Modulation of myosin heavy chains in rat laryngeal muscle

OBJECTIVES: To test the hypothesis that myosin heavy chain (MHC) composition is a biological marker indicative of appropriate and functional reinnervation. STUDY DESIGN: Age-matched adult rats were randomized for prospective study under three experimental conditions. METHODS: In adult rats, three experimental conditions were surgically created, including transient recurrent laryngeal nerve (RLN) crush injury, RLN transection and repair, and cricoarytenoid joint fixation with intact RLN. Animals were survived for 30, 90, and 180 days. At each interval, vocal fold mobility was assessed by rigid microlaryngoscopy. Laryngeal electromyography (EMG) was performed before euthanasia. The thyroarytenoid and posterior cricoarytenoid muscles were then excised, each muscle was processed for sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and MHC composition was determined. RESULTS: Thirty days after nerve crush injury, three of six animals regained vocal fold mobility and normal MHC composition. Impaired vocal fold motion in three of six animals was associated with MHC composition characteristic of denervation. At 90 and 180 days, normal vocal fold motion and normal MHC composition were observed in all animals. Following nerve transection and repair, impaired vocal fold motion and MHC composition characteristic of denervation were observed in all animals, despite evidence of reinnervation on EMG. Following joint fixation, alteration in MHC composition consistent with denervation was observed only at 30 days, as was evident in the nerve crush model. CONCLUSION: Temporary injury and vocal fold immobilization result in transient shifts in MHC composition. Nerve transection and repair result in persistent alteration of MHC composition and vocal fold dysfunction. The expression of normal MHC composition is dependent on the condition of appropriate neural contact and functional reinnervation.     

Vagal stimulation and laryngeal electromyography for recurrent laryngeal reinnervation in children

Ansa-to-recurrent laryngeal nerve (ANSA-RLN) reinnervation procedures are now often first-line treatments for some children with unilateral vocal fold immobility. Although many describe that children with prolonged denervation and true vocal fold atrophy should not undergo this procedure, there has been no gold-standard means of identifying true denervation. Here, we describe a novel technique using evoked vagal electromyography to predict degree of chronic nerve injury prior to recurrent laryngeal nerve reinnervation in children. This is a simple, readily available technique that may play an important role in predicting likelihood of success with pediatric ANSA-RLN reinnervation. Laryngoscope, 130:747–751, 2020     

Spontaneous laryngeal reinnervation after recurrent laryngeal or vagus nerve injury

OBJECTIVES: The status of innervation in patients with laryngeal paralysis is somewhat controversial. Electromyographic activity has been frequently documented in the laryngeal muscles of patients with laryngeal paralysis, and animal experiments report a strong propensity for reinnervation after laryngeal nerve injury. However, a study of intraoperative electromyography performed in patients during reinnervation surgery failed to document activity with stimulation of the recurrent laryngeal nerve (RLN). Noting the long-observed differences in the symptoms of patients with vagus nerve injury and those with RLN injury, I hypothesized that reinnervation is influenced by the site of nerve injury. METHODS: Cats were sacrificed at various intervals after resection of 1 cm of either the RLN or the vagus nerve, without any attempt to repair the nerve. RESULTS: Four months after RLN resection, distal nerve biopsy revealed unmyelinated axons scattered through fibrous tissue. By 6 months, myelinated axons were organized, and electromyographic and histologic examination showed preferential reinnervation of the thyroarytenoid muscle. After vagotomy, the RLN was fibrotic and no axons were present. Both the thyroarytenoid and posterior cricoarytenoid muscles were fibrotic and had no electromyographic activity. CONCLUSIONS: The results confirm the strong propensity for laryngeal reinnervation after RLN injury, but not after vagus nerve injury. Preferential reinnervation of adductor muscles may account for a medial position of the paralyzed vocal fold.     

Configuration of the glottis in laryngeal paralysis. II: Animal experiments

It is widely believed that in isolated recurrent laryngeal nerve paralysis, the paralyzed vocal fold assumes a median or paramedian position, due to the action of the cricothyroid muscle. A review of the literature reveals that support for this theory is not conclusive and, in particular, experiments indicate that the cricothyroid muscle does not appreciably affect vocal fold position in acute paralysis. The research in this study compares the configuration of the glottis in chronic unilateral recurrent laryngeal nerve paralysis in cats, with and without concomitant denervation of the cricothyroid muscle. Results indicate that vocal fold position is not determined by the cricothyroid muscle. Incomplete denervation of intrinsic laryngeal muscles as well as synkinetic reinnervation appear to be significant factors in determining vocal fold position in chronic laryngeal paralysis.     

Intraoperative laryngeal electromyographic assessment of patients with immobile vocal fold.

The differential diagnosis of laryngeal ankylosis versus paralysis is occasionally difficult in patients with immobile vocal folds. Eight patients with acute and chronic evidence of vocal fold immobility were investigated by intraoperative electromyography (IEMG) during planned microlaryngoscopy. Bipolar hook wire electrodes were inserted into the thyroarytenoid muscle, of which the electrical activity was monitored during neuromotor blockade and emergence from anesthesia. The normal side and the side with ankylosis or stenosis showed normal IEMG activity. There was progressive recruitment of larger motor units during recovery from muscle relaxation. Patients with laryngeal paralysis failed to show such recruitment patterns. Thus, IEMG can be used as a diagnostic tool during operative laryngoscopy to differentiate neuromotor injury from anatomic causes of vocal fold immobility. The advantages of IEMG are its ease of application and certainty of electrode position. It can also be used to monitor recurrent nerve integrity and detect early laryngospasm. Further IEMG clinical study is warranted.     

Motion-specific laryngeal reinnervation using muscle-nerve-muscle neurotization

There is no current treatment method that can reliably restore physiologic movement to a paralyzed vocal fold. The purposes of this study were to test the hypotheses that 1) muscle-nerve-muscle (M-N-M) neurotization can be induced in feline laryngeal muscles and 2) M-N-M neurotization can restore movement to a paralyzed vocal fold. Muscle-nerve-muscle neurotization can be defined as the reinnervation of a denervated muscle via axons that are induced to sprout from nerves within an innervated muscle and that then traverse a nerve graft interposed between it and the target denervated muscle. A paralyzed laryngeal muscle could be reinnervated by axons from its contralateral paired muscle, thus achieving motion-specific reinnervation. Eighteen adult cats were divided into sham, hemilaryngeal-denervated, and M-N-M-reinnervated thyroarytenoid muscle groups. Five of the 6 reinnervated animals had histologic evidence of axons in the nerve graft, 4 of the 6 had evoked electromyographic evidence of crossed reinnervation, and 1 of the 6 had a return of appropriately phased adduction. This technique has great potential and should be further investigated.     

Electromyographic findings in recurrent laryngeal nerve reinnervation

Abductor, adductor, and combined reinnervation procedures have been explored with variable success rates. We describe the experience of a tertiary care center with adductor reinnervation procedures, including preoperative and postoperative videostroboscopy and electromyography (EMG) findings. A retrospective chart review was performed from 1997 to 2001 that included 9 patients. Preoperative and postoperative voice comparison was performed by 3 blinded speech pathologists. Clinical comparisons of videostroboscopy findings for vocal fold bulk, tone, position, presence of gap, and movement are elucidated. The preoperative and postoperative EMG findings are described. In all patients, preoperative EMG revealed a dense, complete denervation of the affected recurrent laryngeal nerve. No movement was noted on videostroboscopy with persistent glottic gap. Reinnervation involved a nerve-muscle pedicle or a direct neurorrhaphy of the ansa cervicalis to the recurrent laryngeal nerve. Voice improvement was noted between 60 days and 3 months after reinnervation. Four postoperative EMG studies were performed. An early postoperative EMG study at 5 months revealed activation of the lateral cricoarytenoid muscle and thyroarytenoid muscle with head-lift. Videostroboscopy showed excellent near-midline static positioning of the vocal fold. Late EMG studies, performed 12 to 16 months after reinnervation, revealed "learning" of these muscles, with new activation on "eee" phonation. We conclude that recurrent laryngeal nerve reinnervation procedures belong in the armamentarium of the laryngologist for the treatment of vocal fold paralysis. The EMG findings reported in this study suggest that ongoing reinnervation allows for activation with phonation in matured neuronal anastomoses. Overall, this procedure results in excellent patient acceptance and near-normal vocal quality.     

Aberrant recurrent laryngeal nerve reinnervation as a cause of stridor and laryngospasm

Unilateral vocal fold paralysis rarely presents with symptoms of stridor, laryngospasm, and dyspnea. Abnormal reinnervation of abductor nerve branches into adductor fibers may be one cause. Four patients have been positively identified by laryngeal electromyography. Two patients presented after thyroid surgery for thyroid neoplasm. One presented after cervical disk surgery. One patient presented after herpes simplex infection with multiple cranial nerve involvement. All patients had new onset of dyspnea with exertion many months after the recurrent nerve injury. Fiberoptic laryngoscopy showed the affected vocal fold to be immobile. However, with hyperventilation and deep inspiration, there was paradoxical adductor motion of the paretic vocal fold. Laryngeal electromyography showed evidence of reduced but intact voluntary motor units in the thyroarytenoid muscle. These motor units fired on inspiration and not on phonation. This pattern was not seen on the normal side. These findings are consistent with paradoxical innervation and/or synkinesis. Each patient was managed by Botox injection into the adductor muscle. Periodic reinjections may be necessary to manage the condition.     

Quantitative assessment of laryngeal muscle morphology after recurrent laryngeal nerve injury: right vs. left differences

Objectives/Hypothesis: Reports of laryngeal response to denervation are inconsistent. Some document atrophy and fibrosis in denervated laryngeal muscles, whereas others indicate resistance to atrophy. Spontaneous reinnervation has also been documented. The goal of this study was to clarify the effects of nerve injury and reinnervation on thyroarytenoid (TA) and posterior cricoarytenoid (PCA) muscles. Study Design: Laboratory experiment. Methods: TA and PCA muscles of cats were harvested 5 to 6 months after transecting right or left recurrent laryngeal nerve (RLN). Images of muscle cross‐sections were acquired and studied using an image analysis workstation. Cross‐sectional areas as well as total cross‐sectional area of randomly selected muscle fibers were recorded. Results: TA reinnervation was robust on both sides, but there was less reinnervation of the PCA muscle after left‐sided RLN lesion than after right‐sided injury. Conclusions: Differences in reinnervation after RLN injury could contribute to the higher clinical incidence of left‐ vs. right‐sided laryngeal paralysis.     

Value of laryngeal electromyography in diagnosis of vocal fold immobility

OBJECTIVES: We sought to determine the value of laryngeal electromyography (LEMG) and evoked LEMG in the diagnosis of vocal fold immobility. METHODS: We analyzed 110 cases of vocal fold immobility by their clinical manifestations and LEMG characteristics, including spontaneous potential activity, motor unit potential measurement, recruitment pattern analysis, and evoked LEMG signals. RESULTS: With LEMG, we identified 87 patients with neuropathic laryngeal injuries. Neurogenic vocal fold immobility showed a wide variety of abnormal activity. Fibrillation potentials and positive sharp waves were found in patients with laryngeal nerve injuries. For laryngeal paralysis, there was no reaction with LEMG and evoked LEMG. For incomplete laryngeal paralysis, decreased evoked LEMG signals were also seen with delayed latency (thyroarytenoid muscle, 2.2 +/- 1.0 ms, p < .01; posterior cricoarytenoid muscle, 2.4 +/- 1.0 ms, p < .05) and lower amplitude (thyroarytenoid muscle, 0.9 +/- 0.7 mV, p < .05; posterior cricoarytenoid muscle, 1.2 +/- 1.0 mV, p < .01). Nineteen patients with vocal fold mechanical limitations generally had normal LEMG and evoked LEMG signals. Four patients with neoplastic infiltration of the laryngeal muscles demonstrated abnormal LEMG signals but nearly normal evoked LEMG signals. CONCLUSIONS: We conclude that LEMG and evoked LEMG behavior plays a crucial role in the diagnosis of vocal fold immobility. The decreased recruitment activities on LEMG and the decreased evoked LEMG signals with longer latency and lower amplitude reflect the severity of neuropathic laryngeal injury.     

The cricothyroid muscle does not influence vocal fold position in laryngeal paralysis

The status of the cricothyroid muscle, which is innervated by the superior laryngeal nerve, is believed to influence the vocal fold position in laryngeal paralysis. It is believed that isolated lesions of the recurrent laryngeal nerve generally result in the paralyzed vocal fold assuming a paramedian position but that with lesions of both the superior and recurrent laryngeal nerves, a more lateral (intermediate or cadaveric) vocal fold position can be expected. Twenty-six consecutive patients with unilateral vocal fold paralysis underwent transnasal fiberoptic laryngoscopy (TFL) and laryngeal electromyography (LEMG). By TFL, the vocal fold positions were paramedian in 8 patients, intermediate in 7, and lateral in 11. By LEMG, 13 patients had isolated recurrent laryngeal nerve lesions and 13 patients had combined (superior and recurrent laryngeal nerve) lesions. There was no correlation between the vocal fold position and the status of the cricothyroid muscle, i.e., the status of the cricothyroid muscle by LEMG did not predict the vocal fold position nor did the vocal fold position by TFL predict the site of lesion. In addition, we investigated the possibility that the degree of thyroarytenoid muscle recruitment (tone) might correlate with vocal fold position, but no relation was found. We conclude that 1. the cricothyroid muscle does not predictably influence the position of the vocal fold in unilateral paralysis; 2. thyroarytenoid muscle recruitment (tone) does not appear to influence vocal fold position; and 3. still unidentified and unknown factors may be responsible for determining vocal fold position in laryngeal paralysis.     

Vocal fold paresis and paralysis

Diagnosis and treatment of the immobile or hypomobile vocal fold are challenging for the otolaryngologist. True paralysis and paresis result from vocal fold denervation secondary to injury to the laryngeal or vagus nerve. Vocal fold paresis or paralysis may be unilateral or bilateral, central or peripheral, and it may involve the recurrent laryngeal nerve, superior laryngeal nerve, or both. The physician's first responsibility in any case of vocal fold paresis or paralysis is to confirm the diagnosis and be certain that the laryngeal motion impairment is not caused by arytenoid cartilage dislocation or subluxation, cricoarytenoid arthritis or ankylosis, neoplasm, or other mechanical causes. Strobovideolaryngoscopy, endoscopy, radiologic and laboratory studies, and electromyography are all useful diagnostic tools.     

几丁质管桥接喉返神经缺损的实验研究

为在恢复神经再支配手术治疗喉返神经损伤所致的喉麻痹中,探索一种新的可避免神经误向支配的方法,采用非神经生物材料几了质管,桥接狗喉返神经干的缺损,并用自体神经移植作为对照组.术后5个月,组织学、组织化学和电生理等检测,显示两组动物的喉内肌皆获得神经再支配.实验组术侧声带较明显恢复了和健侧一致的内收和外展运动,甲杓肌(Thyroarytenoid,TAM)和环杓后肌(Posterior cricoarytenoid musdes,PCAM)也分别出现了相应的生理性自发肌电;对照组的术侧声带无明显的运动,TAM和PCAM则在呼气期和吸气期均出现不规则的持续密集自发肌电.提示几丁质桥接喉返神经干缺损可使其定向性再生,选择性支配喉内肌.     

Posterior cricoarytenoid muscle denervation

Vocal fold paralysis most commonly results from injury to the recurrent laryngeal nerve. The length of time required for denervation atrophy would be useful in planning reinnervation procedures. Given the absence of long-term data on the status of the canine posterior cricoarytenoid muscle following denervation, this study was undertaken. The results indicate that muscle atrophy does not occur for at least 1 year following denervation.     

A new electromyographic definition of laryngeal synkinesis

Laryngeal synkinesis involves the misdirected reinnervation of an injured recurrent laryngeal nerve to vocal fold abductor and adductor musculature. The resultant laryngeal dyscoordination can cause vocal fold immobility and airway compromise. Although this entity is sometimes considered in the differential diagnosis, it is only demonstrable with laryngeal electromyography (EMG). We propose a new EMG definition of synkinesis to assist in its identification during workup of vocal fold immobility. A retrospective chart review from 1992 to 1997 in the Voice Disorders Clinic identified 10 patients with laryngeal synkinesis. Five patients had bilateral immobility, and 5 had unilateral immobility. Monopolar EMG was performed on all patients. Fine-wire EMG was performed when monopolar EMG did not elucidate the cause of the immobility. The EMG studies revealed synkinetic reinnervation in all subjects. On the basis of the EMG results, 7 of the 10 patients were treated with botulinum toxin to weaken the undesired reinnervation. Three of the 7 patients had benefit from this therapy. Laryngeal synkinesis should be considered as part of the differential diagnosis of vocal fold immobility. Awake laryngeal EMG is the only method to demonstrate synkinesis of the larynx. The diagnosis of synkinesis is clinically significant in cases of immobility to identify patients who might benefit from botulinum toxin therapy. Additionally, the presence of synkinesis in cases of unilateral immobility may be a contraindication to laryngeal reinnervation procedures. The benefit of botulinum toxin therapy is likely greater in the treatment of bilateral as opposed to unilateral immobility.     

Laryngeal reinnervation for unilateral traumatic recurrent laryngeal nerve injuries]

OBJECTIVE: To investigate 5 procedures of laryngeal reinnervation for unilateral vocal cord paralysis induced by traumatic recurrent laryngeal nerve injury. METHODS: 35 cases were selected for our study, all patients had unilateral recurrent laryngeal nerve injury, including 8 for nerve decompression, 6 for end to end anastomosis of recurrent laryngeal nerve, 16 for main branch of ansa cervicalis anastomosis to recurrent laryngeal nerve, 3 for nerve muscular pedicle and 2 for nerve implantation. All cases have been subjected to preoperative and postoperative voice recording, acoustic analysis, videolaryngoscopy, strobscopy and electromyography. RESULTS: It is found the adductory and abductory motion of the vocal cord restored in 5 cases with less than 4 months course who received nerve decompression. Although functional motion of vocal cord was not seen in two patients who received nerve decompression with a course longer than 4 months and one less than 4 months, and in all cases who received ansa cervicalis anastomosis and end to end anastomosis of recurrent laryngeal nerve, these procedures resulted in medialization of vocal cord and the mass and tension of the reinnervated vocal cord may become much the same as the contralateral normal vocal cord, thus resuming symmetric vibration of the vocal cords and physiological phonation. Nerve muscular pedicle technique and nerve implantation enabled adductory muscles to be reinnervated, thus improving severe hoarseness, but they didn't restore normal voice. CONCLUSIONS: (1) Nerve decompression seems to be the best procedure in laryngeal reinnervation; (2) Main branch of ansa cervicalis technique raises satisfactory reinnervation of adductor muscles; (3) Selection of the laryngeal reinnervation protocols should depend on the course, severity and type of nerve injury.     

Intrinsic laryngeal muscle reinnervation using the muscle-nerve-muscle technique

OBJECTIVES: This study was performed to investigate the muscle-nerve-muscle reinnervation technique in the larynx, in which a nerve conduit implanted into an innervated muscle conducts axonal sprouting into a denervated muscle while maintaining function of the donor muscle. METHODS: In this study, the muscle-nerve-muscle technique was used to direct superior laryngeal nerve axons to reinnervate intrinsic laryngeal muscles by implanting the recurrent laryngeal nerve stump into the cricothyroid muscle in 8 dogs. In 4 of the dogs, the recurrent laryngeal nerve trunk to the adductor muscles was divided so that all axonal sprouting was directed to the posterior cricoarytenoid muscle. Six-month electromyography data were obtained from 6 of the 8 dogs. RESULTS: All 6 dogs showed evidence of successful reinnervation of the thyroarytenoid or posterior cricoarytenoid muscles with action potentials that corresponded to spontaneous respiratory efforts, while the donor cricothyroid muscles retained their phasic contraction. These responses were obliterated when the recurrent laryngeal nerve conduit was divided. Histologic examination of the intrinsic laryngeal muscles demonstrated successful reinnervation. CONCLUSIONS: The results confirm that intrinsic laryngeal muscles may be successfully reinnervated by the superior laryngeal nerve with the muscle-nerve-muscle technique, without sacrifice of function of the cricothyroid muscle. This method offers an alternative source of appropriately firing axons for laryngeal reinnervation procedures.     

Bipolar radiofrequency-induced thermotherapy (rfitt) for the treatment of spasmodic dysphonia. A report of three cases

The symptoms of adductor spasmodic dysphonia are most commonly palliated by periodic botulinum toxin injections. The need for repeated injections, difficulty in obtaining injections and cost make this form of treatment intolerable for some patients. To address these concerns, we propose a new treatment approach utilizing trans-oral recurrent nerve coagulation. The goal is to weaken the force of laryngeal closure during spasms by creating fibrosis of the terminal branches of one recurrent nerve through coagulation. Under general anesthesia without paralysis, an electrical stimulator is used to identify the region within the thyroarytenoid muscle that produces the greatest contraction with minimal stimulation. The radiofrequency laryngeal probe or electrocautery device is introduced into this position, and energy is delivered. The location of the region of maximal stimulation is usually just lateral and anterior to the vocal process of the arytenoids. Between 1989 and 2000, seven patients were treated with electrocautery. To achieve remission of spasms, three patients needed three sessions, four needed two sessions and one only one session. Since 2001, three patients have achieved remission of spasms with a single treatment with radiofrequency during which 80 J was delivered. Voice results are comparable to those obtained with botulinum toxin. Initially, the voice is breathy and laryngeal examination shows complete vocal fold immobility. After 1–2 months, the voice improves and examination reveals unilateral hypomobility. Trans-oral recurrent nerve coagulation is an effective alternative to botulinum toxin injections.     

Potential of laryngeal muscle regeneration using induced pluripotent stem cell-derived skeletal muscle cells

Conclusion Induced pluripotent stem (iPS) cells may be a new potential cell source for laryngeal muscle regeneration in the treatment of vocal fold atrophy after recurrent laryngeal nerve paralysis. Objectives Unilateral vocal fold paralysis can lead to degeneration, atrophy, and loss of force of the thyroarytenoid muscle. At present, there are some treatments such as thyroplasty, arytenoid adduction, and vocal fold injection. However, such treatments cannot restore reduced mass of the thyroarytenoid muscle. iPS cells have been recognized as supplying a potential resource for cell transplantation. The aim of this study was to assess the effectiveness of the use of iPS cells for the regeneration of laryngeal muscle through the evaluation of both in vitro and in vivo experiments. Methods Skeletal muscle cells were generated from tdTomato-labeled iPS cells using embryoid body formation. Differentiation into skeletal muscle cells was analyzed by gene expression and immunocytochemistry. The tdTomato-labeled iPS cell-derived skeletal muscle cells were transplanted into the left atrophied thyroarytenoid muscle. To evaluate the engraftment of these cells after transplantation, immunohistochemistry was performed. Results The tdTomato-labeled iPS cells were successfully differentiated into skeletal muscle cells through an in vitro experiment. These cells survived in the atrophied thyroarytenoid muscle after transplantation.     

Nimodipine improves reinnervation and neuromuscular function after injury to the recurrent laryngeal nerve in the rat

OBJECTIVES: Injury of the recurrent laryngeal nerve (RLN) is associated with a high degree of neuronal survival, but leads to various levels of vocal fold motion impairment or laryngeal synkinesis, which has been attributed to misdirected reinnervation of the target muscles in the larynx or aberrant, competing reinnervation from adjacent nerve fibers. The aim of the present study was to evaluate the impact of the regeneration-promoting agent nimodipine on reinnervation and neuromuscular function following RLN crush injury. METHODS: Sixty adult rats were randomized into nimodipine-treated or untreated groups and then underwent RLN crush injury. Reinnervation of the posterior cricoarytenoid muscle (PCA) was assessed by electrophysiological examination, retrograde tracing of lower motor neurons before and after injury, and quantification of neuromuscular junctions in the PCA muscle. RESULTS: At 6 weeks after injury, the nimodipine-treated animals showed significantly enhanced neuromuscular function and also demonstrated a higher number of motor neurons in the brain stem that had reinnervated the PCA, compared to the untreated animals. The somatotopic organization of ambiguus motor neurons innervating the larynx was similar before injury and after reinnervation. CONCLUSIONS: Nimodipine improves regeneration and neuromuscular function following RLN injury in the adult rat, and could be of use in future strategies following RLN injury.