Mechanosensory units in the hypoglossal nerve of the cat

BRAIN RESEARCH

349

M E C H A N O S E N S O R Y UNITS IN T H E H Y P O G L O S S A L NERVE OF T H E CAT

P. ZAPATA ANDG. TORREALBA* Departamento de Neurofisiologla, Universidad Catdlica de Chile, Santiago (Chile)

(Accepted March 15th, 1971)

INTRODUCTION The presence of sensory fibers from the tongue surface, both tactile and gustatory, has been extensively described in chorda-lingual and glossopharyngeal nerves. However, the existence of proprioceptive fibers from the tongue has not been accepted because only hypoglossal nerve fibers are distributed to tongue muscles, and the XII nerve does not have dorsal roots. Furthermore, there is general agreement that muscle spindles do not occur in the intrinsic or extrinsic musculature of the cat's tongue4,7 -9,26. Moreover, several authors have been unable to record sensorydischarges from either the entire trunk or from dissected filaments of the hypoglossal nerve of the cat3,4,11,19. However,CooperI0and Blom4have reported that afferentimpulsescan be recorded from filamentsof the medialend-branch of the hypoglossalnerve supplying the intrinsic tongue muscles of the cat; but these authors agree that such filaments could be anastomoticbranches from the lingual nerve; in fact, extensiveperipheral connections between hypoglossaland lingual nerves have been described14. The above observations do not extend to humans and monkeys. In these species, muscle spindles have been found in the tongue5,9, and stretch receptor discharges in the hypoglossal nerve have been reported6. Such a marked difference between primates and carnivora prompted us to re-studythe problem in the cat. A preliminary report of these findingshas been made27. METHODS Experiments were performed on adult cats, anesthetized with intraperitoneal injections of pentobarbital (Nembutal, Abbott) 36 mg/kg. Animals were tracheotomized, and the anesthesia was maintained through small doses of pentobarbital administered intravenously. * Postdoctoral trainee, Facultad de Medicina, Universidad Cat61ica de Chile. Brain Research, 32 (1971) 349-367

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The ventral side of the neck was opened by a midline longitudinal incision, and both hypoglossal nerves were exposed for dissection from their exit from the cranium to their terminal branches into the different muscles of the tongue. The dissected nerves were kept in a warm paraffin pool. Excision of the perineurial sheaths and splitting of nerves into fine filaments were performed under a stereomicroscope. For recording, nerves were laid on a pair of fine silver wire electrodes, and the potentials fed to a Tektronix 565 multiple-beam oscilloscope through an AC preamplifier and a Belclere EN-2089 transformer to increase the signal-to-noise ratio 20. The potentials displayed on the oscilloscope screen were photographed, usually on moving film. In addition, the horizontal plates of the oscilloscope were connected to an audiomonitor and to an electronic counter (Hewlett-Packard 5223L) and printer ( H - P 562A) which recorded the number of impulses per second once every second. The tongue was stretched through a thread sutured to its tip. The other end of the thread was attached either to a micromanipulator or to a force-displacement transducer, connected to the second beam of the oscilloscope. The jaws were kept open with a retractor and the canines were excised to avoid contact of the tongue with nearby structures while being stretched. Lateral stretches of the tongue were exerted through threads sutured symmetrically to its lateral borders. Nerve stimulation was performed through a pair of silver electrodes connected to a stimulator and stimulus isolation unit. In a few experiments, the afferent activities of the lingual, glossopharyngeal or superior laryngeal nerves were simultaneously recorded. RESULTS

Extracranial distribution of the hypoglossal nerve A few millimeters after its exit from the cranium, in the submaxillary region, the hypoglossal nerve gives one or two branches ending in the nodose ganglion of the vagus nerve; this anastomosis can be very short but may attain a length of 10-15 mm; this fine nerve filament(s) will be called the 'hypoglosso-nodosal branch' (Fig. 1). Its external diameter is approximately one-fifth that of the main trunk and its perineurium is easily detached. This allows dissections of single fibers with relative ease. Immediately distal to the emergence of the hypoglosso-nodosal branch(es), another branch arises perpendicularly or tangentially to the main trunk of the hypoglossus. Stimulation of this branch produced contraction of the thyrohyoid muscle and corresponded to the 'ramus descendens hypoglossi' commonly described. At approximately 0.5-1.0 cm from the main trunk, the ramus is joined by a fine ascending branch from the nodose ganglion (see Fig. 1). If the mylohyoid muscle is transected and reflected laterally, the 5 terminal branches of the hypoglossal nerve can be studied. In the present paper they will be called: lateral, centro-lateral, central, centro-medial and medial branches. The lateral end-branch is the first to emerge, in a perpendicular way directed to the underlying muscles of the posterior part of the tongue; it is of median calibre. The central endbranch follows the course of the main trunk; it is of large diameter (about two-thirds Brain Research, 32 (1971) 349-367

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Fig. 1. Branching of the hypoglossal nerve (XII) in the cat. X, vagus nerve; HC, hypoglossal canal; HNB, hypoglosso-nodosal branch; JF, jugular foramen; NG, nodose ganglion; RDH, ramus descendens hypoglossi; TH, thyrohyoid muscle.

that of the principal trunk) and is directed toward the tip of the tongue to innervate the intrinsic muscles. The centro-lateral end-branch is a median calibre filament, initially running adjacent to the central end-branch and then diverging more laterally. The medial and centro-medial branches are delicate filaments following a tortuous course and oriented towards the tongue midline. Most papers referring to the innervation of the mammalian tongue describe only a small lateral and a much larger medial divisionX, 14, but they also mention preterminal branches and collaterals. We have had to introduce a new nomenclature based on 5 terminal branches, since this was an ever present picture in our observations and was demanded because different patterns of movements were evoked by electrical stimulation of each of these end-branches, as will be reported elsewhere.

Recordingfrom hypoglossalend-branches The hypoglossus was sectioned at the level where it runs parallel to the lingual artery. Peripheral to this section, the different end-branches of the hypoglossal nerve Brain Research, 32 (1971) 349-367

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Fig. 2. Continuous recordings from a fine strand of the lateral end-branch of the hypoglossal nerve. Between dcts, 20 mm passive protrusion of the tongue. Time, 1 sec. were dissected and prepared for recording under resting conditions and during displacements of the tongue. Sensory discharges were recorded from all 5 terminal branches of the hypoglossal nerve. However, mechanosensory activity was easily detected in the centro-lateral branch, whereas it was difficult to find in the centro-medial and medial branches. Fig. 2 illustrates the electrical activity recorded from a fine filament dissected from the centro-lateral end-branch of the hypoglossal nerve. A fairly constant discharge was observed at rest (A), showing a marked and maintained increase in frequency when the tongue was longitudinally pulled out of the mouth (A-C). A prominent 'silent period' occurred immediately after mechanical stimulation was interrupted (C), followed by a progressive increase in frequency (D) until the resting level of activity was attained (E). Longitudinal stretching of the tongue was the most effective stimulus for evoking an increase in the frequency of discharge, although lateral displacements were also effective. When the tongue was subjected to a moderate degree of protrusion, some sensory units showed respiratory rhythmicity when the anesthetic level was such as to produce slow but deep respiration. These units showed a slight increase in its discharge during inspiration. In these circumstances, an increase in longitudinal tension of the tongue was observed through a suitable mechanical transducer attached to its tip. Respiratory displacements of the tongue and hyoid bone in humans have been observed by Mitchinson and Yoffey is and in rats by AndrewL

Recordings from the hypoglossal main trunk The presence of afferent fibers responsive to tongue stretch in the main trunk of

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Fig. 3. Recordings from peripheral end of cut hypoglosso-nodosal branch. A and B, one experiment, resting activity and response to passive protrusion of the tongue (between dots). C and D, another experiment, resting activity (C) and maximal response (D) to maintained stretch of the tongue. Time, 0.5 sec. the hypoglossus in its traject ventral to the carotid and lingual arteries was also observed in our experiments. The recording of afferent activity from the entire nerve is difficult in this extremely gross segment, mainly composed of afferent fibers and covered by a thick perineurium difficult to detach. However, occasionally it was possible to obtain well-defined sensory discharges after dissection of fine filaments (see below and Fig. 4). We have no adequate information concerning the presence of afferent fibers in Brain Research, 32 (1971) 349-367

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Fig. 4. Discharge frequencies of successive recordings from different segments of the same hypoglossal nerve. The nerve was sectioned immediately after its exit from the cranium, and the ramus descendens was also cut. A, recording from the hypoglosso-nodosal branch sectioned at its entrance to the nodose ganglion; B, recording from the mid-portion; C, recording from the lateral end-branch. Horizontal bars, longitudinal stretching of the tongue through a thread attached to its tip. In A, the tongue displacement produced by a second pull was approximately twice that prodtamd by the first, and the third was about twice the second; the intcmity of this ~ r pull was that ~ inB and C. Ordinates, frequency of sensory discharges in impulse/see. Aba~fat¢, t ~ , the ~ e n t at the end of B corresponding to 10 sec.

the hypoglossal nerve central to the hypoglosso-nodosal branch, since its traject from this point to its emergence from the cranium is too short and no recordings are avail. able from the intracranial roots of the nerve.

Recordings from the hypoglosso-nodosal branch Recording o f afferent activity from the hypoglosso-nodosal branch is relatively easy when this fine nerve has adequate length for recording. Fig. 3 illustrates two o f these cases. In A and B an aperiodic discharge o f low voltage spikes was recorded at rest; during passive protrusion o f the tongue (between dots) there was an increase in the frequency o f these discharges and the appearance o f high voltage spikes. In C and D, obtained f r o m a different experiment, the activity o f a dissected fine filament containing two active units is shown; one o f these units had a regular rate at rest (C) and its frequency increased during forced protrusion o f the tongue (D). Fig. 4 illustrates frequency changes o f sensory discharges recorded successively

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SENSORY FIBERS IN THE HYPOGLOSSAL NERVE

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from different segments of the hypoglossal nerve of the same preparation, at rest, and during longitudinal displacements of the tongue. Electrodes were first placed on the hypoglosso-nodosal branch and the responses to slight, moderate and stronger pulls are illustrated (A); the increases in frequency of sensory units correlate well with the intensities of the mechanical stimuli applied. Then, a fine filament was dissected from the mid-portion of the hypoglossal nerve, peripherally to the emergence of the hypoglosso-nodosal branch but central to the end-branching of the nerve, and changes in frequency were studied (B). Finally, responses from a filament of the lateral endbranch were studied (C); units recorded in this instance showed more rapid adaptation than usual. These recordings show that mechanoreceptor fibers from the tongue are traceable from the hypoglossal end-branches, through the hypoglossal main trunk to the hypoglosso-nodosal branch. The deviation of fibers from the hypoglossus to the vagus nerve through the hypoglosso-nodosal anastomoses does not imply that all mechanoreceptor fibers recorded peripherally enter the cranium through the X nerve. It is possible that some of them make their way through the hypoglossal canal. Most mechanoreceptor units recorded from the hypoglossal end-branches or the hypoglosso-nodosal branch had a basal frequency of discharge when the tongue was at rest; this frequency increased when the tongue was displaced (Figs. 2, 3C-D, 6). Other units were silent at rest but discharged when the tongue was displaced; they became silent when the mechanical stimulus was interrupted (Figs. 3A-B, 9C-F). A few units showed a resting basal discharge but they were silenced during displacements; immediately after interruption of the stimulus they showed a high frequency discharge which returned to the basal activity in a few seconds (Fig. 9A-B). In rhesus monkeys, Bowman and Combs 6 observed that all tongue spindles exhibited a basal spontaneous discharge frequency. When stretch was applied, the units either increased or decreased their discharge. The patterns described by these authors correspond to the first and third group of units described above. Our second group, silent at rest, was not reported by these authors. Since the muscle fibers of the mammalian tongue are oriented longitudinally, vertically or transversally, the entire organ is subjected to combined distortions or displacements in the 3 directions. The possibility arises that the mechanoreceptor endings of the fibers here recorded are specially oriented to detect changes in tension or length in a preferential direction. Fig. 5 illustrates the changes in frequency discharge of a filament of the hypoglosso-nodosal branch. In Fig. 5A the tongue had no longitudinal traction and the displacement of the tongue to the contralateral side (right) was less effective in producing an increase in sensory discharge than the same displacement to the ipsilateral side (left). In Fig. 5B the tongue was subjected to a maintained degree of longitudinal stretch to obtain a maintained higher level of basal discharges; in this condition, ipsilateral displacements were still more effective than contralateral ones for the fibers recorded from this filament. When the directional sensitivity of different units to longitudinal and transversal displacement was studied, it was found that longitudinal stretch was more effective than lateral displacement. With regard to the latter, displacement in one direction was more effective than in the other. Similar findings have been reported by Brain Research, 32 (1971) 349-367

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Fig. 5. Discharge frequencies of recordings from the left hypoglosso-nodosal branch, sectioned at its entrance to the nodose ganglion. The hypoglossal nerve had been sectioned at its exit from the cranium. At A, the tongue was subjected to minimal longitudinal stretch through a thread attached to its tip; at B, the tongue was stretched 10 mm more than in A. Horizontal bars, lateral stretch through threads attached symmetrically to lateral borders of the tongue at the level of its midportion; lateral pulls were produced by 10 mm displacements of the borders to the right (R) or to the left (L) sides. Ordinates, sensory frequency in impulses/sec. Abscissae, each segment corresponding to 10 sec.

Bowman and Combs 6 for proprioceptive fibers in the monkey's tongue. This similarity in response patterns of hypogtossal mechanoreceptor fibers in monkeys and cats must be kept in mind, even though neuromuscular spindles with anulospiral receptors have been observed in the monkey's tongue but seem to be absent in cats (see Introduction). In several of the illustrations already presented, a transient decrease of frequency discharge below the initial baseline follows stimulus discontinuation; in some of them the initial increase of discharge upon mechanical stimulation overshoots the following frequency while the stimulus is still applied. In order to elucidate the importance of the Brain Research, 32 (1971) 349-367

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abruptness of stimulus application and interruption upon 'on' and 'off' discharges, some observations were specially intended to test this factor. Fig. 6 illustrates the discharges obtained from a fine filament of the hypoglosso-nodosal branch in which a single active fiber was periodically discharging in resting conditions, but with a rate slightly increased during inspiratory phases and with a very short silent pause at the end of each of these inspiratory phases. In Fig. 6A, a passive protrusion of the tongue applied in about 1 sec produced a positive acceleration of the sensory discharge. This positive acceleration is more pronounced in E where the stimulus was applied in less than one-fifth of a second, and poorly represented in C where the stimulus was applied in more than 2 sec. The tonic rates of discharge observed while the stimulus was maintained unchanged were usually lower than those attained at the beginning of Brain Research, 32 (1971) 349-367

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Fig. 7. Relation of frequency of sensory discharges in the hypoglosso-nodosal branch to the degree of stretch of the tongue. Longitudinal stretch of the tongue was increased in 5 mm steps from an arbitrary basal tension (O). Each dot is an average of counts during 40 consecutive sec, the initial 10 sec after stimulus application being discarded. Ordinate, average frequency in imp./sec. Abscissa, stretch in mm.

stimulus application. When mechanical stimulation was abruptly interrupted a welldefined silent pause was observed, but when the stimulus was progressively diminished, the silent period was replaced by a progressive decrease in sensory discharges until the baseline rate was re-established. We can conclude from these observations that the rate o f sensory discharge not only signals the intensity o f mechanical stimulation, but also the time course of application and interruption of the stimulus. Fig. 7 gives the relationship between intensity o f mechanical stimulation applied to the tongue and frequency o f sensory discharges recorded from the hypoglosso-nodosal branch. Stretch was progressively applied in 5 m m steps from a point in which the tongue was subjected to no external tension to a maximal longitudinal traction o f 45 m m o f its tip. The l0 sec immediately following an increase in mechanical stimulation were ignored in order to discount the acceleration of discharges seen at 'on'. Then sensory discharges were counted electronically for 40 sec and the average frequency was determined. The curve shows that the discharge frequency was directly related to the intensity of stretch. In order to obtain information on the adaptation rate o f the sensory nerve terminals o f the hypoglosso-nodosal fibers, prolonged stimuli were employed. Fig. 8 Brain Research, 32 (1971) 349-367

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Fig. 8. Time course of sensory discharges in the hypoglosso-nodosal branch during prolonged stretch of the tongue. The circle with vertical bars indicates the average and standard deviation for 15 counts of frequency of discharges immediately preceding the stimulus. At the arrow, a 20 m m stretch was longitudinally applied to the tongue through a thread attached to its tip. Each dot represents 1 count/sec during the first 60 see of stimulation and thereafter it corresponds to an average of counts in 10 sec. Ordinate, frequency of sensory discharges. Abscissa, time of stimulation in seconds, expressed on a logarithmic scale.

illustrates the frequency changes of a preparation in response to such stimulus. The average value and standard deviation for resting conditions were 55.86 -4- 3.53 imp./ sec. Then a 20 mm stretch was longitudinally applied and maintained unchanged for 6 min. The frequency of the sensory discharges increased to 113 impulses during the first second of stimulation, slowly declining to 96 at the tenth second, to 87 at the end of the first minute and to 77 at the end of the sixth minute. In these circumstances, a decay to one-half of the difference between resting conditions and initial maximal activation (i.e., 84 imp./sec) was attained after approximately 2 min of maintained stimulation. Undoubtedly, these receptor terminals must be considered as slowly adapting. The discharge pattern of sensory fibers in the hypoglossal nerve must be compared with that of mechanosensory fibers coursing in the chorda-lingual and glossopharyngeal nerves. A sudden stretch of the tongue of 15 mm or more produced a transient burst of spikes in these nerves lasting less than 1 sec; no discharge was obtained when this was done slowly. A longitudinal traction of 10 mm or less did not usually elicit sensory impulses in these nerves. A 50 g weight deposited on the anterior two-thirds of the tongue surface evoked a high frequency discharge in the lingual Brain Research, 32 (1971) 349-367

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nerve, which subsided in less than 2 sec; the same occurred for the glossopharyngeal nerve fibers when the weight was left on the posterior third of the tongue surface. Furthermore, short bursts of sensory discharges in lingual or glossopharyngeal nerves were evoked by gentle touching of the appropriate areas of the tongue surface with the tip of a human hair. No sensory discharges occurred in the hypoglossonodosal branch when the entire surface of the tongue was tested with the tip of" a plastic probe; only when the probe was deeply indented so as to produce displacements or changes in tension of deep structures (as detected by a transducer attached to the tongue tip), was a change in the rate of discharge of hypoglosso-nodosal fibers observed. We ruled out the possibility that mechanoreceptor fibers of the hypoglossonodosal branch could end on the larynx and upper trachea, being excited by tension changes transmitted to these structures from displacements of the tongue. Thus, stroking the laryngo-tracheal mucosa with a plastic probe, applying tension against the walls of these structures or displacing them did not induce changes in the frequency of sensory discharges recorded from the hypoglosso-nodosal branch. However, the afferent activity in the superior laryngeal nerve increased during application of these stimuli (see also Sampson and Eyzaguirre21). Furthermore, a longitudinal stretch of 35 mm or more was necessary to produce a pronounced upward displacement of the larynx and trachea and an increase in the sensory discharges of the superior laryngeal nerve.

Effects of tongue contractions upon hypoglosso-nodosal discharges The sensory discharges of the hypoglosso-nodosal fibers were modified by movements of the contralateral half of the tongue elicited through stimulation of the corresponding hypoglossal nerve. To study these effects one of the hypoglossal endbranches was stimulated by either single pulses or repetitively. The movements produced were recorded by a mechanical transducer while the afferent activity of the contralateral hypoglosso-nodosal branch was recorded. Electrical stimulation of the central end-branch of one hypoglossal nerve produced protrusion of the tongue and deviation of its tip to the contralateral side (see also Abd-el-Malekl; Torrealba and Zapata, unpublished observations). Recording from the contralateral hypoglosso-nodosal branch showed no change in its resting activity. It must be noted that this observation was done on a hypoglosso-nodosal preparation that gave a more pronounced response to ipsilateral than to contralateral displacement of the tongue. Stimulation of the lateral or centro-lateral end-branch of one hypoglossal nerve produced retraction and ipsilateral deviation of the tip of the tongue. Fig. 9 illustrates the afferent activity recorded from a small filament of the contralateral hypoglossonodosal branch. The tongue retraction elicited by single pulses applied to the hypoglossus produced an upward deflection in the second channel (connected to a transducer attached to the tongue tip). A single fiber discharged at rest (A). Each movement produced a silence (B) or a decrease in frequency of these discharges (C-F), with a

Brain Research, 32 (1971) 349-367

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Fig. 9. Recording from a fine strand of the peripheral end of the left hypoglosso-nodosal branch. Second beam connected to a pressure transducer attached to the tongue tip; upward deflections, retraction of the tongue produced by single pulses applied to the peripheral end of the right hypoglossal nerve. A, basal discharge; B and C, during hypoglossal stimulation at 2.5 V, producing slight retraction of the tongue; D-F, during hypoglossalstimulation at 5.0 V, producing tongue retraction with deviation to the stimulated side. Time, 100 msec.

transient increase during the relaxation phase. However, another fiber, silent at rest, showed a single (C) or a short burst ( D - F ) of high voltage spikes during the contraction phase. These observations show that the discharge of some hypoglosso-nodosal sensory fibers was affected by certain types of tongue movement elicited by stimulation of the contralateral hypoglossal nerve. It is possible that other sensory units may respond to movements that may not affect the fibers here recorded. The complex interlacing of fibers in the intrinsic and extrinsic muscles of the tongue makes possible different patterns of movements and they would affect sensory nerve organs placed and oriented in different planes of the tongue musculature. Fig. 10 illustrates an experiment in which spontaneous tongue movements occurred. In this experiment, the contralateral hypoglossal nerve was not sectioned and the level of anesthesia was light. When a 5 m m stretch was applied to the tongue tip, there was a maintained increase in the frequency of sensory impulses, but this was exceeded by a brief burst of discharges coinciding with a slight spontaneous retraction of the tongue (first arrow). When the stretch applied to the tongue was increased to 10 m m , a further moderate enhancement of sustained activity was observed, but Brain Research, 32 (1971) 349-367

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Fig. I0. Discharge frequency of aVmultiple-fiber preparation from the hypoglosso-nodosal branch, sectioned at its entrance to the nodose ganglion. The main trunk of the hypoglossal nerve had been sectioned at its entrance to the cranium. The contralateral hypoglosaat aerve was intact and the level of an~lahesia was s ~ c i a l during this part of the e x p e r ~ t . Undexl~r~ ~ , tonffu¢ streteh~l 5 ram; double bar, tongue stretched 10 mm. Arrows indicate spontaneous movements of the tongue consisting in retraction and deviation of the tip towards the opposite side of the recorded nerve. Ordinate, sensory frequency in impulses/sec. Abscissa, time in seconds.

again a short burst of high frequency discharges accompanied a new retraction of the tongue (second arrow). The level of anesthesia was then deepened by a small dose of intravenous pentobarbital, and the spontaneous movements of the tongue disappeared. In other experiments, tongue movements were avoided by sectioning the contralateral hypoglossal nerve at the beginning of the experiment. Recording from the rarnus descendens hypoglossi Because of the position of the muscles innervated by the ramus descendens, we assumed that if afferent fibers were present in this branch their discharges would be markedly influenced by tongue position and movements. We proceeded to a previous identification of the ramus descendens observing the contraction of infrahyoid muscles elicited by its electrical stimulation. After section of the branch, we recorded the afferent activity of its peripheral end. The ramus descendens showed a continuous afferent discharge under resting conditions; however, it had cyclic increases in frequency coincident with each inspiratory phase when the animal was breathing spontaneously. Through the dissecting microscope a downward movement of thyroid and cricoid cartilages and upper trachea was evident with each inspiration. We must note that both hypoglossi were cut, since in the intact animal an elevation of hyoid bone and thyroid cartilage is expected z,18. Fig. 11 illustrates the changes in frequency of sensory discharges recorded from a filament of the ramus descendens. A 20 m m longitudinal stretching of the tongue

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Fig. 11. Frequency of discharges recorded from the peripheral end of the ramus descendens hypoglossi, sectioned at its junction with the main trunk of the twelfth nerve; its nodosal origin is also severed. Underlying bars, longitudinal stretching of the tongue. Ordinate, frequency of sensory discharges in impulses/sec. Abscissa, time in seconds.

produced a clear and maintained increase in sensory discharges. This activity was also enhanced by pulling down the trachea with forceps, but only when the tongue was moderately stretched. Local pressure applied through a plastic probe between the thyroid cartilage and hyoid bone in order to separate them was also effective in producing an enhancement of sensory discharges. These observations show that there are afferent fibers in the ramus descendens hypoglossi; these fibers respond to mechanical stimuli and to passive protrusion of the tongue. The finding of these afferent fibers cannot explain the sensory discharges recorded from the hypoglossal nerve trunk or hypoglosso-nodosal anastomose since in those recordings the ramus had been previously sectioned. Recordings in decentralized hypoglossal nerves After 3-6 days of the intracranial section of one hypoglossal nerve performed extradurally at its entrance to the hypoglossal canal, the intrinsic and extrinsic muscles of the ipsilateral side of the tongue were unresponsive to hypoglossal nerve stimulation and showed fibrillary activity. Recording from its end-branches, the ramus descendens or the hypoglosso-nodosal branch showed afferent fibers responsive to tongue stretching. If hypoglossal sensitive fibers had their perikarya in the hypoglossal nuclei of the brain stem or along hypoglossal roots, they would have undergone degeneration and they would not be detected in these circumstances. However, these experiments do not rule out the possible existence of perikarya along the course of the nerve trunk through the hypoglossal canal. In those cats in which vagal rootlets had been sectioned intracranially at their entrance to the jugular foramen 3-5 days before, the recording of the ipsilateral hypoglossal end-branches, ramus descendens or hypoglosso-nodosal branch showed the presence of afferent fibers responding with an increase of their discharges to passive protrusion of the tongue. These experiments are indicative that the perikarya of these sensory fibers are not in the brain stem nuclei or intracranial roots of the Brain Research, 32 (1971) 349-367

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e. Z A P A T A A N D G. T O R R E A L B A

vagus; they suggest that their somata are located extracranially, probably in the nodose ganglion. DISCUSSION

The first objection to the presence of afferent fibers in the hypoglossal nerve in mammals came from the absence of dorsal roots in this nerve. The presence of sensory cells along the hypoglossal roots or trunk is irregular, and when the cells are present they are extremely scarceT,8,16,2~. Furthermore, hypoglossal afferent fibers were still present several days after intracranial section of hypoglossal roots (see Results). In view of the above findings, the cell origin for afferent fibers coursing in the hypoglossal nerve must be sought in neighboring nerves. Hypoglossal afferent fibers ascending from the upper cervical roots have been found in the hedgehog 25 and monkeyl~; however, cervical roots do not contribute to the hypoglossal nerve fibers in the cat11,13,15,24. In the cat, anastomoses are regularly found between the end-branches of hypoglossal and lingual nerves 4,14. These findings raised the possibility that deep mechanoreceptor fibers coursing peripherally with the hypoglossal nerve are conveyed to the brain stem through the trigeminal nerve 19. However, lingual nerve fibers are very rapidly adapting, and as Porter 19 himself concluded, 'stretch of the tongue did not prove to be as efficient in activating the endings as local deformation' produced by stroking the surface with a fine probe. This is in contrast with recordings from hypoglossal end-branches or hypoglosso-nodosal anastomose (see Results), where stroking of the tongue surface was completely ineffective in evoking sensory discharges, and a very deep indentation (as recorded by the transducer attached to the tongue tip) is needed to produce changes in the rate of discharge of hypogtossal fibers. It is perfectly possible that lingual nerve afferents can provide information about changes of tension or abrupt displacements of the tongue muscles, but a sustained discharge continuously signaling the tension and position of the tongue has not been found in these fibers. Our recordings of sensory discharges in the trunk of the hypoglossal nerve far from the terminal anastomoses with the lingual nerve show that the mechanoreceptor fibers of hypoglossal end-branches can travel through most parts of the hypoglossal nerve, at least up to the point of emergence of the hypoglosso-nodosal branch. In the posterior third of the tongue, hypogtossal nerve fibers run near the glossopharyngeal nerve. The possibility occurs that deep mechanoreceptor fibers of hypoglossal end-branches could deviate to the glossopharyngeal trunk and course centrally through this nerve. A similar criticism to that expressed in the preceding paragraph can be made: (1) we have recorded mechano-sensitive fibers in the hypoglossal nerve at a level where no anastomoses with the glossopharyngeal nerve could be shown; and (2) sensory discharges in the glossopharyngeal nerve could easily be evoked by gentle stroking of the posterior third of the lingual surface, but only a brief burst of impulses was recorded when the tongue was stretched abruptly, and no discharge was observed when the same stretch was applied slowly. The possibility of sensory fibers having their perikarya in the nodose ganglion of Brain Research, 32 (1971) 349-367

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the vagus, entering the cranium via the jugular foramen and the brain stem through the vagal dorsal roots, but distributing peripherally with the hypoglossal nerve, is supported by the following findings: (1) the invariable presence of a fine branch between the nodose ganglion and the hypoglossal nerve immediately after its exit from the cranium, according to our observations; (2) the recording of sensory action potentials in this branch in response to tongue stretching; (3) the recording of afferent activity in hypoglossal end-branches and the hypoglosso-nodosal branch in preparations in which hypoglossal or vagal roots had been intracranially severed days before; (4) the observations of Tarkhan and Abou-el-Naga 24 that removal of the ganglion nodosum in the dog led to degeneration of approximately 5 ~ of the fibers in the peripheral trunk of the hypoglossal nerve; (5) the observation of Sauerland and Mizuno z2 that multisynaptic reflexes recorded in the recurrent laryngeal nerve on stimulation of the central end of the cut hypoglossal nerve in the cat can no longer be elicited after section of the vagal rootlets, although remaining unchanged after severance of hypoglossal rootlets; (6) our observations that several reflex effects evoked by stimulation of the central end of the cut hypoglossus are abolished by severance of the hypoglosso-nodosal anastomosis 27. The hypoglossal sensory fibers described in the present paper must terminate in deep receptors of the tongue, since their discharges are not influenced by superficial stroking; otherwise, their discharges are markedly influenced by stretching of the tongue. However, as mentioned in the Introduction, no muscle spindles have been found in the intrinsic or extrinsic musculature of the cat's tongue and from a study of the calibre spectra Blom4 concluded that there are no fibers in the hypoglossal nerve that could be considered as candidates for nuclear bag afferents from muscle spindles. However, a special type of muscle receptor has been described by Law17 in the tongue of the cat, dog and pig; she found, in the intramuscular connective tissue, end-organs formed by a coil of nerve terminals enclosed in an oval or pear-shaped capsule; the structure and position of these organs could result in deformation of the capsule when the adjacent muscles are contracted or lengthened, thus initiating discharges of a proprioceptive nature. A similar structure was found by Weddell et al.Z6 at the attachment of the genioglossus muscle to the mandible. What is the role of the mechanosensory fibers distributed peripherally with the hypoglossal nerve? Several possibilities can be raised, but because of the slow adaptation of these fibers and other characteristics here illustrated, the feasibility that they could subserve kinesthetic sensation from the tongue and also serve as a proprioceptive system for the reflex control of tongue motility seems to be attractive. A full study and discussion on this subject will be published elsewhere (Torrealba and Zapata, in preparation). SUMMARY Sensory fibers were found along the extracranial course of the hypoglossal nerve of the cat: terminal branches, mid-portion of the trunk and ramus descendens hypoglossi. They were also present in a communicating branch between the hypoglossal Brain Research, 32 (1971) 349-367

366

p. ZAPATAAN[) G. TORREALBA

nerve and the nodose ganglion of the vagus, here called the 'hypoglosso-nodosal branch'. These fibers responded with slow adaptation to mechanical stimulation produced by longitudinal or lateral tensions or displacements of the tongue, but they did not respond to tactile stimuli applied to its surface. Most hypoglossal sensory units showed a basal rate of discharge in resting conditions, but some fibers were silent at rest. Their rates of discharge during mechanical stimulation depended on the intensity of the stimulus and its acceleration during application or interruption. Those fibers having a base-line discharge showed a silent pause at off, when the stimuli were abruptly interrupted. The rates of discharge of some hypoglosso-nodosal sensory units were accelerated or reduced by tongue movements elicited through stimulation of some terminal branches of the contralateral hypoglossus. However, other patterns of tongue movement evoked by stimulation of other terminal branches were ineffective. It is suggested that sensory fibers present along most of the extracranial course of the hypoglossus have their perikarya in the nodose ganglion and enter the brain stem through the dorsal roots of the vagus nerve. ACKNOWLEDGEMENTS This investigation was supported by Research Grant No. 68/32 from the Comisi6n Nacional de Investigaci6n Cientifica y Tecnol6gica (CONICYT). The authors express their gratitude to Drs. C. Eyzaguirre and G. Pilar, Department of Physiology, University of Utah, for their valuable comments on the manuscript. Thanks are also due to Mrs. Carolina Zapata and Mr. R. Vega for technical assistance.

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