Age-related changes in rat genioglossus, geniohyoid and masseter muscles.

OBJECTIVES: The aim of this study was to elucidate age-related changes from adult to middle age in the contractile properties of the masseter, genioglossus and geniohyoid muscles of the rat. MATERIALS AND METHODS: We analysed the expressions of myosin heavy chain (MyHC) mRNAs and proteins as indicators of the contractile properties in these muscles obtained from rats at 6, 12, 18 and 24 months of age using real-time PCR and SDS-PAGE. RESULTS: We found no marked age-related changes in the expressions of MyHC mRNAs and proteins in rat masseter and geniohyoid muscles, suggesting that the biological ageing process does not affect contractile properties in these muscles. However, we found a decrease in the expression of MyHC IIb mRNA with ageing in the rat genioglossus muscle, suggesting that biological ageing process induces at least some fast-to-slow myofibre phenotype transition. CONCLUSION: The biological ageing process from adult to middle age appears to differentially affect different types of craniofacial muscles.

Remarkable heterogeneity in myosin heavy-chain composition of the human young masseter compared with young biceps brachii.

Adult human jaw muscles differ from limb and trunk muscles in enzyme-histochemical fibre type composition. Recently, we showed that the human masseter and biceps differ in fibre type pattern already at childhood. The present study explored the myosin heavy-chain (MyHC) expression in the young masseter and biceps muscles by means of gel electrophoresis (GE) and immuno-histochemical (IHC) techniques. Plasticity in MyHC expression during life was evaluated by comparing the results with the previously reported data for adult muscles. In young masseter, GE identified MyHC-I, MyHC-IIa MyHC-IIx and small proportions of MyHC-fetal and MyHC-α cardiac. Western blots confirmed the presence of MyHC-I, MyHC-IIa and MyHC-IIx. IHC revealed in the masseter six isomyosins, MyHC-I, MyHC-IIa, MyHC-IIx, MyHC-fetal, MyHC α-cardiac and a previously not reported isoform, termed MyHC-IIx'. The majority of the masseter fibres co-expressed two to four isoforms. In the young biceps, both GE and IHC identified MyHC-I, MyHC-IIa and MyHC-IIx. MyHC-I predominated in both muscles. Young masseter showed more slow and less-fast and fetal MyHC than the adult and elderly masseter. These results provide evidence that the young masseter muscle is unique in MyHC composition, expressing MyHC-α cardiac and MyHC-fetal isoforms as well as hitherto unrecognized potential spliced isoforms of MyHC-fetal and MyHC-IIx. Differences in masseter MyHC expression between young adult and elderly suggest a shift from childhood to adulthood towards more fast contractile properties. Differences between masseter and biceps are proposed to reflect diverse evolutionary and developmental origins and confirm that the masseter and biceps present separate allotypes of muscle.

Human temporomandibular joint and myofascial pain biochemical profiles: a case-control study.

Neurobiological mechanisms of human musculoskeletal pain are poorly understood. This case-control study tested the hypothesis that biomarkers within temporomandibular muscle and joint disorders (TMJD) subjects' masseter muscles or temporomandibular joint (TMJ) synovial fluid correlate with plasma biomarker concentrations. Fifty subjects were recruited and categorized into TMJD cases (n=23) and pain-free controls (n=27) at the University of Minnesota School of Dentistry. Prior to specimen collection, pain intensity and pressure pain threshold masseter muscles and the TMJs were assessed. We collected venous blood; biopsied masseter muscle; and sampled TMJ synovial fluid on the subjects' side of maximum pain intensity. We assayed these tissues for the presence of nerve growth factor (NGF), bradykinin (BK), leukotreine B(4) (LTB(4) ) and prostaglandin E(2) (PGE(2) ), F(2) -isoprostane (F(2) I) and substance P (SP). The data was analyzed using Spearman Correlation Coefficients. We found that only plasma concentrations of bradykinin statistically correlated with synovial fluid concentrations (ρ=-0·48, P=0·005), but no association was found between pain intensities. The data suggests that biomarkers used to assess TMJD need to be acquired in a site-specific manner. We also discovered that F(2) I concentrations were associated with muscle pain intensity and muscle pressure pain threshold (PTT) (ß=0·4, 95%CI: 0·03-0·8) and joint PPT (ß=0·4, 95%CI: 0·07-0·8) suggesting that muscle oxidative stress is involved in myofascial pain and that F(2) -I may be a biomarker for myofascial pain.

Masseter myosin heavy chain composition varies with mandibular asymmetry.

Human jaw dysmorphologies are frequent and often affect young patients, resulting in malocclusion of teeth and inappropriate jaw relationships. Treatment is performed by means of orthodontics with orthognathic surgery as required. Mandibular asymmetry is one of the most frequent dysmorphologies, but in many cases, the specific cause is unknown.In healthy patients who were undergoing orthognathic surgery for correction of malocclusion, we tested the hypothesis that masseter muscle phenotype composition, which determines contractile properties, was different between sides in patients with mandibular asymmetry but not in those without mandibular asymmetry. After cephalometric analysis, 50 patients from whom we obtained samples of both right and left masseter muscles were separated into 2 groups: with or without mandibular lateral deviation. Samples were immunostained with myosin-isoform-specific antibodies to identify 4 skeletal muscle fiber types, and their fiber areas and proportions were measured. Two-tailed Wilcoxon test for paired samples was used to compare the 4 fiber-type compositions by means of percent occupancy and mean fiber area on both sides. Patients with mandibular asymmetry were associated with a significant increase of type II fiber occupancy (P = 0.0035) on the same side as the deviation. This finding that masseter muscle phenotype is significantly linked to mandibular asymmetry is of relevance to physiotherapeutic and surgical managements of jaw discrepancies and merits further investigation in the light of its possible role in the etiology of this condition.

Interstitial glutamate concentration is elevated in the masseter muscle of myofascial temporomandibular disorder patients.

AIM: To determine if myofascial temporomandibular disorder (TMD) pain patients have elevated interstitial concentrations of glutamate in the masseter muscle. METHODS: Thirteen patients (3 men, 10 women) diagnosed with myofascial TMD pain and 10 (2 men, 8 women) age-matched healthy controls participated in a single microdialysis session. Microdialysis was performed in the patients in the most painful point of the masseter muscle, while in the healthy subjects a standardized point in the muscle was chosen. Two microdialysis samples were collected over 40-minute epochs. A blood sample was also taken for analysis of plasma glutamate concentration. Numeric rating scale (NRS) scores of pain intensity and unpleasantness, McGill Pain Questionnaire data, pain drawing areas, pressure pain thresholds, pressure pain tolerances, maximum voluntary bite force, and maximum voluntary mouth opening were collected as secondary measurements. RESULTS: The median concentration of glutamate in the masseter muscle of the myofascial TMD pain patients (7.5 ± 2.6 ΜM) was significantly higher (P < .023, Mann-Whitney test) than the concentration in healthy controls (0.5 ± 0.4 ΜM). There were, however, no significant correlations between glutamate concentrations in the masseter muscle and NRS pain scores. Plasma concentrations of glutamate were similar in patients and healthy controls. CONCLUSIONS: The present study demonstrates a marked increase in interstitial glutamate concentration in the masseter muscle of myofascial TMD pain patients. These novel findings suggest that peripheral glutamate could be involved in the pathophysiology of myofascial TMD pain.

Histological features of masticatory muscles after botulinum toxin A injection into the right masseter muscle of dystrophin deficient (mdx-) mice.

OBJECTIVE/BACKGROUND: The most frequently used animal model for human DMD (Duchenne muscular dystrophy) research is the mdx mouse. In both species, characteristic histological changes like inflammation, muscle fiber degeneration and fibrosis are the same, but in contrast to humans, in mdx mice, phases of muscle fiber degeneration are compensated by regeneration processes. AIM: Therefore, the interest of this study was to evaluate histological features in masticatory muscles after BTX-A injection into the right masseter muscle of wild type and dystrophic (mdx) mice, illustrating de- and regeneration processes induced by this substance. MATERIAL AND METHODS: The right masseter muscle of 100 days old healthy and mdx mice were selectively paralyzed by a single intramuscular BTX-A injection. Masseter as well as temporal muscle of injection and non-injection side were carefully dissected 21 days and 42 days after injection, respectively, and fiber diameter, cell nuclei position, necrosis and collagen content were analyzed histomorphologically in order to evaluate de- and regeneration processes in these muscles. Statistical analysis was performed using SigmaStat Software and Mann Whitney U-test (significance level: p < 0.05). RESULTS: At both investigation periods and in both mouse strains fiber diameter was significantly reduced and collagen content was significantly increased in the right injected masseter muscle whereas fiber diameters in mdx mice were much smaller, and these differences were even more apparent at the second investigation period. Necrosis and central located nuclei could generally be found in all mdx mice muscles investigated with an amount of centronucleation exceeding 60%, and a significant increase of necrosis six weeks after injection. In wild type mice central located nuclei could primarily be found in the treated masseter muscle with a portion of 2.7%, and this portion decreased after six weeks, whereas in mdx mice a decrease could also be seen in the non-injected muscles. In contrast, in wild type mice necrosis was not apparent at any time and in all muscles investigated. CONCLUSION: From our results it can be concluded that in mdx mice masticatory muscles de- and regeneration processes were extended, triggered by a selective BTX-A injection, or mdx mice at this age, independently of BTX-A treatment, went through another cycle of de- and regeneration as a characteristic of this disease.

Decreased mandibular cortical bone quality after botulinum toxin injections in masticatory muscles in female adults.

This study aimed to clarify how masticatory muscle atrophy induced by botulinum toxin (BTX) injection affects cortical bone quality of the mandible using 3D modeling technology. A total of 39 young (26.9 ± 6.0 years) and 38 post-menopausal (55.3 ± 6.3 years) females were included. Computed tomography (CT) images were obtained before and after 12 months of treatment. Predictor variables were application of a stabilization splint, and/or two times of BTX injection in the bilateral temporalis and masseter muscles within a six-month interval. Outcome variables were changes in average Hounsfield units (HU) and cortical thickness of region of interest (ROI). 3D mandibular models were reconstructed using CT images, and models were used to calculate average HU and cortical thickness of ROIs, including inferior half of the lateral surface of ascending ramus, coronoid process, and temporomandibular joint condyle. Cortical bone quality at muscle insertion site was influenced by decreased muscle thickness but seemed not to be affected by decreased functional loading. Reduced functional loading seemed to influence cortical bone quality of the condyles. These effects were more remarkable in post-menopausal females. Hence, decreased masticatory muscle thickness may lead to alterations of the mandibular cortical structures, especially in post-menopausal females.

Seasonal variation in glucose and neutral amino acid uptake in the estuarine crab Neohelice granulata.

This study investigates the mechanisms of glucose and amino acid transport in gills and jaw muscle of N. granulata collected from an estuarine natural population. The physicochemical parameters of the estuarine environment and of this crustacean's hemolymph were measured during different seasons of the year. In summer, the lagoon water osmolality increased (5-6 times), and hemolymph osmolality decreased. In fall, water pH increased, whereas hemolymph pH decreased markedly. In all seasons, 2-deoxi glucose (DG) uptake in gills was significantly higher than 3-O methyl-glucose (MG) uptake. Phloretin reduced DG uptake in gills; phloretin and phlorizin did not affect MG uptake in this organ. DG and MG uptake was highest in gills during spring and summer. In jaw muscle, MG uptake in winter and spring was higher than DG uptake. In fall, gill methyl aminoisobutyric acid (MeAIB) uptake increased. In jaw muscle, MeAIB uptake was higher in spring. The observed changes in glucose uptake and in the type of glucose and amino acid transporter used in gills and muscle appear to be strategies used by N. granulata to minimize seasonal oscillations in the environmental parameters of their estuarine habitat.

Kinetic analysis of the slow skeletal myosin MHC-1 isoform from bovine masseter muscle.

Several heavy chain isoforms of class II myosins are found in muscle fibres and show a large variety of different mechanical activities. Fast myosins (myosin heavy chain (MHC)-II-2) contract at higher velocities than slow myosins (MHC-II-1, also known as beta-myosin) and it has been well established that ADP binding to actomyosin is much tighter for MHC-II-1 than for MHC-II-2. Recently, we reported several other differences between MHC-II isoforms 1 and 2 of the rabbit. Isoform II-1 unlike II-2 gave biphasic dissociation of actomyosin by ATP, the ATP-cleavage step was significantly slower for MHC-II-1 and the slow isoforms showed the presence of multiple actomyosin-ADP complexes. These results are in contrast to published data on MHC-II-1 from bovine left ventricle muscle, which was more similar to the fast skeletal isoform. Bovine MHC-II-1 is the predominant isoform expressed in both the ventricular myocardium and slow skeletal muscle fibres such as the masseter and is an important source of reference work for cardiac muscle physiology. This work examines and extends the kinetics of bovine MHC-II-1. We confirm the primary findings from the work on rabbit soleus MHC-II-1. Of significance is that we show that the affinity of ADP for bovine masseter myosin in the absence of actin (represented by the dissociation constant K(D)) is weaker than originally described for bovine cardiac myosin and thus the thermodynamic coupling between ADP and actin binding to myosin is much smaller (K(AD)/K(D) approximately 5 instead of K(AD)/K(D) approximately 50). This may indicate a distinct type of mechanochemical coupling for this group of myosin motors. We also find that the ATP-hydrolysis rate is much slower for bovine MHC-II-1 (19 s(-1)) than reported previously (138 s(-1)). We discuss how this work fits into a broader characterisation of myosin motors from across the myosin family.

Transforming growth factor betas are upregulated in the rat masseter muscle hypertrophied by clenbuterol, a beta2 adrenergic agonist.

1. The regulatory mechanism for the hypertrophy of skeletal muscles induced by clenbuterol is unclear. The purpose of the present study was to determine the extent to which transforming growth factor betas (TGFbetas), fibroblast growth factors (FGFs), hepatocyte growth factor (HGF), and platelet-derived growth factors (PDGFs) are involved in the hypertrophy of rat masseter muscle induced by clenbuterol. 2. We measured the mRNA expression levels for TGFbetas, FGFs, HGF, and PDGFs in rat masseter muscle hypertrophied by oral administration of clenbuterol for 3 weeks and determined correlations between the weight of masseter muscle and mRNA expression levels by regression analysis. We determined immunolocalizations of TGFbetas and their receptors (TGFbetaRs). 3. The mRNA expression levels for TGFbeta1, 2, and 3, and for PDGF-B demonstrated clenbuterol-induced elevations and positive correlations with the weight of masseter muscle. In particular, TGFbeta1, 2, and 3 showed strong positive correlations (correlation coefficients >0.6). The mRNA expression levels for PDGF-A, FGF-1 and 2, and HGF showed no significant differences between the control and clenbuterol groups, and no significant correlations. TGFbeta1, 2, and 3 were principally localized in the connective tissues interspaced among myofibers, and TGFbetaRI and II were localized in the periphery and sarcoplasm of the myofibers. 4. These results suggest that paracrine actions of TGFbeta1, 2, and 3 via TGFbetaRI and II could be involved in the hypertrophy of rat masseter muscle induced by clenbuterol. This is the first study to document the involvement of TGFbetas in the hypertrophy of skeletal muscles induced by clenbuterol.

Change of mRNA amount of myosin heavy chain in masseter muscle after orthognathic surgery of patients with malocclusion.

INTRODUCTION: Surgical correction of malocclusion changes the force to moment ratio of masticatory muscles inserting at the mandible caused by shortening, lengthening and rotation of the bone following osteotomy. During muscle adaptation the expression of mRNA for the myosin heavy chain (MyHC) of type I and type II fibres may be changed. MATERIAL AND METHODS: The adaptation of the masseter muscle was investigated at the mRNA level in 10 patients 6 months after orthognathic surgery in the mandible. The competitive polymerase chain reaction (cPCR) is a suitable method for quantification of MyHC mRNA. For application of this minimal invasive method an amount of 35 mg muscle tissue was sufficient. RESULTS: 6 month postoperatively there was a deficiency of about 87% of MyHC mRNA for fibre type I and II in both groups of patients. The deficiency in patients with mesial position of the mandible was higher but not significant different to patients with distal malocclusion. CONCLUSION: Patients should use the postoperative interval for training their masticatory muscles. This improves the stability of treatment result and prevents relapse.

Myosin heavy chain isoform composition of human masseter muscle from subjects with different mandibular plane angles.

AIM: To investigate the presence of myosin heavy chain isoforms in human masseter muscle and to describe any differences in orthognathic surgery patients with different mandibular plane angles. METHOD: Biopsies were obtained from the anterior border of the superficial masseter muscle in 18 patients undergoing various orthognathic procedures. Myosin heavy chain isoforms were isolated and analysed by SDS-PAGE gel electrophoresis. Steiner's mandibular plane angles were measured from pretreatment lateral cephalometric radiographs and used to classify the vertical dimension of each subject. RESULTS: Despite the fact that there was wide individual variation, there appeared to be no direct association between the presence of myosin heavy chain isoforms and specific vertical facial patterns. Type I myosin heavy chain isoform was the most common isoform found in all subjects. More Type IIA myosin heavy chain isoforms were observed in dolichofacial subjects. There were no differences between genders in myosin heavy chain expression. CONCLUSION: A wide variation of myosin heavy chain isoforms exists in the masseter muscle of individuals with different mandibular plane angles. Further investigations involving larger sample sizes and the incorporation of bite-force measurements may help to clarify the relationship between mandibular muscle characteristics and the vertical facial dimension.

Distribution of tropomyosin isoforms in different types of single fibers isolated from bovine skeletal muscles.

To clarify the relationship between myosin heavy chain (MyHC) isoforms and tropomyosin (TPM) isoforms in single fibers, 64 single fibers were isolated from each of bovine three muscles (masseter, semispinalis and semitendinosus). mRNA expressions of MyHC and TPM isoforms were analyzed by real-time PCR. All single fibers from the masseter expressed MyHC-slow. The fibers from the semispinalis expressed both MyHC-slow and 2a. The fibers from the semitendinosus expressed MyHC-slow, 2a and 2x. TPM-1 and TPM-2 were co-expressed in 2a and 2x type fibers, and TPM-2 and TPM-3 were co-expressed in slow type fibers. The expression pattern of TPM isoforms in each fiber type was similar between fibers isolated from different muscles. These results suggest that TPM-1 and TPM-3 isoforms correspond to the function of 2a or 2x type fibers and slow type fibers, respectively, with TPM-2 in common. Furthermore, the patterns of MyHC and TPM isoform combinations did not vary among single fibers isolated from the individual muscles examined.

X-ray microanalysis of elements in the masticatory muscle after paresis of the right masseter.

Muscle activity and function appear to be related to ionic concentrations in the muscle. We investigated whether muscle paresis induced by injection of Botulinum toxin A (Botox) in 16-week-old pigs over a 56-day period is associated with ionic changes in the affected muscles. Tissue samples were taken from the masseter, temporalis, medial pterygoid, and geniohyoid muscles by a standardized method and used for energy-dispersive x-ray microanalysis in an environmental scanning electron microscope. The largest increase in Na(+) was measured in the right and left sides of the masseter muscle in treated animals. Additionally, a significant elevation of Na(+) was measured in the anterior part of the temporalis muscle and in the pterygoid muscle (P < 0.05). In temporalis and pterygoid muscles, an increase in sulfur in both sides of treated pigs' heads was observed. Botox((R)) has an indirect impact on ion concentrations, resulting in changes in muscle functional capacity and adaptive compensation of paretic muscle function by other muscles.

Superficial and deep layer muscle fibre properties of the mouse masseter before and after weaning.

To clarify changes in the properties of the masseter muscle superficial and deep layer muscle fibres, which initiate masticatory movement, myosin heavy chain isoforms were evaluated based on immunohistochemistry at the transcription level in male mice both before and after weaning. In the results, MHC-2b isoforms, the isoforms with the fastest contraction speed, were observed in the superficial layer after weaning. However, MHC-2a isoforms with slower contraction speeds were not apparent. By contrast, in the deep layer, MHC-2a isoforms were present, as were MHC-2b isoforms, however, there were fewer MHC-2b isoforms present than in the superficial layer. The most rapid movement in the mouse mandible was observed anteroposteriorly during mastication. As the superficial layer of the masseter muscle runs parallel to the direction of mandibular movement, the presence of MHC-2b isoforms in it is consistent. The presence of MHC-2a isoforms in the deep layer, lying at right angles to the direction of mastication movement, is consistent with the positional adjustment of the mandible contributed by the deep layer muscle fibres during masticatory movement. We therefore conclude that complicated masticatory movement is achieved by the presence of various muscle bundles within the masseter, each carrying out different roles.

Chronic sleep deprivation alters the myosin heavy chain isoforms in the masseter muscle in rats.

To investigate the changes in myosin heavy chain (MyHC) isoforms of rat masseter muscle fibres caused by chronic sleep deprivation and a possible link with the pathogenesis of disorders of the temporomandibular joint (TMJ). A total of 180 male rats were randomly divided into three groups (n=60 in each): cage controls, large platform controls, and chronic sleep deprivation group. Each group was further divided into three subgroups with different observation periods (7, 14, and 21 days). We investigated he expression of MyHC isoforms in masseter muscle fibres by real-time quantitative polymerase chain reaction (PCR), Western blotting, and immunohistochemical staining. In rats with chronic sleep deprivation there was increased MyHC-I expression in layers of both shallow and deep muscles at 7 and 21 days compared with the control groups, whereas sleep deprivation was associated with significantly decreased MyHC-II expression. At 21 days, there were no differences in MyHC-I or MyHC-II expression between the groups and there were no differences between the two control groups at any time point. These findings suggest that chronic sleep deprivation alters the expression of MyHC isoforms, which may contribute to the pathogenesis of disorders of the TMJ.

Rabbit masseter expresses the cardiac alpha myosin heavy chain gene. Evidence from mRNA sequence analysis.

The presence of myosin alpha heavy chain in the rabbit masseter has been previously suggested at the protein level [(1991) Basic App. Myol. 1, 23-34; (1991) Histochem. J. 23, 160-170]. To confirm this finding, we cloned most of the mRNA corresponding to the myosin heavy chain S2 subfragment. PCR analysis and subsequent nucleotide sequence determination of the amplified cDNA demonstrates the presence of a myosin alpha heavy chain mRNA in rabbit masticatory muscles.

Fetal myosin heavy chain increases in human masseter muscle during aging.

Biochemical, immunohistochemical and molecular biological methods were used to detect fetal myosin heavy chain (MyHC) in the human masseter of elderly and young subjects. Samples from the elderly subjects contained larger amounts of fetal MyHC than those of young adults. Only a very small amount of embryonic MyHC could be detected in both age groups. Embryonic and fetal MyHCs were never detected in the control adult orofacial, limb and trunk muscles. Polymerase chain reaction (PCR) analysis revealed the presence of fetal mRNA sequences in elderly and young masseter muscles. We conclude that fetal MyHC is present in the human masseter throughout the life span and that there is an increase in the relative amount of this protein with age.

Sexually dimorphic expression of myosin heavy chains in the adult mouse masseter.

Little is known regarding the role of androgenic hormones in the maintenance of myosin heavy chain (MHC) composition of rodent masticatory muscles. Because the masseter is the principal jaw closer in rodents, we felt it was important to characterize the influence of androgenic hormones on the MHC composition of the masseter. To determine the extent of sexual dimorphism in the phenotype of masseter muscle fibers of adult (10-mo-old) C57 mice, we stained tissue sections with antibodies specific to type IIa and IIb MHC isoforms. Females contain twice as many fibers containing the IIa MHC as males, and males contain twice as many fibers containing the IIb MHC as females. There is a modest amount of regionalization of MHC phenotypes in the mouse masseter. The rostral portions of the masseter are composed mostly of type IIa fibers, whereas the midsuperficial and caudal regions contain mostly type IIb fibers. Using immunoblots, we showed that castration results in an increase in the expression of type IIa MHC fibers in males. Ovariectomy has no effect on the fiber type composition in females. We conclude that testosterone plays a role in the maintenance of MHC expression in the adult male mouse masseter.

Spatial distribution of myosin heavy-chain isoforms in mouse masseter.

There is a paucity of information regarding the anatomy and muscle fiber phenotype of the masseter. The objective of this study was to characterize the distribution of each myosin heavy-chain (MyHC) isoform within different anatomical regions of male and female mouse masseters. Masseters from male and female CD-1 mice (2-4 months old) were examined for description of the anatomical partitioning of muscle fibers and endplate distribution. The spatial distribution of MyHC isoforms--embryonic, neonatal, slow, alpha-cardiac, IIa, and IIb--was determined within the defined masseter partitions by means of Western blot analysis and immunofluorescent localization. Types IIa, IIx, and IIb were the predominant MyHC isoforms observed. Distinct differences in the spatial distribution of these MyHC isoforms were found between muscle regions and varied between sexes. The regionalization of muscle fiber types in the mouse masseter is consistent with the functional compartmentalization of the masseter observed in other species.