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Guidelines for Botulinum Neurotoxin Injection for Facial Contouring

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DOI: 10.1097/PRS.0000000000009444

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 SPECIAL TOPIC
Guidelines for Botulinum Neurotoxin Injection
for Facial Contouring
Kyu-Ho Yi, M.D.
Hyung-Jin Lee, Ph.D.
Hye-Won Hur, Ph.D.
Kyle K. Seo, M.D., Ph.D.
Hee-Jin Kim, D.D.S., Ph.D.
Seoul, Republic of Korea
Summary: The hypertrophied temporalis and masseter muscles give a muscular
shaped and bulky contour to the face. Botulinum neurotoxin injection methods
are commonly used for facial contouring; however, adverse effects have been
reported owing to a lack of delicate anatomical information. The anatomical con-
siderations when injecting botulinum neurotoxin into the temporalis and masseter
muscles were reviewed in the present study. Current knowledge on the localization
of the botulinum neurotoxin injection point with more recent anatomical dissec-
tion and modified Sihler staining procedures was assessed. The authors found that,
for the muscles, the injection point can be more precisely demarcated. Optimal
injection sites are presented for the temporalis and masseter muscles, and the
injection technique is suggested. The authors propose the optimal injection sites
in relation to external anatomical landmarks for the frequently injected muscles of
the face to facilitate the efficiency of botulinum neurotoxin injections. In addition,
these guidelines would aid in more precise practice without the adverse effects of
botulinum neurotoxin. (Plast. Reconstr. Surg. 150: 562e, 2022.)

B
otulinum neurotoxin hinders neural trans-
mission by stimulating the release of ace-
tylcholine at the neural endplate and
inhibiting muscle contraction.1 Botulinum neuro-
toxins are used extensively for aesthetic objectives
and therapeutic options for the control of brux-
ism, headaches, and myofascial pain syndrome.
In aesthetic clinics, botulinum neurotoxin is
used primarily for wrinkle removal by weakening
the facial expression muscles, such as the orbicu-
laris oculi, zygomaticus major, corrugator super-
cilli, and frontalis muscles. In addition, it is used
for facial contouring, which commonly targets the
temporalis and masseter muscles.2–7 The hypertro-
phic temporalis and masseter muscles contribute to
a thick and stocky contour of the face, and muscu-
larly shaped facial lines are of cosmetic concern for
many people (Fig. 1). Facial contouring allows for
a smoother and slimmer face by injection of botuli-
num neurotoxin into the masticatory muscles.
From the Wonju City Public Health Center; Division in
Anatomy and Developmental Biology, Department of Oral
Biology, Human Identification Research Institute, BK21 FOUR
Project, Yonsei University College of Dentistry; Department of
Anatomy, Catholic Institute for Applied Anatomy, College of
Medicine, Catholic University of Korea; Modelo Clinic; and
Department of Materials Science & Engineering, College of
Engineering, Yonsei University.
Received for publication November 4, 2020; accepted
November 9, 2021.
The first two authors contributed equally to this work.
Copyright © 2022 by the American Society of Plastic Surgeons
DOI: 10.1097/PRS.0000000000009444
When injecting botulinum neurotoxin into the
masseter and temporalis muscles, anatomical struc-
tures should be carefully considered to avoid side
effects, such as unwanted paralysis of nearby mus-
cles, asymmetric smile, and paradoxical bulging.8–10
Today, botulinum neurotoxin injection is recog-
nized as the safest and most effective treatment for
muscle inactivation.11–14 The effects of botulinum
neurotoxin rely on uptake by the presynaptic mem-
branes of motor neurons at the neuromuscular
junction; consequently, injections should be admin-
istered into the neuromuscular junction area.1,15,16
The importance of using neuromuscular junction–
targeted botulinum neurotoxin injections has been
confirmed by clinical studies in the iliopsoas and
biceps brachii muscles. Neuromuscular junction–
focused injections result in a much larger volume
reduction than conventional injections.17,18
Overdoses of botulinum neurotoxin may
cause the toxin to disseminate to adjacent
Disclosure: The authors have no financial interest
to declare in relation to the content of this article.
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 Volume 150, Number 3 • Botulinum Neurotoxin in Face Contouring

Fig. 1. Schematic images. (Left) Hypertrophied temporalis and masseter muscles. (Right)
Contoured facial line after botulinum neurotoxin treatment.

muscles, resulting in their paralysis (e.g., dyspha-
sia).19,20 Some patients have reported temporo-
mandibular joint pain caused by weakened muscle
strength because of botulinum neurotoxin over-
dose in the temporalis and masseter muscles.21–23
Consequently, the dose must be adjusted based
on a person’s muscle volume to prevent severe
muscular weakness. Moreover, repetitive facial
contouring with botulinum neurotoxin must be
avoided because it can result in reduced cortical
bone quality and morphological bony changes.24,25
Furthermore, repetitive injections and
increased doses of botulinum neurotoxin may
create antibodies that could result in inadequate
treatment.19,20,26,27 In addition, an earlier study
reported neuropathy due to mechanical injury
following injection in the nerve trunks.28 Thus,
to reduce side effects and increase efficacy, bot-
ulinum neurotoxin would be more potent and
beneficial if injected near or into arborized areas.
Several studies on intramuscular neural arboriza-
tion of muscles have already been published.29–33
The objective of this study was to propose
effective and safe botulinum neurotoxin injection
points and injective methods for the temporalis
and masseter muscles for facial contouring.
ANATOMY OF THE TEMPORALIS
MUSCLE
Botulinum neurotoxin treatment targeting
the temporalis muscle may be used in patients with
a wide upper face, which results in a prominent
contour. The hypertrophied temporalis muscle
is the key reason for botulinum neurotoxin treat-
ment in this region.
The temporalis is a fan-shaped muscle with two
layers that cover the lateral part of the cranium.
The muscle originates from the temporal line and
inserts into the coronoid process. The deep layer
arises from the temporal fossa, and the superficial
layer arises from the deep temporal fascia. The
anterior part of the muscle runs vertically, whereas
the posterior portion runs horizontally. The deep
temporal fascia is a taut membranous structure
that superficially covers the temporalis muscle.
The muscle is fed by anterior and posterior deep
temporal arteries and is innervated by the anterior,
middle, and posterior deep temporal nerves. The
action of the muscle is to close the mouth by elevat-
ing the mandible.
A study discovered that even in subclinical
doses, botulinum neurotoxin easily passes through
muscle fascia at a rate of 77 percent.34 Once botu-
linum neurotoxin has been injected into these fas-
cial layers, it may diffuse down toward the fascial
layer, resulting in unwanted paralysis and no effect
on muscle atrophy. Consequently, understanding
the fascial layers of the muscles is important when
treating with botulinum neurotoxin (Fig. 2).35
In addition, the thickness of the temporalis
muscle should be considered in deciding the dose
of botulinum neurotoxin.
According to Choi et al.,36 the temporalis ten-
don is fan-shaped, with the most distant point of
the tendon located 45 mm from the zygomatic

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Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
 Plastic and Reconstructive Surgery • September 2022
Fig. 2. A schematic image showing the temporalis muscle cov-
ered by two fasciae: the superficial temporal fascia (STF) and
deep temporal fascia (DTF).
arch. They have recommended that the site of
botulinum neurotoxin injection into the tempo-
ralis muscle should be at least 45 mm from the
zygomatic arch. Sihler staining conducted by
Choi et al.,36 illustrates that the nerve endings are
tightly and evenly dispersed in the upper regions
of the temporalis muscle (Fig. 3).
INJECTION TECHNIQUES
Regarding the anatomical aspect, botulinum
neurotoxin needs to be administered at least
45 mm above the zygomatic arch to avoid injection
into the tendon. In addition, after the needle pen-
etrates to the taut resisting superficial and deep
Fig. 3. Sihler staining of the temporalis muscle demonstrates tightly and
evenly dispersed nerve endings in the upper regions.
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Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
temporal fascia, a retrograde injection should
be administered after the syringe touches the
bone surface to avoid the botulinum neurotoxin
spreading to the fascial layer. This will confirm
that the botulinum neurotoxin is in the neural
arborized area of the muscle belly, providing the
best clinical effect with minimum concentration.
To identify the temporalis muscle belly, an imagi-
nary transverse line passing the superior rim of
the orbit is set up so that we can identify the tem-
poralis muscle belly above the line (Fig. 4). Before
injection, while palpating the temporal area and
clenching their teeth, clinicians can recognize the
hypertrophied muscle bundle (Fig. 5, left).
According to Choi et al.,36 botulinum neuro-
toxin should be injected mainly into the anterior
part of the muscle by setting up an imaginary ver-
tical line passing the tragus. The posterior part
of the temporalis muscle is covered with hair,
which makes it difficult to identify the temporalis
bulging. In addition, injection of botulinum neu-
rotoxin right behind the orbital rim should be
avoided since temporal hollowing may appear,
which exaggerates the prominent zygoma (Fig. 5,
right).37
Previous studies administered botulinum
neurotoxin injections to the temporalis muscle,
with dosages ranging from 20 to 50 U and one
to five injection sites.5,38,39 Singh et al.40 reported
that the thickness of hypertrophied temporalis
was 0.79 cm on average. Botulinum neurotoxins
typically spread up to 2 to 4 cm from the injec-
tion site.15 We recommend injections of 5 U
per 0.2-ml aliquots at four to five distinct sites
according to the anatomical features of the mus-
cle (Fig. 4). When injecting five distinct sites,
since the muscle belly is thicker in the inferior
portion, three injections should be administered
inferiorly.
 Volume 150, Number 3 • Botulinum Neurotoxin in Face Contouring
Fig. 4. Guidance for injecting botulinum neurotoxin into the
temporalis muscle. It is recommended that botulinum neuro-
toxin be injected at four to five injection points over the line of
the eyebrow, with the anterior portion of the vertical line pass-
ing the tragus and 2 cm away from the orbital rim.
ANATOMY OF THE MASSETER MUSCLE
The masseter muscle starts from the zygomatic
arch, running inferiorly to posteriorly, and ends
on the ramus of the mandible and mandibular
Fig. 5. Patient images of the hypertrophied masseter muscles (left) and temporal hollowing (right).
Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
angle. It is known to have three layers: the super-
ficial, middle, and deep layers (Fig. 6). The super-
ficial layer originates from the zygomatic process
of the maxilla, runs inferiorly to posteriorly, and
inserts into the mandibular angle. The middle
layer originates from the lower border of the pos-
terior one-third of the zygomatic arch and the
internal surface of the anterior two-thirds of the
arch; it runs perpendicularly and ends on the lat-
eral surface of the ramus. The deep layer origi-
nates from the internal surface of the zygomatic
arch and inserts into the ramus of the mandible.
The middle and deep layers are separated by the
masseteric branch of the mandibular nerve. Seo
et al. analyzed the thickness of the masseter mus-
cle using computed tomography and reported it
to be 14.9 ± 2.2 mm, on average.5 The inferior por-
tion of the masseter muscle where the superficial,
middle, and deep layers merge is the thickest.
Botulinum neurotoxin injection into the mas-
seter muscle by unskilled providers can cause
facial paralysis. The most frequently reported
change is inability to lift the corner of the mouth
while smiling (Fig. 7). Reports suggest that an
asymmetric smile after masseteric botulinum neu-
rotoxin injection is due to overspread into the
risorius muscle.8
The study by Bae et al.8 divided the muscle
into six different parts by dividing the anterior,
middle, and posterior parts of the muscle into
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 Plastic and Reconstructive Surgery • September 2022
Fig. 6. Illustration of the coronal section of the masseter muscle.
S, superficial layer of the masseter; M, middle layer of the mas-
seter; D, deep layer of the masseter; DIT, deep inferior tendon;
SaM, superficial aponeurosis.
upper and lower. They reported that 53.3 per-
cent of the population had risorius superficially
covering the anterior region (III and VI) of the
Fig. 7. Side effect of asymmetrical smiling after botulinum neurotoxin injection into the masseter muscle. Patient with asymmetri-
cal smile (left) and schematic image (right).
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Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
masseter muscles (Fig. 8). This finding suggests
that should the anterior part of the masseter mus-
cle be deeply injected, botulinum neurotoxin may
affect the risorius muscle, possibly causing iatro-
genic abnormal facial expressions.
In addition, Lee et al.10 discovered a deep
tendinous structure, which was a reason for the
paradoxical masseter bulging (Fig. 9). Previous
articles reported incidences of masseteric bulging
observed in about 0.49 to 18.8 percent and occur-
ring 2 to 4 weeks after a botulinum neurotoxin
injection.41,42 This is caused by restricted and local-
ized dispersion of botulinum neurotoxin partially
into either the superficial or deep muscle belly.
These superficial and deep muscle bellies are
divided by the deep inferior tendon. If the botu-
linum neurotoxin spreads to only one of the bel-
lies, the other belly that is not affected appears
prominently bulged while clenching. The tendon
structure was located deep in the lower third of
the superficial part of the masseter. In cadaveric
and ultrasonographic studies, most of the subjects
had this anatomical structure designated as a deep
inferior tendon (Figs. 6 and 10). As this structure
may evenly block diffusion over the muscle belly,
it is recommended to perform layer-by-layer retro-
grade injections into the superficial, middle, and
deep layers of the masseter.
 Volume 150, Number 3 • Botulinum Neurotoxin in Face Contouring
Fig. 8. The relationship between the risorius and masseter mus-
cles. The areas III and IV are superficially covered by the risorius
muscle.
The intramuscular neural distribution of the mas-
seter muscle was observed by Sihler staining method
(Fig. 11).43 The neural arborization of the masseteric
nerve branches is mostly located in the middle to
lower part of the masseter. The middle lower part
Fig. 9. A patient (left) and schematic view (right) of the paradoxical masseteric bulging after botulinum neurotoxin injection.
Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
of the masseter is recommended as the safest and
most effective botulinum neurotoxin injection site
for masseter muscle hypertrophy. The upper part
of the muscle is inappropriate for injection because
the parotid duct passes through and because of the
existence of the superficial aponeurosis (Figs. 6 and
12).44,45 In addition, the posterior part overlaps with
the parotid gland, which should be avoided.
INJECTION TECHNIQUES
Botulinum neurotoxin should be injected into
two to three separate points in the lower half of
the muscle. If it is injected into the upper part of
the masseter muscle, damage to the parotid duct
and superficial aponeurosis will occur. In addi-
tion, neural arborization is not well distributed
in the upper part of the masseter, which has less
effect than the lower part.
When injecting into the anterior part of the
masseter muscle, superficial injection should be
avoided to prevent botulinum neurotoxin diffu-
sion to the risorius muscle, which may cause asym-
metric smiling.8,46,47 In addition, the facial artery
runs immediately anterior to the masseter muscle;
therefore, the anterior portion of the masseter
muscle should be avoided. The line connecting
the mouth angle and the tragus where the supe-
rior border meets the lower mandibular border is
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 Plastic and Reconstructive Surgery • September 2022

Fig. 10. A schematic view (left) and dissected image (right) of the deep interior tendon after
revealing the superficial muscle belly of the masseter muscle.

known as the inferior border. The muscle should
be divided into upper and lower halves as well as
the anterior, middle, and posterior parts of the
masseter muscle (Fig. 8).
In a clinical study by Seo,5 the volume reduc-
tion of the masseter muscle after botulinum neu-
rotoxin injection in comparison with the initial
volume was 29.4 percent in the 40-U group, 26.8
percent in the 30-U group, and 27.1 percent in
the 20-U group, with no significant differences.
Therefore, 20 U of botulinum neurotoxin is the
minimal dose with effectiveness. Each 5 U should
be injected into four injection points in the lower
anterior and lower middle parts of the masseter
muscle (Fig. 13).5 Deep intramuscular injection
should be performed with the needle positioned
vertically over the skin; the needle should be
inserted until the needle tip touches the perios-
teum. A superficial injection of botulinum neu-
rotoxin should be avoided because of the risk of
superficial spread to the risorius muscle located
anterior to the masseter muscle.7 In addition, the
injection should be performed in a retrograde
manner and layer by layer into the superficial,
middle, and deep layers, as deep inferior tendons
might cause paradoxical masseteric bulging.

Fig. 11. The modified Sihler method conducted in the masseter
muscle. Area V (blue) is the most arborized area for the botuli-
num neurotoxin injection site. Areas III and VI (green) should be
avoided because they overlap with the risorius muscle. Area VI
should also be avoided as the facial artery passes immediately
anterior to area VI. Areas I and II (red) should not be injected to
avoid damage to the masseteric nerve trunk and parotid duct
and gland. Area IV (brown) should be avoided as the parotid
gland overlaps the area.
DISCUSSION
The temporalis and masseter muscles are mas-
ticatory muscles responsible for closing the jaw.
Therefore, hypertrophied temporalis and masseter
muscles are linked directly to chewing habits and
genetic factors that contribute to the disfiguring
condition of the widened face.48–51 These muscles
develop depending on the degree of mastication
and bruxism, which are causes of muscular hyper-
trophy. Today, facial contouring techniques using

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 Volume 150, Number 3 • Botulinum Neurotoxin in Face Contouring

Fig. 12. The tendinous digitation of the superficial part of the masseter muscle. The numbers and
lengths of the tendinous digitation were higher in male (left) than in female subjects (right). Male
subjects had three tendinous digitations, whereas female subjects had two tendinous digitations.

botulinum neurotoxin that reduce muscle volume
in masseter hypertrophy are important aesthetic
procedures. Smooth square jaws are generally
known to be attractive. Considering that Asians have
a prominent malar and mandibular angle, man-
dibular angle resection surgery has been specifically
developed in Asian countries. The aesthetic effects
of botulinum neurotoxin injection in facial contour-
ing start to appear within 2 weeks and move toward
maximum reduction after 3 months, however.5
Mostly, botulinum neurotoxin acts on the
neuromuscular junction. Therefore, broad and
precise anatomical knowledge of the neuro-
muscular arborization patterns of the muscles is
essential for attaining maximum effect with the
lowest possible amount of botulinum neurotoxin.
Even though botulinum neurotoxin proce-
dures are minimally invasive compared to surgi-
cal procedures, there is still a risk of damaging the
nerve trunks. Therefore, a precise knowledge of the

Fig. 13. Botulinum neurotoxin injection points for masseter muscles. The injection should be con-
ducted in region V with two (left) to three (right) injections. The injection points should be 1 cm apart.

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Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
 Plastic and Reconstructive Surgery • September 2022
anatomy of these muscles is essential. Sihler staining
is a whole-mount staining method that dyes myelin
sheaths, providing effective tracking of nerve end-
ings without damaging the nerves.30–33,52 The applica-
tion of Sihler staining to the temporalis and masseter
muscles will facilitate the accurate and comprehen-
sive understanding of the neural distribution.
Currently, there is no standardized injection
point or optimal dose for botulinum neurotoxin
treatment of the temporalis and masseter muscles.
We have reviewed the strategies of previous stud-
ies and divided them into two distinctive methods.
One is injection into a fixed site, and the other
is the palpation of the bulged area. Even with a
fixed-site injection, the location of the injection
point varies in each study.5
As facial contouring via botulinum neurotoxin
is frequently conducted, side effects, such as paraly-
sis of adjacent muscles, asymmetric smile, and para-
doxical bulging, have been reported. In addition,
the amount of botulinum neurotoxin should be
adequate to allow sufficient toxin levels in the arbo-
rized area of neural distribution. These complica-
tions suggest that botulinum neurotoxin should be
delivered in the arborized area of the target muscle
in small doses and with fewer injections.
SUMMARY
The temporal muscle injection should be con-
ducted superior to the transverse line passing the
superior rim of the orbit and anterior to the vertical
line passing the tragus. We recommend injections
of 5 U at five distinct sites according to the ana-
tomical features of the muscle (Fig. 4). In masseter
muscle, botulinum neurotoxin should be injected
into three separate points with 8 U each in the
lower half of the masseter muscle, with the injec-
tion performed in a retrograde manner and layer
by layer (Fig. 4). This study performed an exten-
sive analysis of published research on the anatomy
of the temporalis and masseter muscles to provide
anatomical guidelines for facial contouring.
Hee-Jin Kim, D.D.S., Ph.D.
Division in Anatomy and Developmental Biology
Department of Oral Biology
Yonsei University College of Dentistry
50-1 Yonsei-ro, Seodaemun-gu
Seoul 03722, Republic of Korea
hjk776@yuhs.ac
ACKNOWLEDGMENTS
This work was supported by the Korea Medical Device
Development Fund grant funded by the Korean government
(the Ministry of Science and ICT, the Ministry of Trade,
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Copyright © 2022 American Society of Plastic Surgeons. Unauthorized reproduction of this article is prohibited.
Industry and Energy, the Ministry of Health and Welfare,
and the Ministry of Food and Drug Safety; Project Number:
1711138194, KMDF_PR_20200901_0109-01). This
study was conducted in compliance with the principles set
forth in the Declaration of Helsinki. Consent was received
from the families of the deceased patients before beginning
the dissections. The authors sincerely thank those who
donated their bodies to science so that anatomical research
could be performed. Results from such research can poten-
tially increase mankind’s overall knowledge, which can then
improve patient care. Therefore, these donors and their fami-
lies deserve our highest gratitude. The authors thank Eun-
Byul Yi from Eonbuk Elementary School for illustrations.
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