An excised animal larynx model has been used in many studies to better understand the physiological and anatomical properties of the human larynx. One difference between an ex vivo model and an in vivo model is that ion loss occurs postmortem. To compensate for this in the excised model, researchers most commonly use a preservation method that includes completely submerging the specimen in isotonic saline (0.9% Na+Cl-) and then flash freezing it in liquid nitrogen. The flash freezing method allows researchers to maintain the integrity of the structures while also being able to gather specimens as they become available. Not enough research has been done to understand the effects of a preservation method on the outcomes of the study. Additionally, no common method has been established for preservation across studies to ensure that results are not being influenced by this variable. This prospective, mixed experimental design study includes three groups, a control group and two experimental groups. The control group consisted of 10 bench-mounted porcine larynges that were soaked in isotonic saline and flash frozen with liquid nitrogen. Prior to the experiment, the frozen larynges were thawed overnight before trials. The other two groups consisted of 10 bench-mounted porcine larynges each; these larynges were soaked in either isotonic saline or Ringer’s solution, a balanced fluid used in vivo to counteract dehydration. Larynges from these two groups were kept fresh and stored in a refrigerator overnight before trials. On the day of experimentation, each larynx was mounted on a bench top setup including three micropositioners to stabilize, adduct, and elongate the vocal folds. All the larynges were connected to a pseudolung via the trachea and humidified air was passed through to the vocal folds until phonation was achieved. Phonatory trials consisted of brief phonation followed by 5-minute desiccation intervals until phonation was no longer achieved. Phonation threshold flow (PTF), defined as the flow observed at the onset of phonation, was observed during each phonation trial; and flow values were compared within and between groups. Statistically significant differences were found between the Ringer’s group and the fresh saline group as well as between the Ringer’s group and the frozen saline group, indicating that PTF is influenced by the larynx preservation method.

Animal models are used extensively in voice research to study aspects of phonation, including physiology, kinematics, structure, and histology. Animals such as dog, cow, pig, sheep, deer, monkey, ferret, and rabbit have been used in voice research, with pig being one of the most common models. It is thought that the pig larynx is highly similar to the human larynx and one of the best models used in animal translational research. As with any model, however, the pig larynx does have some limitations. Perhaps a limitation most important to the rationale of this investigation is that pigs are difficult animals to study in vivo. Maintenance for a pig is challenging due to its large size and the variability of phonation use in the animal. Therefore, viable and practical alternatives are needed for in vivo voice research. The current study collected preliminary normative data from an alternate animal model, the rabbit, which has been used more recently in studies to model human phonation. The rabbit model was chosen due to its histological similarities to humans, in vivo phonation patterns, size, and practicality. The rabbit represents a more practical model for some longitudinal designs, as well as ex vivo phonatory models with aerodynamic measures as the primary variables. The current study involved a comparison of two aerodynamic measures, specifically phonation threshold pressure (PTP) and phonation threshold flow (PTF) between two groups, pig and rabbit larynges. The purpose of this study was to determine normative aerodynamic values for rabbits and to compare these with normative values for pigs during excised larynx benchtop phonation. Each group consisted of 15 larynges that were finely dissected to reveal the true vocal folds. Each larynx was then connected to a pseudolung and humidified air was passed through it. Fifteen phonation trials were elicited and the results averaged for each larynx. The results indicated that PTP and PTF were significantly different between the two groups, with PTP and PTF being lower for the rabbit group. Additionally, PTP values for rabbits were closer than pigs to the typical human value; however, some methodological challenges to rabbit benchtop models, including size and structural integrity, also exist. But the results from this study indicate that rabbits should be considered a viable option for voice research that would be more feasible with a small animal option that translates well to humans than a large animal option.

This thesis is part of a larger series of studies being conducted by Kristine Tanner, PhD, Associate Professor in the Department of Communication Disorders at Brigham Young University (BYU). The larger project is funded by the National Institute on Deafness and Other Communication Disorders at the National Institutes of Health. This thesis primarily investigated the effects of combination inhaled corticosteroids (ICs) on aerodynamic measures of the voice. In recent years, an increase in the localized laryngeal side effects from IC treatment, including dysphonia, have been reported. This study employed a between-groups experimental design, with two groups of rabbit larynges having been exposed to either ICs or nebulized isotonic saline two times each day for eight weeks at The University of Utah. For this study, the independent variable is group condition (i.e., IC versus saline) and the dependent variables are two aerodynamic measurements made at the onset of phonation using a benchtop experimental setup, namely phonation threshold pressure (PTP; cmH2O) and phonation threshold flow (PTF; L/min). The results of this study indicate a significant difference in PTP and PTF between vocal folds treated with IC as compared to vocal folds treated with nebulized isotonic saline solution. Implications of this study suggest negative changes in the voice due to IC treatment.

The purpose of this study was to examine external laryngeal oscillation and its potential effects on phonation threshold pressure (PTP; cm H2O) and phonation threshold flow (PTF; L/sec). Measures of PTP and PTF have inherent limitations due to the nonlinear nature of phonation, influencing their clinical and experimental utility. This is true particularly for tracking relatively small changes in voice function because variability in the measure itself can be larger than that resulting from voice change. Elevated PTP and PTF are associated with a variety of voice disorders and correlate with self-reported vocal effort and fatigue. Prior studies involving silicone and excised animal larynges have demonstrated PTP reduction in response to external oscillation. In an extension of this work, this thesis examined external laryngeal oscillation and aerodynamic voice measures in two experiments including a translational benchtop to human approach. Experiment 1 used a within-subjects counterbalanced design to examine PTF in 12 porcine larynges. Larynges were fitted with a custom oscillation device and 30 phonation trials were conducted for each larynx, 15 with external oscillation and 15 without. Although summary statistics indicated that PTF was lower with external oscillation, differences were not significant. Experiment 2 applied a within-subjects counterbalanced design to examine PTP in four healthy adult females and one healthy adult male. Individuals produced repeated syllable strings of /pi/ productions at comfortable pitch with and without external oscillation using an electrolarynx and the second and third syllables were averaged. Descriptive analysis indicated that PTP was lowered for female participants but not the male participant. Taken together, the results of these studies offer preliminary evidence that external oscillation influences voice onset aerodynamic measures. The effects of external oscillation seem to be more evident in PTP. These findings have important clinical and research applications for PTP measurement and the potential positive influence on voice function. These preliminary results indicate the need for further research in this area.

An excised animal larynx model has been used in many studies to better understand the physiological and anatomical properties of the human larynx. One difference between an ex vivo model and an in vivo model is that ion loss occurs postmortem. To compensate for this in the excised model, researchers most commonly use a preservation method that includes completely submerging the specimen in isotonic saline (0.9% Na+Cl-) and then flash freezing it in liquid nitrogen. The flash freezing method allows researchers to maintain the integrity of the structures while also being able to gather specimens as they become available. Not enough research has been done to understand the effects of a preservation method on the outcomes of the study. Additionally, no common method has been established for preservation across studies to ensure that results are not being influenced by this variable. This prospective, mixed experimental design study includes three groups, a control group and two experimental groups. The control group consisted of 10 bench-mounted porcine larynges that were soaked in isotonic saline and flash frozen with liquid nitrogen. Prior to the experiment, the frozen larynges were thawed overnight before trials. The other two groups consisted of 10 bench-mounted porcine larynges each; these larynges were soaked in either isotonic saline or Ringers solution, a balanced fluid used in vivo to counteract dehydration. Larynges from these two groups were kept fresh and stored in a refrigerator overnight before trials. On the day of experimentation, each larynx was mounted on a bench top setup including three micropositioners to stabilize, adduct, and elongate the vocal folds. All the larynges were connected to a pseudolung via the trachea and humidified air was passed through to the vocal folds until phonation was achieved. Phonatory trials consisted of brief phonation followed by 5-minute desiccation intervals until phonation was no longer achieved. Phonation threshold flow (PTF), defined as the flow observed at the onset of phonation, was observed during each phonation trial; and flow values were compared within and between groups. Statistically significant differences were found between the Ringers group and the fresh saline group as well as between the Ringers group and the frozen saline group, indicating that PTF is influenced by the larynx preservation method.

Subglottic stenosis (SGS) is an abnormal narrowing of the airway at the level of the cricoid cartilage, above the first tracheal ring and immediately beneath the vocal folds. Individuals with SGS experience a reduction in their ability to breathe as well as adverse effects on voice function. SGS can result from a variety of causes with the type of treatment depending on stenosis severity. Surgical techniques such as laryngotracheal and cricotracheal reconstruction are beneficial for airway maintenance; however, these procedures have resulted in negative effects on voice production. On the other hand, there are patients with SGS who do not require surgery and still experience voice problems. The purpose of this study was to quantify the effects of SGS on vocal fold vibration using an excised larynx benchtop mechanical model. Using a within-subjects repeated measures design, nine porcine larynges underwent experimental conditions including 0% (i.e., normal airway), 50% and 75% stenosed. The primary outcome measure was phonation threshold flow (PTF), which is the rate of flow observed at the onset of phonation. For all larynges, the normal and stenosed conditions were sampled three times each and averaged. Analysis of the results revealed no statistically significant differences in PTF; however, descriptive data showed decreases in PTF and increased variability in PTF values as percent stenosis increased. These findings lay important groundwork for future research in SGS, specifically those that employ ex vivo methodologies. PTF has emerged as a promising means of quantifying voice function in addition to the traditional onset pressure measures. Future studies should examine a broader range of stenosis conditions with a larger sample size to promote generalization to clinical populations including individuals with SGS.

Subglottic stenosis (SGS) is a narrowing of the airway below the vocal folds and above the trachea. This narrowing may be idiopathic or caused by scarring in the airway due to prolonged endotracheal intubation, radiation therapy, trauma, or gastroesophageal reflux disease. People who present with SGS often experience respiratory difficulty both at rest and during exertion. Breathing difficulty increases with stenosis severity. SGS is also associated with voice problems. Research has identified relationships among stenosis severity, voice function and certain types of surgical management; however, many aspects of these relationships are not fully understood due to the complexities of studying human phonation in this population. The purpose of the present study was to examine the effects of SGS on aerodynamic features of voice function using an excised larynx benchtop mechanical model. Specifically, this research involved the comparison of excised porcine vocal fold vibration at baseline and under experimental conditions of 50% and 75% stenosed. The dependent variable was phonation threshold pressure (PTP), the minimum pressure needed to initiate and maintain vocal fold vibration. PTP was analyzed for nine excised porcine larynges, sampled three times each, at baseline and the two stenosis conditions. The results of this study revealed no differences in PTP based on within-subjects comparisons. Because airflow changes with airway narrowing, this finding might indicate that other factors are responsible for the voice problems associated with SGS that were not accounted for in the current mechanical model. Vocal fold tone is not easily simulated in a benchtop setup and might be an important consideration for future studies. The quantification and manipulation of vocal fold adduction, as well as the study of high-speed imaging, could be useful in future work involving excised larynx mechanical models for the study of SGS. The results from this pilot work represent an important step toward optimizing the experimental setup for studying aerodynamic features of SGS.

Vocal fold hydration is important for efficient oscillation during voice production. Dehydration of the vocal fold surface is believed to produce adverse effects on the voice. Specifically, low environmental humidity, mouth breathing, and certain medical conditions may contribute to laryngeal and vocal fold dehydration. This dehydration effect may be quantified using the observed pressure and flow at the onset of phonation, operationally defined as phonation threshold pressure (PTP) and phonation threshold flow (PTF), respectively. Previous research has documented that nebulized isotonic saline (0.9% Na+Cl-) can reduce PTP. Additionally, the topical application of liquid saline increases vocal fold hydration in excised larynx studies. However, no studies have examined the prevention of vocal fold dehydration using aerosolized saline in an excised larynx mechanical model. The purpose of the current investigation was to determine the preventive effects of aerosolized isotonic saline in a physiologically realistic excised larynx model. Using a prospective, mixed experimental design with a control group, five bench-mounted, excised porcine larynges received 4-min doses of aerosolized saline delivered supraglottally for a total of 24 min. Subsequently, larynges received 1-min doses of desiccated air (<1% relative humidity) delivered supraglottally. A control group of five porcine larynges received only desiccated air. Phonation was attempted following each dose of aerosolized saline or desiccated air. The desiccation doses were repeated for both groups until the larynges were no longer able to phonate. The PTP and PTF were measured at baseline and following each dose of aerosolized saline or desiccated air. Analysis of the results indicated that aerosolized saline significantly delayed the adverse effects of vocal fold dehydration based on the total number of desiccation doses required to cease phonation for experimental versus control groups (p = .002). Trends demonstrated that PTP decreased after aerosolized saline and increased during desiccation trials. The PTF trends were similar during desiccation. The results from this study indicate that aerosolized saline may be used prophylactically to prevent vocal fold dehydration. These findings offer important advances in vocal fold hydration theory and dehydration prevention in a physiologically realistic excised mechanical model.

Previous studies have found vocal fold hydration to be crucial for healthy function of the vocal mechanism. Surface tissue hydration facilitates efficient vocal fold oscillation. The composition of vocal fold surface fluid includes protective water and mucus layers, similar to the fluid that covers the mucosa and epithelia of the upper airway. Laryngeal dehydration has been linked to several factors such as mouth breathing, obstructive sleep apnea, dry air exposure, upper airway hypersensitivity, and certain diseases or behavioral voice use factors. Laryngeal dehydration affects phonation threshold pressure (PTP) and phonation threshold flow (PTF), defined as the pressure and flow observed at the onset of phonation, respectively. The application of topical nebulized isotonic saline (0.9% Na+Cl-) has been shown in previous work to decrease PTP. However, there are no studies examining the effects of aerosolized saline, administered supraglottally, on dehydrated excised porcine larynges. Examining the effects of aerosolized saline in an excised model is essential to determine any independent effects of this treatment in the absence of other physiologic mechanisms such as mucus secretion. This study sought to investigate the effects of aerosolized saline on dehydrated animal vocal folds to determine if the administration of supraglottic aerosolized saline, via a nebulizer, could reverse the adverse effects of laryngeal dehydration. The study included a prospective, mixed experimental design with two groups, one desiccation/aerosolization (A/B) group and a control (A) group, each comprised of five bench-mounted porcine larynges. Larynges in both groups received desiccated air (<1% relative humidity) supraglottally via custom tubing for 1-min doses until the vocal folds ceased audible phonation. Following the desiccation challenge, the A/B group received 2-min doses of aerosolized isotonic saline until phonation began again. The PTP and PTF were measured during phonation trials following each dose of the desiccation or aerosolization treatment. Significant changes in PTP and PTF were observed following both the dehydration and aerosolization treatment. The PTP increased significantly following the dehydration challenge and returned near baseline following the aerosolization treatment. The results of this investigation supported the hypothesis that the administration of aerosolized saline may reverse the adverse effects of vocal fold dehydration. Moreover, in a more physiologically realistic excised model, applying the mechanics of respiration, this study advanced the development of innovative theories related to the reversal of the adverse effects of dehydration, which may prevent the development of voice disorders.

Although combination inhaled corticosteroids (ICs) are known to cause voice disorders in otherwise vocally healthy individuals, it is unknown whether those adverse effects can be reversed by the cessation of treatment. Quantitative aerodynamic measures such as phonation threshold pressure (PTP) and phonation threshold flow (PTF) can be used to identify the development of, and recovery from, vocal pathologies. We examined the effects and reversibility of ICs on laryngeal aerodynamics. This study was a mid-project investigation as part of a larger ongoing project. The 18 larynges were from rabbits that received ICs, a control condition, or no treatment. Experimental group rabbits received ICs twice per day until inflammatory changes (e.g., erythema, edema) became visible through endoscopic observation. One experimental group (i.e., the induction group) received treatment until symptoms were observed and then larynges were harvested. The other experimental group (i.e., the reversibility group) had ICs withdrawn once visible changes were detected via endoscopy; larynges were harvested only after these changes no longer differed from baseline. Both experimental groups had corresponding control rabbits that received twice-daily nebulized saline and followed the same withdrawal and larynx harvest schedule. A final group received no treatment. During benchtop phonation trials, PTP and PTF values were determined. All data were analyzed using descriptive and parametric statistics. No significant between-group differences were observed. Descriptively, however, average PTP and PTF values for the reversibility group were lower than the induction group. Additionally, average PTP and PTF values for the reversibility group were slightly lower than the induction group. Both experimental groups had higher PTP and PTF values than the control larynges. The results of this study indicate a trend in recovery for larynges afforded a recovery period from ICs. Further testing is needed to substantiate these preliminary findings.