Variational anatomy of cervical part of phrenic nerve: An observational human cadaveric study

Abstract

Introduction:

The phrenic nerve (PN) is not an exception to the variations. Recent cadaveric studies report that the third and fifth cervical nerves may be involved. The PN communicating branch might be considered as another option to neurotise the suprascapular nerve. The variants of the relationship between the subclavian vein and the PN should be familiar to anesthesiologists during subclavian vein cannulation to achieve a successful vein approach without causing PN palsy.

Materials and Methods:

The study was carried out on both sides of 42 head and neck regions, which were obtained from the Department of Anatomy, KGMU, Lucknow. The study received Institutional Ethics Committee approval (Ref. code: XIV-PGTSC-IIA/P78).

Result:

Classical three root (C3,4,5), double root with varying contribution from C3 or C5 (C3,4/C4,5), and single root (C4/C5). Classical triple root PN formation was present in 57.68% of cases and was the most common pattern in our study population. The second most common (30.95%) formation was observed for the double root pattern (C4,5) by contribution of distal cervical segment (C5). Double root configuration of C3,4 was observed on 4.76% sides. Out of single root contribution incidence of C4 pattern was higher (4.76%) as compared to C5 (2,14%). One cadaver exhibited a dual C5 root with C4 on the right side in a male cadaver.

Conclusion:

Although researchers have demonstrated that the PN nucleus extends locally from spinal cord segments C3 to C5, histological findings suggest that maximum contribution is derived from the C4 spinal nerve.

INTRODUCTION

The phrenic nerve (PN) is an inspiratory nerve. PN is a mixed spinal nerve that arises mainly from the ventral rami of cervical segments. It originates at the upper part of the lateral border of the scalenus anterior and then passes vertically downward over its anterior surface, posterior to the prevertebral fascia, to pass posterior to the sternocleidomastoid (SCM), the inferior belly of the Omohyoids, the internal jugular vein, the transverse cervical artery, and the suprascapular artery. At the root of the neck, it passes anterior to the second part of the subclavian artery and enters the thorax by crossing medially anterior to the internal thoracic artery. The course of the right and left PNs is different. The pericardiophrenic vessels accompany it along most of its course in the thorax. PN is derived from the neural crest cells in the third week of intrauterine life. The septum transversum, which creates the thoracic diaphragm, descends from the cervical vertebral level to the thoracolumbar vertebral level between the fifth and sixth week. Along with the septum transversum, PN descends, carrying innervation from the ventral rami of C3 to C5.[1] PN is no exception to the variations. Normally, it is formed by the fourth cervical nerve, but cadaveric studies report that the third and fifth cervical nerves may be involved,[2] with a wide variation and associated accessory phrenic nerves (APNs). The diaphragm, the most important respiratory muscle, receives its motor supply mainly from the PN.[3] It helps the surgeons find out the exact cause of paralysis of the diaphragm and helps in sectioning the PN for the nerve graft in the repair of post-cricoarytenoid muscle.[4] It is necessary to be aware of the supraclavicular triangle below Erb’s point during neck dissection procedures.[5] The PN communicating branch might be considered as another option to neutralize the suprascapular nerve.[6] The variants of the relationship between the subclavian vein and the PN should be familiar to anesthesiologists during subclavian vein cannulation to achieve a successful vein approach without causing PN palsy.[7]

MATERIALS AND METHODS

Study settings

The study was conducted in the cadaveric lab of the Department of Anatomy. Ethical Clearance was obtained from Institutional Ethical Committee with (Ref. code: XIV PGTSC IIA/P78).

Principal selection

The study was carried out on 21 head and neck regions of cadavers of either sex, which were obtained from the Department of Anatomy.

Study design

Observational study

Sample size

21 cadavers (42 sides of neck)

Inclusion criteria

• Adult cadavers of both either sex of the adult age group.

Exclusion criteria

• Presence of any surgical marks in the area of interest.

• Presence of any scar in the area of interest.

• Short neck.

The study received Institutional Ethics Committee approval (Ref. code: XIV-PGTSC-IIA/P78).

Dissection protocol

Exposure of the PN on cadaveric specimens was acquired by the following surgical protocol:

Step 1: The procedure began with a vertical skin incision in the midline from the sternal notch to the hyoid bone [Figure 1a], then an oblique incision was made from the hyoid bone toward the mastoid process [Figure 1b].

Figure 1.

(a) Vertical skin incision line in the median plane from the sternal notch to the lower border of the hyoid bone. (b) The oblique incision line from the midpoint on the lower border of the hyoid bone up to the tip of the mastoid process

Step 2: Then, an incision was made along the upper border of the clavicle from the sternum to the anterior border of the trapezius muscle. Then, cutaneous and subplatysmal flaps were directed laterally toward the anterior border of the trapezius muscle.

Step 3: The cutaneous nerves were preserved and traced to their trunks as they penetrated the deep fascia at the midpoint of the posterior border of the SCM.

Step 4: The incision was made through the investing layer of the deep fascia of the neck above the clavicle and along the anterior margin of the SCM to expose the deeper structures of the anterior triangle.

Step 5: The fascia covering the anterior border of the SCM muscle was incised and then the sternal and clavicular heads of the SCM were incised followed by a reflection of the muscle belly toward its mastoid attachment.

Step 6: The carotid sheath was visualized and retracted medially. This helped in visualizing the prevertebral fascia.

Step 7: At the junction of the carotid sheath with the omohyoid muscle, the dissection was directed posteriorly. The vertical fibers of the scalenus anterior muscle were identified to locate the main trunk of the PN, which was located on its medial aspect.

Step 8: The prevertebral fascia was removed and separated to the upper border of the clavicle.

Step 9: The main trunk of the PN was finely traced upward toward the contributing cervical rootlets and variations were noted.

Step 10: Morphometric analysis (measurement of length and diameter) of the PN was performed.

Step 11: Relation of the PN was noted with scalenus anterior, transverse cervical vessels, and subclavian vein at the root of the neck.

Step 12: The cervical part was thoroughly observed for any communicating branches with nearby nerves and the origin of APN.

Step 13: Observe the descent of the PN in relation to the superficial cervical artery and subclavian vein.

Statisticalanalysis

Sample size

21 neck–42 sides

(Rosner B. Fundamentals of Biostatistics. 7^th ed. Boston, MA: Brooks/Cole; 2011.)

Sample Size
Group 1	16
Group 2	16
Total	32

p1, p2 = proportion (incidence) of groups 1 and 2

Ø = |p2 − p1| = absolute difference between two proportions

n1 = sample size for group 1

n2 = sample size for group 2

Study Parameters
Incidence, group 1	60.6%
Incidence, group 2	12.1%
Alpha	0.05
Beta	0.15
Power	0.85

α = probability of type I error (usually 0.05)

β = probability of type II error (usually 0.2)

z = critical Z value for a given α or β

K = ratio of sample size for group 2 to group 1

The frequency of group 1 (phrenic nerve) (i.e., 60.6%) and the frequency of group 2 (Ansa cervicalis) (i.e., 12.1%) from the study. Washout = 10% samples during the study considered. Thus, the total sample was 42.

OBSERVATION AND RESULT

A total of 21 cadavers were examined during the study period, in which a total of 21 (100%) right side, and 21 (100%) left side were examined. Out of 21, a total of 15 (71.43%) cadavers were male and 6 (28.57%) cadavers were female.

Table 1 shows the percentage of contributing roots of PN in males and females according to its side, out of 21, a total of 15 males and 6 females. The percentage of C3,4,5, C3,4, C4,5, C3, C4, C5 and C4, C5, C5 roots of PN were 40.00%, 6.67%, 40.00%, 0.00%, 6.67%, 0.00%, 6.67% in male and 66.67%, 0.00%, 16.67%, 0.00%, 0.00%, 16.67%, 0.00% in female right side whereas 53.33%, 6.67%, 33.33%, 0.00%, 6.67%, 0.00%, 0.00% in male and 83.33%, 0.00%, 16.67%, 0.00%, 0.00%, 0.00%, 0.00% in female left side.

Table 1.

Percentage of contributing roots of PN in male and female according to its side

	Right			Left			Total (42) necks	Percentage of contributing roots (%)
	Male (15)	Female (6)	Total (21)	Male (15)	Female (6)	Total (21)
C3,4,5	6 (40.00%)	4 (66.67%)	10	8 (53.33%)	5 (83.33%)	13	23	54.76
C3,4	1 (6.67%)	0 (0%)	1	1 (6.67%)	0 (0%)	1	2	4.76
C4,5	6 (40.00)	1 (16.67%)	7	5 (33.33%)	1 (16.67%)	6	13	30.95
C3	0 (0%)	0 (0%)	0	0 (0%)	0 (0%)	0	0	0
C4	1 (6.67%)	0 (0%)	1	1 (6.67%)	0 (0%)	1	2	4.76
C5	0 (0%)	1 (16.67%)	1	0 (0%)	0 (0%)	0	1	2.38
C4, C5, C5	1 (6.67%)	0 (0%)	1	0 (0%)	0 (0%)	0	1	2.38

Table 2 shows the frequency of various patterns of roots of the PN. The percentage of C3,4,5, C3,4, C4,5, C3, C4, C5 and C4, C5, C5 roots of PN were 46.66%, 6.6%, 36.66%, 0.00%, 6.6%, 0.00%, 2.38% in male and 75.0%, 0.0%, 16.66%, 0.0%, 0.0%, 8.33%, and 0.00% in female, respectively.

Table 2.

Distribution of frequency of similarity and dissimilarity of Roots of phrenic nerve

	Similar pattern of roots to PN on both sides			Dissimilar pattern of roots to PN on both sides
	Male (15)	Female (6)	Total (42) 21 × 2	Male (15)		Female (6)		Total (42)
				Right	Left	Right	Left
C3,4,5	5 (33.33%)	4 (83.33%)	10 (47.61%)	1 (6.66%)	3 (20%)	0 (0%)	0 (0%)	4 (9.52%)
C3,4	0 (0%)	0 (0%)	0 (0%)	1 (6.66%)	1 (6.66%)	0 (0%)	0 (0%)	2 (4.76%)
C4,5	3 (20.00%)	1 (16.66%)	4 (19.04%)	3 (20%)	2 (13.33%)	0 (0%)	0 (0%)	5 (11.90%)
C3	0 (0%)	0 (0%)	0 (0%)	0 (0%)	0 (0%)	0 (0%)	0 (0%)	0 (0%)
C4	0 (0%)	0 (0%)	0 (0%)	1 (6.66%)	1 (6.66%)	0 (0%)	0 (0%)	2 (4.76%)
C5	0 (0%)	0 (0%)	0 (0%)	0 (0%)	0 (0%)	1 (6.66%)	0 (0%)	1 (2.38%)
C4, C5, C5	0 (0%)	0 (0%)	0 (0%)	1 (6.66%)	0 (0%)	0 (0%)	0 (0%)	1 (2.38%)
Total	8 (53.33%)	5 (83.33%)	14 × 2 (66.66%)	7 (46.66%)	7 (46.66%)	1 (16.66%)	0 (0%)	14 (33.33%)

DISCUSSUION

The literature quotes that the variations in the origin, course, and relations of the cervical part of PN are common and have several clinical implications, even in extremely simple and routine procedures.[8] During therapeutic paralysis of a diaphragm in tubercular cases, clinicians observed significant variations in the origin of PNs.[9] Procedures like supraclavicular block, cannulation of the right subclavian vein, and a routine vascular access technique may lead to ipsilateral diaphragmatic paralysis.[7,10] A thorough knowledge of the anatomy of the PN is essential to preserve the mobility of the diaphragm while the removal of metastatic lymph nodes is done in the neck region.[11] Accessory roots of the PN have also been utilized for re-innervating recurrent laryngeal nerves in cases of vocal cord paralysis.[12] Apart from inadvertent damage of the PN during surgical procedures, clinicians are also utilizing this nerve to free quadriplegic patients from ventilatory support.[13] Therefore, to perform various surgeries safely and effectively in this area, one must have a thorough understanding of the anatomic variations of the PN. In the view that morphological variations exhibit geographical patterns or differences, this descriptive observational study has been aimed to explore the morphology and course of PNs in the cervical region in human cadavers of the Uttar Pradesh region.

According to most popular textbooks of anatomy, the PN is derived from C3, C4, and C5 nerve roots. Suggest that principal PNs were present in 97.1%. On the other six sides, the nerve had a similar origin but passed anterior to the SV and therefore was designated as an APN without PN. In contrast, 52% had only the PN without APN or secondary PNs.[14] According to Clemente CD the PN typically arises from the root of C4, with support from C3 and C5. Formally, the nerve is considered a member of the cervical plexus because it usually emerges from C4.[15] Although researchers have demonstrated that the PN nucleus extends locally from spinal cord segments C3 to C5,[16] histological findings suggest that maximum contribution is derived from C4 spinal nerve.[4]

The literature emphasizes the fact that contributing fibers show a high degree of variations in their origin. A total of eight unique PN rootlet patterns were identified in a cadaveric feasibility study in a North American population.[17] They observed the highest frequency (35%) of triple root PN (C3,4,5) followed by double roots C3,4 (26%), and C4,5 (21%), respectively. Out of single root PN, maximum frequency was observed for C4 (10%) followed by C3 (3%).

On the contrary, in a study conducted in Japan on 208 necks in more than 50% (C4,5) cases, a double root pattern was observed. An extraordinary frequency of single root (43%) PN from the C4 was noted by them in their study population and only 1.5% of cases exhibited a classical three roots (C3,4,5) pattern.[18]

In a study conducted in the southern part of India, a higher occurrence (55%) of double root PN (C3,4) was observed. They noted triple roots (C3.4.5) in 20% of cases, whereas in 10% of their study sample single root (C4) PN was present.[4]

In this study from North India, a triple root pattern (C3,4,5) was observed in maximum cadavers (>50%). The next common findings were the double roots originating from C3 and C4 at a value of around 30%. We did not find single root PN by C4 as a common observation. As far as single root C5 PN is concerned, we observed only one such case on the left side of a male cadaver. Also described a case report of Iranian origin that exhibited the formation of the right PN by C5 alone.[19]

It is clear from the available literature that PN does not follow any specific pattern that can be assigned to a specific geographical area. Studies show that double rootlets may be present. Mendelssohn observed double roots of C4 (8%) and C5 (5%) in PN with C3,4,5 roots. Verin observed double rootlets of C3 in one case (C3,4) out of three cadavers they observed. In this study, we observed double rootlets of C5 in PN contributed by C4,5 in [Image 1.1] and another observation was formation of PN only from C3, C4 as in [Image 1.2]. The morphological significance of such rootlets is still unclear but their presence can be researched for the possibility of being used for neurotization of recurrent laryngeal nerve.[12]

Image 1.1.

Dual contribution from C5 nerve root

Image 1.2.

C3, C4 nerve roots contributing to the formation of PN

Mendelsohn analyzed no difference between side or sex and frequency of pattern in PN.[20] In this study, it was observed that the variation anatomy of PN roots did not exhibit any gender-linked specificity in terms of frequency or side. Secondly, no pattern could be found specific to any side. The occurrence of variation on one side of the neck may not be mirrored as such on the other side. In a single neck, both sides may show similar or different patterns.

Researchers acclaim that PN can receive communicating branches proximally from C1, C2 and distally from C6, C7, C8, and T1.[21] It may also receive contributions from the XI and XII cranial nerves.[7] Reports are available showing connections of PN with the nerve to subclavius, nerve to sternohyoid, Ansa cervicalis, and brachial plexus.[22] In this study, none of such communication was observed. Such communications are looked at as an opportunity in neurotization surgeries. Individual case reports are observed as rare entities and surgeons must learn to confirm ultrasonographic localization of such additional sources before planning restoration procedures.

In the year 1970, Kikuchi defined PN in relation to the subclavian vein.[21] According to him, a nerve derived from cervical roots, descending in front of scalenus anterior, if passes posterior to the SV must be termed as PN and any additional nerve passing in front of SV must be designated as APN. The possibility of PN penetrating the SV has also been mentioned (Last, 1984; Matsumura and Kumaki, 1996). Last (1984) explained that this could occur due to the formation of SV by a coalescence of a rich venous plexus in the embryo. In this study all PN were passing behind the SV at the root of the neck and without exception were bracing the scalenus anterior insertion from its medial side.

In this study, the APN was found in 14% of total neck sides. All nerves contributing to the PN after crossing the anterior scalene were considered to be APNs. Loukas et al. (2016),[22] observed the origin of APN from nerve to subclavius (60.6%), ansa cervicalis (12.1%), and nerve to sternohyoid (7%). Graves et al. (2017), displayed the presence of APN in 36.5% of hemi-necks and around 16.5% arising from ansa cervicalis.[23] They suggested that incidence varies between populations, and within similar geographical areas variations may be attributed to ethnic differences. However, the literature states that APNs may potentially arise from around 11 sites and are acclaimed to have a heterogeneous anatomic organization. The different course of the APN and its connection to other structures of the neck are causally related to its different origins. The different levels of diaphragmatic dysfunction seen in patients with APN lesions could possibly be due to the heterogeneity of origin.[22]

Literature reports that in embalmed cadavers, the morphometry of structures is altered. In this study, we used 12 (57.14%) embalmed and 9 (42.85%) unembalmed cadavers. However, we did not find any significant difference in the mean length or thickness of PN in embalmed or unembalmed cadavers. Mendelsohn et al. (2011)[20] demonstrated a significant difference in the length of PN rootlets in two types of cadavers. This difference in the observation could be attributed to the duration of preservation. Our preserved cadavers were relatively fresh. We assume that the duration of preservation affects the hardness of tissues and thus more the duration, the more will be tissue shrinkage [Table 3].

Table 3.

Various patterns of origin of PN

Author	Geographical area/sample size	Triple-root PN			Double-root PN		Single-root PN			Accessory	Others
		C3,4,5	C3,4++,5	C3,4,5++	C3,4	C4,5	C3	C4	C5	APN
Verin		–	–	–	–	–	–	–	–	–	C3++,4
Abie H. Mendelsohn 2011	North America 111	22%	8%	5%	26%	21%	3%	10%	0%	6%	–
Andrade	South India 20	20%	–	–	7.5%	55%	2.5%	10%	2.5%	–	2.5%
Banneseka	Japan 208	1.5%			2.5%	52%	0%	43.5%	0.5%	–	–
Ahmad Pour	Iran 2016 case report	–	–	–	–	–	–	–	Single C5	–	–
Present study	North India 42 hemi necks	54.76%	–	–	4.76%	30.95%	0%	4.76%	2.38%	7.14%	C4,5++ (2.38%)

Thus, this study highlights the need for a comprehensive understanding of the variations of the PN in the cervical region.

CONCLUSION

This study was carried out to explore the morphology and course of PNs in the cervical region of human cadavers. For this purpose, an observational study was conducted involving a total of 21 cadavers (2 × 21 sides of the neck) during the study period. Three types of origin of PN were observed, classical three root (C3,4,5); double root with varying contribution from C3 or C5 (C3,4/C4,5), and single root (C4/C5). Classical triple root PN formation was present in 57.68% of cases and was the most common pattern in our study population. The second most common (30.95%) formation was observed for the double root pattern (C4,5) by contribution of distal cervical segment (C5). Double root configuration of C3,4 was observed on 4.76% of sides. Out of single root contribution incidence of C4 pattern was higher (4.76%) as compared to C5 (2,14%). Variation in the root pattern was not dominated or linked to gender or the side of the neck. All PNs irrespective of their side or gender exhibited a constant formation site at the lateral border of the scalenus anterior muscle, descended obliquely across its anterior surface to the root of the neck. In only 7.14% of cases, APN was found. The mean length of PN in embalmed male cadavers was 7.68 cm on the right and 7.68 on the left side. The mean length and thickness of PN were almost similar in all cases. One cadaver exhibited a dual C5 root with C4 on the right side in a male cadaver. Spinal cord injuries in the cervical spine 3,4,5 (cervical tetraplegia) are associated with high morbidity and mortality, and artificial ventilation in these patients can lead to long-term infections, atelectasis, and respiratory failure. Because of its function in respiration, the PN is one of the most important nerves in the body. The diaphragm, the most important respiratory muscle, receives its motor supply mainly from the PN. There is one left and one right PN each.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.