muscles is the principal means by which the nasopharynx and

larynx achieve dilation and do not collapse in the face of such negative driving pressures.

31

We observed that the peak and mean electrical activity of the stylopharyngeus muscle
increased significantly with speed, likely because breathing effort and exertion increased
as treadmill speed increased. More negative driving pressures are required to increase
airflow and minute ventilation as the demand for oxygen by the tissues increases.
Negative upper airway pressures have been shown to reflexly augment phasic inspiratory
and tonic EMG activity in many upper airway dilator muscles and to play an important
role in regulating EMG activity of these muscles during breathing (Van Lunteren 1983,
Homer 1991, Van der Touw 1994). This reflex is thought to increase airway patency.
Pharyngeal size and stability are major factors influencing airway collapsibility. Firstly,
the narrower the airway, the more vulnerable it is to closure and the more dependent it is
on pharyngeal dilator tone for patency. Secondly, the narrower the airway the greater the
inspiratory force necessary to achieve an adequate airflow. The resultant high transmural
pressures will favor dynamic pharyngeal collapse during inspiration. Increased
contraction of the stylopharyngeus muscle is therefore necessary to enlarge the upper
airway as the breathing effort increases to prevent dynamic pharyngeal collapse and

upper airway obstruction.

The laryngeal mucosa contains multiple mechanoreceptors that initiate the reflex
pathways involved in recruitment of upper airway muscles (Sant’Ambrogio 1983;
Tsubone 1998). The stimulus capable of eliciting this reflex is subatrnospheric pressure
within the laryngeal lumen (Sant’Ambrogio 1983; Van Lunteren 1983; Kuna 1991). In
people, upper airway dilator muscles are activated throughout inspiration via continued

stimulation of these transducing elements (Akahoshi 2001). These mechanoreceptors

32

respond to negative pressures and their activity increases in face of airway obstruction,
promoting airway patency (Sant’Ambrogio 1983). The internal branch of the superior
laryngeal nerve and the recurrent laryngeal nerve provide the afferent pathways for the
laryngeal mucosal receptors (Widdicombe et al. 1988). Sectioning of the superior
laryngeal nerve decreased the EMG activity of the pharyngeal dilator muscles in response
to upper airway negative pressure, and decreased the level of negative pressure required
to elicit upper airway collapse (Homer 1991). Other pathways are also involved in
recruiting upper airway muscles. In goats, the activity of the stylopharyngeus muscle
increases with augmented inspiratory drive achieved with hypercapnia and then decreases
parallel to a decrease in diaphragmatic muscle activity (Feroah 2000). Locomotion also
influences respiratory frequency and airway muscle activity. It is therefore likely that all
of these mechanisms are involved in modulating the activity of the stylopharyngeus

muscle during exercise in horses.

Desensitization of the laryngeal mechanoreceptors has been shown in horses to induce
nasopharyngeal collapse during exercise (Holcombe et al. 2001). Therefore, further work
is needed to evaluate how the stylopharyngeus muscle responds to these different stimuli
and in particular to the laryngeal mecanoreceptors in order to understand what leads to
failure of stylopharyngeus recruitment in clinical cases of nasopharyngeal collapse in

horses.

33

APPENDIX

34

Figure 1: Illustration of the origin and insertion of the stylopharyngeus muscle. Notice
how the muscles originate on the axial aspects of the stylohyoid bones and insert on the

muscles of the dorsal pharynx.

f": églqhugnfiurs

\ mated: s.

 

35

Fig 2a: Endoscopic image of the guttural pouch. Notice the origin of the stylopharyngeus

muscle (arrow) on the medial aspect of the stylohyoid bone.

a." \ll} l\.'h)'lil\l
- ' .

.\lctli.ll
tonlp.ulnicnl ' ' ‘
I

lateral
C(niuiurlnlcnl

 

Fig 2b: Cadaveric specimen showing the insertion of the stylopharyngeus muscle (white

asterisk) on to the musculature of the dorsal nasopharynx.

 

36

Fig 3: Endoscopic image of the nasopharynx of a horse running on the treadmill
following bilateral glossopharyngeal nerve block. Notice that the dorsal wall of the

nasopharynx has collapsed, obscuring the view of the corniculate processes of the

arytenoids cartilages.

 

Fig 4: Endoscopic view of the nasopharynx of a horse following right glossopharyngeal
nerve block, while the nares are occluded. Notice that only the right hemipharynx has

collapsed, while the left side remains in a normal position.

 

37

Fig 5: Peak inspiratory pressure in 6 horses during treadmill exercise with and without

bilateral glossopharyngeal nerve block. White bars: unblocked trials; Grey bars: with

glossopharyngeal nerve anesthesia.

 

Pressure (cm H20)

 

Peak inspiratory pressure in 6 horses during treadmill
exercise with and without bilateral glossopharyngeal

 

 

 

   

 

 

 

 

 

o nerve block .
-5 : HR HR HR
.10 t
-15 .
-20 .
-25 1
-30 . —1
-351
40 1
-45

 

 

 

Speed (%HRmax)

 

 

38

Figure 6. Recording of upper airway pressure (top tracing) and moving time averaged

EMG activity of the stylopharyngeus muscle. Notice that the EMG activity of the

stylopharyngeus muscle peaks during the early inspiration phase.

 

 

 

Figure 7. Moving time averaged EMG activity of the stylopharyngeus muscle (top
tracing) and raw EMG activity (bottom tracing) recorded at HRmaxso, HRImam 75, and
HRmax. Notice that the activity increases at each speed. Speed = 10 mm/sec

Speed = 10 mm/s

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

39

Figure 8. Graph of the peak EMG activity of the stylopharyngeus muscle in 5 exercising

horses. Notice that the activity increases at each speed for each horse. The thick black

line represents the average of the 5 horses.

 

Percent

 

Mean peak EMG activity of the stylopharyngeus muscle in
5 exercising horses

600 I
500 4
400 4
300 ~

200 - W

100 I I

 

 

50% 75% 100%

Speed (% maximum heart rate)

 

40

 

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41

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