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Excitatory and inhibitory inputs to single neurones in the solitary tract nucleus and adjacent reticular formation
Brain Research, 53 (1973) 319-331
319
© Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
EXCITATORY AND INHIBITORY INPUTS TO SINGLE NEURONES IN THE SOLITARY TRACT NUCLEUS AND ADJACENT RETICULAR FORMATION
B A R R Y J. SESSLE
Division of Biological Sciences, University of Toronto Faculty of Dentistry, Toronto 2 (Canada) (Accepted October 20th, 1972)
SUMMARY
Microelectrode recordings were made from single neurones located in the solitary tract nucleus and adjacent reticular formation of decerebrate or anaesthetized cats. Stimulation of various cranial nerves and cutaneous areas and the cerebral cortex was used to characterize the excitatory and inhibitory inputs to these brain stem regions. Most solitary tract units could be excited by stimulation of only the superior laryngeal nerve (SLN) or glossopharyngeal nerve (IX). In particular the laryngeal input was examined, and both primary afferent fibres and neurones in the nucleus could be activated with a short-latency to SLN stimulation. The neurones could be inhibited for as long as 700 msec by conditioning stimulation especially of SLN and IX and of the infraorbital nerve (IO). Other conditioning stimuli used were rarely effective. Neurones in the reticular formation ventral to the nucleus exhibited an excitatory input from SLN, IX, IO and sometimes other cranial nerves. However, some neurones could be excited only by IO stimulation. The latencies of the neurones to SLN stimulation were longer in general than those noted in solitary tract neurones. The neurones were subject to more widespread inhibitory influences than those in solitary tract neurones, and the responses of some could also be facilitated. On the basis of antidromic activation, m a n y reticular formation neurones were found to project rostrally to the vicinity of the trigeminal m o t o r nucleus. The excitatory and inhibitory influences acting on the solitary tract and reticular formation neurones are discussed in relation to their possible involvement in the perceptual and reflex activities with which the orofacial and pharyngeal-laryngeal areas are concerned.
INTRODUCTION
Although there has been a considerable amount of study of the brain stem nuclei
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that receive information from the face and other cutaneous body surfaces 5,6,x6, the brain stem regions related to laryngeal and oropharyngeal function have not received much attention. Information from the larynx and pharynx is carried primarily in the superior laryngeal nerve (SLN) and glossopharyngeal nerve (IX) to the solitary tract nucleus in the caudal brain stem7, 31. Very little is known of the central neural mechanisms that subsequently are activated, firstly to initiate and coordinate the reflex activities (e.g. swallowing, coughing, gagging) with which the larynx and pharynx are concerned 7,31, and secondly to relay information to higher nervous centres for the conscious perception of laryngeal and pharyngeal stimuli17,18. For this reason, studies have been initiated to shed some light on the brain stem mechanisms involved in laryngeal and oropharyngeal function. Investigations in the dorsal column and trigeminal sensory nuclei (for review, see Darian-Smith 5,6, Mountcastle and Darian-Smith 16, Sessle and Dubner 29) have revealed that neurones in these brain stem nuclei have restricted receptive fields and are susceptible to afferent and corticofugal regulation through facilitatory and inhibitory neural mechanisms. The present study was designed to characterize the input to the solitary tract nucleus and adjacent reticular formation and to determine if similar regulatory processes influence neurones in these regions. METHODS
Preparation of animals The experiments were carried out on 18 adult cats (2.5-4.5 kg) of which 15 were anaesthetized with chloralose (60 mg/kg) and 3 were decerebrated. All animals were paralyzed with gallamine triethiodide and artificially ventilated. Blood pressure and expired Pco2 were continuously monitored and rectal temperature maintained at 37.5 °C. The animal was mounted in a stereotaxic apparatus and the neck carefully dissected from a ventral approach to expose, on the right side, SLN, IX, the pharyngeal branch of the vagus nerve (Phar X), and the hypoglossal nerve (XII). The contralateral SLN was also isolated, and then a midline incision made along the ventral surface of the larynx. The larynx was protected from drying out and held open by a spring clip. After the animal was turned over, the ipsilateral infraorbital nerve (IO) was isolated and the caudal brain stem exposed by an occipital craniotomy. In some cats, a limited frontoparietal craniotomy was also carried out for the placement of stimulating electrodes in the ventrobasal complex of the posterior thalamus and the sensorimotor cerebral cortex. This procedure was omitted for the 3 cats that were decerebrated. Instead, small openings were made in the skull for the stereotaxic insertion of a pair of needles which was used to decerebrate the animals electrolytically at the midcollicular level.
Recording procedures The activity of single neurones in the solitary tract nucleus and adjacent areas was recorded extracellularly with tungsten microelectrodes a0. Neuronal responses were
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amplified (PAR Preamplifier 113) and displayed on a Tektronix D10/5A20N oscilloscope. The responses could also be led to a Tektronix scan converter 4501 which served as a storage oscilloscope and permitted permanent photographic records to be taken of selected responses with a Grass C4 camera. Electrode penetrations were made stereotaxically in 0.5 mm steps in the caudal brain stem in planes 0-3 mm anterior to the obex and 1--4 mm lateral to the midline. In most animals the caudal tip of the cerebellum was removed by suction to afford direct vision of the penetrations. These recording sites and the stimulating loci in the rostral brain stem region (see below) were checked in histological sections (stained with haematoxylin and eosin) of the brain stem that was perfused with 10 ~ formalin at the end of the experiment.
Stimulation procedures Orthodromic activation of brain stem neurones from peripheral sites was tested by bipolar stimulation (0.01-5.0 mA, 0.1 msec) of the exposed nerves. The stimulating sites for SLN and XII were approximately 25 mm distal to the jugular foramen; for IX and Phar X the sites were about 10 mm distal to the foramen. The exposed nerves and silver hook stimulating electrodes were usually covered with petroleum jelly to minimize the possibility of stimulus spread. In two cats the ipsilateral forepaw and isolated lingual nerve were also used as stimulating sites. The responsiveness of brain stem neurones to auditory stimulation was also tested in these two cats by a 1 msec duration click delivered from a loudspeaker. Although the excitatory receptive field of neurones was not routinely tested in this study, the mechanoreceptive field of some neurones was determined with a plastic probe. Two pairs of bipolar electrodes (interpolar distance 0.5-1.0 mm) were placed on the contralateral sensorimotor cortex for subsequent stimulation (0.05-5.0 mA, 0.1 msec, 1-4 pulses at 250/sec) of corticofugal pathways. One pair was located in the region of the maximum evoked response to IO stimulation, the other pair more rostral in the region previously foundS, 9 to have a rich projection to trigeminal brain stem sensory nuclei and adjacent areas. Two pairs of bipolar electrodes for subsequent stimulation were also placed in the vicinity of the ipsilateral trigeminal motor nucleus utilizing stereotaxic coordinates, histological verification of the sites, and observations of short-latency ( < 5 msec)jaw closure elicited by stimulation at this rostral brain stem site and recorded electromyographically. The electrodes were used to determine if neurones recorded in the solitary tract nucleus or adjacent areas could be antidromically activated from this rostral brain stem region, since a study in this laboratory (Sessle et aL, in preparation) had revealed short-latency SLN and IX evoked responses in the vicinity of the trigeminal motor nucleus. In 3 cats another two pairs of bipolar electrodes were located in the posterior thalamic region of the contralateral ventrobasal complex utilizing stereotaxic coordinates and recordings of the maximum evoked response to IO and forepaw stimulation. These electrodes were then used for stimulation and possible antidromic activation of solitary tract~neurones to determine if the ventrobasal complex might serve as a relay for the transfer of laryngeal information to the cerebral cortex. Antidromic
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activation of a neurone from the vicinity of the ventrobasal complex or the rostral brain stem region was based on a consistent, all-or-none response with short-latency to ventrobasal or rostral brain stem stimulation (0.05-5.0 mA, 0.1 msec) at rates of 200/sec or more.
Conditioning procedures Stimuli applied to the various exposed nerves and cerebral cortex were used to test for facilitatory and inhibitory effects on the excitatory input to brain stem neurones. This was done by applying the conditioning nerve or cortical stimulus at various intervals preceding the excitatory (test) stimulus. The time course of an observed effect on a neurone was obtained by comparing, at the various conditioning-test intervals, the total number of spikes evoked in the neurone by 10-20 test stimuli with the total number evoked by a similar number of stimuli in the presence of a conditioning stimulus. In addition to the use of an electrical stimulus of the exposed nerves and cerebral cortex for conditioning stimulation, a mechanical conditioning stimulus (10 msec) applied to the canine tooth by a mechanical stimulator 35 was frequently employed. RESULTS
General aspects Microelectrode recordings were made from 457 neurones in the caudal brain stem. One-third of these neurones was recorded in decerebrate preparations, the remainder from cats anaesthetized with chloralose. There was no apparent difference between the two preparations in the response properties of neurones isolated from the solitary tract nucleus and adjacent reticular formation. The response properties of the neurones (see below) and the subsequent histological localization of the microelectrode penetrations indicated that neuronal responses were recorded, as well as from the solitary tract and its nucleus, from the motor nucleus of X I I and that part of the reticular formation lateral to this nucleus and ventral to the solitary tract nucleus. Some penetrations may also have passed through the dorsal motor nucleus of X, but methods (e.g. stimulation of X to evoke antidromic responses) to identify neurones in this nucleus were not employed. In the more lateral penetrations at depths 0.8-1.5 m m below the brain stem surface, a small field potential evoked by SLN stimulation was encountered. This potential reached its peak amplitude within the next 0.5 m m and was often coextensive with a smaller field potential resulting from IX stimulation (Fig. l, see also ref. 1), particularly at the more rostral planes. Stereotaxic coordinates and histological verification indicated that this region of SEN input was the solitary tract and its nucleus. The SLN evoked field potential typically showed two negative components. The first component had a latency of approximately 1 msec and followed SLN stimulation rates of more than 200/sec. Thus it probably represented the summed response of evoked action potentials in SEN primary afferents in the region. The later phase of negativity had a latency of 3 msec and a duration of 4-5 msec and did not follow
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Fig. 1. Evoked field potentials and neuronal responses recorded in 5 microelectrode penetrations in the caudal brain stem. The penetrations (a-e) were made in 0.5 mm steps mediolaterally in the one anteroposterior plane (2 mm anterior to obex) and are indicated in the transverse section, On the right side of the figure are illustrated the field potentials evoked by superior laryngeal nerve (SLN) and glossopharyngeal nerve (IX) stimulation and recorded at various depths (dotted arrows) of penetration c through the solitary tract and its nucleus (outlined) and the reticular formation ventral to it (vertical depth measurements were determined from readings of the electrode microdrive with an allowance of 2 0 ~ for tissue shrinkage). Single units recorded in the 5 penetrations are indicated by the various symbols on the transverse section. In the lower part of the figure are shown typical responses to SLN stimulation of a solitary tract primary fibre (open circle) at 200/sec stimulation rate, a solitary tract neurone (closed circle), and a neurone located in the adjacent reticular formation. A closed triangle indicates a neurone projecting to the rostral brain stem whereas an open triangle represents a reticular formation neurone that could not be antidromically activated from this region. Also illustrated is the response to infraorbital nerve (IO) stimulation of a reticular formation neurone (V) with only a demonstrated IO input. Negative polarity in this and subsequent figures is upwards; voltage calibration, 0.4 mV; time calibration, 4 msec for all records except the solitary tract fibre and neurone for which it is 2 msec; V, nucleus of the spinal tract of the trigeminal nerve, with the tract lateral to it; X, dorsal motor nucleus of the vagus nerve; XII, hypoglossal motor nucleus.
s t i m u l a t i o n r a t e s o f 50/sec (cf. ref. 1). T h i s s e c o n d p h a s e was also v e r y susceptible to a s p h y x i a a n d m i g h t be a t t r i b u t e d to p o s t s y n a p t i c a c t i v i t y e v o k e d by S L N s t i m u l a t i o n in s o l i t a r y t r a c t n e u r o n e s . T h e s e s h o r t - l a t e n c y S L N e v o k e d field p o t e n t i a l s w e r e p r e s e n t for a b o u t 1 m m a n d t h e n a b r u p t l y r e p l a c e d by a single w a v e o f n e g a t i v i t y w i t h a l a t e n c y o f a b o u t 4 m s e c a n d a d u r a t i o n o f 6 - 7 m s e c (Fig. 1). E v o k e d field p o t e n t i a l s o f a p p r o x i m a t e l y t h e s a m e l a t e n c y to I O , I X a n d s o m e t i m e s to X I I a n d P h a r X s t i m u l a t i o n also w e r e p r e s e n t . T h i s r e g i o n o f m u l t i p l e i n p u t in the r e t i c u l a r f o r m a t i o n h a d a v e r t i c a l e x t e n t
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of 0.5-1.0 mm. Very little activity could be recorded on deeper microelectrode penetration by stimulation of the exposed nerves. However, a field potential evoked by only IO stimulation was sometimes present, particularly in more medial penetrations passing close to the motor nucleus of XII. In these situations an antidromic field potential elicited by XII stimulation was also recorded. The findings of the response properties of single neurones will now be presented, with particular reference to the neurones recorded in the solitary tract and its nucleus, and in the reticular formation adjacent to it.
Solitary tract and its nucleus Input. A total of 167 single units were isolated in this region. All but 6 were characterized as being activated by a single stimulus to either SLN or IX stimulation. The remaining 6 neurones were activated by both SLN and IX stimulation. None of the neurones tested could be activated by auditory stimuli or by stimulation of the other isolated nerves (including the contralateral SLN), the cerebral cortex, or forepaw. Sixty-four of the units were classified as primary fibres on the basis of a shortlatency response that followed stimulation rates greater than 200/sec. The majority of units in this class, although not all (Fig. 1), also showed an initially positive-going spike response, in accordance with findings3,11 for fibres elsewhere in the central nervous system. On the other hand, units classified as brain stem neurones invariably showed negative (Fig. 1) or positive-negative (Fig. 3) spike forms and did not follow stimulation rates greater than 50/sec. Moreover, on the basis of these criteria of stimulation following rate and spike polarity, it was found that the solitary tract units separated into two latency groups (see below and Fig. 2) that corresponded closely to the two components of the SLN evoked field potential (although there was a small amount of overlap between the two latency groups). The possibility that some of the 64 units classified as primary fibres may have actually been nucleus ambiguus motoneurones was ruled out by the superficiality (histologically verified) of the recording sites and their activation by stimulation of just one peripheral nerve or of the larynx. Solitary tract neurones (and primary afferent fibres) with a laryngeal input were particularly sought for in this study, and of the 103 neurones isolated in the nucleus, 78 were driven only by SLN stimulation. Some of these were examined and found to have laryngeal mechanoreceptive fields (e.g. Fig. 3). Neurones activated by IX stimulation, although few (19) in number, tended to occur in the more rostral planes examined in the brain stem. The primary fibres and neurones appeared to be intermingled in the solitary tract nuclear region (Fig. 1) since none of the microelectrode penetrations and recordings provided any clear-cut distinction, on electrophysiological grounds, between the solitary tract and its nucleus. This was not an unexpected finding in view of the comparatively small size of the tract and nucleus and the microelectrode's capability of recording responses from neurones some distance away from the microelectrode tip. Latency and discharge characteristics. The units (57) classed as SLN primary afferent fibres showed response latencies to SLN stimulation which fell within the
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range 1.0-2.9 msec (Fig. 2) with a mean value of 1.8 msec. The conduction path from the SLN stimulation site to caudal brain stem measured 35-40 mm, thus the range of conduction velocities of the SLN primary units recorded in the solitary tract region in this study approximated 12-40 m/sec. In comparison, however, the minimum latency to SLN stimulation of neurones in this region was 2.1 msec; fewer than 2 0 ~ had latencies shorter than 2.9 msec. The mean latency value was 3.5 msec and almost 90 ~ of these solitary tract neurones had latencies less than 5.0 msec (Fig. 2). An interesting feature typical of the discharge of these neurones to SLN stimulation was that there were no more than 2 or 3 spikes evoked by the SLN stimulus. A typical response is shown in the bottom part of Fig. 1. The stimulus threshold for the first spike in most of the neurones was 0.5-1.0 mA, and increasing the stimulus intensity to m a x i m u m rarely resulted in more than two additional spikes. Antidromic activation. Bipolar stimulation in the thalamic ventrobasal complex and in the rostral brain stem was used to determine if there was a direct projection from the solitary tract nucleus to either of these regions. Thalamic stimulation failed to elicit an antidromic response in 16 solitary tract neurones tested. These and another 36 neurones tested for a projection to the rostral brain stem could not be antidromically activated from this area. Conditioning effects. A total of 61 solitary tract neurones was examined to determine if their excitatory input from SLN or IX could be influenced by conditioning stimulation. All but 3 of these neurones showed an inhibition of their excitatory response to SLN or I X stimulation. Neurones with an SLN excitatory input were particularly examined and 45 of 48 tested could be inhibited by a previous conditioning stimulus delivered to SLN. The discharge of the remaining 3 units could not be influenced by any of the conditioning stimuli used. The inhibitory effect of the SLN con-
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Fig. 3. Time course of inhibition produced in a solitary tract neurone with a superior laryngeal nerve (SLN) excitatory input. Only SLN and glossopharyngeal nerve (IX) conditioning stimuli were effective in inhibiting the neurone. The time course was obtained by comparing, at various conditioning-test intervals, the total number of spikes evoked by 10 SLN test stimuli (control) with the number obtained by 10 stimuli in the presence of a conditioning stimulus. The top record illustrates 5 superimposed control responses of the neurone to SLN stimulation. The middle and bottom records show the inhibitory effect, at a 40 msec conditioning-test interval, of IX and SLN conditioning stimuli respectively. Voltage calibration, 0.5 mV; time calibration, 5 msec. The neurone's laryngeal mechanoreceptive field is indicated (arrow) by the black patch in the illustration of the exposed larynx.
ditioning stimulus typically had an onset of 20 msec, a peak at 40 msec, and lasted for 300-400 msec. An example of this time course of inhibition is shown in Fig. 3. However, 20 ~o of the neurones showed an inhibition which lasted as long as 600-700 msec or as short as 150 msec (Fig. 4). A IX conditioning stimulus was also effective in inhibiting the SEN response in all but 5 of the 45 neurones inhibited by SLN conditioning (Figs. 3 and 4). An IO stimulus or a mechanical conditioning stimulus applied to the canine tooth produced inhibition of the SLN response in 10 of 28 neurones tested (Fig. 4). However, Phar X and XII conditioning stimuli were rarely effective and produced inhibition in only 3 of 31 neurones examined. Conditioning stimulation of the cerebral cortex produced a similar ratio of inhibitory effects, and forepaw and auditory stimuli had no conditioning effect in any neurone tested. Thirteen solitary tract neurones with only a IX excitatory input were tested for conditioning influences. All could be inhibited by a preceding IX conditioning stimulus and most by SLN conditioning stimulation as well. A preceding IO or canine stimulus was effective in 5 of 10 neurones tested, but Phar X (or XII) stimulation produced inhibition in only one of these neurones. A facilitatory effect of conditioning was seen in 4 solitary tract neurones. These neurones had a 1X excitatory input, and a preceding IX conditioning stimulus decreased the latency of the initial spike and increased the number of spikes produced by the control (test) IX stimulus. This facilitation lasted for about 150 msec in one neu-
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rone a n d for as long as 600 msec in another. The r e m a i n i n g two n e u r o n e s were facilit a t e d for a b o u t 40 msec.
Adjacent reticularformation Input. A p a r t f r o m 9 m o t o n e u r o n e s (identified by their a n t i d r o m i c response to X I I s t i m u l a t i o n ) l o c a t e d in the m o t o r nucleus o f X I I , the r e m a i n i n g 287 n e u r o n e s rec o r d e d were isolated in the reticular f o r m a t i o n lateral to this nucleus a n d ventral to the solitary t r a c t nucleus. It is extremely unlikely t h a t a significant n u m b e r o f these n e u r o n e s was r e c o r d e d in the d o r s a l m o t o r nucleus o f X, since less t h a n 5 ~ o f t h e m c o u l d be localized directly m e d i a l to the solitary t r a c t in the region o f this nucleus. M o s t o f the reticular f o r m a t i o n n e u r o n e s h a d an i n p u t t h a t usually involved S L N , I X a n d IO, a n d often X I I . Cortical, P h a r X or c o n t r a l a t e r a l S L N , b u t n o t a u d i t o r y or forepaw, s t i m u l a t i o n was also effective in exciting some. A l t h o u g h a wides p r e a d i n p u t was a feature o f these neurones, 69 could be activated only b y s t i m u l a t i o n o f I O (Fig. 1). L i n g u a l nerve s t i m u l a t i o n was p a r t i c u l a r l y effective in activating neurones in the vicinity o f the m o t o r nucleus o f X I I . S t i m u l a t i o n o f S L N a n d I X also p r o d u c e d a s h o r t - l a t e n c y a c t i v a t i o n o f m a n y n e u r o n e s in this region. Latency and discharge characteristics. The response latencies to S L N s t i m u l a t i o n o f the reticular f o r m a t i o n n e u r o n e s with an extensive i n p u t were longer t h a n the range o f l a t e n c y values o f a p p r o x i m a t e l y 90 ~ o f solitary t r a c t neurones. F o r example, the S L N latencies o f those n e u r o n e s t h a t were r e c o r d e d j u s t ventral to the solitary t r a c t nucleus a n d p r o j e c t e d to the r o s t r a l b r a i n stem (see below) r a n g e d f r o m 4 to 12 msec with a m e a n value o f 7 msec. L o n g e r a n d m o r e variable latencies to S L N s t i m u l a t i o n were m o r e c o m m o n deeper in the reticular f o r m a t i o n . The threshold for S L N stimulat i o n was similar to t h a t o f solitary t r a c t neurones. However, the response to a single
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SLN stimulus was a burst of spikes (Fig. 1), the number of spikes being dependent on stimulus intensity. Antidromic activation. Only 22 neurones were tested but none could be antidromically activated by thalamic stimulation. However, approximately 40~o (41 of 107 neurones tested) showed an antidromic response with rostral brain stem stimulation. The majority of these neurones with an extensive input were located close to the ventral extent of the solitary tract and its nucleus (Fig. I). A similar percentage of reticular formation neurones with only an IO input also projected rostrally. Conditioning effects. The reticular formation nemones could be inhibited from many sources. Conditioning stimulation of each of the nerves that excited a neurone would inhibit the neurone's responses to the other inputs. The duration of the inhibition produced in these neurones varied between 150-700 msec. The onset and peak were typically 20 and 40 msec respectively, but in some neurones longer values were noted since an early facilitation, which had an onset of 10 msec and a duration of approximately 40 msec, preceded the inhibition.
DISCUSSION
The present study of single neurones in the brain stem revealed that many units isolated in the caudal part of the solitary tract and its nucleus have a single excitatory input from SLN or IX. In a study of the vagal input to solitary tract and nucleus ambiguus neurones, Porter la indicated that SLN stimulation would evoke unitary activity in both nuclei. L a m and Ogura 14 failed to identify a laryngeal projection to the solitary tract nucleus, but in support of the present findings and those of Porter are studies 2a, 24,28,30 reporting antidromic activation of SLN primary afferent units by stimulation in this nucleus. A IX input to some neurones is supported by the finding a2 that stimulation of oropharyngeal areas will elicit solitary tract neuronal responses. The response latencies of solitary tract neurones to SLN stimulation were in the range of 2.1-5.0 msec for about 90 ~o of the neurones. This range and the mean value of 3.5 msec are compatible with the latencies to SLN stimulation reported by Porter 19. The mean value and range of latencies noted in SLN primary afferent fibres in the region would suggest that most of the neurones were activated directly by the incoming afferents. The estimated conduction velocities of these primary afferent units indicated that more than 90 ~ were small myelinated (A6) nerve fibres, in accordance with findings15, 2s of the conduction velocities of SLN primary afferent units recorded in the nodose ganglion. T h e response of adjacent reticular formation neurones to SLN stimulation was repetitive and more quantitatively related to the stimulus intensity than responses of solitary tract neurones. Most of these neurones displayed an extensive input involving IO, SLN and IX especially. Excitation of neurones in the reticular formation from a variety of sources is an established feature of this area 2,4,25,27, but no previous report has revealed this particular combination of inputs to neurones ventral to the solitary tract nucleus. The latency values of their responses would indicate that many of the
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neurones receive a direct excitatory input from the solitary tract nucleus. Some of these neurones may act as the source of inhibitory interneurones influencing the discharge of solitary tract neurones. But 40 ~o were found to project rostrally to the vicinity of the trigeminal motor nucleus. This projection is supported by findings in this laboratory (Sessle et al., in preparation) of short-latency inputs to this region evoked by SLN and IX stimulation. The functional significance of this relay cannot be defined at this time, but some integrative action in swallowing and mastication or in protective jaw reflexes seems likely. The orofacial input to the reticular formation neurones probably involved at least a relay in the trigeminal sensory nuclear complex. Anatomical studies in man a4, monkey 22, rat aa, and cat12, la have described trigeminal primary afferent fibres passing to the vicinity of the solitary tract nucleus. Sumi az has also reported that some neurones in the rostral part of this nucleus may be activated by tooth stimulation. However, in the present study, no neurones in the caudal part of the nucleus could be activated by IO stimulation. This would indicate that the trigeminal afferents described anatomically in the nucleus either do not terminate here but end on neurones in the adjacent reticular formation or more rostral solitary tract neurones, or carry information (e.g. gustatory) to the nucleus via other branches of the trigeminal nerve. A trigeminal input was also noted for neurones adjacent to the motor nucleus of XII but many of these neurones could also be activated by SLN and IX stimulation. Such neurones have not been described previously in this region, although Lam and Ogura 14 noted in the reticular formation close to this nucleus an evoked field potential with SLN stimulation. The neurones are possibly involved in a reflex regulation of hypoglossal motoneurones in a manner similar to the lingual-hypoglossal reflex2°, 21, since SLN (or IX) stimulation produces a short-latency activation of the tongue musculature and a subsequent, prolonged inhibition of its activity z°. Solitary tract neurones could be inhibited from SLN or IX, but auditory stimulation or stimuli applied to Phar X, XII, forepaw, and cerebral cortex were rarely effective. These findings would indicate that neurones innervating visceral structures display the phenomenon of afferent or surround inhibition that has been described in somatic, auditory and visual relay areas and has been implicated in spatial discrimination 10. However, some neurones could also be inhibited by electrical or natural stimulation of the orofacial region, which is innervated by a somatic nerve, and the following paper z8 indicates that presynaptic depolarization may account largely for these effects. A regulatory role in reflex activities for such orofacial conditioning stimulation is discussed elsewhere ao.
ACKNOWLEDGEMENTS
I thank Mr. M. Kalovsky and Miss B. Holmwood for their technical assistance and Drs. Arthur Storey and Ronald Dubner for critically reading the manuscript. This study was supported by Grant D.G. 73 from the Canadian Medical Research Council.
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REFERENCES 1 BISCOE,T. J., AND SAMPSON,S. R., Field potentials evoked in the brain stem of the cat by stimulation of the carotid sinus, glossopharyngeal, aortic and superior laryngeal nerves, J. Physiol. (Lond.), 209 (1970) 341-358. 2 BiSCOE,T. J., AND SAMlaSON, S. R., Responses of cells in the brain stem of the cat to stimulation of the sinus, glossopharyngeal, aortic and superior laryngeal nerves, J. Physiol. (Lond.), 209 (1970) 359-373. 3 BisHot', P. O., BURKE, W., AND DAVIS, R., The identification of single units in central visual pathways, J. Physiol. (Lond.), 162 (1962) 409-431. 4 BOWSHER,D., Place and modality analysis in caudal reticular formation, J. Physiol.(Lond.), 209 (1970) 473-486. 5 DARIAN-SM1TH,I., Neural mechanisms of facial sensation, Int. Rev. NeurobioL, 9 (1966) 301-395. 6 DARIAN-SMITH,I., Somatic sensation, Ann. Rev. Physiol., 31 (1969) 417-450. 7 DOTY, R. W., Neural organization of deglutition. In C. F. CODE (Ed.), Handbook of Physiology, Vol. IV, Sect. 6, Amer. Physiol. Soc., Washington, 1968, pp. 1861-1902. 8 DUBNER, R., AND SESSLE, B. J., Presynaptic modification of corticofugal fibers participating in a feedback loop between trigeminal brain-stem nuclei and sensorimotor cortex. In R. DUBNERAND Y. KAWAMURA(Eds.), Oral-Facial Sensory and Motor Mechanisms, Appleton-Century-Crofts, New York, 1971, pp. 299-314. 9 DUBNER,R., AND SESSLE, B, J., Presynaptic excitability changes of primary afferent and corticofugal fibers projecting to trigeminal brain stem nuclei, Exp. NeuroL, 30 (1971) 223-238. 10 HUaEL, D. H., Tungsten micro-electrode for recording from single units, Science, 125 (1957) 549550. 1 l HUBEL, O. H., Single unit activity in lateral geniculate body and optic tract of unrestrained cats, J. Physiol. (Lond.), 150 (1960) 91-104. 12 KERR, F. W. L., Structural relation of the trigeminal spinal tract to upper cervical roots and the solitary nucleus in the cat, Exp. Neurol., 4 (1961) 134-148. 13 KERR, F. W. L., The divisional organization of afferent fibres of the trigeminal nerve, Brain, 86 (1963) 721-732. 14 LAM, R. L., AND OGURA, J. H., Afferent projection of the superior laryngeal nerve in the brain stem, Neurology (Minneap.), 4 (1954) 630-632. 15 MEI, N., Disposition anatomique et propri6t6s 61ectrophysiologiques des neurones sensitifs vagaux chez le chat, Exp. Brain Res., 11 (1970) 465-479. 16 MOUNTCASTLE,V. B., ANDDARIAN-SMITH,I., Neural mechanisms in somesthesia. In V. B. MOUNTCASTLE(Ed.), MedicalPhysiology, Mosby, St. Louis, Mo., 1968, pp. 1373-1423. 17 O'BRIEN, J. n., PIMPANEAU, A., AND ALBE-FESSARD,O., Evoked cortical responses to vagal, laryngeal and facial afferents in monkeys under chloralose anaesthesia, Electroenceph. clin. Neurophysiol., 31 (1971) 7-20. 18 OGURA, J. H., AND LAM, R. L., Anatomical and physiological correlations on stimulating the human superior laryngeal nerve, Laryngoscope (St. Louis), 63 (1953) 947-959. 19 PORTER, R., Unit responses evoked in the medulla oblongata by vagus nerve stimulation, J. Physiol. (Lond.), 168 (1963) 717-735. 20 PORTER, R., Synaptic potentials in hypoglossal motoneurones, J. Physiol. (Lond.), 180 (1965) 209-224. 21 PORTER,R., The synaptic basis of a bilateral lingual-hypoglossal reflex in cats, J. Physiol. (Lond.), 190 (1967) 611-627. 22 RHOTON, A. L., O'LEARY, J. L., AND FERGUSON,J. P., The trigeminal, facial, vagal and glossopharyngeal nerves in the monkey, Arch. NeuroL (Chic.), 14 (1966) 536-540. 23 RUDOMIN,P., Presynaptic inhibition induced by vagal afferent volleys, J. Physiol. (Lond.), 30 (1967) 964-981. 24 RUDOMIN, P., Excitability changes of superior laryngeal, vagal and depressor afferent terminals produced by stimulation of the solitary tract nucleus, Exp. Brain Res., 6 (1968) 156-170. 25 SCHEIBEL,M., SCHEIaEL, A., MOLLICA, A., AND MORUZZI, G., Convergence and interaction of afferent impulses on single units of reticular formation, J. Neurophysiol., 18 (1955) 309-331. 26 SCnMITT,A., YU, S.-K. J., AND SESSLE, B. J., Excitatory and inhibitory influences from laryngea and orofacial areas on tongue position in cat, Arch. oral Biol., (1973) in press. 27 SEGUNOO,J. P., TAKENAKA,T., AND ENCABO, H., Somatic sensory properties of bulbar reticular neurones, J. Neurophysiol., 30 (1967) 1221-1238.
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28 SESSLE,B. J., Presynaptic excitability changes induced in single laryngeal primary afferent fibres, Brain Research, 53 (1973) 333-342. 29 SESSLE,B. J., AND DUBNER, R., Presynaptic depolarization and hyperpolarization of trigeminal primary and thalamic afferents. In R. DUBNERAND Y. KAWAMURA(Eds.), Oral-Facial Sensory and Motor Mechanisms, Appleton-Century-Crofts, New York, 1971, pp. 279-298. 30 SESSLE,B. J., AND STOREY,A. T., Periodontal and facial influences on the laryngeal input to the brain stem of the cat, Arch. oral Biol., 17 (1972) 1583-1595. 31 STOgEY, A.T., Extra-trigeminal sensory systems related to oral function. In J. F. BOSMA(Ed.), Symposium on Oral Sensation and Perception, Thomas, Springfield, Ill., 1967, pp. 84-97. 32 SUMI, T., Neuronal mechanisms in swallowing, Pfliigers Arch. ges. Physiol., 278 (1964) 467-477. 33 TORVIK,A., Afferent connections to the sensory trigeminal nuclei, the nucleus of the solitary tract and adjacent structures - - an experimental study in the rat, J. comp. Neurol., 106 (1965) 51-141. 34 WALLENBERG,A., Das dorsale Gebiet der spinalen Trigeminuswurzel und seine Beziehungen zum solit/iren Biindel beim Menschen. Ein Beitrag zur Anatomie und Physiologie des Trigeminus, Dtsch. Z. Nervenheilk., 11 (1897) 391-405. 35 WERNER,G., AND MOUNTCASTLE,V. B., Neural activity in mechanoreceptive cutaneous afferents: stimulus-response relations, Weber functions and information transmission, J. Neurophysiol., 28 (1965) 359-397.
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© Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
EXCITATORY AND INHIBITORY INPUTS TO SINGLE NEURONES IN THE SOLITARY TRACT NUCLEUS AND ADJACENT RETICULAR FORMATION
B A R R Y J. SESSLE
Division of Biological Sciences, University of Toronto Faculty of Dentistry, Toronto 2 (Canada) (Accepted October 20th, 1972)
SUMMARY
Microelectrode recordings were made from single neurones located in the solitary tract nucleus and adjacent reticular formation of decerebrate or anaesthetized cats. Stimulation of various cranial nerves and cutaneous areas and the cerebral cortex was used to characterize the excitatory and inhibitory inputs to these brain stem regions. Most solitary tract units could be excited by stimulation of only the superior laryngeal nerve (SLN) or glossopharyngeal nerve (IX). In particular the laryngeal input was examined, and both primary afferent fibres and neurones in the nucleus could be activated with a short-latency to SLN stimulation. The neurones could be inhibited for as long as 700 msec by conditioning stimulation especially of SLN and IX and of the infraorbital nerve (IO). Other conditioning stimuli used were rarely effective. Neurones in the reticular formation ventral to the nucleus exhibited an excitatory input from SLN, IX, IO and sometimes other cranial nerves. However, some neurones could be excited only by IO stimulation. The latencies of the neurones to SLN stimulation were longer in general than those noted in solitary tract neurones. The neurones were subject to more widespread inhibitory influences than those in solitary tract neurones, and the responses of some could also be facilitated. On the basis of antidromic activation, m a n y reticular formation neurones were found to project rostrally to the vicinity of the trigeminal m o t o r nucleus. The excitatory and inhibitory influences acting on the solitary tract and reticular formation neurones are discussed in relation to their possible involvement in the perceptual and reflex activities with which the orofacial and pharyngeal-laryngeal areas are concerned.
INTRODUCTION
Although there has been a considerable amount of study of the brain stem nuclei
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that receive information from the face and other cutaneous body surfaces 5,6,x6, the brain stem regions related to laryngeal and oropharyngeal function have not received much attention. Information from the larynx and pharynx is carried primarily in the superior laryngeal nerve (SLN) and glossopharyngeal nerve (IX) to the solitary tract nucleus in the caudal brain stem7, 31. Very little is known of the central neural mechanisms that subsequently are activated, firstly to initiate and coordinate the reflex activities (e.g. swallowing, coughing, gagging) with which the larynx and pharynx are concerned 7,31, and secondly to relay information to higher nervous centres for the conscious perception of laryngeal and pharyngeal stimuli17,18. For this reason, studies have been initiated to shed some light on the brain stem mechanisms involved in laryngeal and oropharyngeal function. Investigations in the dorsal column and trigeminal sensory nuclei (for review, see Darian-Smith 5,6, Mountcastle and Darian-Smith 16, Sessle and Dubner 29) have revealed that neurones in these brain stem nuclei have restricted receptive fields and are susceptible to afferent and corticofugal regulation through facilitatory and inhibitory neural mechanisms. The present study was designed to characterize the input to the solitary tract nucleus and adjacent reticular formation and to determine if similar regulatory processes influence neurones in these regions. METHODS
Preparation of animals The experiments were carried out on 18 adult cats (2.5-4.5 kg) of which 15 were anaesthetized with chloralose (60 mg/kg) and 3 were decerebrated. All animals were paralyzed with gallamine triethiodide and artificially ventilated. Blood pressure and expired Pco2 were continuously monitored and rectal temperature maintained at 37.5 °C. The animal was mounted in a stereotaxic apparatus and the neck carefully dissected from a ventral approach to expose, on the right side, SLN, IX, the pharyngeal branch of the vagus nerve (Phar X), and the hypoglossal nerve (XII). The contralateral SLN was also isolated, and then a midline incision made along the ventral surface of the larynx. The larynx was protected from drying out and held open by a spring clip. After the animal was turned over, the ipsilateral infraorbital nerve (IO) was isolated and the caudal brain stem exposed by an occipital craniotomy. In some cats, a limited frontoparietal craniotomy was also carried out for the placement of stimulating electrodes in the ventrobasal complex of the posterior thalamus and the sensorimotor cerebral cortex. This procedure was omitted for the 3 cats that were decerebrated. Instead, small openings were made in the skull for the stereotaxic insertion of a pair of needles which was used to decerebrate the animals electrolytically at the midcollicular level.
Recording procedures The activity of single neurones in the solitary tract nucleus and adjacent areas was recorded extracellularly with tungsten microelectrodes a0. Neuronal responses were
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amplified (PAR Preamplifier 113) and displayed on a Tektronix D10/5A20N oscilloscope. The responses could also be led to a Tektronix scan converter 4501 which served as a storage oscilloscope and permitted permanent photographic records to be taken of selected responses with a Grass C4 camera. Electrode penetrations were made stereotaxically in 0.5 mm steps in the caudal brain stem in planes 0-3 mm anterior to the obex and 1--4 mm lateral to the midline. In most animals the caudal tip of the cerebellum was removed by suction to afford direct vision of the penetrations. These recording sites and the stimulating loci in the rostral brain stem region (see below) were checked in histological sections (stained with haematoxylin and eosin) of the brain stem that was perfused with 10 ~ formalin at the end of the experiment.
Stimulation procedures Orthodromic activation of brain stem neurones from peripheral sites was tested by bipolar stimulation (0.01-5.0 mA, 0.1 msec) of the exposed nerves. The stimulating sites for SLN and XII were approximately 25 mm distal to the jugular foramen; for IX and Phar X the sites were about 10 mm distal to the foramen. The exposed nerves and silver hook stimulating electrodes were usually covered with petroleum jelly to minimize the possibility of stimulus spread. In two cats the ipsilateral forepaw and isolated lingual nerve were also used as stimulating sites. The responsiveness of brain stem neurones to auditory stimulation was also tested in these two cats by a 1 msec duration click delivered from a loudspeaker. Although the excitatory receptive field of neurones was not routinely tested in this study, the mechanoreceptive field of some neurones was determined with a plastic probe. Two pairs of bipolar electrodes (interpolar distance 0.5-1.0 mm) were placed on the contralateral sensorimotor cortex for subsequent stimulation (0.05-5.0 mA, 0.1 msec, 1-4 pulses at 250/sec) of corticofugal pathways. One pair was located in the region of the maximum evoked response to IO stimulation, the other pair more rostral in the region previously foundS, 9 to have a rich projection to trigeminal brain stem sensory nuclei and adjacent areas. Two pairs of bipolar electrodes for subsequent stimulation were also placed in the vicinity of the ipsilateral trigeminal motor nucleus utilizing stereotaxic coordinates, histological verification of the sites, and observations of short-latency ( < 5 msec)jaw closure elicited by stimulation at this rostral brain stem site and recorded electromyographically. The electrodes were used to determine if neurones recorded in the solitary tract nucleus or adjacent areas could be antidromically activated from this rostral brain stem region, since a study in this laboratory (Sessle et aL, in preparation) had revealed short-latency SLN and IX evoked responses in the vicinity of the trigeminal motor nucleus. In 3 cats another two pairs of bipolar electrodes were located in the posterior thalamic region of the contralateral ventrobasal complex utilizing stereotaxic coordinates and recordings of the maximum evoked response to IO and forepaw stimulation. These electrodes were then used for stimulation and possible antidromic activation of solitary tract~neurones to determine if the ventrobasal complex might serve as a relay for the transfer of laryngeal information to the cerebral cortex. Antidromic
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activation of a neurone from the vicinity of the ventrobasal complex or the rostral brain stem region was based on a consistent, all-or-none response with short-latency to ventrobasal or rostral brain stem stimulation (0.05-5.0 mA, 0.1 msec) at rates of 200/sec or more.
Conditioning procedures Stimuli applied to the various exposed nerves and cerebral cortex were used to test for facilitatory and inhibitory effects on the excitatory input to brain stem neurones. This was done by applying the conditioning nerve or cortical stimulus at various intervals preceding the excitatory (test) stimulus. The time course of an observed effect on a neurone was obtained by comparing, at the various conditioning-test intervals, the total number of spikes evoked in the neurone by 10-20 test stimuli with the total number evoked by a similar number of stimuli in the presence of a conditioning stimulus. In addition to the use of an electrical stimulus of the exposed nerves and cerebral cortex for conditioning stimulation, a mechanical conditioning stimulus (10 msec) applied to the canine tooth by a mechanical stimulator 35 was frequently employed. RESULTS
General aspects Microelectrode recordings were made from 457 neurones in the caudal brain stem. One-third of these neurones was recorded in decerebrate preparations, the remainder from cats anaesthetized with chloralose. There was no apparent difference between the two preparations in the response properties of neurones isolated from the solitary tract nucleus and adjacent reticular formation. The response properties of the neurones (see below) and the subsequent histological localization of the microelectrode penetrations indicated that neuronal responses were recorded, as well as from the solitary tract and its nucleus, from the motor nucleus of X I I and that part of the reticular formation lateral to this nucleus and ventral to the solitary tract nucleus. Some penetrations may also have passed through the dorsal motor nucleus of X, but methods (e.g. stimulation of X to evoke antidromic responses) to identify neurones in this nucleus were not employed. In the more lateral penetrations at depths 0.8-1.5 m m below the brain stem surface, a small field potential evoked by SLN stimulation was encountered. This potential reached its peak amplitude within the next 0.5 m m and was often coextensive with a smaller field potential resulting from IX stimulation (Fig. l, see also ref. 1), particularly at the more rostral planes. Stereotaxic coordinates and histological verification indicated that this region of SEN input was the solitary tract and its nucleus. The SLN evoked field potential typically showed two negative components. The first component had a latency of approximately 1 msec and followed SLN stimulation rates of more than 200/sec. Thus it probably represented the summed response of evoked action potentials in SEN primary afferents in the region. The later phase of negativity had a latency of 3 msec and a duration of 4-5 msec and did not follow
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Fig. 1. Evoked field potentials and neuronal responses recorded in 5 microelectrode penetrations in the caudal brain stem. The penetrations (a-e) were made in 0.5 mm steps mediolaterally in the one anteroposterior plane (2 mm anterior to obex) and are indicated in the transverse section, On the right side of the figure are illustrated the field potentials evoked by superior laryngeal nerve (SLN) and glossopharyngeal nerve (IX) stimulation and recorded at various depths (dotted arrows) of penetration c through the solitary tract and its nucleus (outlined) and the reticular formation ventral to it (vertical depth measurements were determined from readings of the electrode microdrive with an allowance of 2 0 ~ for tissue shrinkage). Single units recorded in the 5 penetrations are indicated by the various symbols on the transverse section. In the lower part of the figure are shown typical responses to SLN stimulation of a solitary tract primary fibre (open circle) at 200/sec stimulation rate, a solitary tract neurone (closed circle), and a neurone located in the adjacent reticular formation. A closed triangle indicates a neurone projecting to the rostral brain stem whereas an open triangle represents a reticular formation neurone that could not be antidromically activated from this region. Also illustrated is the response to infraorbital nerve (IO) stimulation of a reticular formation neurone (V) with only a demonstrated IO input. Negative polarity in this and subsequent figures is upwards; voltage calibration, 0.4 mV; time calibration, 4 msec for all records except the solitary tract fibre and neurone for which it is 2 msec; V, nucleus of the spinal tract of the trigeminal nerve, with the tract lateral to it; X, dorsal motor nucleus of the vagus nerve; XII, hypoglossal motor nucleus.
s t i m u l a t i o n r a t e s o f 50/sec (cf. ref. 1). T h i s s e c o n d p h a s e was also v e r y susceptible to a s p h y x i a a n d m i g h t be a t t r i b u t e d to p o s t s y n a p t i c a c t i v i t y e v o k e d by S L N s t i m u l a t i o n in s o l i t a r y t r a c t n e u r o n e s . T h e s e s h o r t - l a t e n c y S L N e v o k e d field p o t e n t i a l s w e r e p r e s e n t for a b o u t 1 m m a n d t h e n a b r u p t l y r e p l a c e d by a single w a v e o f n e g a t i v i t y w i t h a l a t e n c y o f a b o u t 4 m s e c a n d a d u r a t i o n o f 6 - 7 m s e c (Fig. 1). E v o k e d field p o t e n t i a l s o f a p p r o x i m a t e l y t h e s a m e l a t e n c y to I O , I X a n d s o m e t i m e s to X I I a n d P h a r X s t i m u l a t i o n also w e r e p r e s e n t . T h i s r e g i o n o f m u l t i p l e i n p u t in the r e t i c u l a r f o r m a t i o n h a d a v e r t i c a l e x t e n t
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of 0.5-1.0 mm. Very little activity could be recorded on deeper microelectrode penetration by stimulation of the exposed nerves. However, a field potential evoked by only IO stimulation was sometimes present, particularly in more medial penetrations passing close to the motor nucleus of XII. In these situations an antidromic field potential elicited by XII stimulation was also recorded. The findings of the response properties of single neurones will now be presented, with particular reference to the neurones recorded in the solitary tract and its nucleus, and in the reticular formation adjacent to it.
Solitary tract and its nucleus Input. A total of 167 single units were isolated in this region. All but 6 were characterized as being activated by a single stimulus to either SLN or IX stimulation. The remaining 6 neurones were activated by both SLN and IX stimulation. None of the neurones tested could be activated by auditory stimuli or by stimulation of the other isolated nerves (including the contralateral SLN), the cerebral cortex, or forepaw. Sixty-four of the units were classified as primary fibres on the basis of a shortlatency response that followed stimulation rates greater than 200/sec. The majority of units in this class, although not all (Fig. 1), also showed an initially positive-going spike response, in accordance with findings3,11 for fibres elsewhere in the central nervous system. On the other hand, units classified as brain stem neurones invariably showed negative (Fig. 1) or positive-negative (Fig. 3) spike forms and did not follow stimulation rates greater than 50/sec. Moreover, on the basis of these criteria of stimulation following rate and spike polarity, it was found that the solitary tract units separated into two latency groups (see below and Fig. 2) that corresponded closely to the two components of the SLN evoked field potential (although there was a small amount of overlap between the two latency groups). The possibility that some of the 64 units classified as primary fibres may have actually been nucleus ambiguus motoneurones was ruled out by the superficiality (histologically verified) of the recording sites and their activation by stimulation of just one peripheral nerve or of the larynx. Solitary tract neurones (and primary afferent fibres) with a laryngeal input were particularly sought for in this study, and of the 103 neurones isolated in the nucleus, 78 were driven only by SLN stimulation. Some of these were examined and found to have laryngeal mechanoreceptive fields (e.g. Fig. 3). Neurones activated by IX stimulation, although few (19) in number, tended to occur in the more rostral planes examined in the brain stem. The primary fibres and neurones appeared to be intermingled in the solitary tract nuclear region (Fig. 1) since none of the microelectrode penetrations and recordings provided any clear-cut distinction, on electrophysiological grounds, between the solitary tract and its nucleus. This was not an unexpected finding in view of the comparatively small size of the tract and nucleus and the microelectrode's capability of recording responses from neurones some distance away from the microelectrode tip. Latency and discharge characteristics. The units (57) classed as SLN primary afferent fibres showed response latencies to SLN stimulation which fell within the
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range 1.0-2.9 msec (Fig. 2) with a mean value of 1.8 msec. The conduction path from the SLN stimulation site to caudal brain stem measured 35-40 mm, thus the range of conduction velocities of the SLN primary units recorded in the solitary tract region in this study approximated 12-40 m/sec. In comparison, however, the minimum latency to SLN stimulation of neurones in this region was 2.1 msec; fewer than 2 0 ~ had latencies shorter than 2.9 msec. The mean latency value was 3.5 msec and almost 90 ~ of these solitary tract neurones had latencies less than 5.0 msec (Fig. 2). An interesting feature typical of the discharge of these neurones to SLN stimulation was that there were no more than 2 or 3 spikes evoked by the SLN stimulus. A typical response is shown in the bottom part of Fig. 1. The stimulus threshold for the first spike in most of the neurones was 0.5-1.0 mA, and increasing the stimulus intensity to m a x i m u m rarely resulted in more than two additional spikes. Antidromic activation. Bipolar stimulation in the thalamic ventrobasal complex and in the rostral brain stem was used to determine if there was a direct projection from the solitary tract nucleus to either of these regions. Thalamic stimulation failed to elicit an antidromic response in 16 solitary tract neurones tested. These and another 36 neurones tested for a projection to the rostral brain stem could not be antidromically activated from this area. Conditioning effects. A total of 61 solitary tract neurones was examined to determine if their excitatory input from SLN or IX could be influenced by conditioning stimulation. All but 3 of these neurones showed an inhibition of their excitatory response to SLN or I X stimulation. Neurones with an SLN excitatory input were particularly examined and 45 of 48 tested could be inhibited by a previous conditioning stimulus delivered to SLN. The discharge of the remaining 3 units could not be influenced by any of the conditioning stimuli used. The inhibitory effect of the SLN con-
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ditioning stimulus typically had an onset of 20 msec, a peak at 40 msec, and lasted for 300-400 msec. An example of this time course of inhibition is shown in Fig. 3. However, 20 ~o of the neurones showed an inhibition which lasted as long as 600-700 msec or as short as 150 msec (Fig. 4). A IX conditioning stimulus was also effective in inhibiting the SEN response in all but 5 of the 45 neurones inhibited by SLN conditioning (Figs. 3 and 4). An IO stimulus or a mechanical conditioning stimulus applied to the canine tooth produced inhibition of the SLN response in 10 of 28 neurones tested (Fig. 4). However, Phar X and XII conditioning stimuli were rarely effective and produced inhibition in only 3 of 31 neurones examined. Conditioning stimulation of the cerebral cortex produced a similar ratio of inhibitory effects, and forepaw and auditory stimuli had no conditioning effect in any neurone tested. Thirteen solitary tract neurones with only a IX excitatory input were tested for conditioning influences. All could be inhibited by a preceding IX conditioning stimulus and most by SLN conditioning stimulation as well. A preceding IO or canine stimulus was effective in 5 of 10 neurones tested, but Phar X (or XII) stimulation produced inhibition in only one of these neurones. A facilitatory effect of conditioning was seen in 4 solitary tract neurones. These neurones had a 1X excitatory input, and a preceding IX conditioning stimulus decreased the latency of the initial spike and increased the number of spikes produced by the control (test) IX stimulus. This facilitation lasted for about 150 msec in one neu-
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Fig. 4. Effect of various conditioning stimuli on the response of a solitary tract neurone with a superior laryngeal nerve (SLN) excitatory input. The time courses of the inhibition produced by electrical stimulation of the SLN, the glossopharyngeal nerve (IX), and the infraorbital nerve (IO), and by mechanical stimulation of the canine tooth,were obtained as outlined in Fig. 3.
rone a n d for as long as 600 msec in another. The r e m a i n i n g two n e u r o n e s were facilit a t e d for a b o u t 40 msec.
Adjacent reticularformation Input. A p a r t f r o m 9 m o t o n e u r o n e s (identified by their a n t i d r o m i c response to X I I s t i m u l a t i o n ) l o c a t e d in the m o t o r nucleus o f X I I , the r e m a i n i n g 287 n e u r o n e s rec o r d e d were isolated in the reticular f o r m a t i o n lateral to this nucleus a n d ventral to the solitary t r a c t nucleus. It is extremely unlikely t h a t a significant n u m b e r o f these n e u r o n e s was r e c o r d e d in the d o r s a l m o t o r nucleus o f X, since less t h a n 5 ~ o f t h e m c o u l d be localized directly m e d i a l to the solitary t r a c t in the region o f this nucleus. M o s t o f the reticular f o r m a t i o n n e u r o n e s h a d an i n p u t t h a t usually involved S L N , I X a n d IO, a n d often X I I . Cortical, P h a r X or c o n t r a l a t e r a l S L N , b u t n o t a u d i t o r y or forepaw, s t i m u l a t i o n was also effective in exciting some. A l t h o u g h a wides p r e a d i n p u t was a feature o f these neurones, 69 could be activated only b y s t i m u l a t i o n o f I O (Fig. 1). L i n g u a l nerve s t i m u l a t i o n was p a r t i c u l a r l y effective in activating neurones in the vicinity o f the m o t o r nucleus o f X I I . S t i m u l a t i o n o f S L N a n d I X also p r o d u c e d a s h o r t - l a t e n c y a c t i v a t i o n o f m a n y n e u r o n e s in this region. Latency and discharge characteristics. The response latencies to S L N s t i m u l a t i o n o f the reticular f o r m a t i o n n e u r o n e s with an extensive i n p u t were longer t h a n the range o f l a t e n c y values o f a p p r o x i m a t e l y 90 ~ o f solitary t r a c t neurones. F o r example, the S L N latencies o f those n e u r o n e s t h a t were r e c o r d e d j u s t ventral to the solitary t r a c t nucleus a n d p r o j e c t e d to the r o s t r a l b r a i n stem (see below) r a n g e d f r o m 4 to 12 msec with a m e a n value o f 7 msec. L o n g e r a n d m o r e variable latencies to S L N s t i m u l a t i o n were m o r e c o m m o n deeper in the reticular f o r m a t i o n . The threshold for S L N stimulat i o n was similar to t h a t o f solitary t r a c t neurones. However, the response to a single
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SLN stimulus was a burst of spikes (Fig. 1), the number of spikes being dependent on stimulus intensity. Antidromic activation. Only 22 neurones were tested but none could be antidromically activated by thalamic stimulation. However, approximately 40~o (41 of 107 neurones tested) showed an antidromic response with rostral brain stem stimulation. The majority of these neurones with an extensive input were located close to the ventral extent of the solitary tract and its nucleus (Fig. I). A similar percentage of reticular formation neurones with only an IO input also projected rostrally. Conditioning effects. The reticular formation nemones could be inhibited from many sources. Conditioning stimulation of each of the nerves that excited a neurone would inhibit the neurone's responses to the other inputs. The duration of the inhibition produced in these neurones varied between 150-700 msec. The onset and peak were typically 20 and 40 msec respectively, but in some neurones longer values were noted since an early facilitation, which had an onset of 10 msec and a duration of approximately 40 msec, preceded the inhibition.
DISCUSSION
The present study of single neurones in the brain stem revealed that many units isolated in the caudal part of the solitary tract and its nucleus have a single excitatory input from SLN or IX. In a study of the vagal input to solitary tract and nucleus ambiguus neurones, Porter la indicated that SLN stimulation would evoke unitary activity in both nuclei. L a m and Ogura 14 failed to identify a laryngeal projection to the solitary tract nucleus, but in support of the present findings and those of Porter are studies 2a, 24,28,30 reporting antidromic activation of SLN primary afferent units by stimulation in this nucleus. A IX input to some neurones is supported by the finding a2 that stimulation of oropharyngeal areas will elicit solitary tract neuronal responses. The response latencies of solitary tract neurones to SLN stimulation were in the range of 2.1-5.0 msec for about 90 ~o of the neurones. This range and the mean value of 3.5 msec are compatible with the latencies to SLN stimulation reported by Porter 19. The mean value and range of latencies noted in SLN primary afferent fibres in the region would suggest that most of the neurones were activated directly by the incoming afferents. The estimated conduction velocities of these primary afferent units indicated that more than 90 ~ were small myelinated (A6) nerve fibres, in accordance with findings15, 2s of the conduction velocities of SLN primary afferent units recorded in the nodose ganglion. T h e response of adjacent reticular formation neurones to SLN stimulation was repetitive and more quantitatively related to the stimulus intensity than responses of solitary tract neurones. Most of these neurones displayed an extensive input involving IO, SLN and IX especially. Excitation of neurones in the reticular formation from a variety of sources is an established feature of this area 2,4,25,27, but no previous report has revealed this particular combination of inputs to neurones ventral to the solitary tract nucleus. The latency values of their responses would indicate that many of the
SOLITARY TRACT NUCLEUS AND RETICULAR FORMATION
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neurones receive a direct excitatory input from the solitary tract nucleus. Some of these neurones may act as the source of inhibitory interneurones influencing the discharge of solitary tract neurones. But 40 ~o were found to project rostrally to the vicinity of the trigeminal motor nucleus. This projection is supported by findings in this laboratory (Sessle et al., in preparation) of short-latency inputs to this region evoked by SLN and IX stimulation. The functional significance of this relay cannot be defined at this time, but some integrative action in swallowing and mastication or in protective jaw reflexes seems likely. The orofacial input to the reticular formation neurones probably involved at least a relay in the trigeminal sensory nuclear complex. Anatomical studies in man a4, monkey 22, rat aa, and cat12, la have described trigeminal primary afferent fibres passing to the vicinity of the solitary tract nucleus. Sumi az has also reported that some neurones in the rostral part of this nucleus may be activated by tooth stimulation. However, in the present study, no neurones in the caudal part of the nucleus could be activated by IO stimulation. This would indicate that the trigeminal afferents described anatomically in the nucleus either do not terminate here but end on neurones in the adjacent reticular formation or more rostral solitary tract neurones, or carry information (e.g. gustatory) to the nucleus via other branches of the trigeminal nerve. A trigeminal input was also noted for neurones adjacent to the motor nucleus of XII but many of these neurones could also be activated by SLN and IX stimulation. Such neurones have not been described previously in this region, although Lam and Ogura 14 noted in the reticular formation close to this nucleus an evoked field potential with SLN stimulation. The neurones are possibly involved in a reflex regulation of hypoglossal motoneurones in a manner similar to the lingual-hypoglossal reflex2°, 21, since SLN (or IX) stimulation produces a short-latency activation of the tongue musculature and a subsequent, prolonged inhibition of its activity z°. Solitary tract neurones could be inhibited from SLN or IX, but auditory stimulation or stimuli applied to Phar X, XII, forepaw, and cerebral cortex were rarely effective. These findings would indicate that neurones innervating visceral structures display the phenomenon of afferent or surround inhibition that has been described in somatic, auditory and visual relay areas and has been implicated in spatial discrimination 10. However, some neurones could also be inhibited by electrical or natural stimulation of the orofacial region, which is innervated by a somatic nerve, and the following paper z8 indicates that presynaptic depolarization may account largely for these effects. A regulatory role in reflex activities for such orofacial conditioning stimulation is discussed elsewhere ao.
ACKNOWLEDGEMENTS
I thank Mr. M. Kalovsky and Miss B. Holmwood for their technical assistance and Drs. Arthur Storey and Ronald Dubner for critically reading the manuscript. This study was supported by Grant D.G. 73 from the Canadian Medical Research Council.
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REFERENCES 1 BISCOE,T. J., AND SAMPSON,S. R., Field potentials evoked in the brain stem of the cat by stimulation of the carotid sinus, glossopharyngeal, aortic and superior laryngeal nerves, J. Physiol. (Lond.), 209 (1970) 341-358. 2 BiSCOE,T. J., AND SAMlaSON, S. R., Responses of cells in the brain stem of the cat to stimulation of the sinus, glossopharyngeal, aortic and superior laryngeal nerves, J. Physiol. (Lond.), 209 (1970) 359-373. 3 BisHot', P. O., BURKE, W., AND DAVIS, R., The identification of single units in central visual pathways, J. Physiol. (Lond.), 162 (1962) 409-431. 4 BOWSHER,D., Place and modality analysis in caudal reticular formation, J. Physiol.(Lond.), 209 (1970) 473-486. 5 DARIAN-SM1TH,I., Neural mechanisms of facial sensation, Int. Rev. NeurobioL, 9 (1966) 301-395. 6 DARIAN-SMITH,I., Somatic sensation, Ann. Rev. Physiol., 31 (1969) 417-450. 7 DOTY, R. W., Neural organization of deglutition. In C. F. CODE (Ed.), Handbook of Physiology, Vol. IV, Sect. 6, Amer. Physiol. Soc., Washington, 1968, pp. 1861-1902. 8 DUBNER, R., AND SESSLE, B. J., Presynaptic modification of corticofugal fibers participating in a feedback loop between trigeminal brain-stem nuclei and sensorimotor cortex. In R. DUBNERAND Y. KAWAMURA(Eds.), Oral-Facial Sensory and Motor Mechanisms, Appleton-Century-Crofts, New York, 1971, pp. 299-314. 9 DUBNER,R., AND SESSLE, B, J., Presynaptic excitability changes of primary afferent and corticofugal fibers projecting to trigeminal brain stem nuclei, Exp. NeuroL, 30 (1971) 223-238. 10 HUaEL, D. H., Tungsten micro-electrode for recording from single units, Science, 125 (1957) 549550. 1 l HUBEL, O. H., Single unit activity in lateral geniculate body and optic tract of unrestrained cats, J. Physiol. (Lond.), 150 (1960) 91-104. 12 KERR, F. W. L., Structural relation of the trigeminal spinal tract to upper cervical roots and the solitary nucleus in the cat, Exp. Neurol., 4 (1961) 134-148. 13 KERR, F. W. L., The divisional organization of afferent fibres of the trigeminal nerve, Brain, 86 (1963) 721-732. 14 LAM, R. L., AND OGURA, J. H., Afferent projection of the superior laryngeal nerve in the brain stem, Neurology (Minneap.), 4 (1954) 630-632. 15 MEI, N., Disposition anatomique et propri6t6s 61ectrophysiologiques des neurones sensitifs vagaux chez le chat, Exp. Brain Res., 11 (1970) 465-479. 16 MOUNTCASTLE,V. B., ANDDARIAN-SMITH,I., Neural mechanisms in somesthesia. In V. B. MOUNTCASTLE(Ed.), MedicalPhysiology, Mosby, St. Louis, Mo., 1968, pp. 1373-1423. 17 O'BRIEN, J. n., PIMPANEAU, A., AND ALBE-FESSARD,O., Evoked cortical responses to vagal, laryngeal and facial afferents in monkeys under chloralose anaesthesia, Electroenceph. clin. Neurophysiol., 31 (1971) 7-20. 18 OGURA, J. H., AND LAM, R. L., Anatomical and physiological correlations on stimulating the human superior laryngeal nerve, Laryngoscope (St. Louis), 63 (1953) 947-959. 19 PORTER, R., Unit responses evoked in the medulla oblongata by vagus nerve stimulation, J. Physiol. (Lond.), 168 (1963) 717-735. 20 PORTER, R., Synaptic potentials in hypoglossal motoneurones, J. Physiol. (Lond.), 180 (1965) 209-224. 21 PORTER,R., The synaptic basis of a bilateral lingual-hypoglossal reflex in cats, J. Physiol. (Lond.), 190 (1967) 611-627. 22 RHOTON, A. L., O'LEARY, J. L., AND FERGUSON,J. P., The trigeminal, facial, vagal and glossopharyngeal nerves in the monkey, Arch. NeuroL (Chic.), 14 (1966) 536-540. 23 RUDOMIN,P., Presynaptic inhibition induced by vagal afferent volleys, J. Physiol. (Lond.), 30 (1967) 964-981. 24 RUDOMIN, P., Excitability changes of superior laryngeal, vagal and depressor afferent terminals produced by stimulation of the solitary tract nucleus, Exp. Brain Res., 6 (1968) 156-170. 25 SCHEIBEL,M., SCHEIaEL, A., MOLLICA, A., AND MORUZZI, G., Convergence and interaction of afferent impulses on single units of reticular formation, J. Neurophysiol., 18 (1955) 309-331. 26 SCnMITT,A., YU, S.-K. J., AND SESSLE, B. J., Excitatory and inhibitory influences from laryngea and orofacial areas on tongue position in cat, Arch. oral Biol., (1973) in press. 27 SEGUNOO,J. P., TAKENAKA,T., AND ENCABO, H., Somatic sensory properties of bulbar reticular neurones, J. Neurophysiol., 30 (1967) 1221-1238.
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28 SESSLE,B. J., Presynaptic excitability changes induced in single laryngeal primary afferent fibres, Brain Research, 53 (1973) 333-342. 29 SESSLE,B. J., AND DUBNER, R., Presynaptic depolarization and hyperpolarization of trigeminal primary and thalamic afferents. In R. DUBNERAND Y. KAWAMURA(Eds.), Oral-Facial Sensory and Motor Mechanisms, Appleton-Century-Crofts, New York, 1971, pp. 279-298. 30 SESSLE,B. J., AND STOREY,A. T., Periodontal and facial influences on the laryngeal input to the brain stem of the cat, Arch. oral Biol., 17 (1972) 1583-1595. 31 STOgEY, A.T., Extra-trigeminal sensory systems related to oral function. In J. F. BOSMA(Ed.), Symposium on Oral Sensation and Perception, Thomas, Springfield, Ill., 1967, pp. 84-97. 32 SUMI, T., Neuronal mechanisms in swallowing, Pfliigers Arch. ges. Physiol., 278 (1964) 467-477. 33 TORVIK,A., Afferent connections to the sensory trigeminal nuclei, the nucleus of the solitary tract and adjacent structures - - an experimental study in the rat, J. comp. Neurol., 106 (1965) 51-141. 34 WALLENBERG,A., Das dorsale Gebiet der spinalen Trigeminuswurzel und seine Beziehungen zum solit/iren Biindel beim Menschen. Ein Beitrag zur Anatomie und Physiologie des Trigeminus, Dtsch. Z. Nervenheilk., 11 (1897) 391-405. 35 WERNER,G., AND MOUNTCASTLE,V. B., Neural activity in mechanoreceptive cutaneous afferents: stimulus-response relations, Weber functions and information transmission, J. Neurophysiol., 28 (1965) 359-397.