Effect of betel nut chewing on the otolithic reflex system

Clinical Neurophysiology 128 (2017) 138–146

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Effect of betel nut chewing on the otolithic reflex system Chuan-Yi Lin a, Yi-Ho Young b,⇑ a b

Department of Otolaryngology, Lo Tung Poh-Ai Hospital, Ilan, Taiwan Department of Otolaryngology, National Taiwan University Hospital Taipei, Taiwan

a r t i c l e

i n f o

Article history: Accepted 14 September 2016 Available online 13 October 2016 Keywords: Areca nut Betel nut Cervical vestibular-evoked myogenic potential Ocular vestibular-evoked myogenic potential Vestibulo-ocular reflex Vestibulo-sympathetic reflex

h i g h l i g h t s  Chewing betel nuts induced a transient loss of the otolithic reflex systems in fresh chewers.  Absent oVEMP and cVEMP may resolve to normal 20 min after chewing betel nut in fresh chewers.  Loss of otolithic reflexes may become permanent in habitual chewers.

a b s t r a c t Objective: This study investigated the effect of betel nut chewing on the otolithic reflex system. Methods: Seventeen healthy volunteers without any experience of chewing betel nut (fresh chewers) and 17 habitual chewers underwent vital sign measurements, ocular vestibular-evoked myogenic potential (oVEMP), and cervical VEMP (cVEMP) tests prior to the study. Each subject then chewed two pieces of betel nut for 2 min (dosing). The same paradigm was repeated immediately, 10 min, and 20 min after chewing. On a different day, 10 fresh chewers masticated chewing gum as control. Results: Fresh chewers exhibited significantly decreased response rates of oVEMP (53%) and cVEMP (71%) after dosing compared with those from the predosing period. These abnormal VEMPs returned to normal 20 min after dosing. In contrast, 100% response rates of oVEMP and cVEMP were observed before and after masticating chewing gum. In habitual chewers, the response rates of oVEMP and cVEMP were 32% and 29%, respectively, 20 min after dosing. Conclusion: Chewing betel nuts induced a transient loss of the otolithic reflexes in fresh chewers but may cause permanent loss in habitual chewers. Significance: Chewing betel nuts can cause a loss of otholitic reflex function. This creates a risk for disturbed balance and malfunction, for instance, during driving. Ó 2016 International Federation of Clinical Neurophysiology. Published by Elsevier Ireland Ltd. All rights reserved.

1. Introduction Betel nut (also called areca nut) is obtained from betel palm tree (Areca catechu), which is widely cultivated in tropical and subtropical areas, including India, Southeast Asia, East Africa, and New Guinea (Deng et al., 2001; Garg et al., 2014). A betel nut preparation conventionally comprises three ingredients: betel fruit, leaf of the betel pepper (Piper betle Linn.; a climbing shrub not related to the betel palm), and slaked lime paste obtained from shells, coral, or limestone (Nelson and Heischober, 1999). However, betel nut preparation ingredients may vary with demography. For instance, tobacco ingredients are added into the betel nut prepara⇑ Corresponding author at: Department of Otolaryngology, National Taiwan University Hospital, 1 Chang-te St., Taipei, Taiwan. Fax: +886 2 23946774. E-mail address: [email protected] (Y.-H. Young).

tion in most countries of Southeast Asia, whereas the commonest type of betel nut preparation in Taiwan, called ‘‘Ching-a” (Fig. 1), is made by wrapping together an unripe betel fruit, inflorescence of P. betle slaked with lime paste, and a piece of betel leaf. All three parts of ‘‘Ching-a” are chewed together. Although it is the fruit that is chewed, we will refer to it as the betel nut in this article. Betel nut chewing has become widespread in Taiwan during the past two decades, with the estimated number of habitual consumers being approximately 2.4 million, accounting for 11.4% of the total population in Taiwan (Chu, 2001). Most of them are indigenous people and blue-collar workers who chew it to reduce tension and provide a means for social interaction. Consequently, oral cancer became the fifth leading cause of death among males in Taiwan, with up to 90% of these attributable to betel nut chewers (Yang et al., 2001; Chen et al., 2008).

http://dx.doi.org/10.1016/j.clinph.2016.09.016 1388-2457/Ó 2016 International Federation of Clinical Neurophysiology. Published by Elsevier Ireland Ltd. All rights reserved.

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Fig. 1. The commonest type of betel nut preparation in Taiwan called ‘‘Ching-a” is made by wrapping together an unripe betel fruit (1), inflorescence of Piper betle Linn. slaked with lime paste (2), and a piece of betel leaf (3). All three parts are chewed together.

Betel nut chewing may cause a series of symptoms such as euphoria, palpitation, salivation, sweating, diaphoresis, heightened alertness, warm sensation in the body, and increased capacity to work. These psychological and neurological effects of betel nut may be attributed to the presence of arecoline, with parasympathomimetic properties acting on both muscarinic and nicotinic receptors (von Euler and Domeij, 1945; Chu, 2002). In addition, betel nut chewing increases the plasma concentrations of norepinephrine and epinephrine and thus may affect both the central and autonomic nervous systems (Chu, 2001). Autonomic response and vestibular symptoms are closely related. Patients with acute vertigo often have concomitant distressing autonomic symptoms such as nausea, vomiting, pallor, and sweating, implying that an efferent vestibular influence markedly affects the sympathetic outflow (Lin et al., 2012). Vomiting during caloric irrigation is common in those with anxiety/panic symptoms, which implies that the vestibulo-ocular reflex (VOR) shares synapses with the vestibulo-sympathetic reflex (Yates and Bronstein, 2005). By stimulating the ear with loud sound or bone vibration, the vestibular-evoked myogenic potential (VEMP) can be recorded on contracted neck muscles, termed cervical VEMP (cVEMP), and on the extraocular muscles, termed ocular VEMP (oVEMP) (Colebatch et al., 1994; Rosengren et al., 2005). These two emerging tests have been widely adopted in clinical practice for assessing the dynamic otolithic function (Young, 2013). The oVEMP primarily originates from the utricular macula, through the crossed VOR, along the superior vestibular nerve to the opposite extraocular muscles (Curthoys, 2010). In contrast, the cVEMP is evoked by air-conducted sound and primarily assesses the sacculo-collic reflex, whereas bone vibration-evoked cVEMPs probably assess both saccular and utricular vestibulo-collic projections. Although the psychological and neurological effects of betel nut have been substantially investigated (Chu, 2002), the impact of betel nut chewing on the otolithic function has never been explored. This study performed oVEMP and cVEMP tests to investigate the effect of betel nut chewing on the otolithic reflex system. 2. Methods 2.1. Participants Initially, 22 healthy volunteers without a history of betel nut chewing participated in this study. Contamination from other psychoactive drugs or alcohol was excluded. Subjects who failed to complete the examination were also excluded. Finally, 17 healthy

volunteers were enrolled and assigned to the fresh chewer group (15 males and 2 females; mean age = 28 years, range 24–39). None of the participants had any systemic disease or ear disorders and were further checked by otoscopy and audiometry. In the predosing period, baseline vital signs including body temperature, heart rate, and blood pressure were measured in each volunteer. Thereafter, each subject was administered with two pieces (one by one) of betel nut, called ‘‘Ching-a” (Fig. 1), for chewing for 2 min. Each piece of betel nut weighed 3.8 ± 0.6 g. One gram of betel nut contained four major alkaloids: arecoline (7.5 mg), arecaidine (1.5 mg), guvacoline (2.0 mg), and guvacine (2.9 mg) (Wang et al., 1997). The rationale for chewing two pieces of betel nut is that facial flushing is noted in all volunteers (100%) after chewing two pieces (one by one) but in only 24% of the volunteers after chewing one piece. Immediately after chewing (dosing period), vital sign measurements followed by oVEMP and cVEMP tests were repeated. Then, all the subjects rested in a quiet room, and 10 and 20 min after dosing (postdosing I and II, respectively), all the subjects underwent the same paradigm. Fig. 2 illustrates the flow chart of the testing procedure. On a different day, 10 subjects from the fresh chewer group repeated the same paradigm; however, this time betel nut was replaced by chewing gum (control group). Each subject underwent oVEMP and cVEMP tests before and after dosing. This study was approved by the institutional review board of the university hospital, and each subject signed the informed consent form before participation. 2.2. oVEMP test The subject was seated and gazed upward at a fixed target that was >2 m from the eyes during recording (Smart EP 3.90; Intelligent Hearing Systems, Miami, FL). Two active surface electrodes were attached 1 cm below the lower eyelids. The other two reference electrodes were 1–2 cm below the active ones, and a ground electrode was placed on the sternum. Each response had duration of 50 ms for analysis, with a stimulation rate of 5/s, and 30 responses were averaged for each run. The operator held the vibrator (Minishaker 4810; Bruel & Kjaer Co., Nærum, Denmark) and tapped on the subject’s skull on the forehead. The input signal was 500 Hz sine wave, with the initial peak driving voltage about 144 dB force level. The initial negative–positive biphasic waveforms were termed waves nI and pI, respectively. Consecutive runs were undertaken to confirm the reproducibility of the nI–pI waveform, and oVEMPs

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Fig. 2. Flow chart of the testing procedure in betel nut chewers. HR: heart rate, oVEMP: ocular vestibular-evoked myogenic potential; cVEMP: cervical vestibular-evoked myogenic potential.

were deemed to be present. The norm for the latency of peak nI at our laboratory was 11.4 ± 0.8 ms. Alternatively, if oVEMP was not induced, then the ipsilateral mastoid site (2 cm behind the opening of external ear canal) was tapped. Ears with absent oVEMP from forehead tapping but present oVEMP from mastoid tapping were also interpreted as having reduced responses (Tseng et al., 2012; Lin et al., 2013). 2.3. cVEMP test All subjects were in a supine position with elevated head during testing. Two active electrodes were attached bilaterally on the midpoint of the sternocleidomastoid muscles; one reference electrode was placed on the suprasternal notch, and a ground electrode was placed on the forehead. The other settings were the same as in the oVEMP test, except that the vibrator was tapped on the subject’s head at the inion (Tseng et al., 2013). To measure background muscle activity, subjects were given feedback on the level of electromyographic (EMG) activity in their sternocleidomastoid muscles during data collection. The electromyographic signals were amplified, and actual mean rectified background muscle activity for cVEMPs was measured (Chang et al., 2007). A total of 50 responses were averaged and recorded bilaterally. The first positive and second negative polarities of the biphasic waveform were termed waves p13 and n23, respectively. Consecutive runs were undertaken to confirm the reproducibility of p13–n23 waveform, and cVEMPs were termed to be present. The norm for the latency of p13 at our laboratory was 14.4 ± 1.3 ms. 2.4. Long-term effects of betel nut chewing Seventeen habitual betel nut chewers (16 males and 1 female; mean age = 52 years, range 35–59) also underwent heart rate measurement, oVEMP test, and cVEMP test before and after chewing betel nut for studying the long-term effects of betel nut chewing (Fig. 2). The mean duration of betel nut chewing was 12 years (range 5–20 years), and the amount of betel nut chewing was approximately 10 pieces (1 pack) daily. No systemic diseases or middle/inner ear disorders were noted, and each subject was further checked by otoscopy. 2.5. Statistical methods The data of vital signs with respect to betel nut dosing were analyzed using one-way repeated-measures analysis of variance

(ANOVA) with Bonferroni-adjusted paired t-test for multiple comparisons. Responses of oVEMPs or cVEMPs to dosing periods were compared using Cochran Q test, followed by McNemar test. The response rates of VEMP responses between the two groups were compared using Fisher’s exact test. The characteristic parameters of oVEMP or cVEMP among the four periods were investigated using one-way repeated-measures ANOVA with Bonferroniadjusted paired t-test for multiple comparisons. A p value of <0.05 was considered to be significant. 3. Results 3.1. Clinical manifestation After chewing betel nut, flushing was observed in all the 17 (100%) fresh chewers, followed by dizziness in 10 subjects (59%), nausea with gastric upset in 8 subjects (47%), palpitation in 7 subjects (41%), and sweating in 7 subjects (41%). All these symptoms were transient and subsided 20 min after dosing. None of them had rotatory vertigo, nystagmus, or abnormal head impulse test. 3.2. Vital signs in betel nut dosing Vital signs comprising heart rate, body temperature, and blood pressure were measured in the predosing, dosing, and postdosing I and II periods (Table 1). The mean heart rate per minute was 71 ± 12 beats in the predosing period, which elevated to 98 ± 8 beats in the dosing period, and then declined to 80 ± 9 and 73 ± 11 beats in the postdosing I and II periods, respectively, exhibiting a significant relationship with betel nut dosing (p < 0.001, one-way repeated-measures ANOVA test; Table 1). The heart rate in the dosing period, but not in the postdosing I and II periods, was significantly higher than that in the predosing period (p < 0.0001, Table 1), indicating that betel nut has a transient effect (<10 min) on the heart rate. In contrast, relationship between betel nut dosing and other vital signs, i.e., body temperature, systolic pressure, and diastolic pressure was not observed (p > 0.05, Table 1). 3.3. oVEMPs in betel nut dosing All the 17 fresh chewers (34 ears; 100%) had clear oVEMPs, bilaterally, in the predosing period (Fig. 3). However, immediately after chewing betel nut, the response rate of oVEMP declined to 53% of the ears, which then increased to 76% in postdosing I period

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C.-Y. Lin, Y.-H. Young / Clinical Neurophysiology 128 (2017) 138–146 Table 1 Changes in vital signs with respect to betel nut dosing. N Heart rate (/min) Body temperature (°C) Systolic pressure (mmHg) Diastolic pressure (mmHg)

17 17 17 17

Predosing

Dosing

71 ± 12 36.2 ± 0.5 126 ± 15 74 ± 12

a

98 ± 8 36.4 ± 0.5 131 ± 18 76 ± 11

Postdosing I b

80 ± 9 36.3 ± 0.4 133 ± 11 80 ± 8

Postdosing II b

73 ± 11 36.2 ± 0.5 127 ± 14 78 ± 10

p value <0.001 0.631 0.646 0.463

The data are expressed as mean ± SD; Postdosing I and II correspond to 10 and 20 min after dosing, respectively. p value: repeated-measures one-way ANOVA test with Bonferroni-adjusted paired t test. a p < 0.001. b p > 0.05 compared with the predosing period.

Fig. 3. The ocular vestibular-evoked myogenic potential (oVEMP; nI–pI waveform) responses with respect to betel nut chewing. (A) Predosing, normal oVEMPs are shown, bilaterally. (B) In dosing period, bilateral oVEMPs are not elicited. (C) 20 min after dosing, normal oVEMPs are shown bilaterally.

and resolved and returned to 100% in postdosing II period. A significant relationship was identified between the response rates of oVEMP and betel nut dosing (p < 0.001, Cochran Q test; Table 2).

Furthermore, compared to the predosing period, significant differences existed in the response rates of oVEMP in the dosing (p < 0.0001) and postdosing I (p = 0.0133) periods but not in the

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Table 2 Comparison of characteristic parameters of oVEMP with respect to betel nut dosing. N (ears) Response rate nI latency (ms) pI latency (ms) nI–pI amplitude (lV)

34 18 18 18

Predosing 100% 8.2 ± 0.5 11.7 ± 0.8 14.2 ± 9.0

Dosing 53% (18/34) 8.5 ± 0.6 12.1 ± 0.9 19.4 ± 11.2

Postdosing I a

76% (26/34) 8.4 ± 0.7 12.1 ± 0.9 15.2 ± 8.1

b

Postdosing II

p value

100% 8.4 ± 0.7 12.1 ± 0.7 14.1 ± 8.7

<0.001 0.614 0.359 0.403

The data are expressed as mean ± SD; oVEMP: ocular vestibular-evoked myogenic potential. Postdosing I and II correspond to 10 and 20 min after dosing, respectively. p value: Cochran Q test or one-way ANOVA test. a p < 0.0001. b p = 0.0133, McNemar test, compared with the predosing period.

postdosing II period (p > 0.05, McNemar test; Table 2), indicating that the effect of betel nut on oVEMPs is also transient (<20 min) and reversible. However, the characteristic parameters of oVEMPs, i.e., nI latency, pI latency, and nI–pI amplitude, did not significantly differ among the four periods (p > 0.05, one-way ANOVA test; Table 2).

identified between the response rates of cVEMP and betel nut dosing in habitual chewers (p < 0.01, Cochran Q test, Table 5). Furthermore, like in the fresh chewers, the characteristic parameters of oVEMPs/cVEMPs did not significantly differ among the four periods (p > 0.05). In summary, more than 60% of the ears showed loss of oVEMP (68%) and cVEMP (71%) in habitual chewers.

3.4. cVEMPs in betel nut dosing 4. Discussion The mean rectified background muscle activities for cVEMPs in the predosing, dosing, and postdosing I and II periods were 66 ± 20, 60 ± 15, 58 ± 17, and 61 ± 13 lV, respectively (p > 0.05, one-way ANOVA test). The cVEMP test (Fig. 4) showed a relationship between the response rates of cVEMP and betel nut dosing (p < 0.001, Cochran Q test; Table 3), which was similar to the oVEMP test. Compared to the predosing period (100%), there were significant differences in the response rates of cVEMP in the dosing (71%) and postdosing I (82%) periods (p < 0.05) but not in the postdosing II period (p > 0.05, McNemar test; Table 3), indicating that the response for cVEMP is also reversible. Again, the characteristic parameters of cVEMP, i.e., p13 latency, n23 latency, and p13–n23 amplitude, were not significantly different among the four periods (p > 0.05, Table 3). 3.5. Chewing gum group On a different day, 10 subjects from the fresh chewer group masticated chewing gum to serve as a control group. None of them experienced facial flushing, dizziness, nausea with gastric upset, palpitation, sweating, and so on after chewing. All the subjects showed 100% response rates of oVEMP and cVEMP regardless of predosing, dosing, and postdosing periods. The characteristic parameters of oVEMPs and cVEMPs did not significantly differ among these periods (p > 0.05, one-way ANOVA test; Table 4). 3.6. Long-term effects of betel nut chewing Seventeen habitual chewers were also enrolled in this study to investigate the long-term effects of betel nut chewing. The mean heart rate per minute was 68 ± 12 beats in the predosing period, which elevated to 89 ± 8 beats in the dosing period, and then declined to 75 ± 9 and 70 ± 11 beats in the postdosing I and II periods, respectively, exhibiting a significant relationship with betel nut dosing like in fresh chewers (p < 0.001, one-way repeatedmeasures ANOVA test; Table 5). In the predosing period, present oVEMPs were noted in 11 ears (32%) and absent oVEMP in 23 ears. The response rate of oVEMPs in habitual chewers declined to 18% in the dosing period and then recovered to 21% and 32% in the postdosing I and II periods, respectively, exhibiting a significant relationship between the response rates of oVEMP and betel nut dosing like in fresh chewers (p < 0.01, Cochran Q test; Table 5). A similar relationship was also

The top four consumed psychoactive substances in the world are from nicotine, alcohol, caffeine, and betel nut (Garg et al., 2014). Previously, Lin and Young (2001) reported that intractable vertigo is related to smoking behavior but is unrelated to smoking years and pack years. The latter was defined as daily smoking amount (pack/day) multiplied by smoking years (number of years smoked). Next, Chiang and Young (2007) studied the influence of alcohol on the vestibular system and revealed that the deterioration of the VOR performance correlates with 0.25 mg/L breath alcohol concentration (BrAC), whereas the sacculo-collic reflex and vestibulo-cerebellar interaction were affected by alcohol at BrAC of <0.25 mg/L. At a maximum BrAC of 0.15 mg/L, Rosengren et al. (2014) reported that alcohol consumption had a selective dampening effect on oVEMP amplitude. Furthermore, impairment of the VOR evoked by high-acceleration head impulses was identified after alcohol consumption (Roth et al., 2014). With regard to the betel nut, although it is consumed by approximately 10% of the world population, its impact on the otolithic reflex system has never been explored. Conventionally, the main constituents of betel nut are arecoline (alkaloids of areca) and arecaidine (a hydrolyzed derivative of arecoline) that may affect many systems such as the nervous, cardiovascular, gastrointestinal, respiratory, and endocrine systems. Therefore, a variety of symptoms such as facial flushing, dizziness, nausea with gastric upset, palpitation, and sweating were experienced in the betel nut chewers in our study. Notably, arecoline has been shown to devastate the oral tissues by diffusing through the oral mucosal tissues, resulting in leukoplakia, submucous fibrosis, and verrucous lesion, which are precancerous lesions with an incidence of 1–18% progression to oral cancer (Reibel, 2003; Javed et al., 2010). During the past two decades, betel nut chewing has become a serious public health problem in Taiwan. The prevalence of oral cancer increased from 10.79 per 100,000 males in 1991 to 15.47 per 100,000 males in 1995. That is a 43% increase in oral cancer in Taiwan within four years (Yang et al., 2001). 4.1. Change in vital signs In addition to causing oral cancer, chewing betel nut may affect the cardiovascular system. This study identified a significant relationship between betel nut chewing and heart rate but not body temperature and blood pressure (Table 1). Compared with the pre-

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Fig. 4. The cervical vestibular-evoked myogenic potential (cVEMP; p13–n23 waveform) responses with respect to betel nut chewing. (A) Predosing. (B) Bilateral cVEMPs are absent immediately after chewing two pieces of betel nut but are present 20 min after chewing betel nut (C).

dosing period (mean heart rate, 71 beats/min), significantly increased mean heart rate was noted in the dosing period (mean, 98 beats/min), which then resolved to 73 beats/min 20 min after dosing, indicating that the effect of betel nut chewing on heart rate is transient and reversible. This finding was compatible with the literature that cardioacceleratory effect begins within 2 min of chewing, reaches a maximal effect within 4–6 min, and lasts for

an average of 16.8 min, implying that arecoline exerts a cardiovascular effect through the central cholinergic mechanism (Chu, 2001). However, the effect of betel nuts on blood pressure did not exhibit a statistically significant relationship with betel nut dosing, which is consistent with a previous report (Lin et al., 2002). Similarly, body temperature also showed no significant relationship

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Table 3 Comparison of characteristic parameters of cVEMP with respect to betel nut dosing. N (ears) Response rate p13 latency (ms) n23 latency (ms) p13–n23 amplitude (lV)

34 24 24 24

Predosing

Dosing

100% 15.0 ± 1.5 21.7 ± 2.2 69.6 ± 23.6

Postdosing I

71% (24/34) 14.8 ± 1.5 22.0 ± 2.5 70.6 ± 18.6

a

82% (28/34) 13.7 ± 1.6 21.6 ± 2.4 64.4 ± 15.0

b

Postdosing II

p value

100% 14.3 ± 1.3 22.0 ± 2.5 68.7 ± 16.6

<0.001 0.098 0.925 0.890

The data are expressed as mean ± SD; cVEMP: cervical vestibular-evoked myogenic potential. Postdosing I and II correspond to 10 and 20 min after dosing, respectively. p value: Cochran Q test or one-way ANOVA test. a p = 0.0044. b p = 0.0412, McNemar test, compared with the predosing period.

Table 4 Comparison of characteristic parameters of oVEMP and cVEMP with respect to chewing gum dosing.

Response rate nI latency (ms) pI latency (ms) nI–pI amplitude (lV) p13 latency (ms) n23 latency (ms) p13–n23 amplitude (lV)

N (ears)

Predosing

Dosing

Postdosing I

p value

20 20 20 20 20 20 20

100% 8.3 ± 0.4 11.9 ± 0.6 10.8 ± 5.8 13.6 ± 1.3 21.2 ± 1.5 111 ± 44

100% 8.0 ± 0.4 11.9 ± 0.9 12.6 ± 4.9 13.6 ± 1.5 20.9 ± 2.2 115 ± 42

100% 8.1 ± 0.5 11.9 ± 1.0 12.4 ± 4.7 13.6 ± 1.4 22.0 ± 1.9 106 ± 27

0.149 0.981 0.541 0.973 0.916 0.944

The data are expressed as mean ± SD; p value: one-way ANOVA test; postdosing I corresponds to 10 min after dosing.

Table 5 Change in the heart rate and VEMPs in habitual chewers with respect to betel nut dosing.

Predosing Dosing Postdosing I Postdosing II p value

N

Heart rate (beats/min)

N (ears)

oVEMP Response rate

cVEMP Response rate

17 17 17 17

68 ± 12 89 ± 8a 75 ± 9b 70 ± 11b <0.001*

34 34 34 34

32% 18% 21% 32% <0.01#

29% 15% 24% 29% <0.01#

The data are expressed as mean ± SD; Postdosing I and II correspond to 10 and 20 min after dosing, respectively. * repeated-measures one-way ANOVA test with Bonferroni-adjusted paired t-test. a p < 0.001. b p > 0.05 compared with the predosing period. # Cochran Q test.

with betel nut dosing, although facial flushing (warm sensation) is the most prominent symptom after chewing betel nut. Chu (1995) proposed that facial flushing immediately after betel nut chewing was caused by heat production from hydrolysis of lime, whereas others suggested that betel nut induces adrenal chromaffin cells to release catecholamine (Boucher and Mannan, 2002). In summary, betel nut may affect the central and autonomic nervous systems because of its arecoline content, which possesses parasympathomimetic properties that stimulate both muscarinic and nicotinic receptors (Garg et al., 2014). A network of vestibulo-autonomic projections in the brainstem and cerebellum, i.e., vestibulo-autonomic regions in vestibular nuclei, were demonstrated to direct descending pathways to brainstem circuitry and ascending pathways to the parabrachial nucleus (Balaban and Porter, 1998). Thus, dizziness and nausea after chewing betel nut may be mediated by vestibular autonomic regulation.

among the four periods, indicating that betel nut chewing has a transient and reversible impact on the otolithic-ocular reflex in fresh chewers, possibly acting as a form of reversible conduction block rather than interruption by the nausea or ill sensation (autonomic dysfunction) after betel nut chewing. Furthermore, it is hypothesized that patients with motion sickness, i.e., nausea/vomiting frequently combined with hypotension, have a compensatory response of increased muscular sympathetic nerve activity, as evidenced by the hyperactive caloric responses (Furman et al., 1998; Lin et al., 2012). Because both caloric and oVEMP tests partly share the VOR system, absent oVEMP instead of augmented oVEMP in fresh chewers is unrelated to an autonomic effect. Comparing the transient interval, the oVEMP (20 min), which is modulated by a network of vestibulo-autonomic interaction, takes a longer time than the heart rate (10 min) to recover to its predosing condition, possibly because two systems are involved, namely cardiovascular and otolithic-ocular reflex systems.

4.2. Change in oVEMPs 4.3. Change in cVEMPs In this study, the response rates of present oVEMP in fresh chewers was 100% in the predosing period, 53% in the dosing period, and 76% 10 min after dosing, returning to 100% 20 min after dosing (Fig. 3), showing a significant relationship to betel nut dosing (Table 2). However, a significant difference was not observed in the characteristic parameters (latencies and amplitude) of oVEMP

Similar to the oVEMP test, the response rate of present cVEMPs was 100% in the predosing period and 71% in the dosing period and returned to 100% 20 min after dosing (Fig. 4), showing a significant relationship between cVEMPs and betel nut dosing (Table 3). Similarly, characteristic parameters did not significantly differ among

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the four periods, implying that betel nut also affects the sacculocollic reflex transiently and reversibly. Afferent fibers from otolithic organs, which trigger the otolithicocular, sacculo-collic, and vestibulo-sympathetic reflexes, synapse at the vestibular nuclei (Wilson et al., 1995). Both otolithic-ocular (assessed by the oVEMP test) and vestibulo-collic signals (assessed by the cVEMP test) transit through the vestibular nuclei; in addition, vestibulo-sympathetic reflexes synapse at the vestibular nuclei. It is our premise that betel nut chewing produces a central sympathetic effect because of arecoline as evidenced by the accelerated heart rate and a conduction block on the otolithic reflex, which resulted in temporarily absent oVEMP (47%) and cVEMP (29%) in fresh chewers immediately after dosing. 4.4. Effect of mastication One may argue that mastication may have altered the VEMP responses because mastication can induce significant vertigo and nystagmus in patients with vestibulopathy but not in healthy subjects (Park et al., 2014). Thus, a control group that masticated chewing gum was included, and 100% responses rates of oVEMP and cVEMP were observed before or after chewing (Table 4). Therefore, it is the betel nut itself rather than mastication that affects the otolithic reflex systems. 4.5. Effect of habitual betel nut chewing A temporary hearing threshold shift after short-term noise exposure may become a permanent hearing threshold shift after long term-noise exposure (Hsu et al., 2008); similarly, transiently absent oVEMP and cVEMP in fresh chewers may become a permanent absence as evidenced by the >60% loss of oVEMPs and cVEMPs in habitual chewers (Table 5). However, the interval for the recovery of the heart rate and VEMP responses after dosing did not significantly differ between the fresh and habitual chewers, which is compatible with the results of previous studies (Chu, 2001). The reason may be that arecoline has a neurotoxic effect on the otolithic reflexes by increasing oxidative stress and suppressing the antioxidant system. Thus, the accumulated toxic effects of arecoline may result in permanent loss of otolithic reflexes (Shih et al., 2010). In this study, the mean age of the habitual chewers (52 years) was higher than that of fresh chewers (28 years) partly because the former had a duration of betel nut chewing of 5–20 years. Therefore, one may question regarding the deterioration of VEMPs due to aging because the effect of aging (>60 years) on the VEMPs includes reduced response rate, prolonged latencies, and decreased amplitude (Tseng et al., 2010). However, because all the betel nut chewers in our study were below the age of 60 years, the effect of aging on the response rate of VEMPs is less as decline in the numbers of vestibular hair cells and afferent neurons occurs in subjects aged >60 years (Tseng et al., 2010). Thus, >60% loss of oVEMP and cVEMP were mainly attributed to the neurotoxic effect of betel nuts rather than aging. 4.6. Clinical relevance Before the era of oVEMP test, the classical subjective visual horizontal (SVH) or vertical (SVV) test was used to evaluate utricular function. However, the SVH test failed to gain popularity mainly because of its multiple confounding factors, i.e., it is affected by the head position relative to the gravity and by linear acceleration forces acting on the gravity vector (Bohmer and Rickenmann, 1995). Lin and Young (2011) correlated the static SVH test with the dynamic oVEMP test in healthy and pathological ears and confirmed that both tests may, at least in part, share the same utricular

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reflex pathway. In addition, the Romberg quotient of sway area on foam posturography has been associated with the oVEMP test results (Lin et al., 2013). However, it is the acceleration of the head or tilt that activates the otolithic organs (from their steady state) and not standing still on a foam pad. Thus, the dynamic oVEMP test is superior to static posturography or the SVH/SVV test for evaluating the utricular function. Because cVEMP is related to head control (Chen et al., 2007) and oVEMP is related to independent walking in small children (Young, 2015), loss of otolithic reflexes may affect the balance function in a moving environment. Therefore, engagement in driving, craft, or precise machine work immediately after chewing betel nuts may create a potential risk of imbalance.

5. Conclusion Chewing betel nuts induced a transient and reversible effect on the otolithic-ocular reflex and vestibulo-collic reflex systems in fresh chewers, as evidenced by transiently absent oVEMP and cVEMP responses that resolved and returned to normal 20 min after chewing betel nut. However, loss of otolithic reflexes may become permanent in habitual chewers.

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