Anat Cell Biol.  2024 Sep;57(3):392-399. 10.5115/acb.24.052.

Development and cross-sectional morphology of the recurrent laryngeal nerves in human fetuses

• Baratela MC
• Mayer WP
• Baptista JdS

• KMID: 2559756
• DOI: http://doi.org/10.5115/acb.24.052

Abstract

The recurrent laryngeal nerve is a bilateral branch of the vagus nerve that is mainly associated with the motor innervation of the intrinsic muscles of the larynx. Despite its bilateral distribution, the right and left recurrent laryngeal nerves display unequal length due to embryological processes related to the development of the aortic arches. This length asymmetry leads to theories about morphological compensations to provide symmetrical functions to the intrinsic muscles of the larynx. In this study we investigated the developmental and cross-sectional morphometrics of the recurrent laryngeal nerves in human fetuses. Fifteen stillbirth fetuses donated to anatomical and medical research were used for investigation. Fetuses had intrauterine age ranging from 30 to 40 weeks estimated by biometry methods. Specialized anatomical dissection of the visceral block of the neck was performed to prepare histological samples of the recurrent laryngeal nerves in its point of contact with the larynx, and morpho-quantitative techniques were applied to evaluate the epineurium and perineural space of the recurrent laryngeal nerves. No statistical difference in the cross-sectional morphology of the epineurium and perineural space between right and left recurrent laryngeal nerves intra-individually was confirmed, however, we found evidence that these structures are under greater development in the left recurrent laryngeal nerve during 30 to 40 weeks of intrauterine life. Our data suggest that the nerves are under morphological development that possibly set the stage for accommodation of larger diameter and myelinization of the left recurrent laryngeal nerve during post-natal life.

Keyword

Recurrent laryngeal nerve; Nerve tissue; Fetal development; Larynx

Figure

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  Fig. 1 Diagram of the external anatomy of the larynx, trachea, pharyngeal muscles, and esophagus in lateral view. The plane of the study for histological preparations is indicated in pink. Note that the cricopharyngeus muscle covers the recurrent laryngeal nerve in its final trajectory within the tracheoesophageal groove.

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  Fig. 2 Recurrent laryngeal nerve in human fetuses. (A, B) Photomacrography of the vagal detachment of the right and left recurrent laryngeal nerves. Red, aorta and its branches. Turquoise, phrenic nerve. Blue, pulmonary trunk. Purple, ductus arteriosus. Yellow, vagus nerve. Green, recurrent laryngeal nerve. Tr, trachea; Lg, right lung; Hr, heart. Scale bar=1 cm.

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  Fig. 3 Histological preparation and cross-sectional morphometry of the recurrent laryngeal nerve. (A) Photomicrography of the right recurrent laryngeal nerve (RLN) superficially to the cricoid cartilage (CC) and deep to the cricopharyngeus muscle (CPM). Scale bar=100 µm. (B) Inset of the RLN showing the analyzed area of histology slides for epineurium (black line), and perineural space (yellow shadow). Note the presence of two vasa nervorum within the epineurium and nerve axons enveloped by the perineural epithelium. Scale bar=50 µm. (C) Cross-sectional area of the epineurium (I) and perineural space (II) from the right (R) and left (L) recurrent laryngeal nerves. R and L scores from the same subjects are connected in the plot. ns, non significant P>0.05.

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  Fig. 4 Development of the epineurium and perineural space from 30 to 40 weeks. (A) Scatter plot showing the relationship between gestational age and cross-section morphometry of the perineurium in right (I) and left (II) recurrent laryngeal nerves. (B) Scatter plot showing the relationship between gestational age and cross-section morphometry of the perineural space in right (I) and left (II) recurrent laryngeal nerves. Linear regression equation, correlation coefficient and probability values are represented by y, R, and P within the plots.

Reference

References

1. Damste PH. 1965; The larynx as a wind-instrument. Pract Otorhinolaryngol (Basel). 27:94–5. DOI: 10.1159/000274637. PMID: 14302934.

2. Saito S, Fukuda H, Kitahara S, Kokawa N. 1978; Stroboscopic observation of vocal fold vibration with fiberoptics. Folia Phoniatr (Basel). 30:241–4. DOI: 10.1159/000264132. PMID: 730090.

3. Gardner ED, O'Rahilly R, Müller F. Gardner-Gray-O'Rahilly anatomy: a regional study of human structure. 5th ed. Saunders;1986.

4. Dalley AF, Agur AMR. Moore's clinically oriented anatomy. 9th ed. Wolters Kluwer Health;2021.

5. Maranillo E, de Blas CS, Górriz MC, Quinones S, Verdú E, Quer M, León X, Vázquez T, Sañudo JR, Konschake M. 2021; Comparative study of the length of human laryngeal nerves and their variations: functional and clinical considerations. Eur J Anat. 25:653–63.

6. Prades JM, Dubois MD, Dumollard JM, Tordella L, Rigail J, Timoshenko AP, Peoc'h M. 2012; Morphological and functional asymmetry of the human recurrent laryngeal nerve. Surg Radiol Anat. 34:903–8. DOI: 10.1007/s00276-012-0999-7. PMID: 23150169.

7. Valenzuela-Fuenzalida JJ, Baeza-Garrido V, Navia-Ramírez MF, Cariseo-Ávila C, Bruna-Mejías A, Becerra-Farfan Á, Lopez E, Orellana Donoso M, Loyola-Sepulveda W. 2023; Systematic review and meta-analysis: recurrent laryngeal nerve variants and their implication in surgery and neck pathologies, using the anatomical quality assurance (AQUA) checklist. Life (Basel). 13:1077. DOI: 10.3390/life13051077. PMID: 37240722. PMCID: PMC10221453.

8. Atkins JP Jr. 1973; An electromyographic study of recurrent laryngeal nerve conduction and its clinical applications. Laryngoscope. 83:796–807. DOI: 10.1288/00005537-197305000-00015. PMID: 4702478.

9. Pascual-Font A, Merchán A, Maranillo E, Brillas A, Sañudo JR, Valderrama-Canales FJ. 2006; Morphometry of the recurrent laryngeal nerves of the rat. Acta Otorrinolaringol Esp. 57:435–40. Spanish. DOI: 10.1016/S0001-6519(06)78744-1. PMID: 17228641.

10. Patel RR, Awan SN, Barkmeier-Kraemer J, Courey M, Deliyski D, Eadie T, Paul D, Švec JG, Hillman R. 2018; Recommended protocols for instrumental assessment of voice: American Speech-Language-Hearing Association expert panel to develop a protocol for instrumental assessment of vocal function. Am J Speech Lang Pathol. 27:887–905. DOI: 10.1044/2018_AJSLP-17-0009. PMID: 29955816.

11. Harrison DF. 1981; Fibre size frequency in the recurrent laryngeal nerves of man and giraffe. Acta Otolaryngol. 91:383–9. DOI: 10.3109/00016488109138519. PMID: 7023176.

12. Shin T, Rabuzzi DD. 1971; Conduction studies of the canine recurrent laryngeal nerve. Laryngoscope. 81:586–96. DOI: 10.1288/00005537-197104000-00010. PMID: 5553125.

13. March MI, Warsof SL, Chauhan SP. 2012; Fetal biometry: relevance in obstetrical practice. Clin Obstet Gynecol. 55:281–7. DOI: 10.1097/GRF.0b013e3182446e9b. PMID: 22343244.

14. Streeter GL. 1920; Weight, sitting height, head size, foot length, and menstrual age of the human embryo. Carnegie Inst Wash. 11:143–79.

15. Kinoshita H, Umezawa T, Omine Y, Kasahara M, Rodríguez-Vázquez JF, Murakami G, Abe S. 2013; Distribution of elastic fibers in the head and neck: a histological study using late-stage human fetuses. Anat Cell Biol. 46:39–48. DOI: 10.5115/acb.2013.46.1.39. PMID: 23560235. PMCID: PMC3615611.

16. Mallory FB. 1900; A contribution to staining methods: I. A differential stain for connective-tissue fibrillae and reticulum. II. Chloride of iron haematoxylin for nuclei and fibrin. III. Phosphotungstic acid haematoxylin for neuroglia fibres. J Exp Med. 5:15–20. DOI: 10.1084/jem.5.1.15. PMID: 19866932. PMCID: PMC2117995.

17. Sadler TW. Langman's medical embryology. 15th ed. Wolters Kluwer;2024.

18. World Health Organization. c2024. Stillbirth [Internet]. World Health Organization;Available from: https://www.who.int/health-topics/stillbirth. cited 2024 Apr 4.

19. Aminu M, Bar-Zeev S, van den Broek N. 2017; Cause of and factors associated with stillbirth: a systematic review of classification systems. Acta Obstet Gynecol Scand. 96:519–28. DOI: 10.1111/aogs.13126. PMID: 28295150. PMCID: PMC5413831.

20. Naidu L, Lazarus L, Partab P, Satyapal KS. 2014; Laryngeal nerve "anastomoses". Folia Morphol (Warsz). 73:30–6. DOI: 10.5603/FM.2014.0005. PMID: 24590520.

21. Henry BM, Pękala PA, Sanna B, Vikse J, Sanna S, Saganiak K, Tomaszewska IM, Tubbs RS, Tomaszewski KA. 2017; The anastomoses of the recurrent laryngeal nerve in the larynx: a meta-analysis and systematic review. J Voice. 31:495–503. DOI: 10.1016/j.jvoice.2016.11.004. PMID: 27939121.

22. Ellwanger JH, da Costa Rosa JP, dos Santos IP, da Rosa HT, Jotz GP, Xavier LL, de Campos D. 2013; Morphologic evaluation of the fetal recurrent laryngeal nerve and motor units in the thyroarytenoid muscle. J Voice. 27:668–73. DOI: 10.1016/j.jvoice.2013.07.004. PMID: 24128892.

23. Jotz GP, de Campos D, Rodrigues MF, Xavier LL. 2011; Histological asymmetry of the human recurrent laryngeal nerve. J Voice. 25:8–14. DOI: 10.1016/j.jvoice.2009.06.007. PMID: 20083375.

24. Harding R. 1984; Function of the larynx in the fetus and newborn. Annu Rev Physiol. 46:645–59. DOI: 10.1146/annurev.ph.46.030184.003241. PMID: 6370121.

25. Fleming JC, Gibbins N, Ingram PJ, Harries M. 2011; An anatomical study of the myelination of human laryngeal nerves. J Laryngol Otol. 125:1263–7. DOI: 10.1017/S0022215111001939. PMID: 21854693.

26. Woźniak W, O'Rahilly R. 1981; Fine structure and myelination of the developing human vagus nerve. Acta Anat (Basel). 109:218–30. DOI: 10.1159/000145387. PMID: 7257721.

27. Tanaka S, Mito T, Takashima S. 1995; Progress of myelination in the human fetal spinal nerve roots, spinal cord and brainstem with myelin basic protein immunohistochemistry. Early Hum Dev. 41:49–59. DOI: 10.1016/0378-3782(94)01608-R. PMID: 7540130.

28. Standring S, Gray H. Gray's anatomy: the anatomical basis of clinical practice. 42nd ed. Elsevier;2021. p. 1588.

29. Lanigan LG, Russell DS, Woolard KD, Pardo ID, Godfrey V, Jortner BS, Butt MT, Bolon B. 2021; Comparative pathology of the peripheral nervous system. Vet Pathol. 58:10–33. DOI: 10.1177/0300985820959231. PMID: 33016246.

30. Brown IS. 2016; Pathology of perineural spread. J Neurol Surg B Skull Base. 77:124–30. DOI: 10.1055/s-0036-1571837. PMID: 27123388. PMCID: PMC4846404.

31. Crowe TP, Greenlee MHW, Kanthasamy AG, Hsu WH. 2018; Mechanism of intranasal drug delivery directly to the brain. Life Sci. 195:44–52. DOI: 10.1016/j.lfs.2017.12.025. PMID: 29277310.

32. Shanthaveerappa TR, Bourne GH. 1966; Perineural epithelium: a new concept of its role in the integrity of the peripheral nervous system. Science. 154:1464–7. DOI: 10.1126/science.154.3755.1464. PMID: 5921613.