516

Brain Research, 93 (1975) 516-524 © Elsevier ScientificPublishing Company, Amsterdam - Printed in The Netherlands

Thalamic, callosal and reticular converging inputs to parietal association cortex in cat

ANNE K1TSIKIS ANDMIRCEA STERIADE Laboratoire de Neurophysiologie, Ddparternent de Physiologie, Facultd de Mddecine, Universitd Laval, Quebec (Canada)

(Accepted May 5th, 1975)

In view of studying changes in excitatory-inhibitory sequences of association cortex neurons during different levels of vigilance, it was first necessary to explore the input-output organization of the investigated area. This introductory paper therefore deals with the synaptic inputs of parietal associative cortex in cat, as revealed by surface and depth responses to stimulation of appropriate thalamic nuclei, homotopic points in the contralateral cortex and mesencephalic reticular formation (RF). A great deal of experiments undertaken in the last two decades revealed the cat's suprasylvian gyrus to be the site of heterogenous sensory interactions and emphasized the unequal convergence of various peripheral inputs on different recorded areas (see review in ref. 2). Much of this work was conducted under chloralose anesthesia which artificially enhances the evoked responses and, as a rule, natural sensory stimuli were used to determine the projection modalities. It should be stressed that in an analytical study on fluctuations in temporal patterns of cortical responses during the sleep-waking cycle it is undoubtedly preferable to employ testing shocks applied to central pathways just before the investigated synapses since, with peripheral stimuli, competitive alterations occurring at multiple intercalated subcortical relays may obscure the knowledge on cortical excitability. In the few studies using thalamic stimulation and suprasylvian cortical recording, the thalamic site was designated as the lateralis posterior (LP) nucleus or, even more vaguely, as the 'pulvinar-lateralis posterior' (Pul-LP) complex. Recent anatomical work by Graybiel a emphasized, however, the considerable regional distribution within this thalamic territory and demonstrated the specific pattern of projection of various regions of the pulvinar-posterior system upon the cerebral hemisphere. Particularly relevant to the results of the present paper, she revived a 40-year-old thalamic systematization 7,9 and showed that the rostral and dorsal part of the thalamic region, usually labeled LP, constitutes an entity (lateralis intermedius, LI) with distinct projections to the cortex. The present experiments essentially show that direct synaptic inputs from LI and homotopic contralateral cortical points converge with oligosynaptic projections arising in the mesencephalic tegmentum on layers II[ and IV of the crown of areas 5b and 7.

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Fig. 1. Distribution of cortical responses to single shock stimulation of LI-LP and VB thalamic nuclei. Two thalamic descents: A9.5, L5, D6-1; and A8, L5, D6-1. Surface cortical recordings in the primary somatosensory area (SI), parietal associative areas 5 and 7 (points 1,2 and 3) and middle suprasylvian gyrus (points 4, 5 and 6). Fifty averaged responses. Note: prevalent distribution of the rostrally located LI nucleus (descent at A9.5, according to frontal plane coordinates indicated by Ingram et al. 6) to anterior suprasylvian areas 5 and 7; more posterior thalamic points in the LI-LP nuclei (A8) project heavily to the middle suprasylvian gyrus when stimulating at D6 4, while clearcut but smaller amplitude responses appear also in anterior areas 5 and 7 by stimulating the LP at D3; lack of LI- and LP-evoked potentials posterior to the middle suprasylvian point 5; elective occurrence of VB-evoked (at ventral points of A9.5, and at A8, D1) responses in S1 with typical spike-like positive deflections. In this and subsequent figures, positivity downwards.

518 Experiments were carried out on locally anesthetized, enc6phale isol6 cats, with control of the CO2 content of the expired air at 3.8-4 ~. Stimulation was applied through coaxial electrodes (the tip, 40 #m in diameter, was 0.7 mm apart from the de-insulated ring, resulting in impedances of 40-60 kf~) inserted in the LI-LP and the underlying ventrobasal (VB) thalamic nuclei (A8-9.5, L4-6, D6-1), the deep layers of the symmetrical contralateral cortical areas or white matter just below, and the RF (A3, L2.5, D - - 1). Stimuli (0.1 msec) were adjusted not to exceed 0.2 mA in the case of thalamic and transcallosal activation. Up to this intensity, physical spread of current did not exceed 1 ram, as shown by disappearance of anterior and middle suprasylvian responses when leaving LI nucleus (A9.5, L5, D6-4) and entering the VB complex (see Fig. 1). RF stimulation was applied with brief (40-70 msec) trains (250-350/sec) or single shocks (pulse duration 0.1 msec) at 0.04-0.06 mA, an intensity which represents about 80 ~ of that required to elicit EEG activation and pupillary dilatation. The location of deep stimulating electrodes was established after routine histological verification. Surface recordings were performed with small (1 mm) silver balls placed on the anterior marginal, middle and anterior suprasylvian, and posterior sigmoid-coronal gyri, as indicated in Fig. 1. For depth recordings of evoked responses, semi-microelectrodes of platinum blacked stainless steel (5-10/~m at the tip) were inserted perpendicularly to the crown cortical surface, avoiding rim areas. The depth was estimated from readings on the micromanipulator combined with histological verification following small lesions through the microelectrode advanced into the white matter. Fifty evoked responses were averaged by means of an Intertechnique Didac 800 analyzer. The few data on unit activity reported here were obtained by using fine (1 #m) platinum blacked stainless steel or tungsten microelectrodes. All recordings were monopolar and were performed with a bandwidth of 1-3,000 Hz. Fig. 1 shows that the rostrally located (A9.5) LI nucleus preferentially projects to the anterior parts of the suprasylvian and lateral gyri (points 1, 2 and 3) which correspond to areas 5 and 7 in the cortical map by Hassler and Muhs-Clement 5. Stimulation along a more caudal track (A8) elicited maximal responses in the middle suprasylvian gyrus (points 4 and 5). Thalamically evoked potentials exhibited the features of primary specific responses: initial surface-positivity, with wave initiation at 0.7-1.2 msec and peak latency at around 3-3.5 msec. A second positivity, with initiation at 8-10 msec and peak latency at 13-15 msec, appeared in area 7. This secondary wave further developed into augmenting responses with repetitive stimuli at 8-12/sec. In agreement with morphological data a, ventral points in the A9.5 descent belonging to the VB complex did not project to suprasylvian gyrus; the small amplitude deflections seen at points 1-3 were presumably volume conductor recordings from the huge, classical spike-like evoked potential obtained in the neighbouring SI, and they contrasted with clear-cut responses of the same suprasylvian points evoked by stimulating more dorsal (LI) thalamic areas (D4-6). It is worth noting in Fig. 1 that specific thalamocortical responses in areas 5-7 and middle suprasylvian gyrus lack the three initial spike-like surface-positive deflections characterizing SI (and visual) cortical responses to VB (and lateral geniculate,

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Fig. 2. Laminar distribution of responses evoked in area 5 by thalamic LI, transcallosal and mesencephalic RF stimulation. Evoked potentials (50 averaged sweeps) were recorded by means of a semimicroelectrode at different indicated depths along a track on the crown of the anterior suprasylvian gyrus (A14, L10). Single shocks (arrows) delivered to the LI thalamic nucleus (A9.5, L5, D6) and depth layers of the homotopic contralateral point (Call.); a 60 msec train of 330/sec pulses (horizontal bar) was applied to the RF (A3, L2.5, D - - 1). See text. LG) thalamic nuclei, thus resembling the motor cortical mass response to ventrolateral (VL) thalamic stimulation which also lacks these successive rapid deflections 1°. Responses with initial spike-like deflections in suprasylvian areas, mentioned by some authors, were only obtained in our experiments by increasing the stimulation intensity above 0.2 mA and probably causing spread of current from LI-LP nuclei to the VB complex and recording in volume conductor from SI area. These dissimilar response patterns (precruciate and suprasylvian v e r s u s SI and visual areas) likely reflect a different synaptic organization of thalamocortical pathways. Interestingly enough, thalamocortical suprasylvian evoked potentials, like VL-evoked motor precruciate responses 10, strikingly diminished in amplitude during RF-elicited E E G activation (see Fig. 4), thus contrasting with the well-known enhancement of VB- and LGevoked responses in SI and visual cortices under the same experimental conditions. The synaptic inputs of thalamic, contralateral cortical and RF origin converge onto the same pool of anterior suprasylvian neurons. This is shown in Fig. 2 depicting the LI-, transcallosally- and RF-evoked responses along a semi-microelectrode track

520 in an area which corresponds to the cytoarchitectonic division 5b 5. The salient features of LI and transcallosally elicited excitatory-inhibitory sequences may be summarized as follows. (1) The depth-reversal of the LI-evoked primary surface-positive potential began at 0.5 mm and the primary depth-negativity, reflecting the initial excitation, was found to be maximal from 0.7 to 1 mm. Complete reversal of thetranscallosally elicited primary potential was already seen at 0.4 mm (and, in some instances, even at 0.3 mm; see Fig. 4), more superficially than for LI, but maximum amplitude of the depth-negativity was found at around 0.7 mm. This suggests that termination of the LI-fugal fibers mostly concentrates in layer IV and only to some extent in the lower part of layer III and in layer V, while callosal fibers extensively ramify from the lower part of layer II and upper part of layer III to a maximum condensation within layer IV (correlations established by comparing the depths of the present recordings to the layer disposal in area 5b depicted by Hassler and Muhs-Clement ~ in their Figs. 30 and 31). The laminar distribution indicated here fits well with the pattern of anterograde degeneration of thalamofugal fibers to areas 5 and 7 in the morphological study by Graybiel 4. (2) Other features of thalamically and callosally evoked responses appeared when analyzing the late events, subsequent to the initial excitation. A long-lasting (around 100 msec), slow positive wave was observed to follow, in both types of responses, the primary depth-negative response. This positive component, associated with silent periods of firing in unit recordings, presumably reflects hyperpolarizing potentials. It had the same time course as the intracellularly recorded inhibitory potentials seen in anterior suprasylvian cells following LP thalamic stimulation 3. It can be therefore postulated that the slow positive wave recorded deeper than 0.4 mm and maximal in layer IV is generated by inhibitory synaptic actions exerted there on neurons receiving primary afferents. (3) The inhibitory component of the LI-evoked response was followed by a high-amplitude, depth-negative wave (horizontal arrow in Fig. 2), superimposed by spike discharges. This wave occurred at 0.5 mm, was maximal at 0.7 mm, and diminished progressively below this depth. This component was 40-150~o higher in amplitude than the early depth-negative wave at 0.5 mm, while at a depth of 1.3 mm it had only 80-90~ of the amplitude of the early component. No such late potential occurred following transcallosal stimulation, in spite of the fact that the hyperpolarizing slow positive shift following the callosally elicited initial excitation was at least as developed as in the case of thalamic LI stimulation. This may lead to the assumption that the post-inhibitory excitatory potential does not evolve as a mere consequence of preceding inhibition (the so-called 'rebound'), but requires a source of excitatory synaptic drives which are brought into action by afferent thalamic, and not by transcallosal, stimulation. In the VL thalamus, the spike barrages of units 'rebounding' sooner than relay cells have been considered as belonging to excitatory interneurons; these local elements have been envisaged to exert a powerful excitatory drive on projection cells when the long-lasting hyperpolarization of the latter declines xt. Similarly, the VB-evoked late depolarizing (negative) focal waves

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Fig. 3. Mass and unit responses evoked in cortical area 7 by RF stimulation at A3, L2.5. A: evoked potentials (50 averaged sweeps) recorded at a cortical depth of 0.9 mm; RF stimulation at different indicated depths in the mesencephalon with a 45 msec duration, 300/see train (horizontal bar). B: another experiment; simultaneous recordings of two single units in area 5b (a and b), both activated by a single RF shock (dot). Superimposition of several traces and vertical display of 4 sweeps with reduced amplitude. See text.

following long-lasting positive shifts in the depth of SI area were observed to occur in close time relation with immediately preceding high-frequency bursts of putative excitatory interneurons (Steriade, Yossif and Oakson, in preparation). These results suggest that the LI-evoked huge negative wave at 0.5 - 0.7 m m (Fig. 2, arrow) represents more the synchronous activation of local excitatory internuncial cells than that of their pyramidal targets. This may well be the case, taking into account that the late negative wave, which far exceeds in amplitude the primary depth-negative component at 0.5-1 mm, was scarcely reflected at the cortical surface and often appeared abruptly at 0.5 ram. Such a distribution may be attributable to the fact that interneurons are not vertically oriented. The RF-elicited response illustrated in Figs. 2 and 3 deserves special consideration as it not only disclosed the commonly observed long-latency, slow negative shift, but also a localized, short-latency ( < 10 msec) excitation. This early excitation is particularly evident in Fig. 3A, showing two distinct, successive negative wavelets which can be clearly dissociated at a depth of 0.7 - 1 mm. Fig. 2 reveals that above 0.7 m m the first negative component gave way to a low-amplitude, fast positivity as the electrode is moved towards the surface (see 0.5, 0.4 and 0 ram). The presence of an early activation was also confirmed in unit recordings which disclosed that some associative neurons were excited by a single R F shock at latencies as short as 3.5-5.5 msec (Fig. 3B). Such an early activation with evoked discharges concentrated within a bin of 1 msec implies either direct excitation through slow conducting (5-7 m/sec) pathways or bisynaptic activation through an intercalated relay in LI-LP neurons.

522 The latter have actually been found to be a target of a monosynaptic, high security reticulofugal pathway, in view of their ability to follow 100-150/sec R F shocks at latencies of 1.5-2 msec (Steriade, Diallo and White-Guay, in preparation). The second component of the RF-elicited response is a slow negative wave which is maximal between 0.7 m m and 0.5 ram, gradually decreasing by 30-65~o at recording sites situated nearer the surface (Fig. 2). This negative slow component does not change its polarity when recorded near the surface, thus indicating a wide distribution of corticipetal fibers of reticular origin. The duration of the RF-evoked negative wave did not outlast that of the pulse-train (Fig. 2), but in some experiments the negative wave was twice as long as the stimulating train (Fig. 3A). Similarly to the phenomena elicited by the other (LI and callosal) synaptic inputs, the excitatory response evoked by RF was followed by a long-lasting (around 100 msec) slow positive wave. The presence of this positive shift subsequent to the negative component indicates that, like the thalamic and callosal afferents, the ascending fibers arising in the RF succeed in generating a long-lasting hyperpolarization following the initial depolarizing event• Being well aware that mesencephalic R F stimulation may lead to spread of current to the superior colliculus which sends projections to the LP thalamic nucleus, special care was taken to prevent this eventuality. We already mentioned in the methods that with the type of electrodes and the stimulation intensities used, the spread of current did not exceed 1 m m in the case of thalamic stimulation (see again Fig. 1); in the case of R F stimulation the intensity applied was only one-fourth (around 0.05 mA) of that employed for thalamic stimulation (0.2 mA). Moreover, by stimulating several points along a track at A3, L2.5, the optimal site for eliciting LI

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Fig. 4. Effects of RF stimulation on LI and transcallosally evoked responses. Responses (50 averaged sweeps) were recorded along a semi-microelectrode track in area 5b; only potentials evoked at the surface and a depth of 0.3 mm are illustrated. The first traces, at both 0 and 0.3 mm, represent control responses evoked by single shocks to LI and contralateral homotopic cortical area (Call.). The second traces depict a 320/sec pulse train of 50 msec duration applied to the mesencephalic RF preceding the LI and Call-evoked response (at right the response to RF alone). Comments in text.

523 a maximal evoked response in the depth of areas 5b or 7 was found at a depth o f - - 1 in the mesencephalic tegmentum; the amplitude of responses evoked by stimulating 1 mm above this point, therefore closer to the superior colliculus, was largely reduced (Fig. 3A). RF stimulation applied prior to a testing LI or contralateral cortical shock invariably diminished the amplitude of the thalamically or transcallosally evoked mass potential (Fig. 4). This striking response decrement was observed at all (surface and depth) recording points along a track, regardless of differences between the reversal points of the two types of evoked potentials. Fig. 4 depicts the RF-induced reduction in amplitude of testing responses at a depth of 0.3 mm where the callosally evoked response was already reversed while the LI-evoked potential was still initially positive. The diminished amplitude of the early (depolarizing) event triggered by thalamic or transcallosal stimulation when preceded by an RF shock-train, which induces by itself a focal negative (depolarizing) shift, is not surprising. It is well known that the early component of specific thalamocortical evoked responses mainly consists of summated EPSPs elicited in apical dendrites by corticipetal fibers. As suggested in Fig. 2, the site of generation of depolarizing events is shared by specific thalamic, callosal and reticular inputs. Shunting effects may be found in such cases leading to non-linear summation of EPSPs, as described for spinal motoneurons 1. Important non-linearity was predicted when strong dendritic depolarization (as likely produced by RF stimulation, see Figs. 2-4) causes a significant decrease of the effective synaptic driving potential produced by other inputs reaching closely neighboring dendritic branches z. This explanation, based on large diminution of EPSP amplitude to a testing shock of the less polarized subsynaptic dendritic membrane, may also account for the RF-induced marked decrease in amplitude of the VL-motor cortex evoked potential 10. When recording unit activity, the much less spectacular decreased probability of VL-evoked synaptic discharges in a particular class of PT neurons during RF stimulation was interpreted as a partial compensation of the decrease in dendritic EPSP amplitude by the RF-induced increase of soma excitation, with the result that neuron polarization is closer to the firing level lz. Similar findings have been obtained in an analysis of RF-elicited effects on LI- and LP-evoked synaptic discharges in single units recorded from cortical areas 5 and 7 (in preparation). Supported by Grants from the Medical Research Council (MT-3689) and the Minist~re de l'Education du Gouvernement du Qu6bec (subvention pour formation de chercheurs).

1 BURKE, R. E., Composite nature of the monosynapticexcitatory postsynaptic potential, J. NeurophysioL, 30 (1967) 1114-1137. 2 BUSER,P., AND BIGNALL, K. E., Nonprimary sensory projections on the cat neocortex, Int. Rev. Neurobiol., 10 (1970) 111-165. 3 DUBNER, R., AND RUTLEDGE, L. T., Intracellular recording of the convergence of input upon

neurons in cat association cortex, Exp. Neurol., 12 (1965) 349-369.

524 4 GRAYBIELA. M., Some ascending connections of the pulvinar and nucleus lateralis posterior of the thalamus in the cat, Brain Research, 44 (1972) 99-125. 5 HASSLER, R., UND MUllS-CLEMENT, K., Architektonischer Aufbau des sensomotorischen und parietalen Cortex der Katze, J. Hirnforsch., 6 (1964) 377-420. 6 HEATH, C. J., AND JONES, E. G., The anatomical organization of the suprasylvian gyrus of the cat, Ergebn. Anat. Entwickl.-Gesch., 45 (1971) 5-64. 7 INGRAM,W. R., HANNETT,F. I., ANDRANSON,S. W., The topography of the nuclei of the diencephalon of the cat, J. comp. NeuroL, 55 (1932) 333-394. 8 RALL, W., Distinguishing theoretical synaptic potentials computed for different soma-dendritic distributions of synaptic input, J. Neurophysiol., 30 (1967) 1138-1168. 9 RIOCH, D. McK., Studies on the diencephalon of carnivora. L The nuclear configuration of the thalamus, epithalamus, and hypothalamus of the dog and cat, J. comp. Neurol., 49 (1929) 1-119. 10 STER1ADE,M., Ascending control of motor cortex responsiveness, Electroenceph. clin. Neurophysiol., 26 (1969) 25-40. 11 STERIADE,M., WYZINSKI,P., AND APOSTOL, V., Corticofugal projections governing rhythmic thalamic activity. In T. L. FRIGYESI)E. RINVIK AND M. D. YAHR (Eds.), Corticothalamic Projections and Sensorimotor Activities, Raven Press, New York, 1972, pp. 221-272. 12 STERIADE, M., WYZINSKI) P., AND APOSTOL, V., Differential synaptic reactivity of simple and complex pyramidal tract neurons at various levels of vigilance, Exp. Brain Res., 17 (1973) 87-110.