Larynx is a conduit between the pharynx and trachea. It contributes to the functions of breathing, vocalization and deglutition (Fulton et al., 2012). This organ comprises several cartilages viz., epiglottis, thyroid, arytenoid and cricoid, amongst which the arytenoid is a paired one (Sisson et al., 1975). Complexity of laryngeal anatomy is the result of curious relationships that exist among the five laryngeal cartilages (Michaels, 1990). It has been well stated that “if one wishes to become acquainted with the structure and form of any organ, there is perhaps no better way than to trace its development step by step, back into the early embryonic stages (Shikinami, 1926). The development of larynx is divided into embryonic and fetal periods of which the former occurs during first 8 weeks of intrauterine life. Development starts as a ventral groove in the pharynx known as laryngotracheal groove during the 4^th week of gestation which fuses caudo-cranially. The cranial part of the laryngeo- tracheal groove develops into the larynx. Major portions of the larynx including the epiglottis, thyroid, cricoid and arytenoid cartilages are formed from the fourth and sixth pharyngeal arches in domestic animals (Hyttel et al., 2010).

The present study was conducted for the period of three years from 2020 to 2023 at the Department of Veterinary Anatomy, College of Veterinary Science, Rajendranagar,  Hyderabad. The approximate age of prenatal specimens was calculated by adopting the equation of Gall et al., (1994).
 

Y = 2.74 X + 30.15

 
‘Y’ = Age of the fetus (in days).
‘X’ = Crown rump length (CRL) of the fetus (in cm).
       
CRL of fetal specimens was measured in ‘cm’ with the help of a thread and scale to assess the approximate age of embryos and fetuses in days (Y). Prenatal specimens were categorized into three groups based on their crown rump length (CRL) viz., Group I (1 cm to 4.6 cm) i.e., day 33 to 43 days, Group II (6 cm - 22 cm) aged between 47 to 90 days, whereas specimens of Group III (22.5 - 42.0 cm) ranged from 92 to 146 days respectively (Kadu and Kaikeni, 1987). Gross morphological features of larynx and its constituent cartilages were studied in fetuses aged 47 days (6 cm CRL) and above. Specimens were dissected along with intact neck carefully to the extent possible to observe and record the gross features of laryngeal cartilages and their associated structures. Group II larynges were dissected by using a dissecting microscope while the length and width of larynx of prenatal specimens as a whole and laryngeal cartilages of Group II and Group III were measured using digital Vernier calipers (Harjeet et al., 2010).

In the present study, 1.5 cm CRL i.e., 34 days of age, fetal specimens of group I showed incompletely differentiated pre-cartilaginous larynx with mesenchymal condensation which was wide anteriorly and narrow at posterior end (Fig 1A). Fetal larynx was located within the inter-mandibular space which by 41 days of age was extended till the anterior end of the second cervical vertebrae. Grossly, chondrification was evident by 41 days (3.8 cm CRL) wherein thyroid and cricoid cartilages were demarcated on ventral aspect of the neck. However, during group I period all cartilages were not yet formed, but an immature thyro-hyoideus muscle was seen (Fig 1B). The primitive larynx of 43 days (4.6 cm CRL) fetus consisted of epiglottis, thyroid, arytenoid and cricoid cartilages among which epiglottis and arytenoid swellings were still in pre-cartilagenous state and also primitive stylo-thyrohyoideus and crico-arytenoideus dorsalis muscles were evident (Fig 2). In the present study, all the laryngeal cartilages were appreciable in 48 days foetus (6.6 cm CRL, Group II specimens), wherein they resembled the shape of adult cartilages but were thin flap like and flaccid from which few cartilaginous nodules were isolated from the arytenoid cartilage (Fig 3). The present findings were in accordance with Rodrigues et al., (2014), whose studies revealed that the development of larynx was evident in the equines at 26 days of gestation, where they noticed the epiglottic protrusions with the evidence of thyroid and cricoid cartilage by day 30 of gestation, which were completely developed by 45 days.






       
In group II fetuses, the epiglottis differentiated into a leaf shaped cartilage with an apex and base out of which the apex was pre-cartilaginous and the latter presented a ventral growth i.e., (fat pad) projecting ventrally. However, the petiole of epiglottis was not yet developed at this stage.       

The dorsal surface of epiglottis was curved and the base was much wider. The epiglottis in early prenatal stage was softer to touch (Fig 4A) because of it was composed mostly of embryonic tissue, whereas from middle age group onwards its touch became more resilient and its shape was becoming more definite with developing fat pad and petiole extending onto the dorsal surface of thyroid cartilage similar to the findings in adult Zebu (Kalita et al., 2003) (Fig 4B and C).


       
Arytenoid cartilage attained the shape as in adult with all the processes differentiated except the corniculate process which was not evident by 64 days (Fig 5A). The medial surface of the arytenoid revealed the elevated convex hyaline cartilage. The muscular process gradually attained the shape and size by the mid prenatal stage. Except for the elevated portion, the rest of the structure consisted of undifferentiated cartilage. Corniculate process with tuberculum corniculatum was developed by 71 days of gestation (Fig 5B and C). However, the curvature of the corniculate process was not yet formed. Above findings showed that development of goat Larynx was more advanced than human fetal larynges where chondrification progressed in 10th to 12th weeks (Harjeet et al., 2010 and Raina et al., 2013). According to the latter author, maturity of laryngeal cartilages advanced in caudocranial order, while in the present study vocal cord was last to be appreciated grossly.


       
Laryngeal prominence of thyroid cartilage was developed, although less prominent in 48 days fetus which showed early fusion compared to humans where it was seen by 16 weeks of intrauterine life (Raina et al., 2013). The anterior cornu was short and thin while the posterior one was long, broad and more curved than that of adult larynx. The anterior fissure was in the form of a shallow notch while the posterior one was like a fissure (Fig 6A and B). Anterior cornu of thyroid cartilage increased in size and attained feature of adult larynx by 64 days. Cranial fissure was well appreciated while the cartilage at the ventral laryngeal prominence was thin than the other part of the thyroid cartilage where it constituted ventral diverticulum centrally (Fig 6C) which was prominent in 88 days old fetal larynx.




In 48 day fetal larynx, the dorsal lamina of cricoid was developed whereas the lateral arches were incompletely formed while the ventral arch was not yet progressed (Fig 3).                 

Cricoid cartilage was thin, signet ring shaped with minimal chondrification at 64 days of age (Fig 7A). Since this age, the dorsal lamina was advanced in growth than the ventral arch and it acquired muscular process by 71 days (14.3 cm CRL) (Fig 8), whereas by 74 days the cricoid cartilage resembled the signet ring shape with a defined lamina and ventral arches (Fig 7B). By 71 days (14.3 cm CRL), Intrinsic muscles such as cricoarytenoids dorsalis and lateralis, thyro-arytenoideus and crico-thyroideus were better developed, which later increased in size but not in structural features which is in akin with the findings of Alli et al., (2013) in rats, who observed the immature structures of the larynx and the skeletal muscle at E17 stage.




       
Recurrent laryngeal nerve was identified in 84 days (19.7 cm CRL) fetus (Fig 9A). Wherein the shape and structure of laryngeal cartilages and muscles imprinted the adult larynx (Fig 9B). As per the present studies, the laryngeal cavity was closed at the glottic area as the vocal tract was close to each other in early stages and was not continuous with infra-glottic cavity. Cavity was well defined with lumen that continued posteriorly into the infra-glottic area and trachea by 60 (14 cm CRL) days beyond which it gradually widened towards 90 days (21.8 cm CRL) (Fig 10A and B respectively). Vocal cords increased in mass by 105 days of age and by last trimester of gestation i.e., 42.0 cm CRL stage (146 days), all the laryngeal cartilages were similar in form and nature to the adult cartilages (Fig 11).






       
Morphometrical studies of samples revealed that there was gradual increase in the dimensions of larynx in proportion with advancing age in development. Mean length, breadth (transverse diameter) and height (antero-posterior diameter) of Group I (34 to 43 days) to Group III (95-146 days) larynges were in range of 1.86±0.22 mm to 13.78±0.96 mm, 1.61±0.22 mm to 11.39±0.79 mm and 1.02±0.16 to 12.11±0.81 mm respectively (Table 1). Mean length, breadth and height of thyroid cartilage of group II to Group III ranged from 6.08 ± 0.61 mm to 13.02±1.09 mm, 4.79±0.48 mm to 10.70±0.73 mm and 5.56±0.65mm to 12.05±1.09 mm respectively while that of cricoid were 4.30±0.71 mm to 10.62±0.89 mm, 2.69±0.43 mm to 7.28±0.58 and 5.43±0.61 to 10.10±1.07 respectively. Mean length and breadth of prenatal arytenoid cartilages ranged from 3.02-±0.46 mm to 6.43±0.33 mm and 1.80±036 mm to 5.35±0.41 mm whereas that of epiglottis was in range of 4.81±0.59 mm to 11.33±0.77 and 2.51±0.45 to 6.77±0.42 mm respectively (Table 2). Similar inference was drawn in humans by Harjeet et al., (2010) and in rats by Alli et al., (2013). They justified that the dimensions of larynx, epiglottis and cricoid cartilage showed positive correlation with CRL.

By the early second term of gestation, the larynx has a fully established basic framework, including the major cartilages “thyroid, cricoid, arytenoids and epiglottis”. Developmental studies of the larynx in goats conclude that the organ undergoes significant structural maturation during the fetal period, resulting in a complex, functional structure ready for postnatal life. In summary, this research contributes significant data to the anatomical understanding of the caprine larynx and highlights the clinical importance of unimpeded prenatal development for maintaining a patent airway for optimal respiration in goats after birth. Coordinated neuromuscular development results in complex pathways that facilitate vital reflexes for swallowing, breathing, protecting the airway and phonation.

The present study was supported by Department of Veterinary Anatomy and Histology, College of Veterinary Science, Rajendranagar, PVNR TVU, Hyderabad and Department of Veterinary Anatomy and Histology, Veterinary College, Hebbal, KVAFSU, Bidar.
 
Disclaimers
 
The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
 
Informed consent
 
All animals for the present study were procured from slaughter house. Hence, permission was not obtained from the Animal ethics committee. Though all the standard procedures were followed in the work done.

The authors declare that there are no conflicts of interest regarding the publication of this article.