Projection of taste nerve afferents to anterior opercular- insular cortex in squirrel monkey (Saimiri sciureus)

BRAIN RESEARCH

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P R O J E C T I O N OF T A S T E N E R V E A F F E R E N T S TO A N T E R I O R O P E R C U L A R I N S U L A R C O R T E X I N S Q U I R R E L M O N K E Y (SAIMIRI SCIUREUS}

ROBERT M. B E N J A M I N AND H A R O L D BURTON*

Laboratory of Neurophysiology and Department of Physiology, University of Wisconsin Medical Center, Madison, Wis. (U.S.A.) (Accepted August 1st, 1967)

INTRODUCTION

The background for these studies was elaborated in the preceding paper 7. The purpose of the experiments was twofold: first, to locate potential taste areas by mapping the distribution of cortical responses evoked by electrical stimulation of the nerves that innervate the tongue; secondly, to ablate these areas to determine which would cause retrograde degeneration of the thalamic relay for taste. This paper will describe the results obtained from opercular-insular cortex; the preceding paper described those from somatic sensory area I. PROCEDURE

Most of the techniques used in these experiments were described in the preceding paper 7. Differences and additions will be noted here. Steel microelectrodes 9 were used exclusively for recording in depth. Critical sites were marked with microlesions (75 # A DC for 10 sec, electrode negative). The electrophysiological data were recorded in 17 preparations. Ten animals were used in the thalamic degeneration studies. Survival times ranged from 30 to 40 days. RESULTS

Electrophysiological Both of the taste nerves (the chorda tympani and the lingual-tonsilar branch of the glossopharyngeal) were found to have ipsilateral projections to anterior opercular-insular cortex. The data from six chorda tympani experiments were combined in Fig. 1. In all cases the intact ipsilateral nerve was stimulated electrically in the middle ear, the response in somatic sensory area I (S I) was monitored continuously * National Institutes of General Medical Sciences Predoctoral Fellow (I FI GM-29,882-01).

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Fig. 1. Reconstruction of the electrode tracks from six experiments with ipsilateral stimulation of the chorda tympani nerve. The level of the five standard coronal sections is marked on the outline drawing and the boundaries of the insula have been projected on to the surface. The segments of punctures which were responsive are located on the upper set of sections and the unresponsive sites on the lower set. The claustrum is outlined. with a gross surface electrode and the cortex buried within the sylvian fissure was probed with a microelectrode. The locations of the responsive sites were plotted on the upper set of five coronal sections (Fig. 1). The most rostral responses (section 2) were subjacent to the rostral extension of the claustrum. In sections 2 and 3, the active cortex gradually becomes thicker before the most rostral part of the superior limiting sulcus finally develops in section 4. At that level, the responses were confined to the inside of the operculum a n d did n o t extend to insular cortex as defined in terms of gross morphology. Before the fissure forms, the distinction between opercular and insular cortex is n o t clear and the location of the active region may be best Brain Research, 7 (1968) 221-231

TONGUE PROJECTIONSTO OPERCULUM AND INSULA

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described as the c3rtex which caps or wraps around the rostral part of the superior limiting sulcus. The total A - P extent of the active cortex was approximately 3 mm. The one active puncture in section 5 marks the anterior border of the chorda tympani projection area in S I. The lower set of sections plots (Fig. 1) punctures or segments of punctures which were unresponsive. They completely surround and isolate the active focus. The location of the electrode within the cortical layers could often be predicted from the polarity of the response. For example, the first response encountered in the upper three punctures of section 4 (Fig. 1) was negative and, as the electrode penetrated more deeply toward the surface of the cortex inside the operculum, the polarity reversed to positive. The most ventral of the two responsive tracks in section 2 was 12345 IPSILATERAL IX

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positive throughout its extent and predictably confined to the upper layers of the cortex. The other track, dorsal to the first in the deeper cortical layers, was negative throughout. The largest negative response usually had the shortest latency. In five experiments the values ranged from 8 to 11 msec; the mean was 9.0 msec. The latency of the largest surface positive response from S I in the same preparations averaged 6.4 msec. The latency of the opercular-insular response was always 2-3 msec longer than the S I surface response. The results from electrical stimulation of the lingual-tonsilar branch of the IXth nerve are plotted in Fig. 2. The active region did not extend quite as far rostrally as the chorda tympani projection (cf. section 2) and reached further dorsally and medially in its posterior extent (cf. section 4). Whether insular cortex was involved is equivocal. In two experiments the latencies for the largest negative responses were 9.0 and 10 msec respectively, 2.0 and 3.0 msec longer than the largest surface positive response in S I. No other tongue nerve projections to this anterior opercular-insular cortex were found. Electrical stimulation of the contralateral chorda tympani and IXth nerves and the contralateral and ipsilateral lingual nerves, which presumably contain no taste fibers, produced only negative results in a total of 8 experiments. It is 1234

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quite unlikely that any substantial projection from these nerves was missed. The region active to ipsilateral taste nerve stimulation and the surrounding cortex were thoroughly explored. A surface monitor in S I assured that the nerves were functioning. In some experiments the ipsilateral chorda tympani was stimulated simultaneously. Ipsilateral responses were always located indicating that the cortex was functional. Mechanical stimulation of the tongue was always tested in experiments involving electrical stimulation of the intact chorda tympani in the middle ear. In this type of preparation all the tongue afferents were still connected to the central nervous system. No responses to mechanical stimulation were recorded from buried cortex, but were easily evoked from the tongue projection in S I. The most reasonable conclusion that can be drawn from these data is that only the two ipsilateral taste nerves project to anterior opercular-insular cortex and that the projections are not from mechanoreceptors. A composite map (Fig. 3) combining all the responsive sites for the two nerves (Figs. 1 and 2) was constructed to serve as a guide for ablation studies. The solid area in the upper diagram is the opercular-insular projection drawn on the lateral surface of the brain. The stipples demark the equivalent composite map for the S I responses, but also includes the contra- and ipsilateral lingual areas 7. A wedge of cortex (sections 3 and 4) separates the two responsive regions. The only possible connection, if one exists, would have to be just caudal to section 4.

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Fig. 4. The sylvian fissure has been spread open to expose the buried cortical surfaces. See text. Other cortex within the sylvian fissure was explored extensively. In Fig. 4 the fissure has been spread open to expose the five enfolded cortical surfaces. Proceeding dorsoventrally these are: the upper bank of the sylvian fissure or the lip of the frontoparietal operculum; the cortex overlying the dorsal half of the insula or the inside of the operculum; the insula proper; the temporal cortex overlying the ventral half of the insula; and finally, the lower bank of the sylvian fissure or superior temporal plane. The approximate location of the anterior opercular-insular taste nerve focus is marked, as are somatic sensory area II (S II) as defined by potentials evoked from Brain Research, 7 (1968) 221-231

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mechanical stimulation of the body surface 5 and auditory cortex including A I as defined by potentials evoked by click stimulation 10. Twelve experiments were devoted to exploration of two of these enfolded cortical surfaces, the upper bank of the sylvian fissure (opercular lip) and the insula. They were exposed by aspiration of either temporal or parietal tissue and recording accomplished with a 27 gauge stainless steel surface electrode. N o responses from electrical stimulation of the two taste nerves or the ipsilateral lingual nerve were found. A strong projection from the contralateral lingual was demonstrated (60-201) and is marked by parallel lines on the diagram (Fig. 4). The responsive region included the appropriate part of S !I, but extended more rostrally on the upper bank o f the sylvian fissure. The extension beyond S II may be due to the relative efficiency o f an electrical as compared to a mechanical stimulus. When parietal c~rtex was spared, S I responses were monitored as previously and S II was proved to be functional with mechanical stimulation of the body surface.

Thalamic degeneration Successful unilateral aspiration of the responsive opercular-insular cortex was accomplished in 3 animals. All of the region was removed with minimal damage to overlying tissue and minimal invasion of S I or its projection fibers. A reconstruction of a representative case is presented in Fig. 5. There was no observable cell loss 65-275-L

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Fig. 5. Reconstruction of a lesion of the opercular-insular cortex. The level of the five pairs of coronal sections is marked on the outline drawing. The right section of each pair is a tracing from the experimental brain, the left section from a normal brain with dots approximating the removed tissue. or gliosis in the ventrobasal or ventromedial thalamic nuclei. Thus the thalamic relay for taste 2 in the ventromedial complex was preserved, as it was after removal of the tongue nerve projection areas in S I (ref. 7).

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Fig. 6. Left: Reconstruction of a lesion removing both tongue nerve projection areas. See Fig. 5 for description. Right: Resulting thalamic retrograde degeneration in the ventrobasal (VB) and ventromedial (VM) nuclear complexes subjacent to the centre median (CM). The four coronal sections are spaced approximately 0.5 mm apart, most anterior level at the top, most posterior at the bottom.

Fig. 7. Photomicrograph at the same level as thalamic reconstruction, section 3, Fig. 6. R e t r o g r a d e d e g e n e r a t i o n o f t h a l a m i c ' t a s t e ' n e u r o n s does occur if b o t h the o p e r c u l a r - i n s u l a r a n d S I p r o j e c t i o n areas are r e m o v e d in the same p r e p a r a t i o n . O n e case has been r e c o n s t r u c t e d in Fig. 6. T h e lesion was deficient in one respect. T h e m o s t caudal part o f the o p e r c u l a r - i n s u l a r p r o j e c t i o n was spared (cf. cortical

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section 4 of Fig. 6 with section 4 of Fig. 3). In spite of this, almost all of the ventromedial complex was degenerated (see Fig. 7). At the time this lesion was performed the opercular-insular taste nerve projection had not been discovered. Its inclusion in the lesion was merely a fortuitous result of poor surgery. This case is the only available one in which the two projection areas were lesioned in isolation without damage to other cortex. Fourteen other, more extensive lesions, of the same vintage provide evidence supporting the degeneration results of this single case. The intent was to ablate all of the S I taste nerve projection areas and, in addition, to remove all of the frontoparietal operculum to the caudal border of S 1I (see Fig. 4). In most cases the underlying insular cortex was also aspirated. Most of the lesions were bilateral and most of the animals were tested behaviorally for taste discrimination deficits. The S I taste nerve areas were successfully ablated in all cases. Seven lesions included essentially all of the opercularinsular projection as well, resulting in almost complete degeneration of the ventromedial complex. The other seven lesions spared varying amounts of the opercularinsular projection with variable degeneration of the ventromedial complex. None of the animals had deficits in taste discrimination, but no animal had complete bilateral destruction of all of the projection areas. DISCUSSION

Cortical responses evoked by electrical stimulation of the chorda tympani nerve have been mapped in the rat 4, the cat s,14 and the dog as. Only one cortical projection area was described for each of these species. Unpublished experiments on the rhesus monkey 6, however, have located two spatially distinct projections quite analogous to those in the squirrel monkey. Is the double projection a unique characteristic of primates or has a second area been overlooked in these other animals ? The chorda tympani nerve projection described for the cat is certainly homologous to the S I projections in the squirrel monkey. It lies in the appropriate part of the somatotopic sequence of the feline S I area. It probably does not include the equivalent of the second projection, that is, the opercular-insular projection overlying the claustrum, because, as the authors carefully point out, it lies rostral to claustral cortex 14. If a second projection exists in the cat, it is most likely to be located in buried claustral cortex near the confluence of the rhinal fissure and the presylvian sulcus. The relationships should be similar for the dog. The rat presents a problem. The single taste nerve area has some characteristics of both of the monkey projection areas. It has the appropriate spatial relationships with S 1, but part overlies the poorly developed rat claustrum 3. Perhaps this single area contains both projections, undifferentiated by evoked potential technique in this very small brain. The observation that lesions of the area result in transitory taste deficits and degeneration of the rat thalamic taste relay provides suggestive, though not conclusive evidence for this speculation 3. The functional significance of the double projection is obscure at best. Either area can be removed without demonstrable effect on the squirrel monkey's taste Brain Research, 7 (1968) 221-231

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sensitivity as measured by the preference technique 7. Unfortunately, we have no behavioral data on the effects of bilateral destruction of both areas together. Substantial impairment of taste discrimination should resultL Both areas must be removed to cause retrograde degeneration of the thalamic taste relay in the ventromedial complex. Individual lesions had no discernable effect. By definition then, thalamic 'taste' neurons have only sustaining, not essential, projections to squirrel monkey cortex. The same relationships were found between somatic sensory area II and auditory area I and the posterior thalamic nuclear complex (PO) of the cat 16. Removal of both cortical areas was necessary to cause retrograde degeneration of PO neurons. Much of the opercular and insular cortex of the rhesus monkey receives only sustaining thalamic projections~2,15. The anatomical substrate accounting for sustaining projections is not known. The simplest hypothesis evokes a bifurcating axon, one branch to each cortical area. The thalamic cell body will not degenerate unless both branches are damaged. This model has been diagrammed in Fig. 8. The bifurcating axon of a

Fig. 8. Diagram of hypothetical thalamo-cortical connections. See text. thalamic 'taste' neuron sends one branch to terminate in S I and the other to opercularinsular cortex. Presumably, the bifurcation could occur at any point along the axon. S I has essential projections from the more laterally situated ventrobasal complex which is responsive to thermal and mechanical stimulation of the tongue 2. The model does not explain why responses to electrical stimulation of the taste nerves are 2-3 msec longer in the opercular-insular area than in S I . Differences in fiber diameter is one possibility, but extra synapses in the bulbo-cortical pathway cannot be discounted. If a 'pure' cortical taste area does exist, it is not likely to be in S I. All of the taste nerve areas there were also responsive to mechanical stimulation of intra- or perioral structures. In the chorda tympani S I cortex of the cat, single units responsive to gustatory stimulation of the tongue were found intermingled with neurons responsive to thermal and mechanical stimulationS, 11. The logical candidate for a pure taste area uncontaminated by other lingual modalities is the opercular-insular projection. It was responsive only to electrical stimulation of the two taste nerves and not to mechanical stimulation of the tongue. The projection was ipsilateral. All of the thalamic neurons in the squirrel monkey responded only to gustatory stimulation of the ipsilateral side of the tongue 2. A recent report describes ipsilateral taste deficits from cortical lesions in man 13. Brain Research, 7 (1968) 221-231

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A subsequent p a p e r describes the cyto- and myeloarchitectonic characteristics o f these two areas and speculates on their phylogenetic d e v e l o p m e n t iv. SUMMARY

Cortex buried within the sylvian fissure o f deeply anesthetized squirrel m o n k e y s was p r o b e d with steel microelectrodes to record slow wave responses evoked by electrical s t i m u l a t i o n o f the three nerves innervating the tongue. A responsive locus was located in the most a n t e r i o r o p e r c u l a r - i n s u l a r cortex. Only s t i m u l a t i o n o f the two ipsilateral taste nerves (the c h o r d a t y m p a n i and the lingual-tonsilar branch o f the glossopharyngeal) was effective. Electrical s t i m u l a t i o n o f the contralateral taste nerves, the c a n t r a - a n d ipsilateral lingual nerves and mechanical s t i m u l a t i o n o f the tongue evoked no responses in this region. A b l a t i o n o f the responsive region did not cause any r e t r o g r a d e d e g e n e r a t i o n o f the t h a l a m i c taste relay in the v e n t r o m e d i a l nuclear complex. If the o t h e r taste nerve p r o j e c t i o n areas located in somatic sensory area I were included in the lesion, c o m p l e t e degeneration o f t h a l a m i c taste neurons resulted. It was concluded that the taste system has only sustaining projections to the neocortex. ACKNOWLEDGEMENTS

We wish to express our a p p r e c i a t i o n to Miss Helen Brandemuehl, Mrs. Isabel Lucey and Mrs. Jo A n n Ekleberry for p r e p a r a t i o n o f the histological material and to Mr. Terrill Stewart for excellent p h o t o g r a p h y . This investigation was s u p p o r t e d by G r a n t s NB1932, NB5326 and NB06225 f r o m N a t i o n a l Institutes o f Health.

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10 HIND, J. E., BENJAMIN, R. M., AND WOOLSEY>C. N., Auditory cortex of squirrel monkey (Saimiri sciureus), Fed. Proc., 17 0958) 71. 11 LANDGREN, S., Convergence of tactile, thermal, and gustatory impulses on single cortical cells, Actaphysiol. scand., 40 (1957) 210-221. 12 LOCKE, S., Thalamic connections to insular and opercular cortex of monkey, J. comp. Neurol., 129 (1967) 219-240. 13 MOTTA, G., I centri corticali del gusto, Boll. Sci. reed., 131 (1959) 480-493. 14 PATTON, H. O., AND AMASS1AN, V. E., Cortical projection zone of chorda tympani nerve in cat, J. Neurophysicl., 15 0952) 245-250. 15 ROBERTS, T. S., A~,!DAKERT, K., Insular and opercular cortex and its thalamic projection in Macaca mulatta, Schweiz. Arch. Neurol. Neurochir. Psychiat., 92 (1963) 1-43. 16 ROSE, J. E., AND WOOLSEY, C. N., Cortical connections and functional organization of the thalamic auditory system of the cat. In H. F. HARLOW AND C. N. WOOLSEY (Eds.), Biological and Biochemical Bases of Behavior, Univ. Wisconsin Press, Madison, Wisc., 1958, pp. 127-150. 17 ~At',IDES, F., The architecture of the cortical taste nerve areas in squirrel monkey (Saimiri sciureus) and their relationships t3 insular, sensorimotor and prefrontal regions, Brain Research, in press. 18 SANTIBANEZ, G., TARNECI
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