Combined effects of starvation and butyrate on autophagy-dependent gingival epithelial cell death.

BACKGROUND AND OBJECTIVE: Bacteria in the dental biofilm surrounding marginal gingival grooves cause periodontal diseases. Numerous bacteria within the biofilm consume nutrients from the gingival crevicular fluid. Furthermore, some gram-negative bacteria in mature dental biofilms produce butyrate. Thus, gingival epithelial cells in close proximity to mature dental biofilms are at risk of both starvation and exposure to butyrate. In the present study, we determined the combined effects of starvation and butyrate exposure on gingival epithelial cell death and the underlying mechanisms. MATERIAL AND METHODS: The Ca9-22 cell line was used as an in vitro counterpart of gingival epithelial cells. Cell death was measured as the amount of total DNA in the dead cells using SYTOX Green dye, which penetrates through membranes of dead cells and emits fluorescence when it intercalates into double-stranded DNA. AMP-activated protein kinase (AMPK) activity, the amount of autophagy, and acetylation of histone H3 were determined using western blot. Gene expression levels of microtubule-associated protein 1 light chain 3b (lc3b) were determined using quantitative reverse transcription-polymerase chain reaction. RESULTS: Butyrate-induced cell death occurred in a dose-dependent manner whether cells were starved or fed. However, the induction of cell death was two to four times higher when cells were placed under starvation conditions compared to when they were fed. Moreover, both starvation and butyrate exposure induced AMPK activity and autophagy. While AMPK inactivation resulted in decreased autophagy and butyrate-induced cell death under conditions of starvation, AMPK activation resulted in butyrate-induced cell death when cells were fed. Combined with the results of our previous report, which demonstrated butyrate-induced autophagy-dependent cell death, the results of this study suggest that the combination of starvation and butyrate exposure activates AMPK inducing autophagy and subsequent cell death. Notably, this combination markedly induced LC3B production and the induction was attenuated by AMPK inhibition. LC3B knockdown, in turn, significantly decreased butyrate-induced cell death. Therefore, AMPK-dependent LC3B induction apparently plays an important role in butyrate-induced cell death. There was a lack of correspondence between the levels of AMPK activation and LC3B induction; this may reflect the histone deacetylase-inhibitory capacity of butyrate on histone proteins. CONCLUSION: Taken together, starvation and butyrate exposure promote autophagy via AMPK signaling, while the histone deacetylase-inhibitory effects of butyrate alter chromatin to transcriptionally active state, resulting in strong LC3B induction and subsequent cell death. These findings may help improve the understanding of the cellular processes underlying periodontal disease initiation.

Does migraine-associated vertigo share a common pathophysiology with Meniere's disease? Study with vestibular-evoked myogenic potential.

To clarify if migraine-associated vertigo (MAV) and Meniere's disease (MD) share a common pathophysiology, vestibular-evoked myogenic potentials (VEMP) were measured in 11 patients with MAV, 11 with unilateral MD and eight healthy subjects. As acoustic stimuli, tone bursts (TB; 250, 500, 1000 and 2000 Hz) were presented. In healthy subjects, 500-Hz TB evoked the largest amplitude. To quantify this tendency, 500-1000 VEMP slope was calculated, and 500-1000 VEMP slope was the smallest on the affected side of MD patients. Among the 11 MD patients, five had significantly decreased 500-1000 VEMP asymmetry (shift of the tuning to 1000 Hz). Three of the 11 MAV patients also showed a significantly decreased 500-1000 VEMP slope. This finding suggests that MAV might share a common pathophysiology with MD. In addition to this finding, four of the other eight MAV patients showed prolonged p13 latencies. This suggests that MAV could consist of patients with different lesion sites.

The role of the superior vestibular nerve in generating ocular vestibular-evoked myogenic potentials to bone conducted vibration at Fz.

OBJECTIVE: The n10 component (n10) of the ocular vestibular evoked myogenic potential (oVEMP) to brief bone conducted vibration (BCV) of the forehead at Fz is probably caused by the vibration selectively activating vestibular otolithic receptors. If the n10 is due primarily to utricular activation then diseases which affect only the superior division of the vestibular nerve (SVN) should reduce or eliminate n10. METHODS: The n10 component of the oVEMP was measured in 13 patients with unilateral SVN but with inferior vestibular nerve function preserved. RESULTS: We compared the n10 to BCV of these 13 SVN patients to previously published data for healthy subjects and patients after complete unilateral vestibular loss. We found that in 12 out of the 13 patients with SVN, n10 was markedly reduced or absent under the contralesional eye. CONCLUSION: Since all utricular afferents course in the superior vestibular nerve and in 12/13 of these patients the n10 was reduced we conclude that the n10 component of the oVEMP to BCV is probably mediated by the superior vestibular nerve and probably due to activation of mainly utricular receptors. SIGNIFICANCE: The n10 appears to be a simple new test of superior vestibular nerve and probably mainly utricular function.

Myelin-deficiency in the cochlear nerve of the 'bt' mutant hamster.

In a previous report, we showed abnormal auditory evoked potentials in the mutant hamster, 'black tremor (bt)', with significantly prolonged wave latencies of auditory brainstem responses and prolonged N1 latencies of compound action potentials, but normal cochlear microphonics. In this report, we present the results of morphological studies supporting the results of our electrophysiological studies of the auditory pathway in bt. Observation by transmission electron microscopy revealed an abnormal myelin sheath surrounding the spiral ganglion cells, and a thinner compact myelin sheath surrounding the axons in bt than in normal hamsters. The bt hamster has a myelin deficiency not only in the brainstem, but also in the cochlear nerve.

Vestibular evoked myogenic potentials in patients with idiopathic bilateral vestibulopathy. Report of three cases.

Idiopathic bilateral vestibulopathy (IBV) is an acquired bilateral peripheral vestibular disorder of unknown cause. Three patients diagnosed as IBV by neuro-otological examination were reported. They underwent vestibular evoked myogenic potential (VEMP) testing which reflects the functionality of the sacculo-collic pathway. As a result, 2 of the 3 patients showed bilateral absence of VEMPs and one showed unilateral absence. The VEMPs of the 3 patients revealed that IBV affects not only the superior but also the inferior vestibular nerve systems. As previously reported in the cases of vestibular neuritis, VEMP could be useful for classifying IBV according to the function of the inferior vestibular nerve.

Click- and short tone burst-evoked myogenic potentials in cerebellopontine angle tumors.

We report results of vestibular-evoked myogenic potentials (VEMPs) in patients with cerebellopontine angle tumors and compare results obtained using clicks with those obtained using 500 Hz short tone bursts (STB). We reviewed the records of 87 patients with cerebellopontine angle tumors. Clicks (0.1 ms, 95 dB nHL) were presented to all patients and STB (500 Hz, rise/fall time 1 ms, plateau time 2 ms, 95 dB nHL) were presented to 27 patients. Click-evoked VEMPs were abnormal in 69/87 patients (79%; no response in 55 patients, decreased response in 14 patients, normal response in 18 patients). STB-evoked VEMPs were abnormal in 22/27 patients (82%; no response in 18 patients, decreased response in 4 patients, normal response in 5 patients). Click- and STB-evoked VEMPs were identical in 23/27 patients (85%). Two patients showed normal STB-evoked VEMPs and decreased click- evoked VEMPs, and 2 patients showed decreased STB-evoked VEMPs and absent click- evoked VEMPs. These results confirm our previous study in a small number of patients. Vestibular afferents seem to respond better to 500 Hz STBs than to clicks.

The outcome of the canalith repositioning procedure for benign paroxysmal positional vertigo: are there any characteristic features of treatment failure cases?

To demonstrate the success rate of the canalith repositioning procedure (CRP) in our clinic and to establish any characteristic features of cases of treatment failure, we reviewed clinical records of 62 patients who were diagnosed with posterior semicircular canal-benign paroxysmal positional vertigo (BPPV) and treated with the CRP. The basic strategy of the CRP was to rotate the involved canal slowly in the plane of gravity so that free-floating materials could migrate into the utricle only once. After the procedure we instructed patients to keep their heads upright for 10 h and not to sleep on the affected ear for 2 weeks. After the initial treatment, successful results were obtained in 51 of the 62 patients (82.2%). After the second treatment, 56 patients (90.3%) experienced success. Six patients (9.7%) did not obtain resolution even after the second treatment. While 46 patients were diagnosed with idiopathic BPPV, in 16 patients a different diagnosis was determined (head injury in 7 patients, Ménière's disease in 2, vestibular neuritis in 2 and unilateral sensorineural hearing loss in 5). We categorized these 16 patients as having secondary BPPV. Patients with idiopathic BPPV showed a significantly higher success rate with CRP than those with secondary BPPV. Patients with secondary BPPV may have quantitatively or qualitatively different lesions than those with idiopathic BPPV.

The effect of rise/fall time on vestibular-evoked myogenic potential triggered by short tone bursts.

Vestibular-evoked myogenic potentials (VEMPs) are used more and more frequently as a clinical tool to test if the sacculocollic reflexes are intact in patients. They can be evoked by short tone bursts (STBs) as well as by clicks. Although most previous studies used traditional clicks to generate VEMP responses, our clinical experience showed that STBs were prone to evoke them more clearly, at least in some patients. Four kinds of STB stimulation patterns in a random order were used to test 22 ears using changing rise/fall times (0.3, 1, 3 and 10 ms). VEMP responses (p13/n23) triggered by these patterns were clearly observed in all 22 ears. When the rise/fall time was prolonged from 0.3 to 10 ms, the p13 latency was prolonged in parallel. There was a similar trend for the n23 latency, although a significant difference was not attained between 0.3 ms and 1 ms rise/fall times. Considering the p13 and n23 latencies for the 4 rise/fall times, the variances were smallest for the 1 ms stimulation, meaning that it caused the smallest interaural latency differences. The amplitude or relative amplitude in the individual ears tested was lowest for the 10 ms stimulation, being comparable among the other 3 rise/fall times. In conclusion, the 1 ms rise/fall time was a remarkable stimulation pattern because its VEMP responses were simultaneously more constant and conspicuous.

Diagnostic value of prolonged latencies in the vestibular evoked myogenic potential.

BACKGROUND: As a parameter for the evaluation of the vestibular evoked myogenic potential (VEMP), amplitude has been used clinically. However, the significance of latency has not been considered. OBJECTIVE: To clarify the diagnostic value of latencies of the VEMP. DESIGN: We reviewed records of the VEMP of patients with various diseases and compared them with records of healthy volunteers. SETTING: Data were collected from patients in an outpatient clinic of a tertiary care center and healthy volunteers. SUBJECTS: Clinical records of 134 patients (61 men and 73 women, aged 20-75 years) were reviewed. Diagnoses were Meniere disease in 43 patients, acoustic neuroma in 62 patients, vestibular neuritis in 23 patients, and multiple sclerosis in 6 patients. Also, 18 healthy volunteers (13 men and 5 women, aged 25-38 years) were enrolled. INTERVENTION: Diagnostic. MAIN OUTCOME MEASURES: Click-evoked myogenic potentials were recorded with surface electrodes over each sternocleidomastoid muscle. Latencies and amplitudes of responses were measured. RESULTS: Vestibular evoked myogenic potentials were absent or decreased in 51% of patients with Meniere disease (n = 22), 39% with vestibular neuritis (n = 9), 77% with acoustic neuroma (n = 48), and 25% with multiple sclerosis (3 of 12 sides of 6 patients). Concerning latency, patients with Meniere disease or vestibular neuritis hardly showed any latency prolongation. Four patients with acoustic neuroma showed prolonged p13; all had large tumors. All patients with multiple sclerosis showed prolonged p13. CONCLUSION: Prolonged latencies of the VEMP suggest lesions in the retrolabyrinthine, especially in the vestibulospinal tract.

Glycerol affects vestibular evoked myogenic potentials in Meniere's disease.

OBJECTIVES: to show that abnormal vestibular evoked myogenic potentials on the sternocleidomastoid muscle (SCM) in patients with unilateral Meniere's disease are caused by endolymphatic hydrops. SUBJECTS: six normal volunteers and 17 patients with unilateral Meniere's disease were examined. METHODS: click-evoked myogenic potentials were recorded with surface electrodes over each SCM. Responses evoked by clicks recorded after oral administration of glycerol (1.3 g/kg body weight) were compared with those recorded before administration. RESULTS: the change rate of the p13-n23 amplitude was calculated. The mean+standard deviation (S.D.) of the change rate was 3.52+14.6% in normal subjects. On the unaffected side of patients the change rates were within the normal range (within the mean+/-2S.D.) in 13 patients, and three ears showed significant decrease. Only one ear showed significant increase. On the affected side, five ears showed significant increase of the amplitude while two ears showed significant decrease after oral administration of glycerol. Effects on evoked myogenic potentials were independent of those on pure tone hearing. CONCLUSION: vestibular evoked myogenic potentials in some patients with unilateral Meniere's disease were improved by oral administration of glycerol. This result suggests that abnormal vestibular evoked myogenic potentials in patients with unilateral Meniere's disease could result from endolymphatic hydrops.

Sensorineural hearing loss as the initial manifestation of polyarteritis nodosa.

A 74-year-old male was referred for the sudden onset of bilateral sudden deafness. The patient had no history of any disease or trauma to the head. Pure tone audiometry revealed bilateral moderate, to severe, sensorineural hearing loss. Auditory brain stem responses (ABRs) showed normal peak and interpeak latencies. These audiological findings suggested that his hearing loss could be attributed to inner ear lesions. However, we felt an alternative explanation for this sudden deafness was likely to exist because the patient also had a month-long fever of unknown origin (FUO) and weight loss of 5 kg/month. Using the criteria of The American College of Rheumatology, we made the diagnosis of polyarteritis nodosa (PAN). Serum MPO-ANCA was positive (x 661). For treatment, the patient was begun on prednisolone and cyclophosphamide. Nine months later, fever, hypertension, nephritis, pneumonitis, and arthritis had completely resolved, the MPO-ANCA became negative (MPO-ANCA < x 10). Furthermore, his hearing improved.

Narrow internal auditory meatus: an idiopathic case confirming the origin and pathway of vestibular evoked myogenic potentials in humans.

OBJECTIVE: To confirm the origin and pathway of vestibular evoked myogenic potentials (VEMPs) in humans. DESIGN: Case study. SETTING: University hospital. PATIENT: A patient with a narrow internal auditory meatus (IAM). MAIN OUTCOME MEASURES: Imaging studies and functional studies concerning the seventh and eighth cranial nerves. RESULTS: Of the 4 nerves in the IAM, all but the cochlear nerve had normal function and normal courses, despite the pronounced narrowing of the IAM. The facial nerve had a normal diameter, but the vestibular nerves were thinner. Imaging revealed that the cochlear nerve was absent or extremely thinned. Both the cochlea and the cochlear nerve showed no function in the affected ear, although the VEMPs were evoked normally. CONCLUSION: Our results definitively support the vestibular origin of VEMPs in humans.

The effect of sternocleidomastoid electrode location on vestibular evoked myogenic potential.

OBJECTIVE: The purpose of this study was to investigate the effect of a sternocleidomastoid (SCM) electrode array on the vestibular evoked myogenic potential (VEMP) and the most optimal recording site for clinical use. METHODS: Fifteen normal adults (10 men and 5 women, aged 18 to 38 years) were tested. We placed electrodes at four different locations over the SCM muscle: the upper part of the SCM muscle at the level of mandibular angle, the middle part of the muscle, and immediately above sternal and clavicular origins of the SCM muscle. Sound evoked myogenic potentials in response to monoaurally delivered short tone-bursts (500 Hz at 95 dBnHL, rise/fall time=1 ms and plateau=2 ms) were recorded with surface electrodes over the isometrically contracting SCM muscle. RESULTS: On the clavicle, the upper and middle parts of SCM from all subjects, air-conducted short tone burst evoked biphasic responses (p13-n23). VEMPs recorded at the upper part of the muscle showed the largest amplitude, followed by that at the middle part. However, the latency of the first peaks (p13-n23) was not constant in the upper part. Recording from the middle part of SCM muscle were more consistent. CONCLUSION: Our findings suggest that the middle part of the SCM muscle is the optimal location for recording vestibular evoked myogenic potential.

The effects of plateau time on vestibular-evoked myogenic potentials triggered by tone bursts.

Vestibular-evoked myogenic potentials (VEMPs) can be triggered by acoustic, vibratory or galvanic stimuli. However, each method has drawbacks for studying if the vestibulocollic reflex is intact in the patients tested. We used air-conducted VEMPs as a screening test to examine the integrity of the sacculocollic reflex. In a previous study, we defined the optimal rise/fall time of short tone bursts (STBs) to evoke VEMPs. In this paper, we studied the optimal plateau time of tone bursts to evoke VEMPs. Four different plateau times (1, 2, 5 and 10 ms) were used in a random order to test 26 normal ears. VEMP responses (p13/n23) triggered by the tone bursts were clearly observed in all ears. When the plateau time was increased in order from 1 to 10 ms, the latencies (p13, n23) and interval (p13-n23) were also increased in parallel, although significant differences were not observed between some plateau times. Considering the latencies and interval together for the four plateau times, the variances were smallest for the 2 ms plateau time, meaning that it caused the smallest interaural VEMP differences. The amplitude or relative amplitude in individual ears was lowest for the 1 ms plateau time, while it was comparable for the other three plateau times. In conclusion, we recommend that the ideal stimulation pattern for evoking STB VEMPs is as follows: frequency 500 Hz; stimulation repetition rate 5 Hz; rise, fall time 1 ms; and plateau time 2 ms. The waveform morphology of the VEMP responses observed with this stimulation pattern was simultaneously the most constant and marked.

Bone-conducted evoked myogenic potentials from the sternocleidomastoid muscle.

The aim of this study was to show that bone-conducted clicks and short tone bursts (STBs) can evoke myogenic potentials from the sternocleidomastoid muscle (SCM) and that these responses are of vestibular origin. Evoked potential responses to bone-conducted auditory stimuli were recorded from the SCMs of 20 normal volunteers and from 12 patients with well-defined lesions of the middle or inner ear or the VIIIth cranial nerve. The subjects, who had various labyrinthine and retro-labyrinthine pathologies, included five patients with bilateral profound conductive hearing loss, two with bilateral acoustic neuroma post-total neurectomy and five with bilateral sensorineural hearing loss. Air- and bone-conducted evoked myogenic potentials in response to clicks and STBs were recorded with surface electrodes over each SCM of each subject. In normal subjects, bone- and air-conducted clicks and STBs evoked biphasic responses from the SCM ipsilateral to the stimulated ear. The bone-conducted clicks evoked short-latency vestibular-evoked myogenic potential (VEMP) responses only in young subjects or in subjects with conductive hearing loss. STBs evoked VEMPs with higher amplitude and better waveform morphology than clicks with the same acoustic intensity. Patients with total VIIIth cranial nerve neurectomy showed no responses to air- or bone-conducted click or STB stimuli. Clear VEMP responses were evoked from patients with conductive or sensorineural hearing loss. It is concluded that loud auditory stimuli delivered by bone- as well as air conduction can evoke myogenic potentials from the SCM. These responses seem to be of vestibular origin.

Isolated directional preponderance of caloric nystagmus: I. Clinical significance.

OBJECTIVES: To determine the clinical significance of an isolated directional preponderance (DP) on bithermal caloric testing. An isolated caloric DP was defined as a DP, calculated according to the standard Jongkees formula, of > or = 40%, with a spontaneous nystagmus (SN) in darkness of < or = 2 degrees/s and a canal paresis (unilateral weakness) of < or = 25%. STUDY DESIGN: A retrospective analysis of all 15,542 bithermal caloric tests performed in the authors' department in the previous 10 years to identify all tests with an isolated DP of > or = 40%. This was followed by a review of the clinical data on the 144 patients identified with such a result and then by a telephone or postal follow-up study of these patients. The study group eventually comprised 114 patients; these were patients in whom a clinical diagnosis could be made at the time the caloric test was done, or who responded to requests for follow-up information. The 34 patients in whom a clinical diagnosis could not be made at the time of the caloric test, and who did not respond to requests for follow-up information, were excluded. STUDY SETTING: A balance disorders clinic in a tertiary referral hospital. INTERVENTION: All patients underwent standard bithermal caloric testing. Some of the patients also underwent rotational testing. OUTCOME MEASURES: A clinical diagnosis for the cause of the isolated DP, made either at the time of the caloric test or on the basis of information supplied at follow-up by the patient or by the referring physician. RESULTS: Of 114 patients, 39 had benign paroxysmal positioning vertigo, 14 had Ménière's disease, and 5 had migrainous vertigo. Five patients had central nervous system (CNS) disorders, and this was clinically apparent at the time of the caloric test in 4, so that only 1 patient with an isolated DP developed evidence of a CNS disorder after the caloric test was done. In the other 54 patients, no definite diagnosis could be made, but 41 of these 54 were either completely well or much better at follow-up. CONCLUSIONS: An isolated DP on caloric testing is usually a transient, benign disorder. About half the patients with an isolated DP have either Ménière's disease or benign paroxysmal positioning vertigo; in most of the other half, no definite diagnosis is made but most of these patients will do well. Only approximately 5% have a CNS lesion and in almost all this is apparent at the time the caloric test is done. In a relapsing-remitting peripheral vestibular disorder such as benign paroxysmal positioning vertigo or Ménière's disease, the mechanism of an isolated DP could be enhanced dynamic gain of ipsilesional medial vestibular nucleus neurons, perhaps as a result of intermittent hyperfunction of primary semicircular canal vestibular afferents. The authors postulate that an isolated DP reflects a gain asymmetry between neurons in the medial vestibular nucleus on either side, caused either by increased sensitivity on one side or by reduced sensitivity on the other, perhaps as an adaptive change in response to abnormal input. In an accompanying article, the authors implement a realistic neural network model in which it is possible to simulate an isolated DP by adjusting the dynamic sensitivity of type 1 medial vestibular nucleus neurons on one side or of type 2 medial vestibular nucleus neurons on the other.

Periodic alternating nystagmus and congenital nystagmus: similarities in possibly inherited cases.

In this paper, 2 patients, a daughter with periodic alternating nystagmus and a mother with congenital fixation nystagmus, are presented, and the similarities of the two disorders are discussed, not only in the eye movements, but also in their underlying abnormalities. The literature was reviewed, and the link between periodic alternating and congenital fixation nystagmus, which had been suggested based on circumstantial evidence, may be confirmed by our cases. The possible evidence of inheritance suggests that the two disorders share common abnormalities in visual processing.

Vestibular evoked myogenic potentials in patients with spinocerebellar degeneration.

To estimate vestibulo-collic reflexes in patients with spinocerebellar degeneration (SCD), vestibular evoked myogenic potentials (VEMPs) were recorded in 16 patients with SCD and in 9 normal subjects. The patients with SCD were classified into three groups: those with olivo-ponto-cerebellar ataxia (OPCA; n = 10); those with cortical cerebellar atrophy (CCA; n = 3); and those with Machado-Joseph disease (MJD; n = 3). While all of the patients with OPCA and CCA showed biphasic responses (p13-n23), 2 of the 3 patients with MJD showed abnormal VEMPs bilaterally. Three sides in these 2 patients showed an absence of VEMPs, and one side showed a remarkably delayed p13. These two patients showed little response in caloric tests. These results suggest that the vestibulo-collic reflex as well as the vestibulo-ocular reflex could be damaged in MJD patients but preserved in OPCA and CCA patients.

Vestibular function in auditory neuropathy.

Auditory neuropathy is characterized by mild-to-moderate pure-tone hearing loss, poor speech discrimination out of proportion with this loss, absent or abnormal auditory brainstem responses and normal outer hair cell function as measured by otoacoustic emissions and cochlear microphonics. We followed three patients in our clinic whom we classified as auditory neuropathy patients. These patients also complained of balance disorders and we report our auditory and vestibular system analyses of these patients. The data presented herein include results of audiometric tests (serial pure-tone audiometry and speech discrimination tests), otoacoustic emissions, auditory-evoked brainstem responses and vestibular function tests (clinical tests of balance, electronystagmography, damped rotation tests and vestibular-evoked myogenic potentials). In all patients, pure-tone audiometry revealed mild-to-moderate sensorineural hearing loss, markedly poor speech discrimination scores and absent auditory-evoked brainstem responses, all in the presence of normal otoacoustic emissions. Balance tests (caloric tests and damped rotation test) were abnormal. Saccades, smooth pursuit eye movements and optokinetic nystagmus were normal in all patients. Neurological and motor system evaluations were normal in all patients. These three auditory neuropathy patients manifest a disorder of cochlear nerve function in the presence of normal outer hair cell activity. They additionally manifest a disorder of the vestibular nerve and its end organs. We conclude that, in patients with isolated auditory neuropathy, the vestibular branch of the VIIIth cranial nerve and its innervated structures may also be affected. We suggest the use of the term "cochlear neuropathy" to characterize those patients with involvement of only the auditory branch of the VIIIth cranial nerve and its innervation.