Journal of Plastic, Reconstructive & Aesthetic Surgery (2014) 67, 1663e1668

The topographic anatomy of the massetericnerve: A cadaveric study with an emphasison the effective zone of botulinum toxinA injections in masseter

B. Kaya a, N. Apaydin b,*, M. Loukas c, R.S. Tubbs d

a Department of Plastic and Reconstructive Surgery, Ankara University Faculty of Medicine,Ankara, Turkeyb Department of Anatomy, Ankara University Faculty of Medicine, Ankara, Turkeyc Department of Anatomical Sciences, St. George’s University, St. George’s, Grenadad Pediatric Neurosurgery Children’s Hospital, Birmingham, AL, USA

Received 8 November 2013; accepted 29 July 2014

KEYWORDSMasseter muscle;Masseteric nerve;Motor nerve entrypoint;Masseterichypertrophy;Botulinum toxin;Facial reanimation

* Corresponding author. Ankara Univ5958248, þ90 312 3103010x246; fax: þ

E-mail address: napaydin@gmail.c

http://dx.doi.org/10.1016/j.bjps.2014.01748-6815/ª 2014 British Association of P

Summary Introduction: Botulinum toxin injections are previously reported to be a noninva-sive alternative method for treating masseteric hypertrophy. However, there is a debate onfinding an ideal place for injection. The aim of this study is to document the anatomical land-marks for defining the motor nerve entry points (MNEPs) of the masseteric nerve in themasseter for effective botulinum toxin injections.Materials and methods: Twelve sides from six adult fixed cadavers were used for this study.The MNEPs of the masseteric nerve were defined according to standard landmark linesincluding the orbitomeatal line (OML) and the line (VL), which intersects the mid-distanceof the OML to the tip of the angle of the mandible.Results: All MNEPs were located 4.4 cm inferior to the OML. In addition, the average anteriordistance of the MNEPs to the VL was 1.4 cm and the average posterior distance was 0.6 cm.Conclusion: The ideal site of Botox injection into the masseter is a rectangular area: 5 cm infe-rior to the OML, 1 cm anterior and posterior to the VL, and just above the periosteum. Based onthe data of our study, injections to the parotid gland and branches of the facial nerve such asthe marginal mandibular and buccal can be avoided. The masseteric nerve can easily be found

ersity Faculty of Medicine, Department of Anatomy, 06100 Sihhiye, Ankara, Turkey. Tel.: þ90 31290 312 3105001.

om (N.Apaydin).

7.043lastic, Reconstructive and Aesthetic Surgeons. Published by Elsevier Ltd. All rights reserved.

1664 B. Kaya et al.

approximately 1.0e1.5 cm inferior to the zygomatic arch, 1 cm medial to the temporomandib-ular joint capsule, and 1 cm superior to mandibular notch, which makes its use for facial re-animations more efficient.ª 2014 British Association of Plastic, Reconstructive and Aesthetic Surgeons. Published byElsevier Ltd. All rights reserved.

Introduction

The masseter is one of the four principal muscles ofmastication which covers the angle of the mandible andmandibular ramus. Its main function is elevation of themandible and it has a small effect on side-to-side move-ments.1 It also has an important morphological role indelineating the lower facial contour, which is aestheticallyimportant.2 Masseteric hypertrophy is a rare disorder thatmay present as either a unilateral or bilateral swelling inthe region of the angle and the ramus of the mandible.Patient complaints are usually aesthetic in nature, butthere may also be pain in the region and limitation of mouthopening.3,4

The surgical treatment of this anomaly involves resec-tion of a portion of the masseter muscle.5 However, thereare several complications associated with surgery. Thesecomplications include severe bleeding, hematoma, pro-longed edema, facial nerve injury, trismus, and bad scar-ring. To avoid these complications, noninvasive modalitiessuch as occlusal adjustment, splint therapy, tranquilizers,and antidepressants have been used for treatment.6 Botu-linum toxin type A (BTX-A) injection is an alternative,relatively noninvasive, and effective treatment formasseter muscle hypertrophy.7e9

BTX-A is a potent bacterial neurotoxin produced by ananaerobic bacterium Clostridium botulinum. It causesmuscle paralysis, atrophy, and weakness by chemicaldenervation. In muscle, it blocks the neuromusculartransmission through rapid and strong attachment to thepresynaptic nerve membrane. It inhibits the release ofvesicle-bound acetylcholine at the neuromuscularjunction.10

Previous studies indicate the importance of defining themotor nerve entry points (MNEPs) of a muscle for aneffective, selective, and safe BTX injection.7,11e13 In 2010,Hu et al.6 established the safest injection sites for BTX in-jection into the masseter muscle by a well-designed study.They showed that the safest injection point is the centralcompartment of the muscle so as to avoid injections intothe parotid gland or into the marginal mandibular branch ofthe facial nerve; however, they did not investigate theMNEPs to the muscle.

There are numerous studies targeting the differentmuscles in the body investigating MNEPs13,14; however,the MNEPs of the masseter muscle have not been welldefined according to standard anatomical landmarks. Theaim of this study, therefore, is to document the anatom-ical landmarks for defining the MNEPs of the massetericnerve in the masseter for effective botulinum toxininjections.

Materials and methods

Twelve sides from six adult fixed cadavers were used forthis study. Of these, four were male and two were femaleand ranged in age from 56 to 69 years (mean 63 years) atdeath. None of the cadavers showed any gross pathology orsigns of surgical procedures in the examined area. Eachcadaver was placed prone and the skin and the subcu-taneous tissue over the masseter were cut and removedcompletely. The branches of the facial nerve were carefullypreserved.

Using a surgical microscope, the masseteric nerve wasexposed at the mandibular notch and its branches werefollowed to its entrance into the masseter muscle. Thedorsal part of the inferior attachment of the muscle to themandible was detached in order to observe the entry site ofeach branch.

The masseteric nerve and distribution of its brancheswere determined in reference to the following bonylandmarks

Determination of the origin of the masseteric nervethrough the mandibular notch

The closest distances between the origin of the nervethrough the mandibular notch to the most medial part ofthe temporomandibular joint (TMJ), zygomatic arch, andmandibular notch were measured. The TMJ landmark waslocated on the medial surface of the mandibular condyle. Inaddition, the measurements concerning the mandibularnotch were made on its distal edge.

Determination of the distribution of the branchesof the masseteric nerve

The number of branches of the masseteric nerve at thelevel of the mandibular notch, as well as the MNEPs of eachnerve branch, was recorded. Next, the MNEPs were definedaccording to standard landmark lines. These standard linesincluded the orbitomeatal line (OML) and the line (VL)which intersects the mid-distance of the OML to the tip ofthe angle of the mandible. The OML was classically deter-mined as the line that connects the lateral palpebralcommissure to the tragus. The distance of each motor entrypoint of masseter nerve branches to these lines wasmeasured by taking the OML as X and the VL as Y landmarklines. These landmark lines were expressed in absolutedistances, and based on the Y landmark line, the regionanterior to it was determined as the (þ) Y landmark line

The topographic anatomy of the masseteric nerve 1665

while the region posterior to it was (�) Y landmark line(Figure 1).

All measurements were carried out using digital calipers(Vernier LCD Digital Caliper Measuring, USA). In order tocorrect for individual examiner variability, each specimenwas examined by two members of the team (BK, NA). Eachobserver made single independent measurements and themean of all measurements was recorded.

Results

The masseteric nerve was observed to travel through themandibular notch in all cases. The nerve and its motorbranches coursed just above the periosteum in the majorityof the specimens (eight cases) and it traveled between thesuperficial and deep layers of the masseter in the remainingspecimens (four cases). The branches of the massetericnerve were accompanied by branches of the massetericartery and had no close relation with other nerve branches.The origin of the nerve through the mandibular notch was13.8 � 4.5 mm inferior to the zygomatic arch,10.6 � 2.7 mm medial to the TMJ capsule, and7.8 � 2.0 mm superior to the mandibular notch (Figure 2).

The mean length of the OML was measured as8.4 � 1.8 cm and the VL as 8.6 � 1.5 cm. There were threeto seven branches (mean five) of the masseteric nerveseparately innervating the deep and superficial layers of

Figure 2 Figure showing the determination of the origin ofthe masseteric nerve through the mandibular notch (asterisk).The closest distances between the origin of the nerve throughthe mandibular notch to the most medial part of the tempo-romandibular joint (TMJ), zygomatic arch, and mandibularnotch (notch) were measured.

Figure 1 Figure showing the reference line from the lateralpalpebral commissure to the tragus, namely, the OML and theline (VL) which intersects the mid-distance of the OML to thetip of the angle of mandible. Based on these reference lines,the OML was determined as the X landmark line and the VL asthe Y landmark line. The region anterior to it was determinedas the (þ) Y landmark line while the region posterior to it wasthe (�) Y landmark line.

the masseter. It was also observed in all cases that theupper half of the muscle mass received more fibers than thelower half of the muscle.

The evaluation of the location of the MNEPs showed thatall MNEPs were located inferior to the OML (X landmarkline). The average distance on the X landmark line for allregions of the MNEPs was 4.4 cm (minimum: 3.1 cm,maximum: 7.5 cm). The average distance of the MNEPs onthe (þ) Y landmark line was measured to be 1.4 cm (mini-mum: 1.3 cm, maximum: 2.0 cm) whereas on the (�) Ylandmark line, the average distance was 0.6 cm (minimum:0.2 cm, maximum: 1.1 cm) (Figure 3).

Discussion

BTX-A is commonly used for treating a variety of neuro-muscular disorders such as strabismus, blepharospasm,hemifacial spasm, and torticollis.13,15 It has also been usedas a noninvasive treatment for patients with masseterichypertrophy.8,9 The anatomy of the masseter muscle hasbeen well defined in classical anatomy textbooks1 and inrecent reports.2 However, the innervation patterns and thedistribution of nerve branches into the muscle are not wellinvestigated. Hence, there is still debate in determining theideal site of injection into the masseter muscle.

Figure 3 Figure showing the determination of the distribu-tion of the branches of the masseteric nerve. There are threemotor branches (arrowheads) seen in this figure; all are locatedbelow the X landmark line (orbitomeatal line) and on the (þ) Ylandmark line. The superficial and deep layers of the masseterand the point of nerve origin through the mandibular notch arealso marked on the figure.

1666 B. Kaya et al.

Cotrufo et al.16 have revealed a “masseteric area”where the main trunk of the masseteric nerve can be easilydefined. Thus, they have suggested an optimized techniqueof harvesting it as a donor nerve for facial reanimation. Thebranching pattern of the masseteric nerve has also beeninvestigated by Brenner and Schoeller,17 and it wasrevealed that the masseteric nerve may have variantnumbers of branches after leaving the infratemporal fossa.Although our anatomical landmarks are different, we alsosuggest that the main trunk of the masseteric nerve can befound in a so-called “masseteric area” which is located13.8 � 4.5 mm inferior to the zygomatic arch,10.6 � 2.7 mm medial to the temporomandibular jointcapsule, and 7.8 � 2.0 mm superior to the mandibularnotch.

Previous studies have attempted to define an ideal siteof BTX-A injection to the masseter. Injection into themuscle was first defined by von Lindern et al.7 and the in-jection was suggested to be applied on part of the zygo-matic arch and mandibular angle. However, this methodwas associated with the risk of invading the parotid gland.In a study by Hu et al.,6 the surface of the masseter wascompartmentalized into eight areas and numbered from I toVIII. Areas I, III, V, and VII were assigned to represent the

upper four compartments and areas II, IV, VI, and VIII wereassigned to represent the lower four compartments. Thecenter of compartment VI was suggested to be the safestand most efficient injection site for BTX-A into themasseter muscle to avoid injury to the parotid gland, whichusually occupies compartments I and II, and the marginalmandibular branch of the facial nerve, which was located amean of 7.4 mm superior to the inferior mandibular margin.Although this study provided information about the locationof the nerve branches with respect to the structures thatshould be avoided during injection, the exact locations ofMNEPs were not defined. Additionally, as the patients havemasseteric hypertrophy, the compartmentalization of themuscle mass may not be standardized for each individual.We suggest that defining standard anatomical landmarks,which is not dependent on personal characteristics, ismandatory.

The exact localization of MNEPs of the masseteric nervehas been investigated by Kim et al.18 These authorsdissected twelve masseter muscles and observed theinnervation pattern of masseteric nerve in the superficial,middle, and deep layers of the muscle. They also dividedthe muscle into compartments and investigated the distri-bution of the nerve branches in different compartments.They used the anterior margin of the middle layer of themuscle as a reference, and according to this referencepoint, they divided the masseteric nerve into four groups:posterosuperior, posteroinferior, anterosuperior, andanteroinferior. Although they were able to show theinnervation patterns in each segment and revealed that themasseteric nerve branches were mostly confined to thelower middle third of the whole masseter muscle, they didnot suggest an ideal place for injection of the muscle.

In this study, we defined standard and easily definableanatomical landmarks including the OML and the line (VL),which intersects the mid-distance of the OML to the tip ofthe angle of the mandible. These lines can be used forexact localization of the injection point of BTX-A into themuscle. As these references are individually based, wesuggest that they can be used reliably in any patient. Thesereference lines can be considered as X and Y landmark linesas was defined previously by Lee et al.13 for the sterno-cleidomastoid muscle. Accordingly, our results revealedthat all MNEPs were located 4.4 cm (minimum: 3.1 cm,maximum: 7.5 cm) below the OML (X landmark line). Theaverage distance of the MNEPs on the (þ) Y landmark linewas 1.4 cm (minimum: 1.3 cm, maximum: 2.0 cm) whereason the (�) Y landmark line, it was 0.6 cm (minimum:0.2 cm, maximum: 1.1 cm). These results were addressingthe ideal site of Botox injection into the masseter is anarea, approximately 5 cm inferior to the OML (minimum:3.1 cm, maximum: 7.5 cm), 1 cm anterior and posterior tothe VL (Figure 4). This location can also be considered asthe half distance of the VL. This region was free of theparotid gland, marginal mandibular nerve, and otherbranches of the facial nerve such as its buccal branch.

Once BTX is injected into the muscle, it is known toimmediately diffuse into the muscle within a few centi-meters of the needle tip. When a higher volume is injected,the area of diffusion seems to increase; however, inaccu-rate injection and excessive diffusion of the toxin can leadto systemic adverse effects or unwanted weakness of the

Figure 4 Figure demonstrating the position of the injection.OML: orbitomeatal line; VL: vertical line which intersects themid-distance of the OML to the tip of the angle of mandible,gray square: suggested position of injection.

The topographic anatomy of the masseteric nerve 1667

neighboring muscles.18,19 We suggest that a single injectionto an ideal site would be enough for diffusion of the toxinthrough MNEPs of other branches, and thus systemicadverse effects or unwanted weakness of neighboringmuscles may avoided. Concerning the dosage, there is nostandardized dose of BTX-A for effective treatment ofmasseteric hypertrophy. Although Lee et al.20 have statedthat 30 IU BTX-A injection is effective for masseteric hy-pertrophy, Kim et al.21 had proposed that >30 IU BTX-A maybe required to achieve an adequate result. In addition tothis, Andrade and Deshpande22 have stated they had suc-cessfully managed masseter muscle hypertrophy byinjecting 40 IU of BTX-A.

The branches of the masseteric nerve are increasinglybeing used for facial reanimation procedures.17 However, itmay not always be very easy to locate the nerve due to avariety of described branching patterns and variability offacial measurements on which some surgical approachesare based upon. Borschel et al.23 have studied the relationof the masseteric nerve to some surgical landmarks such asthe auricular tragus and the zygomatic arch on eight ca-davers and found the nerve to lie 3 cm anterior to thetragus at a level 1 cm inferior to the zygomatic arch. Collaret al.24 dissected 10 cadavers and identified the massetericnerve in the “subzygomatic triangle” which was formed bythe zygomatic arch superiorly, the TMJ posteriorly, and thefrontal branch of the facial nerve inferiorly and anteriorly.Cheng et al.25 presented a technique to identify the nervein a 1.5 cm2 area defined by constant anatomical landmarkssuch as the zygomatic arch, condyle, coronoid process, andmandibular notch. Our study revealed that the nerve was1.4 cm inferior to the zygomatic arch, which is consistentwith these studies. Therefore, we also suggest that the

masseteric nerve can easily be found approximately1.0e1.5 cm inferior to the zygomatic arch, 1 cm medial tothe TMJ capsule and 1 cm superior to the mandibular notchwhich makes its use for facial reanimations more efficient.

The masseter muscle has significant applications forhead and neck reconstruction and for muscle transpositionflap for facial nerve paralysis.26e28 Therefore, it is impor-tant to know the course and the location of branches of themasseteric nerve in relation to standard anatomical land-marks not only for BTX injections but also to provide a safeand efficient approach for these applications. Hontanillaand Qui27 had reported a constant course with a distribu-tion pattern that shows a tree-like anatomy in six cadavers.Hwang et al.28 examined the course of the masseteric nervein relation to the masseteric muscle and found that thenerve ran anteriorly and inferiorly between the deep andthe middle layers. The nerve was observed at 33 � 5.6 mmfrom the inferior border of the muscle on the anterior thirdvertical line of the masseter muscle and at 47 � 5.5 mm inthe posterior third. The results of our study also revealedthat the nerve was coursing between the superficial anddeep layers of the muscle. However, we found its motorbranches to distribute widely among muscle fibers and theupper half of the muscle mass received more fibers than thelower half of the muscle.

The limitation of this study is the fact that it is anexperimental study conducted on cadavers. All cadaverstudies have the limitation that it is difficult to reproduce.However, a clinical study would need to be undertaken toproperly determine the effects of this procedure.

Conclusion

According to the results of our study, the ideal site of Botoxinjection into the masseter is a rectangular area, 5 cminferior to the OML, 1 cm anterior and posterior to the VL,and just above the periosteum. These landmarks will avoidinjection into the parotid gland, marginal mandibularnerve, and other branches of the facial nerve such as itsbuccal branch. We also suggest that the masseteric nervecan easily be found approximately 1.0e1.5 cm inferior tothe zygomatic arch, 1 cm medial to the TMJ capsule and1 cm superior to mandibular notch which makes its use forfacial reanimations more efficient.

Conflict of interest

The authors declare that there is no conflict of interest.

References

1. Standring S. Head and neck. In: Standring S, editor. Gray’sanatomy. 40th ed. New York: Elsevier; 2008. p. 538.

2. Lee JY, Kim JN, Yoo JY, et al. Topographic anatomy of themasseter muscle focusing on the tendinous digitation. ClinAnat 2012;25:889e92.

3. Roncevi�c R. Masseter muscle hypertrophy. Aetiology andtherapy. J Maxillofac Surg 1986;14:344e8.

4. Tabrizi R, Ozkan BT, Zare S. Correction of lower facial widenessdue to masseter hypertrophy. J Craniofac Surg 2010;21:1096e7.

1668 B. Kaya et al.

5. Kim SK, Han JJ, Kim JT. Classification and treatment ofprominent mandibular angle. Aesthet Plast Surg 2001;25:382e7.

6. Hu KS, Kim ST, Hur MS, et al. Topography of the massetermuscle in relation to treatment with botulinum toxin type A.Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010;110:167e71.

7. von Lindern JJ, Niederhagen B, Apple T. Type A botulinumtoxin for the treatment of hypertrophy of the masseter andtemporal muscles: an alternative treatment. Plast ReconstrSurg 2001;107:327e32.

8. Castro WH, Gomez RS, Da Silva Oliveira J, Moura MD,Gomez RS. Botulinum toxin type A in the management ofmasseter muscle hypertrophy. J Oral Maxillofac Surg 2005;63:20e4.

9. Choe SW, Cho WI, Lee CK, Seo SJ. Effects of botulinum toxintype A on contouring of the lower face. Dermatol Surg 2005;31:502e7.

10. Schantz EJ, Johnson EA. Properties and use of botulinum toxinand other microbial neurotoxins in medicine. Microbiol Rev1992;56:80e99.

11. Collins A, Jankovic J. Botulinum toxin injection for congenitalmuscular torticollis presenting in children and adults.Neurology 2006;67:1083e5.

12. Durazzo MD, Furlan JC, Teixeira GV, et al. Anatomic landmarksfor localization of the spinal accessory nerve. Clin Anat 2009;22:471e5.

13. Lee JH, Lee BN, Han SH, An XC, Chung RH. The effective zoneof botulinum toxin A injections in the sternocleidomastoidmuscle. Surg Radiol Anat 2011;33:185e90.

14. Apaydin N, Loukas M, Kendir S, et al. The precise localizationof distal motor branches of the tibial nerve in the deep pos-terior compartment of the leg. Surg Radiol Anat 2008;30:291e5.

15. Traba Lopez A, Esteban A. Botulinum toxin in motor disorders:practical considerations with emphasis on interventionalneurophysiology. Neurophysiol Clin 2001;31:220e9.

16. Cotrufo S, Hart A, Payne AP, Sjogren A, Lorenzo A, Morley S.Topographic anatomy of the nerve to masseter: an anatomical

and clinical study. J Plast Reconstr Aesthet Surg 2011;64:1424e9.

17. Brenner E, Schoeller T. Masseteric nerve: a possible donor forfacial nerve anastomosis? Clin Anat 1998;11:396e400.

18. Kim DH, Hong HS, Won SY, et al. Intramuscular nerve distri-bution of the masseter muscle as a basis for botulinum toxininjection. J Craniofac Surg 2010;21:588e91.

19. Hsu TS, Dover JS, Arndt KA. Effect of volume and concentrationon the diffusion of botulinum exotoxin A. Arch Dermatol 2004;140:1351e4.

20. Lee DH, Jin SP, Cho S, et al. RimabotulinumtoxinB versusonabotulinumtoxinA in the treatment of masseter hypertro-phy: a 24-week double-blind randomized split-face study.Dermatology 2013;226:227e32.

21. Kim HJ, Yum KW, Lee SS, Heo MS, Seo K. Effects of botulinumtoxin type A on bilateral masseteric hypertrophy evaluatedwith computed tomographic measurement. Dermatol Surg2003;29:484e9.

22. Andrade NN, Deshpande GS. Use of botulinum toxin (botox) inthe management of masseter muscle hypertrophy: a simplifiedtechnique. Plast Reconstr Surg 2011;128:24ee6e.

23. Borschel GH, Kawamura DH, Kasukurthi R, Hunter DA, Zuker RM,Woo AS. The motor nerve to the masseter muscle: an anatomicand histomorphometric study to facilitate its use in facialreanimation. J Plast Reconstr Aesthet Surg 2012;65:363e6.

24. Collar RM, Byrne PJ, Boahene KD. Plast Reconstr Surg 2013;132:183e8. http://dx.doi.org/10.1097/PRS.0b013e318290f6dc.

25. Cheng A, Audolfsson T, Rodriguez-Lorenzo A, Wong C, Rozen S.J Plast Reconstr Aesthet Surg 2013;66:1438e40. http://dx.doi.org/10.1016/j.bjps.2013.04.009.

26. Manktelow RT, Tomat LR, Zuker RM, Chang M. Smile recon-struction in adults with free muscle transfer innervated by themasseter motor nerve: effectiveness and cerebral adaptation.Plast Reconstr Surg 2006;118:885e99.

27. Hontanilla B, Qiu SS. Transposition of the hemimassetericmuscle for dynamic rehabilitation of facial paralysis. J Cra-niofac Surg 2012;23:203e5.

28. Hwang K, Kim YJ, Chung IH, Song YB. Course of the massetericnerve in masseter muscle. J Craniofac Surg 2005;16:197e200.