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New Animal Model for Studying Mastication in Oral Motor Disorders
1 Division of Oral Physiology, Correspondence: *corresponding author, yamada{at}dent.niigata-u.ac.jp
To identify the basic parameters of oral behavior in mice, we recorded the three-dimensional jaw movement trajectories and masseter and digastric muscle activities in freely behaving mice eating foods of various textures. Results showed that: (1) there are characteristic jaw movement patterns for food intake and mastication; (2) the pattern in a chewing cycle may be divided into opening, closing, and protruding (power) strokes; and (3) food texture affects basic patterns of jaw movement, muscle activities, and chewing rhythms. The oral motor behavior of mice appears identical to those of other experimental animals, so mice are appropriate animal models for the study of mastication.
Key Words: freely behaving mouse • masticatory jaw movement • masticatory muscle activity • chewing rhythm • food texture
Mastication is one of the most common rhythmic behaviors, along with respiration and locomotion, in mammals. The motor command for the basic pattern of rhythmic jaw movements is commonly accepted to originate from a central pattern generator (Lund, 1991; Nakamura and Katakura, 1995). Although the basic motor timing could be programmed in the generator, sensory inputs arising from the movements modify the rhythmic movements reflexively to adapt to environmental demands, such as changes in food texture. To elucidate peripheral control mechanisms, investigators have described oral motor behaviors in the cat (Thexton et al., 1980), rat (Hiiemae, 1968), guinea pig (Byrd, 1981), and rabbit (Morimoto et al., 1985; Schwartz et al., 1989; Yamada and Yamamura, 1996). In contrast, many novel genes have been identified from human genome and other studies. The next important step is to identify the function of a particular gene and its link to human diseases. Gene-targeted mice are useful models for studying the consequences of overexpression, underexpression, and complete inactivation of a particular gene. Such animal models have been developed recently to study genetic links to Alzheimer’s disease (Hsiao et al., 1996) and Huntington’s disease (Carter et al., 1999). Among gene-targeted animals are serotonin receptor-deficient mice (Tecott et al., 1995) that exhibit eating disorders and epilepsy. Serotonin has also been suggested to be linked with oral dyskinesias (Eberle-Wang et al., 1996) that often result from dysfunction of the basal ganglia. Therefore, genetically modified mice are expected to enhance our understanding of neural and motor disorders in the craniomandibular system. Little research has been conducted on this system other than to describe the basic parameters of chewing in the mouse (Kobayashi et al., 2002). The present study describes precise ingestive movements in freely behaving mice from recordings of three-dimensional jaw movement trajectories and masticatory muscle activities during the chewing of foods of different consistencies.
In this study, 5 CH3 male wild-type mice (weighing from 23 to 28 g each) were used. The experimental procedure was reviewed and approved by the Niigata University Intramural Animal Care and Use Committee. We recorded jaw movement trajectories in both frontal and sagittal planes and electromyograms (EMGs) of the masseter (MAS), which closes the jaw, and the digastric (DIG), which opens the jaw. Electrodes were implanted into mice under general anesthesia with sodium pentobarbital (25 mg/kg). Bipolar electrodes were placed on the superficial MAS muscles bilaterally and on the DIG muscle unilaterally, due to space limitations. The animals were allowed to recover for 3 days before we recorded them chewing bread (soft food) or pellets (hard food). A detailed methodology for making recordings from freely moving animals has been previously published (Yamada et al., 1988; Koga et al., 2001). To synchronize the jaw movement and EMG signals, we stored all of the analog signals in computer memory through a 12-bit analog/digital (A/D) converter. Because the sampling rate was fixed at 100 Hz for the jaw-tracking system (Koga et al., 2001), which was not enough to identify the duration of muscle bursts, we used EMG recordings to evaluate the amplitude of muscle bursts. For this, EMG signals were high-pass-filtered (cut-off frequency of 100 Hz), full-wave-rectified, and smoothed before the signals were forwarded to the A/D converter. For each animal, analyses were carried out on 20 chewing cycles in which foods were crushed between the upper and lower molars. The average and standard error of the data from all the animals tested were obtained from the averages of the 20 cycles of each animal (i.e., means’ mean and SEM). Parametric data were analyzed by paired t tests. A p value of less than 0.05 was considered to be statistically significant.
A typical masticatory sequence of a mouse chewing chow pellets is shown in Fig. 1  . The masticatory sequence began with the mouse opening its jaw widely to ingest the food. The animal held an adequate amount of food between its forefeet, then stabilized its head in the upright position for further food processing. The sequence could be divided into two motor behaviors (food intake and mastication) according to the jaw movement orbits. During food intake, the jaw was lowered, protruded, and then withdrawn with little vertical fluctuation, and this stage was characterized by relatively low MAS activity. Food intake then changed to mastication, apparent by large antero-posterior movements. Cycles in mastication (the so-called masticatory series) were characterized by large anterior movements and regular jaw movement trajectories. At the end of a masticatory series, the jaw lowered as seen in the beginning of the sequence for food intake.
Movement orbits of five successive chewing cycles in a masticatory series were reconstructed in the frontal and sagittal planes as each mouse was fed chow pellets and bread (Fig. 2 ). A large anterior movement was obvious in the sagittal plane, and a subsequent lateral movement was obvious in the frontal plane. Chewing movements in the sagittal plane showed three distinct strokes with directional changes. Stroke 1 began at the most protrusive jaw position and ended at the maximum opening position in a single masticatory cycle. The jaw shifted laterally toward the chewing side. After stroke 1, stroke 2 closed the jaw. The jaw then moved laterally toward the centric occlusion. After stroke 2, stroke 3 protruded the jaw. In the middle of stroke 1, a small anterior movement could be seen during bread chewing (Fig. 2B) that was not apparent during pellet chewing (Fig. 2A).
The gape size (mean ± SEM, n = 5) varied between 1020 ± 96 µm (hard food) and 1580 ± 194 µm (soft food) during chewing. Similarly, the anterior movements varied between 380 ± 119 µm (soft food) and 670 ± 133 µm (hard food); the lateral movements varied between 440 ± 192 µm (hard food) and 690 ± 425 µm (soft food). The relative gape size was 35% larger during the chewing of soft food, and the amount of lateral excursion was 37% larger during the chewing of soft food, whereas the amount of anterior excursion was 43% larger during the chewing of hard food. There were significant differences in the gape size (p < 0.01) and the anterior movement (p < 0.05) between soft and hard foods. When individual stroke durations were closely observed, the mean duration of stroke 3 varied significantly (p < 0.01) between 38 ± 5 msec (mean ± SEM, n = 5) (soft food) and 64 ± 5 msec (hard food). In contrast, the mean duration of stroke 1 was significantly shorter (p < 0.05) for the hard food: The difference was 33 msec between soft food (93 ± 20 msec) and hard food (60 ± 10 msec). In contrast, the mean duration of stroke 2 showed no significant differences between foods: The difference was only 9 msec. The mean total cycle (TC) duration of chewing varied between 188 ± 19 msec (hard food) and 204 ± 37 msec (soft food), but there were no significant differences.
Shown in Fig. 3
To investigate factors that may affect the TC duration of chewing, we carried out regression analyses between the TC duration and the duration in each of strokes 1, 2, and 3 (Table ). With the light load on the jaw from bread chewing, only the duration of stroke 1 proved to be a factor affecting TC duration (r = 0.90, slope = 1.33). With the heavy load on the jaw from pellet chewing, only the duration of stroke 2 was determined to be a factor affecting TC duration (r = 0.73, slope = 1.06).
Although the mouse has been used as a transgenic model of many diseases, and its central neuronal networks localized within the hindbrain have been studied (Jacquin et al., 1996; Picciotto and Wickman, 1998), its oral motor behavior remains unclear. In this study, we described precise three-dimensional jaw movement orbits and masticatory muscle activities during chewing in the freely behaving mouse. The results suggest that the mouse can be used, along with a newly developed recording system (Koga et al., 2001), as a model for studying masticatory disorders. Patterns of jaw movements are different among species. Fundamental characteristics of jaw movements of the rat are antero-posterior movements with few lateral excursions (Hiiemae, 1968). In the rabbit, however, jaw movements show large unilateral excursions (Morimoto et al., 1985; Schwartz et al., 1989; Yamada and Yamamura, 1996). Jaw movements in the mouse were found, by two-dimensional analysis (Kobayashi et al., 2002), to be essentially the same as those in the rat, but our study, by three-dimensional analysis, clarified the antero-posterior movements and found a great deal of lateral movement. It is novel that mice have lateral jaw movements similar to those of humans. We could distinguish a mouse’s masticatory sequence, from one food intake event to the next, by the pattern of jaw movements. We could divide the chewing cycle into three strokes on the basis of directional changes of the movement in the sagittal plane. Stroke 1 was an opening movement, stroke 2 was a closing movement, and stroke 3 was a protruding movement. Because the protruding stroke coincided with great MAS activity and lasted longer during the chewing of hard food, it was considered to be comparable with the so-called power stroke of the human chewing cycle. In the middle of the opening stroke, a small anterior movement could be seen during bread chewing, in which the food texture was soft and somewhat sticky, but that same movement was not apparent during pellet chewing. This additional movement may account for the significantly longer duration of the opening stroke during the chewing of soft food than of hard food. Both MAS and DIG muscle activities were affected by food consistency. The closing muscle was more active when the animals were chewing the pellet than while they chewed the bread. The hard food consistency may have required more power activity from the MAS. In contrast, the gape size was greater when they chewed the bread than when they chewed the pellet. The soft food consistency may have required more jaw opening activity from the DIG. The change in the TC duration or the chewing rhythm may not necessarily vary equally over several chewing cycles. In the rabbit, the duration of the opening stroke is reportedly correlated with the TC duration during the chewing of soft food, while the duration of the closing stroke is correlated with the TC duration when hard food is chewed (Yamada and Yamamura, 1996). Although the jaw movement pattern during chewing, especially in stroke 3 (the power stroke), in the mouse showed some difference from that in the rabbit, the present results suggest that the sensory feedback mechanism controlling the masticatory movement may work in the mouse as has been suggested in the rabbit. Our study demonstrated that although a mouse is small, it can be used as a model for studying masticatory disorders for two basic reasons: (a) jaw motor behaviors could be identified from the movement orbit and muscle activity, and (b) the contribution of sensory feedback on jaw behaviors could be evaluated. Making comparisons between the normal mouse and the transgenic mouse with a behavioral dysfunction will greatly assist in the investigation of brain-behavior relationships and may facilitate our understanding of the molecular genetic pathology of oral motor disorders.
We thank Dr. Yoshiyuki Koga and Mr. Hidetoshi Hirano for their technical assistance in making the amplifier and Dr. Jayantha K.C. Amarasena for her valuable help in preparing the manuscript. Received for publication December 6, 2001. Revision received September 23, 2002. Accepted for publication January 23, 2003.
Journal of Dental Research, Vol. 82, No. 4,
318-321 (2003)
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