Extrinsic Laryngeal Muscle Activity With Different Diameters and Water Depths in a Semi-Occluded Vocal Tract Exercise.

PURPOSE: Surface electromyography (sEMG) has been used to evaluate extrinsic laryngeal muscle activity during swallowing and phonation. In the current study, sEMG amplitudes were measured from the infrahyoid and suprahyoid muscles during phonation through a tube submerged in water. METHOD: The sEMG amplitude values measured from the extrinsic laryngeal muscles and the electroglottographic contact quotient (CQ) were obtained simultaneously from 62 healthy participants (31 men, 31 women) during phonation through a tube at six different depths (2, 4, 7, 10, 15, and 20 cm) while using two tubes with different diameters (1 and 0.5 cm). RESULTS: With increasing depth, the sEMG amplitude for the suprahyoid muscles increased in men and women. However, sEMG amplitudes for the infrahyoid muscles increased significantly only in men. Tube diameter had a significant effect on the suprahyoid sEMG amplitudes only for men, with higher sEMG amplitudes when phonating with a 1.0-cm tube. CQ values increased with submerged depth for both men and women. Tube diameter affected results such than CQ values were higher for men when using the wider tube and for women with the narrower tube. CONCLUSIONS: Vocal fold vibratory patterns changed with the depth of tube submersion in water for both men and women, but the patterns of muscle activation differed between the sexes. This suggests that men and women use different strategies when confronted with increased intraoral pressure during semi-occluded vocal tract exercises. In this study, sEMG provided insight into the mechanism for differences between vocally normal individuals and could help detect compensatory muscle activation during tube phonation in water for people with voice disorders.

Speaking without vocal folds using a machine-learning-assisted wearable sensing-actuation system.

Voice disorders resulting from various pathological vocal fold conditions or postoperative recovery of laryngeal cancer surgeries, are common causes of dysphonia. Here, we present a self-powered wearable sensing-actuation system based on soft magnetoelasticity that enables assisted speaking without relying on the vocal folds. It holds a lightweighted mass of approximately 7.2 g, skin-alike modulus of 7.83 × 105 Pa, stability against skin perspiration, and a maximum stretchability of 164%. The wearable sensing component can effectively capture extrinsic laryngeal muscle movement and convert them into high-fidelity and analyzable electrical signals, which can be translated into speech signals with the assistance of machine learning algorithms with an accuracy of 94.68%. Then, with the wearable actuation component, the speech could be expressed as voice signals while circumventing vocal fold vibration. We expect this approach could facilitate the restoration of normal voice function and significantly enhance the quality of life for patients with dysfunctional vocal folds.

Restraining vocal fold vertical motion reduces source-filter interaction in a two-mass model.

Previous experimental studies suggested that restraining the vocal fold vertical motion may reduce the coupling strength between the voice source and vocal tract. In this study, the effects of vocal fold vertical motion on source-filter interaction were systematically examined in a two-dimensional two-mass model coupled to a compressible flow simulation. The results showed that when allowed to move vertically, the vocal folds exhibited subharmonic vibration due to entrainment to the first vocal tract acoustic resonance. Restraining the vertical motion suppressed this entrainment. This indicates that the vertical mobility of the vocal folds may play a role in regulating source-filter interaction.

Brainstem control of vocalization and its coordination with respiration.

Phonation critically depends on precise controls of laryngeal muscles in coordination with ongoing respiration. However, the neural mechanisms governing these processes remain unclear. We identified excitatory vocalization-specific laryngeal premotor neurons located in the retroambiguus nucleus (RAmVOC) in adult mice as being both necessary and sufficient for driving vocal cord closure and eliciting mouse ultrasonic vocalizations (USVs). The duration of RAmVOC activation can determine the lengths of both USV syllables and concurrent expiration periods, with the impact of RAmVOC activation depending on respiration phases. RAmVOC neurons receive inhibition from the preBötzinger complex, and inspiration needs override RAmVOC-mediated vocal cord closure. Ablating inhibitory synapses in RAmVOC neurons compromised this inspiration gating of laryngeal adduction, resulting in discoordination of vocalization with respiration. Our study reveals the circuits for vocal production and vocal-respiratory coordination.

Twin vocal folds as a novel evolutionary adaptation for vocal communications in lemurs.

Primates have varied vocal repertoires to communicate with conspecifics and sometimes other species. The larynx has a central role in vocal source generation, where a pair of vocal folds vibrates to modify the air flow. Here, we show that Madagascan lemurs have a unique additional pair of folds in the vestibular region, parallel to the vocal folds. The additional fold has a rigid body of a vocal muscle branch and it is covered by a stratified squamous epithelium, equal to those of the vocal fold. Such anatomical features support the hypothesis that it also vibrates in a manner like the vibrations that occur in the vocal folds. To examine the acoustic function of the two pairs of folds, we made a silicone compound model to demonstrate that they can simultaneously vibrate to lower the fundamental frequency and increase vocal efficiency. Similar acoustic effects are achieved using different features of the larynx for the other primates, e.g., by vibrating multiple sets of ventricular folds in several species and further by an evolutionary modification of enlarged larynx in howler monkeys. Our multidisciplinary approaches found that these functions were acquired through a unique evolutionary adaptation of the twin vocal folds in Madagascan lemurs.

Vocal fold fibroblasts and exposure to vibration in vitro: Does sex matter?

Studies have shown that certain vocal fold pathologies are more common in one sex than the other. This is often explained by differences in the composition of the lamina propria and anatomical differences between female and male vocal folds, resulting in e.g. different fundamental frequencies. Here, we investigated a potential sex-specific voice frequency effect in an in vitro setting using vocal fold fibroblasts from one male and one female donor with and without cigarette smoke extract (CSE) addition. After exposure to either male or female vibration frequency with or without CSE, cells and supernatants were harvested. Gene and protein analysis were performed by means of qPCR, western blot, ELISA and Luminex. We found that exposure of cells to both male and female vibration pattern did not elicit significant changes in the expression of extracellular matrix-, inflammation-, and fibrosis-related genes, compared to control cells. The addition of CSE to vibration downregulated the gene expression of COL1A1 in cells exposed to the female vibration pattern, as well as induced MMP1 and PTGS2 in cells exposed to both female and male vibration pattern. The protein expression of MMP1 and COX2 was found to be significantly upregulated only in cells exposed to CSE and female vibration pattern. To conclude, different vibration patterns alone did not cause different responses of the cells. However, the female vibration pattern in combination with CSE had a tendency to elicit/maintain more pro-inflammatory responses in cells than the male vibration pattern.

Histologic Examination of Vocal Fold Mucosal Wave and Vibration.

OBJECTIVES: Despite gross anatomic and histologic differences between human and canine vocal folds, similar wave patterns have been described yet not fully characterized. We reconstructed vocal fold (VF) vibration in a canine hemilarynx and performed histologic examination of the same vocal fold. We demonstrate comparable wave patterns while exploring the importance of certain anatomic architectures. METHODS: An in vivo canine hemilarynx was phonated against a glass prism at low and high muscle activation conditions. Vibration was captured using high-speed video, and trajectories of VF medial surface tattooed landmarks were 3D-reconstructed. The method of empirical eigenfunctions was used to capture the essential dynamics of vibratory movement. Histologic examination of the hemilarynx was performed. RESULTS: Oscillation patterns were highly similar between the in vivo canine and previous reports of ex vivo human models. The two most dominant eigenfunctions comprised over 90% of total variance of movement, representing opening/closing and convergent/divergent movement patterns, respectively. We demonstrate a vertical phase difference during the glottal cycle. The time delay between the inferior and superior VF was greater during opening than closing for both activation conditions. Histological examination of canine VF showed not only a thicker lamina propria layer superiorly but also a distinct pattern of thyroarytenoid muscle fibers and fascicles as described in human studies. CONCLUSIONS: Histologic and vibratory examination of the canine vocal fold demonstrated human vocal fold vibratory patterns despite certain microstructural differences. This study suggests that the multilayered lamina propria may not be fundamental to vibratory patterns necessary for human-like voice production. LEVEL OF EVIDENCE: NA (Basic science study) Laryngoscope, 134:264-271, 2024.

Reconstruction of Vocal Fold Medial Surface 3D Trajectories: Effects of Neuromuscular Stimulation and Airflow.

INTRODUCTION: Analysis of medial surface dynamics of the vocal folds (VF) is critical to understanding voice production and treatment of voice disorders. We analyzed VF medial surface vibratory dynamics, evaluating the effects of airflow and nerve stimulation using 3D reconstruction and empirical eigenfunctions (EEF). STUDY DESIGN: In vivo canine hemilarynx phonation. METHODS: An in vivo canine hemilarynx was phonated while graded stimulation of the recurrent and superior laryngeal nerves (RLN and SLN) was performed. For each phonatory condition, vibratory cycles were 3D reconstructed from tattooed landmarks on the VF medial surface at low, medium, and high airflows. Parameters describing medial surface trajectory shape were calculated, and underlying patterns were emphasized using EEFs. Fundamental frequency and smoothed cepstral peak prominence (CPPS) were calculated from acoustic data. RESULTS: Convex-hull area of landmark trajectories increased with increasing flow and decreasing nerve activation level. Trajectory shapes observed included circular, ellipsoid, bent, and figure-eight. They were more circular on the superior and anterior VF, and more elliptical and line-like on the inferior and posterior VF. The EEFs capturing synchronal opening and closing (EEF1) and alternating convergent/divergent (EEF2) glottis shapes were mostly unaffected by flow and nerve stimulation levels. CPPS increased with higher airflow except for low RLN activation and very dominant SLN stimulation. CONCLUSION: We analyzed VF vibration as a function of neuromuscular stimulation and airflow levels. Oscillation patterns such as figure-eight and bent trajectories were linked to high nerve activation and flow. Further studies investigating longer sections of 3D reconstructed oscillations are needed. LEVEL OF EVIDENCE: N/A, Basic Science Laryngoscope, 134:1249-1257, 2024.

Effect of Testosterone on the Thyroarytenoid Muscle and Lamina Propria of Female Rat Vocal Folds.

OBJECTIVE: To analyze the effects of androgen therapy on the thyroarytenoid (TA) muscle, expression of androgen receptors (ARs) and hyaluronic acid (HA) concentration in the vocal folds (VFs) of adult female rats. METHODS: Twenty-one adult female Wistar rats were divided into experimental and control groups. The experimental group received weekly intramuscular injections of nandrolone decanoate for 9 weeks. Following euthanasia and dissection of the VFs, histomorphometric analysis of the TA muscle, immunohistochemical evaluation of ARs, and measurement of HA concentration using the ELISA-like fluorimetric method were performed. RESULTS: The experimental group exhibited a significantly larger mean fiber cross-sectional area in the TA muscle compared to the control group (434.3 ± 68.6 µm2 versus 305.7 ± 110.1 µm2; p = 0.029), indicating muscle hypertrophy. There was no significant difference in the number of muscle fibers. The experimental group showed higher expression of ARs in the lamina propria (62.0% ± 30.3% versus 22.0% ± 22.8%; p = 0.046) and in the TA muscle (45.0% ± 22.6% versus 18.3% ± 9.8%; p = 0.024). There was no significant difference in the concentration of HA. CONCLUSION: Exposure of adult female rats to androgen therapy resulted in hypertrophy of the TA muscle and increased expression of ARs in the VFs. The TA muscle seems to be the primary target of testosterone action in the VF, and the up-regulation of ARs might contribute to the persistent deepening of the voice. LEVEL OF EVIDENCE: NA Laryngoscope, 134:2316-2321, 2024.

Vocal communication: The enigmatic production of low-frequency purrs in cats.

Cat purring, the unusual, pulsed vibration that epitomizes comfort, enjoys a special status in the world of vocal communication research. Indeed, it has long been flagged as a rare exception to the dominant theory of voice production in mammals. A new study presents histological and biomechanical evidence that purring can occur passively, without needing muscle vibration in the larynx controlled by an independent neural oscillator.

Flow-induced oscillations of vocal-fold replicas with tuned extensibility and material properties.

Human vocal folds are highly deformable non-linear oscillators. During phonation, they stretch up to 50% under the complex action of laryngeal muscles. Exploring the fluid/structure/acoustic interactions on a human-scale replica to study the role of the laryngeal muscles remains a challenge. For that purpose, we designed a novel in vitro testbed to control vocal-folds pre-phonatory deformation. The testbed was used to study the vibration and the sound production of vocal-fold replicas made of (i) silicone elastomers commonly used in voice research and (ii) a gelatin-based hydrogel we recently optimized to approximate the mechanics of vocal folds during finite strains under tension, compression and shear loadings. The geometrical and mechanical parameters measured during the experiments emphasized the effect of the vocal-fold material and pre-stretch on the vibration patterns and sounds. In particular, increasing the material stiffness increases glottal flow resistance, subglottal pressure required to sustain oscillations and vibratory fundamental frequency. In addition, although the hydrogel vocal folds only oscillate at low frequencies (close to 60 Hz), the subglottal pressure they require for that purpose is realistic (within the range 0.5-2 kPa), as well as their glottal opening and contact during a vibration cycle. The results also evidence the effect of adhesion forces on vibration and sound production.

The influence of source-filter interaction on the voice source in a three-dimensional computational model of voice production.

The goal of this computational study is to quantify global effects of vocal tract constriction at various locations (false vocal folds, aryepiglottic folds, pharynx, oral cavity, and lips) on the voice source across a large range of vocal fold conditions. The results showed that while inclusion of a uniform vocal tract had notable effects on the voice source, further constricting the vocal tract only had small effects except for conditions of extreme constriction, at which constrictions at any location along the vocal tract decreased the mean and peak-to-peak amplitude of the glottal flow waveform. Although narrowing in the epilarynx increased the normalized maximum flow declination rate, vocal tract constriction in general slightly reduced the source strength and high-frequency harmonic production at the glottis, except for a limited set of vocal fold conditions (e.g., soft, long vocal folds subject to relatively high pressure). This suggests that simultaneous laryngeal and vocal tract adjustments are required to maximize source-filter interaction. While vocal tract adjustments are often assumed to improve voice production, our results indicate that such improvements are mainly due to changes in vocal tract acoustic response rather than improved voice production at the glottis.

Domestic cat larynges can produce purring frequencies without neural input.

Most mammals produce vocal sounds according to the myoelastic-aerodynamic (MEAD) principle, through self-sustaining oscillation of laryngeal tissues.1,2 In contrast, cats have long been believed to produce their low-frequency purr vocalizations through a radically different mechanism involving active muscle contractions (AMC), where neurally driven electromyographic burst patterns (typically at 20-30 Hz) cause the intrinsic laryngeal muscles to actively modulate the respiratory airflow. Direct empirical evidence for this AMC mechanism is sparse.3 Here, the fundamental frequency (fo) ranges of eight domestic cats (Felis silvestris catus) were investigated in an excised larynx setup, to test the prediction of the AMC hypothesis that vibration should be impossible without neuromuscular activity, and thus unattainable in excised larynx setups, which are based on MEAD principles. Surprisingly, all eight excised larynges produced self-sustained oscillations at typical cat purring rates. Histological analysis of cat larynges revealed the presence of connective tissue masses, up to 4 mm in diameter, embedded in the vocal fold.4 This vocal fold specialization appears to allow the unusually low fo values observed in purring. While our data do not fully reject the AMC hypothesis for purring, they show that cat larynges can easily produce sounds in the purr regime with fundamental frequencies of 25 to 30 Hz without neural input or muscular contraction. This strongly suggests that the physical and physiological basis of cat purring involves the same MEAD-based mechanisms as other cat vocalizations (e.g., meows) and most other vertebrate vocalizations but is potentially augmented by AMC.

An Euler-Bernoulli-type beam model of the vocal folds for describing curved and incomplete glottal closure patterns.

Incomplete glottal closure is a laryngeal configuration wherein the glottis is not fully obstructed prior to phonation. It has been linked to inefficient voice production and voice disorders. Various incomplete glottal closure patterns can arise and the mechanisms driving them are not well understood. In this work, we introduce an Euler-Bernoulli composite beam vocal fold (VF) model that produces qualitatively similar incomplete glottal closure patterns as those observed in experimental and high-fidelity numerical studies, thus offering insights into the potential underlying physical mechanisms. Refined physiological insights are pursued by incorporating the beam model into a VF posturing model that embeds the five intrinsic laryngeal muscles. Analysis of the combined model shows that co-activating the lateral cricoarytenoid (LCA) and interarytenoid (IA) muscles without activating the thyroarytenoid (TA) muscle results in a bowed (convex) VF geometry with closure at the posterior margin only; this is primarily attributed to the reactive moments at the anterior VF margin. This bowed pattern can also arise during VF compression (due to extrinsic laryngeal muscle activation for example), wherein the internal moment induced passively by the TA muscle tissue is the predominant mechanism. On the other hand, activating the TA muscle without incorporating other adductory muscles results in anterior and mid-membranous glottal closure, a concave VF geometry, and a posterior glottal opening driven by internal moments induced by TA muscle activation. In the case of initial full glottal closure, the posterior cricoarytenoid (PCA) muscle activation cancels the adductory effects of the LCA and IA muscles, resulting in a concave VF geometry and posterior glottal opening. Furthermore, certain maneuvers involving co-activation of all adductory muscles result in an hourglass glottal shape due to a reactive moment at the anterior VF margin and moderate internal moment induced by TA muscle activation. These findings have implications regarding potential laryngeal maneuvers in patients with voice disorders involving imbalances or excessive tension in the laryngeal muscles such as muscle tension dysphonia.

Effect of functional electric stimulation on phonation in an ex vivo aged ovine model.

With age, the atrophy of the thyroarytenoid muscle (TAM), and thus atrophy of the vocal folds, leads to decreased glottal closure, increased breathiness, and a loss in voice quality, which results in a reduced quality of life. A method to counteract the atrophy of the TAM is to induce hypertrophy in the muscle by functional electric stimulation (FES). In this study, phonation experiments were performed with ex vivo larynges of six stimulated and six unstimulated ten-year-old sheep to investigate the impact of FES on phonation. Electrodes were implanted bilaterally near the cricothyroid joint. FES treatment was provided for nine weeks before harvesting. The multimodal measurement setup simultaneously recorded high-speed video of the vocal fold oscillation, the supraglottal acoustic signal, and the subglottal pressure signal. Results of 683 measurements show a 65.6% lower glottal gap index, a 22.7% higher tissue flexibility (measured by the amplitude to length ratio), and a 473.7% higher coefficient of determination (R2) of the regression of subglottal and supraglottal cepstral peak prominence during phonation for the stimulated group. These results suggest that FES improves the phonatory process for aged larynges or presbyphonia.

Modeling the influence of the extrinsic musculature on phonation.

Neck muscles play important roles in various physiological tasks, including swallowing, head stabilization, and phonation. The mechanisms by which neck muscles influence phonation are not well understood, with conflicting reports on the change in fundamental frequency for ostensibly the same neck muscle activation scenarios. In this work, we introduce a reduced-order muscle-controlled vocal fold model, comprising both intrinsic muscle control and extrinsic muscle effects. The model predicts that when the neck muscles pull the thyroid cartilage in the superior-anterior direction (with a sufficiently large anterior component), inferior direction, or inferior-anterior direction, tension in the vocal folds increases, leading to fundamental frequency rise during sustained phonation. On the other hand, pulling in the superior direction, superior-posterior direction, or inferior-posterior direction (with a sufficiently large posterior component) tends to decrease vocal fold tension and phonation fundamental frequency. Varying the pulling force location alters the posture and phonation biomechanics, depending on the force direction. These findings suggest potential roles of particular neck muscles in modulating phonation fundamental frequency, with implications for vocal hyperfunction.

Predicting 3D soft tissue dynamics from 2D imaging using physics informed neural networks.

Tissue dynamics play critical roles in many physiological functions and provide important metrics for clinical diagnosis. Capturing real-time high-resolution 3D images of tissue dynamics, however, remains a challenge. This study presents a hybrid physics-informed neural network algorithm that infers 3D flow-induced tissue dynamics and other physical quantities from sparse 2D images. The algorithm combines a recurrent neural network model of soft tissue with a differentiable fluid solver, leveraging prior knowledge in solid mechanics to project the governing equation on a discrete eigen space. The algorithm uses a Long-short-term memory-based recurrent encoder-decoder connected with a fully connected neural network to capture the temporal dependence of flow-structure-interaction. The effectiveness and merit of the proposed algorithm is demonstrated on synthetic data from a canine vocal fold model and experimental data from excised pigeon syringes. The results showed that the algorithm accurately reconstructs 3D vocal dynamics, aerodynamics, and acoustics from sparse 2D vibration profiles.

Amount and spatial arrangement of muscle fibers in the human laryngeal Musculus ventricularis.

Textbooks and atlases of human macroscopic and microscopic anatomy of the larynx generally provide, if at all, only sparse information on the laryngeal Musculus ventricularis. However, several studies indicate that this muscle takes over the function of vestibular (ventricular) fold phonation after denervation of the Musculus vocalis. In the present study, 29 laryngeal specimens were coronally dissected at different levels, i.e. the anterior (L1), middle (L2), and posterior third of the vestibular fold (L3), and they underwent histological analysis. In all specimens the vestibular folds of both hemi-larynxes contained striated muscle bundles in variable amounts, representing a ventricularis muscle. These muscle bundles obviously originated from the lateral (external) and thyroepiglottic part of the thyroarytenoid muscle and the aryepiglottic part of the oblique arytenoid muscle, as has been described by other authors. The areas of vestibular folds and their amounts of ventricularis muscle bundles were measured using image analysis software (imageJ) by manual tracing. The mean area of the vestibular folds of both hemi-larynxes was 27.9 mm2 (SD [standard deviation] ± 9.17), and the area occupied by fibers of the ventricularis muscle was 1.5 mm2 (SD ± 1.78). Statistical analysis comparing the areas of both hemi-larynxes and levels resulted in no significant differences, except for the levels 2 and 3. In level 2, significantly more muscle fibers (2.0 mm2 ; SD ± 2.21) were detectable within the vestibular fold than in level 3 (0.9 mm2 ; SD ± 1.43). Level 1 also contained more muscle fibers (1.1 mm2 ; SD ± 1.06) than level 3, however, without significance. In conclusion, the laryngeal ventricularis muscle is present in the majority of reported cases. Since the muscle is of clinical relevance, it should be included in anatomical textbooks by default.

Closed-loop network of skin-interfaced wireless devices for quantifying vocal fatigue and providing user feedback.

Vocal fatigue is a measurable form of performance fatigue resulting from overuse of the voice and is characterized by negative vocal adaptation. Vocal dose refers to cumulative exposure of the vocal fold tissue to vibration. Professionals with high vocal demands, such as singers and teachers, are especially prone to vocal fatigue. Failure to adjust habits can lead to compensatory lapses in vocal technique and an increased risk of vocal fold injury. Quantifying and recording vocal dose to inform individuals about potential overuse is an important step toward mitigating vocal fatigue. Previous work establishes vocal dosimetry methods, that is, processes to quantify vocal fold vibration dose but with bulky, wired devices that are not amenable to continuous use during natural daily activities; these previously reported systems also provide limited mechanisms for real-time user feedback. This study introduces a soft, wireless, skin-conformal technology that gently mounts on the upper chest to capture vibratory responses associated with vocalization in a manner that is immune to ambient noises. Pairing with a separate, wirelessly linked device supports haptic feedback to the user based on quantitative thresholds in vocal usage. A machine learning-based approach enables precise vocal dosimetry from the recorded data, to support personalized, real-time quantitation and feedback. These systems have strong potential to guide healthy behaviors in vocal use.

Impact of Instructed Laryngeal Manipulation on Acoustic Measures of Voice-Preliminary Results.

BACKGROUND: Control of laryngeal muscles is required to manipulate pitch, volume, and voice quality. False vocal fold activity (FVFA) refers to the constriction and release of constriction of the false vocal folds. True vocal fold mass (TVFM) represents the cross-sectional thickness of the vocal folds. Larynx height (LH) refers to the vertical position of the larynx in the neck. To date, studies of voice control have examined the effects of these parameters separately. No study has investigated the impact of instructed systematic manipulation of these parameters on acoustic voice measures in vocally healthy trained subjects. AIMS: This study examined the effects of systematically manipulating FVFA, TVFM, and LH on several acoustic voice measures. METHOD: Twelve vocally trained speakers were instructed to use specific techniques to achieve experimental conditions of constriction and release of constriction of FVFA, thicker and thinner TVFM, and normal and low LH. Each condition was implemented in combination with manipulating the other parameters. Voice recordings of sustained vowel /a/ and Rainbow Passage were obtained for all laryngeal manipulation conditions and underwent acoustic analyses for fundamental frequency (F0), signal typing, harmonics-to-noise ratio (HNR), cepstral peak prominence (CPP), and vocal relative intensity. RESULTS: Constricted FVFA caused more aperiodicity in the signals, lower CPP, and lower vocal relative intensity than release of constriction. Thicker TVFM resulted in significantly higher CPP and vocal relative intensity than thinner TVFM. Modifying TVFM did not affect F0 and HNR. Low LH had significantly lower F0 but did not impact on HNR, CPP, and intensity. CONCLUSIONS: The effects of systematic manipulation of each laryngeal parameter resulted in independent acoustic effects without measurable interaction. Release of constriction of FVFA, thicker TVFM, and low LH were configurations that resulted in more optimal acoustic signals.