Subcutaneous Neurotophin 4 Infusion Using Osmotic Pumps or Direct Muscular Injection Enhances Aging Rat Laryngeal Muscles

Laryngeal dysfunction in the elderly is a major cause of disability, from voice disorders to dysphagia and loss of airway protective reflexes. Few, if any, therapies exist that target age-related laryngeal muscle dysfunction. Neurotrophins are involved in muscle innervation and differentiation of neuromuscular junctions (NMJs). It is thought that neurotrophins enhance neuromuscular transmission by increasing neurotransmitter release. The neuromuscular junctions (NMJs) become smaller and less abundant in aging rat laryngeal muscles, with evidence of functional denervation. We explored the effects of NTF4 for future clinical use as a therapeutic to improve function in aging human laryngeal muscles. Here, we provide the detailed protocol for systemic application and direct injection of NTF4 to investigate the ability of aging rat laryngeal muscle to remodel in response to NTF4 application. In this method, rats either received NTF4 either systemically via osmotic pump or by direct injection through the vocal folds. Laryngeal muscles were then dissected and used for histological examination of morphology and age-related denervation

Enhancement of Aging Rat Laryngeal Muscles with Endogenous Growth Factor Treatment

Clinical evidence suggests that laryngeal muscle dysfunction is associated with human aging. Studies in animal models have reported morphological changes consistent with denervation in laryngeal muscles with age. Life‐long laryngeal muscle activity relies on cytoskeletal integrity and nerve–muscle communication at the neuromuscular junction (NMJ). It is thought that neurotrophins enhance neuromuscular transmission by increasing neurotransmitter release. We hypothesized that treatment with neurotrophin 4 (NTF4) would modify the morphology and functional innervation of aging rat laryngeal muscles. Fifty‐six Fischer 344xBrown Norway rats (6‐ and 30‐mo age groups) were used to evaluate to determine if NTF4, given systemically (n = 32) or directly (n = 24), would improve the morphology and functional innervation of aging rat thyroarytenoid muscles. Results demonstrate the ability of rat laryngeal muscles to remodel in response to neurotrophin application. Changes were demonstrated in fiber size, glycolytic capacity, mitochondrial, tyrosine kinase receptors (Trk), NMJ content, and denervation in aging rat thyroarytenoid muscles. This study suggests that growth factors may have therapeutic potential to ameliorate aging‐related laryngeal muscle dysfunction

Vocalization subsystem responses (Croake et al., 2018)

<div><b>Purpose: </b>The purpose of this study is to quantify the interactions of the 3 vocalization subsystems of respiration, phonation, and resonance before, during, and after a perturbation to the larynx (temporarily induced unilateral vocal fold paralysis) in 10 vocally healthy participants. Using dynamic systems theory as a guide, we hypothesized that data groupings would emerge revealing context-dependent patterns in the relationships of variables representing the 3 vocalization subsystems. We also hypothesized that group data would mask important individual variability important to understanding the relationships among the vocalization subsystems.</div><div><b>Method: </b>A perturbation paradigm was used to obtain respiratory kinematic, aerodynamic, and acoustic formant measures from 10 healthy participants (8 women, 2 men) with normal voices. Group and individual data were analyzed to provide a multilevel analysis of the data. A 3-dimensional state space model was constructed to demonstrate the interactive relationships among the 3 subsystems before, during, and after perturbation.</div><div><b>Results:</b> During perturbation, group data revealed that lung volume initiations and terminations were lower, with longer respiratory excursions; airflow rates increased while subglottic pressures were maintained. Acoustic formant measures indicated that the spacing between the upper formants decreased (F3–F5), whereas the spacing between F1 and F2 increased. State space modeling revealed the changing directionality and interactions among the 3 subsystems.</div><div><b>Conclusions: </b>Group data alone masked important variability necessary to understand the unique relationships among the 3 subsystems. Multilevel analysis permitted a richer understanding of the individual differences in phonatory regulation and permitted subgroup analysis. Dynamic systems theory may be a useful heuristic to model the interactive relationships among vocalization subsystems.</div><div><br></div><div><b>Supplemental Material S1.</b> Intraclass correlation coefficients (ICCs) with 95% confidence intervals (CIs).</div><div><br></div><div><b>Supplemental Material S2. </b>Descriptive statistics: group means, standard deviations, and ranges by condition (pre recurrent laryngeal nerve [RLN] block [PRE], during RLN block [DUR], and post-RLN block recovered [REC]).</div><div><br></div><div><b>Supplemental Material S3.</b> Repeated measures analysis of variance (ANOVA). Tukey’s honestly significant difference test was applied to determine significant differences between conditions. **<i>p</i> < .05. </div><div><br></div><div>Croake, D. J., Andreatta, R. D., & Stemple, J. C. (2018). Vocalization subsystem responses to a temporarily induced unilateral vocal fold paralysis. <i>Journal of Speech, Language, and Hearing Research, 61,</i> 479–495. https://doi.org/10.1044/2017_JSLHR-S-17-0227</div

Reliability of Clinical Office-Based Laryngeal Electromyography in Vocally Healthy Adults

Objective: This study aimed to conduct a 3-session reliability assessment of the laryngeal electromyography (LEMG) signal in healthy participants during intensity controlled vocalization tasks. We hypothesized that vocal intensity level and testing session would affect LEMG measures. Methods: This prospective study used a 2-factor repeated measures design. Seven participants underwent bipolar needle LEMG of the right thyroarytenoid muscle. Data were collected over 3 testing sessions using vocalization tasks performed with visually guided intensity feedback targets (65 and 75 dB SPL). Root mean square amplitudes in microvolts were analyzed for within-session and between-session reliability. Results: The main effect for intensity was found to approach significance (F = 5.71, P = .054). However, intraclass correlation coefficients (ICCs) using a 2-factor mixed random effect model indicated poor to fair signal reliability between testing sessions (ICC = 0.56 at 65 dB, 0.40 at 70 dB). Intraclass correlation coefficients for within-session data indicated excellent reliability for all testing conditions (0.84–0.98). Conclusion: Using a quantitative analysis protocol to inform an essentially qualitative technique, our results indicated that there was generally poor to fair reliability in the LEMG signal over testing sessions. Vocal intensity was an important variable that affected LEMG signal reliability. Standardization of LEMG protocols using vocal control parameters and quantitative analyses may help improve LEMG reliability in clinical settings