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Response properties and visual receptive fields of climbing and mossy fibers terminating in the flocculus of the monkey
EXPERIMENTAL
NEUROLOGY
95,455-47 l(l987)
Response Properties and Visual Receptive Fields of Climbing and Mossy Fibers Terminating in the Flocculus of the Monkey HIROHARU NODA, TATEO
WARABI, AND MIK~O OHNO’
Deparfment of Visual Sciences, School of Optometry, Indiana University, Bloomington, Indiana 47405 Received August 4, I986 Response properties and visual receptive fields of climbing fibers and mossy fibers terminating in the cerebellar flocculus were studied in monkeys trained to fixate a stationary visual target. Among a total of 429 climbing fiber-related units (climbing fibers and complex spikes of Purkinje cells), 20 (4.9%) showed cyclic modulations in firing in response to sinusoidal retinal-slip velocities. Their receptive fields always included the fovea. Among 485 mossy fibers, 64 (13%) responded to the visual stimulation. Of the 64 visually responsive mossy fiber units, 39 (6 1%) responded exclusively to the retinal-slip velocity (visual mossy fibers and the remaining 25 mossy fibers (39%) responded also to the eye and head velocities (visuomotor mossy fibers). Recep tive fields for 17 visual mossy fibers (17/39 or 44%) were within lo” of fixation and those for 22 others (56%) were in the periphery. Receptive fields for all 25 visuomotor mossy fibers were in the periphery. Each mossy fiber unit had a unique velocitytuning curve and, therefore, the response patterns of individual mossy fibers were different depending on the range of their velocity sensitivity and on the retinal-slip Velocity applied. 0 1987 Academic Rey, Inc.
INTRODUCTION Single-unit studies of the cerebellar flocculus in alert macaques has revealed a variety of responses related to eye and head velocity during smoothpursuit eye movements or during suppression of the vestibuloocular reflex Abbreviations: Bkg-random-dot pattern, CF-climbing hber, DLPN-dorsolateral pontine nucleus, MF-mossy fiber, P-Purkinje. i This study was supported by National Institutes of Health grant EYO4063. The authors are grateful to Dr. Nobuo Suzumura for his assistance during various phases of the research, to Mr. Jacque Kubley for photography of figures, and to Mr. Takashi Aso for assistance in computer analysis. 455 0014-4886/87 $3.00 Copyrigltt 8 1987 by Academic Pres, Inc. AU rights of reproduction in any form reserved.
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(11, 17, 18, 22). The flocculus has also been implicated in oculomotor control by improving the function of the oculomotor system during visual and combined visual-vestibular stimulation (34, 35). Lesions of the flocculus produce deficits in smooth pursuit, impairments in optokinetic responses, and a loss of adaptive modification of the vestibuloocular reflex (24,25, 3 1, 36,38). In spite of the variety of functions of the flocculus that require visual information, the properties of visual input signals to the flocculus are practically unknown. Mossy-fiber discharges recorded from the primate have so far demonstrated only the existence of retinal-slip signals in the flocculus (18, 20,34). The present experiment was designed to study the retinal-slip signals which are brought in and processed by the flocculus. In the study of Purkinjecell (P-cell) responses, we discovered that a class of P cells (comprising 39% of the visually responsive P cells) had unique receptive field properties (23). This class of P cells received visual inputs from two functionally distinct portions of the retina. Impulses from one receptive field, situated within 10 of the fovea, caused excitatory responses in these P cells whereas those from the other receptive field, located in the periphery, resulted in suppression of the P cell activity. Both the excitatory and inhibitory responses were stimulus-velocity dependent but they were not related to the direction of the stimulus movement; hence they were bidirectional. In order to explain how such P-cell responses were brought about, the following questions were asked in the present study: (i) Are the impulses from these receptive fields transmitted by mossy cells (MFs) or climbing cells (CFs)? (ii) Are the same input fibers responsible for both the excitatory and inhibitory responses? (iii) Are the signals excitatory or inhibitory at the level of input fibers? (iv) If the signals are transmitted by different input fibers, are there any differences in their response features? (v) Are there any cues to explain why impulses originating from different retinal portions result in antagonistic effects on the P cell activity? Keeping these questions in mind, we studied response properties and receptive field organizations of CFs and MFs terminating in the flocculus of the monkey. The response properties of a group of visually responsive MFs were complex because of interactions among visual, oculomotor, and vestibular inputs. Because different experimental paradigms were needed to study this group of MFs, they were investigated in another series of experiments and have been reported in a separate paper (21). A preliminary report on some visual MFs was presented elsewhere (20). METHODS Ten pig-tailed macaques (Mucucu nemestrina) were used. The flocculi of both sides were studied in six monkeys and only the right flocculus was stud-
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ied in the remaining four monkeys. The surgical preparations, procedures for training monkeys, experimental conditions, the methods of recording single-unit activity and monitoring eye positions with a magnetic search-coil technique were the same as described in preceding papers (2 1,23). To study retinal-slip signals, the monkey was trained to maintain a stable fixation even when the entire visual background was moved. Among several methods tried, the most reliable paradigm was to train the animal to respond to a change in color. First, the monkey was conditioned to press a lever in response to a warning buzzer. When the animal pressed the lever, the sound stopped and simultaneously a red target appeared on the tangent screen. The visual target was a beam of a helium-neon laser, dimmed by a series of neutral density filters. After a preprogrammed interval (which was randomized by a microprocessor to minimize the predictability), the spot turned from red to green. The size and luminance between red and green were matched. If the animal succeeded in releasing the lever within a set time (0.3 to 0.5 s, controlled also by the microprocessor based on the levels of the animal’s performance) after the green light appeared, it was rewarded with a drop of water or fruit juice. The release of the lever before or after the rewarding green period was not rewarded. This behavioral task required unbroken vigilance on the part of the animal and could not be performed without maintaining the foveal fixation. During experiments, the animal was placed in a small room facing a window with a rear-projection screen. When the animal was placed 57 cm from it, the screen subtended 1 lo” of the visual angle horizontally and 90” vertically. A background random-dot pattern (interspot luminance: 0.035 f&L) was projected on and filled the tangent screen. Typically, the visual stimulus was moved sinusoidally in the horizontal plane between 10“ right and lo” left at frequencies from 0.1 to 1 Hz. The movements were produced by rotating a front-surface mirror mounted on the core of a galvanometer which was operated by a function generator through power amplifications. To study the size and location of the visual receptive fields of the axons, the pattern was projected on a limited area of the screen. In Figs. 3,6, and 7, when the animal fixated on the red spot (indicated in the figures by a dot in a circle), the moving pattern was projected through the various sizes of windows so that the stimulus movements could be seen only in the dotted area. The diameter of the circular windows or disks in the insets of the figures correspond to lo”, and the rectangle surrounding a circle represents the entire tangent screen (90° x 1 lo”). The visual responses of CFs and MFs were examined by on-line analysis by constructing phase histograms with a time histogram memory control unit (Ortec, models 4620 and 4621). When the units responded to visual stimulations, spike data, eye positions, and stimulus signals were recorded
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on magnetic tapes, using a 1Cchannel magnetic tape recorder (Ampex PR2200). The responses were analyzed by making phase histograms with a PDP 1 l/23 computer, constructed from 10 or 20 responses selected on the basis of the stability of the eyes on the target. A bin-width of either 20 ms or 25 ms was most commonly used. To evaluate the peak firing rates, the histograms were smoothed by computing a running average over five consecutive bins. The sensitivity to the stimulus velocity was evaluated by two methods. In the first, the peak firing rates in phase histograms made for different frequencies were measured from the smoothed histograms. The velocity sensitivity was expressed as the difference in the peak firing rates divided by the difference in the peak velocities. The second method used more direct measurements from instantaneous activity as illustrated in Fig. 1. It was based on cross-correlations between the spike-density function and the retinal-slip velocity function. The spikedensity function was obtained by replacing each impulse with a Gaussian function (width = 25 ms), as originally described by MacPherson and Aldridge ( 12). The retinal-slip velocity function was obtained by subtracting eye velocity (if any) from stimulus velocity. To estimate the conduction time (TIME LAG) for the signal from the retina to the flocculus, a cross correlogram was made and the peak corresponded to the latency. When we needed the discharge rate of the unit in response to, for example, the retinal-slip velocity 2O”/s to the right, we found the values in the spike-density function (shifted for the time lag) corresponding to this velocity in the retinal-slip function. The velocity-sensitivity plot compiled the averages and the standard deviations of these values at each velocity point which is shown in the abscissae. RESULTS
IdentiJication of Mossy and Climbing Fiber Units. Three kinds of axonalspike units were recordable in the white matter: they were mossy fibers (MFs), climbing fibers (CFs), and the axons of Purkinje cells (P cells). Discharges of P cells were characterized by the presence of complex spikes interspersed among tonic simple spikes. In a separate series of experiments, we discovered that the majority of visually responsive P cells were so-called gaze velocity neurons and that their responses to sinusoidal retinal-slip velocities were unique (23). It is generally agreed that the complex spikes of a P cell are elicited by impulses transmitted by a CF. A CF makes an extensive all-or-none excitatory synaptic connection with the dendrites of a P cell. Whenever the CF discharges, the postsynaptic P cell also discharges. It is also well known that
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FIG. 1. The method used to evaluate sensitivity of neurons to retinal-slip velocity. A-Ecomputer simulation to obtain a smooth spike-density function by replacing each impulse with a Gaussian function (width = 25 ms), as originally described by MacPherson and Aldridge ( 12). G-a cross-correlation between the spike-density function (E) and the stimulus-velocity function (F) to estimate the conduction time (TIME LAG) from the retina to the flocculus in individual input fibers. H-to evaluate the firing rate of a fiber in response to a certain retinal-slip velocity, we measured the value in the spike-density function that corresponded to the velocity after shifting the stimulus velocity curve by the time lag. I-the averages and standard deviations of discharge rates were plotted against the retinal-slip velocity. In actual computations, the retinal-slip velocity was evaluated by subtracting the eye velocity (if any) from the stimulus velocity.
a CF shows characteristic low-frequency discharges. From the characteristic discharges patterns and behaviors of P cells and CFs, identification of these units is not difficult. Thus, MF units were identified by eliminating CF and P-cell units from the axonal-spike units recorded from the white matter of the flocculus. Climbing Fiber Responsesto Sinusoidal Retinal Slip. While the monkey fixated a stationary visual target, the random-dot pattern (Bkg) was moved sinusoidally at 0.5 Hz in the horizontal plane (+ lo” amplitude). Twenty of 429 CF-related units (4.9%) showed cyclic modulations in tiring. Eleven of 232 units (4.7%) were CFs recorded in the white matter; three of 44 units (6.8%) were evaluated from the complex-spike discharges of P cells; and seven of 153 units (4.5%) were CF responses recorded in the molecular layer.
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FIG. 2. Discharges of a climbing fiber (CF) recorded in the white matter of the flocculus during spontaneous saccades and fixation (A), during sinusoidal movements of the full-field randomdot pattern at 0.5 Hz (B), and at 0.3 Hz (C), + lo” amplitude. Calibrations for Sp (spike amplitude), H (horizontal eye position), V (vertical eye position), and Bkg (background movement) represent I mV (positive up), 20”, 20’, and 20”, respectively. D-phase histograms for the same climbing fiber. Open columns, for 0.5 Hz stimulation; stippled columns for 0.3 Hz stimulation. In this and subsequent figures, a single cycle has been repeated to facilitate visualization of the periodic events. The time base is shown in radians to normalize the abscissae for different frequencies of stimulus movements.
As illustrated in Fig. 2A, discharges of CF units were characterized by a low rate of spontaneous firing. The resting firing rates evaluated during primary fixation (fixation of the central target at eye level) ranged from 0.4 to 2.1 (average 1.3) spikes/s for the total sample. Firing rate of the CF increased during sinusoidal movements of Bkg at 0.5 Hz (Fig. 2B) and 0.3 Hz (Fig. 2C). The unit discharged only during ipsilateral Bkg movements (right). Peak firing rates were evaluated from smoothed phase histograms by computing a running average over five consecutive bins. At frequencies of 0. I, 0.3,0.5, and 0.7 Hz (+ lo“), the peak firing rates of the CF units were 3.4, 7.7, 11.8, and 15.6 spikes/s, respectively. The velocity sensitivity (evaluated by dividing the difference in peak firing rates by the
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difference in peak stimulus velocities) was 0.3 1 spikes. s-‘/deg . s-’ for the CF unit. The velocity sensitivity for seven CF units (tested with three or more stimulus frequencies) ranged from 0.03 to 0.44 (average 0.13) spikess-‘/deg . s-‘. The receptive field properties seen in the CF units illustrated in Fig. 3 were commonly observed in all 11 CFs. They were characterized by (i) relatively large size (> lo”) and inclusion of the fovea, (ii) excitatory responses during retinal slip in one direction and none in the other, and (iii) velocity-dependent peaks. All CF units in those tested had receptive fields in corresponding locations of both eyes (not illustrated). As seen in Fig. 3, sinusoidal movements of the Bkg presented within 10 of primary fixation (A), outside lo” (B), and on the entire screen (C) caused similar cyclic modulations in firing. As the peak firing rate was higher when
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both the central retina and periphery were simultaneously stimulated, the CF unit received visual inputs from a relatively large retinal portion. Mossy-Fiber Responses to Sinusoidal Retinal Slip. Among 485 MF units, identified by eliminating CF and P-cell units, 64 MF units (or 13%) responded to sinusoidal Bkg movements in the horizontal plane. Of these visually responsive MFs, 39 units (6 1%) responded exclusively to visual inputs (visual MFs), and the remaining 25 units (39%) responded also to oculomotor and vestibular inputs (visuomotor MFs). While the monkey maintained stationary fixation, the responses of both classes of MFs were almost identical. When retinal slip was accompanied either by eye or head movements, however, the responses of visuomotor MFs were markedly different from those of visual MFs. The details of response properties of visuomotor MFs in relation to retinal-slip, eye, and head velocities have already been described (23). In brief, the visuomotor MFs showed almost identical responses to Bkg movements regardless of whether the eyes were held stationary (causing retinal-slip velocity) or moved with the Bkg (replacing the retinal-slip velocity with eye velocity). The visuomotor MFs responded to the algebraic sum of retinal-slip and eye velocities. In contrast, the visual MFs in the present study responded only to retinal slip. When suppression of optokinetic nystagmus was incomplete, their discharges were unrelated to the sinusoidal Bkg movements. Responses of a visual MF during sinusoidal retinal slip (A) and during constant-velocity triangular oscillation of the Bkg (B) are shown in Fig. 4. As indicated by flat horizontal (H) and vertical (V) components of eye position shown in the excerpts of film records, the eyes were well maintained upon a stationary target. Under such conditions, the Bkg velocity corresponds to the retinal-slip velocity (in the opposite direction). The MF showed a relatively smooth cyclic modulation in response to the sinusoidal retinal slip (A). The peak activity appeared slightly before the maximum retinal-slip velocity. This lead reflected an accommodation of the neuron that is seen also in the slowly decaying response during the constant-velocity retinal slip (B). Among the 39 visual MF units, 26 (67%) showed velocity-related responses as exemplified by the MF of Fig. 4. Discharges in the remaining 13 MF units (33%) increased abruptly following the Bkg turnaround into the preferred direction and showed flat (or gradually decaying) responses thereafter. An example for each type of MF unit is shown in Fig. 5. Comparing the responses (phase histograms) of the two MF units with those of Fig. 4, one readily notices that MF708 (C) showed the flat response which was observed during constant-velocity stimulation (see Fig. 4B). Discharges of MF109 showed a cyclic modulation with peak activity near the maximum stimulus velocity (Fig. 5A). These MFs were sensitive not only to the direction but
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FIG. 4. Responses of a MF unit during sinusoidal Bkg movements (A) and during constantvelocity triangular oscillations (B). H, V, and Bkg in the excerpt of record for both A and B indicate horizontal eye position, vertical eye position, and Bkg movement signals, respectively. Calibration marks for Sp, H, V, and Bkg represent 1mV (positive up), 5’, 5’, and lo’, respectively. Lower panels for both A and B, phase histograms constructed from 20 stimulus cycles during stationary fixation periods. Bin width, 50 ms. The excerpts of records and the phase histograms are in register. Time base (in both seconds and radians) is shown at the bottom of B.
also to the velocity of Bkg movements (velocity MFs). On the other hand, with the stimulus velocity used in the present study (peak velocities ranged from 10 to 500/s), MF708 was sensitive only to the direction but not to the velocity of movements (direction-only MFs). It is inferred that the velocity of Bkg movements exceeded the sensitive range for the latter group of MFs. This inference (drawn from the phase histograms) was substantiated by computing the velocity sensitivity for each unit (B and D in Fig. 5), using the method ilhtstrated in Fig. 1. Visual Receptive Fields of Mossy Fiber Units. In contrast to the large receptive fields of the CF units that always included the fovea, those of MF units were found in two functionally distinct retinal regions. Receptive fields
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FIG. 5. Velocity- and direction-sensitive MF and direction-sensitive MF. The two types of MF are compared by showing phase histograms (A and C) and velocity-sensitive plots (B and D). Bin width for the phase histograms, 25 ms. Both MFs were stimulated with sinusoidal Bkg movements (the phase is shown by sinusoidal dashed lines) at 0.45 Hz, *lo’ amplitude. In response to the sinusoidal stimulus movements, discharges of MF109 were roughly sinusoidal and reached a peak slightly after maximum stimulus velocity whereas those of MF708 responded rather abruptly 100 to 125 ms after the turnaround. It did not show any obvious peak and the response decayed slowly thereafter. The velocity-sensitivity plots (B and D) were computed by evaluating retinal-slip velocity (the difference between stimulus velocity and eye velocity, if any) and smoothed spike density function (computed as illustrated in Fig. 1). The cross correlogram between these functions (inset in B) indicated that the effect of retinal-slip velocity reached maximum at 80 ms. The averages and standard deviations of the values found in the spike density function 80 ms after each velocity point of retinal slip velocity in the abscissae are shown as velocity sensitivity plots.
of 17 visual MFs (17/39 or 44%) were small and confined within lo” of fixation whereas those of 22 visual MFs (22/39 or 56%) were relatively large and were found in the periphery. In the peripheral group, receptive fields for 12 MFs were in the contralateral, those for 5 MFs were in the ipsilateral, and those for the remaining 5 MFs were in bilateral hemiretinae, all in both eyes. The numbers of visual MFs in relation to the range of sensitivity and to the location of the receptive field are summarized in Table 1. Although a slightly larger number of MFs was sensitive only to the direction of Bkg
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The Number of Visual Mossy Fibers in Relation to the Location of the Receptive Field and to the Range of Velocity Sensitivity Receptive field
Sensitive to stimulus velocity used (peak velocity IO-W/s) Responses saturated, not sensitive to the velocity range used
Foveal region
Periphery
12 units 5 units
14 units 8 units
movements among the units activated from the periphery, this tendency was not significant (P z=-0.05, x2 test). Phasehistograms for a representative MF unit which was activated exclusively from the central lo” of fixation are shown in Fig. 6. When the Bkg movements were confined to the central retina (A), the MF showed a flat response.When the central lo’ area of the screenwas occluded and the Bkg was moved outside the occluded area, the MF did not respond (B). Changing the stimulus frequency (not illustrated) did not affect the
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FIG. 6. A MF with receptive field in the central retina. A-phase histogram during sinusoidal movements of Bkg at 0.6 Hz. The stimulus was presented within lo” of primary fixation. Bphase histogram for the same MF when the stimulus was presented outside the central lo”. C and D-phase histograms for the same MF when the presentation of the pattern was confined to the right (C) and left half(D) of the central 1o”, respectively.
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FIG. 7. A MF with receptive field in the periphery. A-phase histogram when the presentation of sinusoidally moving pattern was confined to the central lo” of primary fixation. B-phase histogram for the same MF when the stimulus was presented outside the central Lo’. C and Dphase histograms for the same MF when only the left (C) or right half(D) of the tangent screen was projected with the moving pattern, respectively.
height of responses. Interestingly, the receptive field encompassed both sides of the vertical meridian in all three MFs, showing flat responses (tested with the semicircular areas of the central retina open). This encompassing was not caused by unstable fixation. Phase histograms for a representative MF unit which was activated by visual inputs from the periphery are shown in Fig. 7. The stimulation of the centA retina did not evoke a response (A) whereas that of the periphery evoked responses. During stimulation of the entire screen (not illustrated), the left half of the periphery (C), and the right half of the periphery (D) yielded similar responses, indicating that the MF received excitatory visual inputs from bilateral hemiretinae (of both eyes). Not all units were tested with monocular stimulations, but every unit tested showed responses to stimulation of both eyes. Binocular interactions were not tested in the present study. DISCUSSION Visual Climbing-I;iber Units. The visual climbing fiber (CF) input to the flocculus in the rabbit is we11known ( 14- 16) and arises from the dorsal cap
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of Kooy of the inferior olive (1). The activity of the CF can be well modulated by slowly moving visual stimuli (3, 26-28). In spite of the extensive studies on the rabbit, visual CFs in the primate have not yet been reported. In the present study, 11 CFs and 10 complex-spike units from P-cell discharges responded in a speed- and direction-selective manner to a large textured pattern. These numbers corresponded to less than 5% of the CFs (including the CF responses from P cells) examined. It is likely that the low sampling rate of visual CFs in the primate was caused by the difference in the animal preparation rather than by the species difference. Whether or not the responses could be tested with a slow movement of the stimulus appears to be of critical importance. All rabbits used in the previous reports had been deeply anesthetized and immobilized whereas our monkeys were alert and eye movements were suppressed only by the effort made by the monkeys to fixate on a stationary visual target. According to Simpson and Alley (26), the great majority of CFs in the rabbit were optimally responsive to movements of less than 1O/s.In our study, the range of the peak stimulus velocities was typically from 10 to BY/s. The CFs were stimulated with horizontal movements presented only in the central 1 lo” of fixation (frontal-plane stimulation); all units were activated binocularly; the preferred directions were always the same for both eyes. In the rabbit, however, the receptive fields of dorsal cap neurons extended from the nose (00) to 160” posteriorly along the horizon (29), and were quite different from the present observation. For example, some neurons were modulated only from the contralateral eye and were best excited by horizontal movement in the temporonasal direction and inhibited in the nasotemporal direction. There were such neurons showing receptive fields in both eyes but the preferred direction for one eye was different from that for the other eye (29). If receptive field organizations of dorsal cap neurons in the primate bear any resemblance to those in the rabbit, our binocular and frontal-plane stimulation might have activated only a small percentage of dorsal cap neurons. Visually Responsive Mossy-Fiber Units. Two distinct classes of visually responsive mossy-fiber (MF) units were recorded from the flocculus. Depending on whether or not the eye and head were stationary, their visual responses were markedly different. Visual MF units responded only when the eye and head were stationary, hence responding exclusively to retinalslip velocity. Visuomotor MF units reflected the stimulus movement itself, regardless of eye position; hence the activity was related to the algebraic sum of retinal-slip, eye, and head (in the opposite direction) velocities. The visual MFs originate from different structures in the brain stem. It is most unlikely that their parent cells are vestibular neurons because they did not show any sign of vestibular input. The possibility of being prepositus
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hypoglossi neurons may also be excluded for the same reason because these neurons receive vestibular inputs (2). It is possible that the MFs which received visual inputs from the central retina originate in the dorsolateral pontine nucleus (DLPN). Suzuki and Keller (30) and Mustari et al. (19) have recorded visually responsive units in the DLPN of macaque monkeys. The response characteristics of these neurons were very similar to those of visual MFs recorded in the flocculus of the macaque (20). The receptive fields of these neurons always included the fovea (19). Projections are known from various visual cortices to this pontine nucleus (7, 8), including the superior temporal motion area MT (5, 6, 32, 33). The lateral margin of the DLPN has been identified as a major floccular projecting portion of the pontine nuclei (10). The origin of the MFs receiving visual inputs from the peripheral retina is as yet unclear. Possible sources of visual MFs have been suggested by experiments on lower mammals. The flocculus receives visual information via MFs arising from the nucleus reticularis tegmenti pontis in the rabbit ( 13, 16) and in the cat (9). Neurons in the medial terminal nucleus of the rabbit also respond to slowly moving textured patterns (28). Although the primary projection from this nucleus to the flocculus is via CFs arising from the dorsal cap of the inferior olive, it is possible, but not yet confirmed, that the neurons in the medial terminal nucleus project directly to the flocculus as a MF input (4,371. Comparison between Climbing and Mossy Fiber Visual Units. A striking difference in receptive field organizations between CFs and MFs was that CFs had large and homogeneous receptive fields which always included the fovea, whereas MFs had receptive fields either in the foveal or peripheral region. When the stimulus was confined either to the fovea1 or peripheral retina, MFs tended to show more prominent responses. On the other hand, CFs received similar visual inputs from both the foveal and peripheral regions. When both regions were stimulated simultaneously, the responses tended to be larger in many CFs. Receptive fields of neurons in the flocculus-related brain stem structures have not yet been studied in the primate or in other mammals, except for the dorsal cap neurons in the rabbit (29). Depending on whether or not the animal uses binocular and frontal vision, receptive field organizations of visual neurons would be considerably different. In the present experiments, in addition to the range of stimulus velocities already discussed, there were two other limitations: one was the direction of stimulus movements that were always horizontal, and the other was the area of stimulus presentation that had been limited to only the central 1 lo”. In spite of the limitations in the experimental paradigms, there were several features in the receptive field organization in the primate: (i) both CFs and MFs, in those tested, had almost
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identical receptive fields in corresponding loci of the two eyes; (ii) they had the same preferred directions; and (iii) the two eyes yielded similar responses with respect both to the gain and to the phase-relation to the stimulus. Functional Significance of the Visual Input. The most important finding in the present study was that the flocculus received visual inputs from two functionally distinct portions of the retina via different channels of MFs. One group of MFs conveyed visual inputs only from the foveal region and the other group of MFs was responsible for the visual inputs from the periphery. The signals carried by these two groups of MFs were transmitted through different visual pathways, gave different effects on the P cells in the flocculus, and implied different functional roles. When stimulated with sinusoidally moving patterns, a large number of P cells in the flocculus were bidirectionally depressed (23). Among these cells, approximately three-quarters had two visual receptive fields, one in the foveal region and the other in the periphery 30 to 70” away from the fovea. When visual stimulation was confined to within lo” of fixation, the P cells were excited. The responses were velocity-dependent. When the moving stimulation Bkg was presented in the periphery, the discharge rates of the same P cells were decreased to 40 to 60% of the spontaneous activity. As the visual receptive fields of CFs were large and always included the fovea in the primate, it is unlikely that these fibers are responsible for the different responses in the same P cells. The excitatory and inhibitory responses in the P cells, therefore, must have been caused by the visual inputs transmitted by the MFs which had receptive fields in the foveal and peripheral regions of the retina, respectively. On the other hand, the inhibitory visual input from the periphery must have been conveyed by a group of visual MFs with peripheral receptive fields and the visuomotor MFs. All visuomotor MFs have their visual receptive fields in the periphery. The parent cells of the fibers are most likely in the vestibular nuclei (2 1). The origin of the visual MFs with peripheral receptive fields must be some brain stem nuclei such as the reticularis tegmenti pontis and the medial terminal nucleus which are known to send MFs to the flocculus. The pathway for the inhibitory visual input to the floccular P cells would be, therefore, related to the direct projection from the retina to the brain stem visual nuclei which are responsible for the generation of the optokinetic nystagmus. It is generally agreed that the flocculus serves to control both smooth pursuit and optokinetic nystagmus in the primate. The visual input arising in the foveal region, possibly transmitted through the cerebropontinecerebcllar pathway to the flocculus, may serve the smooth-pursuit function while the input from the peripheral retina, possibly taking the direct retino brain stem pathway, may be related to the optokinetic function. The visually responsive
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MFs discovered by the present study may serve to transmit these signals to the flocculus. REFERENCES 1. ALLEY, K., R. BAKER, AND J. I. SIMPSON. 1975. Afferents to the vestibulo-cerebellum and the origin of the visual climbing fibers in the rabbit. Bruin Res. 98: 582-589. 2. BAKER, R. 1977. The nucleus prepositus hypoglossi. Pages 145-178 in B. A. BROOKS AND F. J. BAJANDAS, Eds., Eye Movements. Plenum, New York/London. 3. BARMACK, N. H., AND D. T. HESS. 1980. Multiple-unit activity evoked in dorsal cap of inferior olive of the rabbit by visual stimulation. J. Neurophysiol. 43: 15 I - 164. 4. BRAUTH, S. E., AND H. J. KARTEN. 1977. Direct accessory optic projections to the vestibulo-cerebellum. A possible channel for oculomotor systems.Exp. Brain Res. 27: 73-84. 5. FRIES, W. 198 1. The projection from striate and prestriate visual cortex onto the pontine nuclei in the macaque monkey. Sot. Neurosci. Abstr. 7: 762. 6. GALLETTI, C., M. G. MAIOLI, S. !~QUATITRO,AND P. F. BATTAGLINI. 1982. Cortico-pontine projections from the visual area of the superior temporal sulcus in the macaque monkey. Arch. Ital. Biol. 120: 411-416. 7. GLICKSTEIN, M., AND J. MAY. 1982. Visual control of movement: the circuits which link visual and motor areas of the brain with special reference to the pons and cerebellum. Pages 103-145 in W. D. NEFF, Ed., Contributions to Sensory PhysioZogy. Academic Press, New York. 8. GLICKSTEIN, M., J. MAY, AND B. MERCER. 1985. Cortico-pontine projection in the macaque: the distribution oflabelled cortical cells after large injections of horseradish peroxidase in the pontine nuclei. J. Comp. Neural. 235: 343-359. 9. KAWASAKI, T., Y. SATO, AND I. KATO. 1980. The visual input pathway to the cerebellar tlocculus. Equilib. Res. 39: l-6. IO. LANGER, T., A. F. FUCHS, C. A. SCUDDER, AND M. C. CHUBB. 1985. Afferents to the flocculus of the cerebellum in the rhesus macaque as revealed by retrograde transport of horseradish peroxidase. J. Comp. Neural. 235: I-25. 11. LISBERGER, S. G., AND A. F. FUCHS. 1978. Role of primate flocculus during rapid behavioral modification of vestibulo-ocular reflex. I. Purkinje cell activity during visually guided horizontal smooth pursuit eye movements and passive head rotation. J. Neurophysiol. 41: 733-763. 12. MACPHERSON, J. M., AND J. W. ALDRIDGE. 1979. A quantitative method of computer analysis of spike train data collected from behaving animals. Brain Res. 175: 183- 187. 13. MAEKAWA, K., M. KIMURA, AND T. TAKEDA. 198 1. Mossy fiber activation ofthe cerebellar flocculus from the visual system.Ann. N. Y. Acad. Sci. 374: 476-490. 14. MAEKAWA, K., AND J. I. SIMPSON. 1973. Climbing fiber response evoked in vestibulocerebellum of rabbit from visual system.J. Neurophysiol. 36: 649-666. 15. MAEKAWA, K., AND T. TAKEDA. 1976. Electrophysiological identification of the climbing and mossy fiber pathways from the rabbit’s retina to the contralateral cerebellar flocculus. Brain Res. 109: 169- 174. 16. MAEKAWA, K., AND T. TAKEDA. 1977. Afferent pathways from the visual system to the cerebellar flocculus of the rabbit. Pages 187-196, in R. BAKER AND A. BERTHOZ, Eds., Control of Gaze by Brain Stem Neurons, Elsevier/North-Holland, Amsterdam/New York. 17. MILES, F. A., AND J. H. FULLER. 1975. Visual tracking and the primate flocculus. Science 189: 1000-1002.
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18. MILES, F. A., J. H. FULLER, D. J. BRAITMAN, AND B. M. Dow. 1980. Long-term adaptive changes in primate vestibule-ocular reflex. III. Electrophysiological observations in flocculus of normal monkeys. J. Neurophysiol. 43: 1437-1476. 19. MUSTARI, M. J., A. F. FUCHS, AND J. WALLMAN. 1984. Smooth-pursuit-related units in the dorsolateral pons ofthe rhesus macaque. Sot. Neurosci. Abstr. 10: 987. 20. NODA, H. 198 1. Visual mossy fiber inputs to the flocculus of the monkey. Ann. N. Y. Acud. Sci. 374: 465-475. 2 1. NODA, H. 1986. Mossy fibres sending retinal-slip, eye, and head velocity signals to the flocculus of the monkey. J. Physiol. (London) 379: in press. 22. NODA, H., AND D. A. SUZUKI. 1979. The role of the flocculus of the monkey in fixation and smooth pursuit eye movements. J. Physiol. (London) 294: 335-348. 23. NODA, H., AND T. WARABI. 1986. Discharges ofPurkinje cells in monkey’s 8occulus during smooth-pursuit eye movements and visual stimulus movements. Exp. Neurol. 93: 390403. 24. OPTICAN, L. M., D. S. ZEE, F. A. MILES, AND S. G. LISBERGER. 1980. Oculomotor deficits in monkeys with flocculus lesions. Sot. Neurosci. Abstr. 6: 474. 25. ROBINSON, D. A. 1976. Adaptive gain control of vestibulo-ocular reflex by the cerebellum. J. Neurophysiol. 39: 954-969. 26. SIMPSON, J. I., AND K. E. ALLEY. 1974. Visual climbing fiber input to rabbit vestibulocerebellum: a source ofdirection-specific information. Bruin Res. 82: 301-308. 27. SIMPSON, J. I., AND R. HESS. 1977. Complex and simple visual messages in the floccuhts. Pages 351-360. in R. BAKER AND A. BERTHOZ, Eds. Control of Gaze by Brain Stem Neurons. Elsevier/North-Holland, Amsterdam/New York. 28. SIMPSON, J. I., R. E. SOODAK, AND R. HESS. 1979. The accessory optic system and its relation to the vestibulocerebellum. Prog. Bruin Res. 50: 7 15-724. 29. SIMPSON, J. I., W. GRAF, AND C. LEONARD. 198 1. The coordinate of visual climbing fibers to the floccuhts. Pages 475-484, in A. F. FUCHSAND W. BECKER, Eds., Progress in Oculomotor Research. Elsevier/North-Holland, Amsterdam, New York. 30. SUZUKI, D. A., AND E. L. KELLER. 1983. Visual signals in the dorso-lateral pontine nucleus of the alert monkey: their relationship to smooth-pursuit eye movements. Exp. Brain Res. 53: 473-478. 3 1. TAKEMORI, S., AND B. COHEN. 1974. Loss of suppression of vestibular nystagmus after flocculus lesions. Brain Res. 72: 2 13-222. 32. UNGERLEIDER, L. G., R. DESIMONE, T. GALKIN, AND M. MISHKIN. 1984. Subcortical projection ofarea MT in the macaque. J. Comp. Neurol. 223: 363-386. 33. VAN ESSEN,D. C., AND J. H. R. MAUNSELL. 1983. Hierarchical organization and functional streams in the visual cortex. Trends Neurosci. 6: 370-375. 34. WAESPE, W., U. B~.~T~NER,AND V. HENN. 198 1. Visual-vestibular interaction in the flocculus ofthe alert monkey. I. Input activity. Exp. Brain Res. 43: 337-348. 35. WAESPE, W., AND V. HENN. 198 1. Visual-vestibular interaction in the flocculus of the alert monkey. II. Purkinje cell activity. Exp. Bruin Res. 43: 349-360. 36. WAESPE, W., B. COHEN, AND T. RAPHAN. 1983. Role of the tlocculus and paraflocculus in optokinetic nystagmus and visual-vestibular interactions: effects of lesions. Exp. Bruin Res. 50: 9-33. 37. WINFXLD, J. A., A. HENDRICKSON, AND A. K~MM. 1978. Anatomical evidence that the medial terminal nucleus ofthe accessory optic tract in mammals provides a visual mossy fiber input to the flocculus. Bruin Res. 151: 175-182. 38. ZEE, D. S., A. YAMAZAKI, P. H. BUTLER, AND G. GUCER. 198 1. Effects of ablation of flocculus and paraflocculus on eye movements in primate. J. Neurophysiol. 46: 878-899.
NEUROLOGY
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Response Properties and Visual Receptive Fields of Climbing and Mossy Fibers Terminating in the Flocculus of the Monkey HIROHARU NODA, TATEO
WARABI, AND MIK~O OHNO’
Deparfment of Visual Sciences, School of Optometry, Indiana University, Bloomington, Indiana 47405 Received August 4, I986 Response properties and visual receptive fields of climbing fibers and mossy fibers terminating in the cerebellar flocculus were studied in monkeys trained to fixate a stationary visual target. Among a total of 429 climbing fiber-related units (climbing fibers and complex spikes of Purkinje cells), 20 (4.9%) showed cyclic modulations in firing in response to sinusoidal retinal-slip velocities. Their receptive fields always included the fovea. Among 485 mossy fibers, 64 (13%) responded to the visual stimulation. Of the 64 visually responsive mossy fiber units, 39 (6 1%) responded exclusively to the retinal-slip velocity (visual mossy fibers and the remaining 25 mossy fibers (39%) responded also to the eye and head velocities (visuomotor mossy fibers). Recep tive fields for 17 visual mossy fibers (17/39 or 44%) were within lo” of fixation and those for 22 others (56%) were in the periphery. Receptive fields for all 25 visuomotor mossy fibers were in the periphery. Each mossy fiber unit had a unique velocitytuning curve and, therefore, the response patterns of individual mossy fibers were different depending on the range of their velocity sensitivity and on the retinal-slip Velocity applied. 0 1987 Academic Rey, Inc.
INTRODUCTION Single-unit studies of the cerebellar flocculus in alert macaques has revealed a variety of responses related to eye and head velocity during smoothpursuit eye movements or during suppression of the vestibuloocular reflex Abbreviations: Bkg-random-dot pattern, CF-climbing hber, DLPN-dorsolateral pontine nucleus, MF-mossy fiber, P-Purkinje. i This study was supported by National Institutes of Health grant EYO4063. The authors are grateful to Dr. Nobuo Suzumura for his assistance during various phases of the research, to Mr. Jacque Kubley for photography of figures, and to Mr. Takashi Aso for assistance in computer analysis. 455 0014-4886/87 $3.00 Copyrigltt 8 1987 by Academic Pres, Inc. AU rights of reproduction in any form reserved.
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(11, 17, 18, 22). The flocculus has also been implicated in oculomotor control by improving the function of the oculomotor system during visual and combined visual-vestibular stimulation (34, 35). Lesions of the flocculus produce deficits in smooth pursuit, impairments in optokinetic responses, and a loss of adaptive modification of the vestibuloocular reflex (24,25, 3 1, 36,38). In spite of the variety of functions of the flocculus that require visual information, the properties of visual input signals to the flocculus are practically unknown. Mossy-fiber discharges recorded from the primate have so far demonstrated only the existence of retinal-slip signals in the flocculus (18, 20,34). The present experiment was designed to study the retinal-slip signals which are brought in and processed by the flocculus. In the study of Purkinjecell (P-cell) responses, we discovered that a class of P cells (comprising 39% of the visually responsive P cells) had unique receptive field properties (23). This class of P cells received visual inputs from two functionally distinct portions of the retina. Impulses from one receptive field, situated within 10 of the fovea, caused excitatory responses in these P cells whereas those from the other receptive field, located in the periphery, resulted in suppression of the P cell activity. Both the excitatory and inhibitory responses were stimulus-velocity dependent but they were not related to the direction of the stimulus movement; hence they were bidirectional. In order to explain how such P-cell responses were brought about, the following questions were asked in the present study: (i) Are the impulses from these receptive fields transmitted by mossy cells (MFs) or climbing cells (CFs)? (ii) Are the same input fibers responsible for both the excitatory and inhibitory responses? (iii) Are the signals excitatory or inhibitory at the level of input fibers? (iv) If the signals are transmitted by different input fibers, are there any differences in their response features? (v) Are there any cues to explain why impulses originating from different retinal portions result in antagonistic effects on the P cell activity? Keeping these questions in mind, we studied response properties and receptive field organizations of CFs and MFs terminating in the flocculus of the monkey. The response properties of a group of visually responsive MFs were complex because of interactions among visual, oculomotor, and vestibular inputs. Because different experimental paradigms were needed to study this group of MFs, they were investigated in another series of experiments and have been reported in a separate paper (21). A preliminary report on some visual MFs was presented elsewhere (20). METHODS Ten pig-tailed macaques (Mucucu nemestrina) were used. The flocculi of both sides were studied in six monkeys and only the right flocculus was stud-
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ied in the remaining four monkeys. The surgical preparations, procedures for training monkeys, experimental conditions, the methods of recording single-unit activity and monitoring eye positions with a magnetic search-coil technique were the same as described in preceding papers (2 1,23). To study retinal-slip signals, the monkey was trained to maintain a stable fixation even when the entire visual background was moved. Among several methods tried, the most reliable paradigm was to train the animal to respond to a change in color. First, the monkey was conditioned to press a lever in response to a warning buzzer. When the animal pressed the lever, the sound stopped and simultaneously a red target appeared on the tangent screen. The visual target was a beam of a helium-neon laser, dimmed by a series of neutral density filters. After a preprogrammed interval (which was randomized by a microprocessor to minimize the predictability), the spot turned from red to green. The size and luminance between red and green were matched. If the animal succeeded in releasing the lever within a set time (0.3 to 0.5 s, controlled also by the microprocessor based on the levels of the animal’s performance) after the green light appeared, it was rewarded with a drop of water or fruit juice. The release of the lever before or after the rewarding green period was not rewarded. This behavioral task required unbroken vigilance on the part of the animal and could not be performed without maintaining the foveal fixation. During experiments, the animal was placed in a small room facing a window with a rear-projection screen. When the animal was placed 57 cm from it, the screen subtended 1 lo” of the visual angle horizontally and 90” vertically. A background random-dot pattern (interspot luminance: 0.035 f&L) was projected on and filled the tangent screen. Typically, the visual stimulus was moved sinusoidally in the horizontal plane between 10“ right and lo” left at frequencies from 0.1 to 1 Hz. The movements were produced by rotating a front-surface mirror mounted on the core of a galvanometer which was operated by a function generator through power amplifications. To study the size and location of the visual receptive fields of the axons, the pattern was projected on a limited area of the screen. In Figs. 3,6, and 7, when the animal fixated on the red spot (indicated in the figures by a dot in a circle), the moving pattern was projected through the various sizes of windows so that the stimulus movements could be seen only in the dotted area. The diameter of the circular windows or disks in the insets of the figures correspond to lo”, and the rectangle surrounding a circle represents the entire tangent screen (90° x 1 lo”). The visual responses of CFs and MFs were examined by on-line analysis by constructing phase histograms with a time histogram memory control unit (Ortec, models 4620 and 4621). When the units responded to visual stimulations, spike data, eye positions, and stimulus signals were recorded
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on magnetic tapes, using a 1Cchannel magnetic tape recorder (Ampex PR2200). The responses were analyzed by making phase histograms with a PDP 1 l/23 computer, constructed from 10 or 20 responses selected on the basis of the stability of the eyes on the target. A bin-width of either 20 ms or 25 ms was most commonly used. To evaluate the peak firing rates, the histograms were smoothed by computing a running average over five consecutive bins. The sensitivity to the stimulus velocity was evaluated by two methods. In the first, the peak firing rates in phase histograms made for different frequencies were measured from the smoothed histograms. The velocity sensitivity was expressed as the difference in the peak firing rates divided by the difference in the peak velocities. The second method used more direct measurements from instantaneous activity as illustrated in Fig. 1. It was based on cross-correlations between the spike-density function and the retinal-slip velocity function. The spikedensity function was obtained by replacing each impulse with a Gaussian function (width = 25 ms), as originally described by MacPherson and Aldridge ( 12). The retinal-slip velocity function was obtained by subtracting eye velocity (if any) from stimulus velocity. To estimate the conduction time (TIME LAG) for the signal from the retina to the flocculus, a cross correlogram was made and the peak corresponded to the latency. When we needed the discharge rate of the unit in response to, for example, the retinal-slip velocity 2O”/s to the right, we found the values in the spike-density function (shifted for the time lag) corresponding to this velocity in the retinal-slip function. The velocity-sensitivity plot compiled the averages and the standard deviations of these values at each velocity point which is shown in the abscissae. RESULTS
IdentiJication of Mossy and Climbing Fiber Units. Three kinds of axonalspike units were recordable in the white matter: they were mossy fibers (MFs), climbing fibers (CFs), and the axons of Purkinje cells (P cells). Discharges of P cells were characterized by the presence of complex spikes interspersed among tonic simple spikes. In a separate series of experiments, we discovered that the majority of visually responsive P cells were so-called gaze velocity neurons and that their responses to sinusoidal retinal-slip velocities were unique (23). It is generally agreed that the complex spikes of a P cell are elicited by impulses transmitted by a CF. A CF makes an extensive all-or-none excitatory synaptic connection with the dendrites of a P cell. Whenever the CF discharges, the postsynaptic P cell also discharges. It is also well known that
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FIG. 1. The method used to evaluate sensitivity of neurons to retinal-slip velocity. A-Ecomputer simulation to obtain a smooth spike-density function by replacing each impulse with a Gaussian function (width = 25 ms), as originally described by MacPherson and Aldridge ( 12). G-a cross-correlation between the spike-density function (E) and the stimulus-velocity function (F) to estimate the conduction time (TIME LAG) from the retina to the flocculus in individual input fibers. H-to evaluate the firing rate of a fiber in response to a certain retinal-slip velocity, we measured the value in the spike-density function that corresponded to the velocity after shifting the stimulus velocity curve by the time lag. I-the averages and standard deviations of discharge rates were plotted against the retinal-slip velocity. In actual computations, the retinal-slip velocity was evaluated by subtracting the eye velocity (if any) from the stimulus velocity.
a CF shows characteristic low-frequency discharges. From the characteristic discharges patterns and behaviors of P cells and CFs, identification of these units is not difficult. Thus, MF units were identified by eliminating CF and P-cell units from the axonal-spike units recorded from the white matter of the flocculus. Climbing Fiber Responsesto Sinusoidal Retinal Slip. While the monkey fixated a stationary visual target, the random-dot pattern (Bkg) was moved sinusoidally at 0.5 Hz in the horizontal plane (+ lo” amplitude). Twenty of 429 CF-related units (4.9%) showed cyclic modulations in tiring. Eleven of 232 units (4.7%) were CFs recorded in the white matter; three of 44 units (6.8%) were evaluated from the complex-spike discharges of P cells; and seven of 153 units (4.5%) were CF responses recorded in the molecular layer.
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As illustrated in Fig. 2A, discharges of CF units were characterized by a low rate of spontaneous firing. The resting firing rates evaluated during primary fixation (fixation of the central target at eye level) ranged from 0.4 to 2.1 (average 1.3) spikes/s for the total sample. Firing rate of the CF increased during sinusoidal movements of Bkg at 0.5 Hz (Fig. 2B) and 0.3 Hz (Fig. 2C). The unit discharged only during ipsilateral Bkg movements (right). Peak firing rates were evaluated from smoothed phase histograms by computing a running average over five consecutive bins. At frequencies of 0. I, 0.3,0.5, and 0.7 Hz (+ lo“), the peak firing rates of the CF units were 3.4, 7.7, 11.8, and 15.6 spikes/s, respectively. The velocity sensitivity (evaluated by dividing the difference in peak firing rates by the
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difference in peak stimulus velocities) was 0.3 1 spikes. s-‘/deg . s-’ for the CF unit. The velocity sensitivity for seven CF units (tested with three or more stimulus frequencies) ranged from 0.03 to 0.44 (average 0.13) spikess-‘/deg . s-‘. The receptive field properties seen in the CF units illustrated in Fig. 3 were commonly observed in all 11 CFs. They were characterized by (i) relatively large size (> lo”) and inclusion of the fovea, (ii) excitatory responses during retinal slip in one direction and none in the other, and (iii) velocity-dependent peaks. All CF units in those tested had receptive fields in corresponding locations of both eyes (not illustrated). As seen in Fig. 3, sinusoidal movements of the Bkg presented within 10 of primary fixation (A), outside lo” (B), and on the entire screen (C) caused similar cyclic modulations in firing. As the peak firing rate was higher when
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both the central retina and periphery were simultaneously stimulated, the CF unit received visual inputs from a relatively large retinal portion. Mossy-Fiber Responses to Sinusoidal Retinal Slip. Among 485 MF units, identified by eliminating CF and P-cell units, 64 MF units (or 13%) responded to sinusoidal Bkg movements in the horizontal plane. Of these visually responsive MFs, 39 units (6 1%) responded exclusively to visual inputs (visual MFs), and the remaining 25 units (39%) responded also to oculomotor and vestibular inputs (visuomotor MFs). While the monkey maintained stationary fixation, the responses of both classes of MFs were almost identical. When retinal slip was accompanied either by eye or head movements, however, the responses of visuomotor MFs were markedly different from those of visual MFs. The details of response properties of visuomotor MFs in relation to retinal-slip, eye, and head velocities have already been described (23). In brief, the visuomotor MFs showed almost identical responses to Bkg movements regardless of whether the eyes were held stationary (causing retinal-slip velocity) or moved with the Bkg (replacing the retinal-slip velocity with eye velocity). The visuomotor MFs responded to the algebraic sum of retinal-slip and eye velocities. In contrast, the visual MFs in the present study responded only to retinal slip. When suppression of optokinetic nystagmus was incomplete, their discharges were unrelated to the sinusoidal Bkg movements. Responses of a visual MF during sinusoidal retinal slip (A) and during constant-velocity triangular oscillation of the Bkg (B) are shown in Fig. 4. As indicated by flat horizontal (H) and vertical (V) components of eye position shown in the excerpts of film records, the eyes were well maintained upon a stationary target. Under such conditions, the Bkg velocity corresponds to the retinal-slip velocity (in the opposite direction). The MF showed a relatively smooth cyclic modulation in response to the sinusoidal retinal slip (A). The peak activity appeared slightly before the maximum retinal-slip velocity. This lead reflected an accommodation of the neuron that is seen also in the slowly decaying response during the constant-velocity retinal slip (B). Among the 39 visual MF units, 26 (67%) showed velocity-related responses as exemplified by the MF of Fig. 4. Discharges in the remaining 13 MF units (33%) increased abruptly following the Bkg turnaround into the preferred direction and showed flat (or gradually decaying) responses thereafter. An example for each type of MF unit is shown in Fig. 5. Comparing the responses (phase histograms) of the two MF units with those of Fig. 4, one readily notices that MF708 (C) showed the flat response which was observed during constant-velocity stimulation (see Fig. 4B). Discharges of MF109 showed a cyclic modulation with peak activity near the maximum stimulus velocity (Fig. 5A). These MFs were sensitive not only to the direction but
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also to the velocity of Bkg movements (velocity MFs). On the other hand, with the stimulus velocity used in the present study (peak velocities ranged from 10 to 500/s), MF708 was sensitive only to the direction but not to the velocity of movements (direction-only MFs). It is inferred that the velocity of Bkg movements exceeded the sensitive range for the latter group of MFs. This inference (drawn from the phase histograms) was substantiated by computing the velocity sensitivity for each unit (B and D in Fig. 5), using the method ilhtstrated in Fig. 1. Visual Receptive Fields of Mossy Fiber Units. In contrast to the large receptive fields of the CF units that always included the fovea, those of MF units were found in two functionally distinct retinal regions. Receptive fields
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FIG. 5. Velocity- and direction-sensitive MF and direction-sensitive MF. The two types of MF are compared by showing phase histograms (A and C) and velocity-sensitive plots (B and D). Bin width for the phase histograms, 25 ms. Both MFs were stimulated with sinusoidal Bkg movements (the phase is shown by sinusoidal dashed lines) at 0.45 Hz, *lo’ amplitude. In response to the sinusoidal stimulus movements, discharges of MF109 were roughly sinusoidal and reached a peak slightly after maximum stimulus velocity whereas those of MF708 responded rather abruptly 100 to 125 ms after the turnaround. It did not show any obvious peak and the response decayed slowly thereafter. The velocity-sensitivity plots (B and D) were computed by evaluating retinal-slip velocity (the difference between stimulus velocity and eye velocity, if any) and smoothed spike density function (computed as illustrated in Fig. 1). The cross correlogram between these functions (inset in B) indicated that the effect of retinal-slip velocity reached maximum at 80 ms. The averages and standard deviations of the values found in the spike density function 80 ms after each velocity point of retinal slip velocity in the abscissae are shown as velocity sensitivity plots.
of 17 visual MFs (17/39 or 44%) were small and confined within lo” of fixation whereas those of 22 visual MFs (22/39 or 56%) were relatively large and were found in the periphery. In the peripheral group, receptive fields for 12 MFs were in the contralateral, those for 5 MFs were in the ipsilateral, and those for the remaining 5 MFs were in bilateral hemiretinae, all in both eyes. The numbers of visual MFs in relation to the range of sensitivity and to the location of the receptive field are summarized in Table 1. Although a slightly larger number of MFs was sensitive only to the direction of Bkg
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FIBERS TO FLOCCULUS TABLE I
The Number of Visual Mossy Fibers in Relation to the Location of the Receptive Field and to the Range of Velocity Sensitivity Receptive field
Sensitive to stimulus velocity used (peak velocity IO-W/s) Responses saturated, not sensitive to the velocity range used
Foveal region
Periphery
12 units 5 units
14 units 8 units
movements among the units activated from the periphery, this tendency was not significant (P z=-0.05, x2 test). Phasehistograms for a representative MF unit which was activated exclusively from the central lo” of fixation are shown in Fig. 6. When the Bkg movements were confined to the central retina (A), the MF showed a flat response.When the central lo’ area of the screenwas occluded and the Bkg was moved outside the occluded area, the MF did not respond (B). Changing the stimulus frequency (not illustrated) did not affect the
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FIG. 6. A MF with receptive field in the central retina. A-phase histogram during sinusoidal movements of Bkg at 0.6 Hz. The stimulus was presented within lo” of primary fixation. Bphase histogram for the same MF when the stimulus was presented outside the central lo”. C and D-phase histograms for the same MF when the presentation of the pattern was confined to the right (C) and left half(D) of the central 1o”, respectively.
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FIG. 7. A MF with receptive field in the periphery. A-phase histogram when the presentation of sinusoidally moving pattern was confined to the central lo” of primary fixation. B-phase histogram for the same MF when the stimulus was presented outside the central Lo’. C and Dphase histograms for the same MF when only the left (C) or right half(D) of the tangent screen was projected with the moving pattern, respectively.
height of responses. Interestingly, the receptive field encompassed both sides of the vertical meridian in all three MFs, showing flat responses (tested with the semicircular areas of the central retina open). This encompassing was not caused by unstable fixation. Phase histograms for a representative MF unit which was activated by visual inputs from the periphery are shown in Fig. 7. The stimulation of the centA retina did not evoke a response (A) whereas that of the periphery evoked responses. During stimulation of the entire screen (not illustrated), the left half of the periphery (C), and the right half of the periphery (D) yielded similar responses, indicating that the MF received excitatory visual inputs from bilateral hemiretinae (of both eyes). Not all units were tested with monocular stimulations, but every unit tested showed responses to stimulation of both eyes. Binocular interactions were not tested in the present study. DISCUSSION Visual Climbing-I;iber Units. The visual climbing fiber (CF) input to the flocculus in the rabbit is we11known ( 14- 16) and arises from the dorsal cap
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of Kooy of the inferior olive (1). The activity of the CF can be well modulated by slowly moving visual stimuli (3, 26-28). In spite of the extensive studies on the rabbit, visual CFs in the primate have not yet been reported. In the present study, 11 CFs and 10 complex-spike units from P-cell discharges responded in a speed- and direction-selective manner to a large textured pattern. These numbers corresponded to less than 5% of the CFs (including the CF responses from P cells) examined. It is likely that the low sampling rate of visual CFs in the primate was caused by the difference in the animal preparation rather than by the species difference. Whether or not the responses could be tested with a slow movement of the stimulus appears to be of critical importance. All rabbits used in the previous reports had been deeply anesthetized and immobilized whereas our monkeys were alert and eye movements were suppressed only by the effort made by the monkeys to fixate on a stationary visual target. According to Simpson and Alley (26), the great majority of CFs in the rabbit were optimally responsive to movements of less than 1O/s.In our study, the range of the peak stimulus velocities was typically from 10 to BY/s. The CFs were stimulated with horizontal movements presented only in the central 1 lo” of fixation (frontal-plane stimulation); all units were activated binocularly; the preferred directions were always the same for both eyes. In the rabbit, however, the receptive fields of dorsal cap neurons extended from the nose (00) to 160” posteriorly along the horizon (29), and were quite different from the present observation. For example, some neurons were modulated only from the contralateral eye and were best excited by horizontal movement in the temporonasal direction and inhibited in the nasotemporal direction. There were such neurons showing receptive fields in both eyes but the preferred direction for one eye was different from that for the other eye (29). If receptive field organizations of dorsal cap neurons in the primate bear any resemblance to those in the rabbit, our binocular and frontal-plane stimulation might have activated only a small percentage of dorsal cap neurons. Visually Responsive Mossy-Fiber Units. Two distinct classes of visually responsive mossy-fiber (MF) units were recorded from the flocculus. Depending on whether or not the eye and head were stationary, their visual responses were markedly different. Visual MF units responded only when the eye and head were stationary, hence responding exclusively to retinalslip velocity. Visuomotor MF units reflected the stimulus movement itself, regardless of eye position; hence the activity was related to the algebraic sum of retinal-slip, eye, and head (in the opposite direction) velocities. The visual MFs originate from different structures in the brain stem. It is most unlikely that their parent cells are vestibular neurons because they did not show any sign of vestibular input. The possibility of being prepositus
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hypoglossi neurons may also be excluded for the same reason because these neurons receive vestibular inputs (2). It is possible that the MFs which received visual inputs from the central retina originate in the dorsolateral pontine nucleus (DLPN). Suzuki and Keller (30) and Mustari et al. (19) have recorded visually responsive units in the DLPN of macaque monkeys. The response characteristics of these neurons were very similar to those of visual MFs recorded in the flocculus of the macaque (20). The receptive fields of these neurons always included the fovea (19). Projections are known from various visual cortices to this pontine nucleus (7, 8), including the superior temporal motion area MT (5, 6, 32, 33). The lateral margin of the DLPN has been identified as a major floccular projecting portion of the pontine nuclei (10). The origin of the MFs receiving visual inputs from the peripheral retina is as yet unclear. Possible sources of visual MFs have been suggested by experiments on lower mammals. The flocculus receives visual information via MFs arising from the nucleus reticularis tegmenti pontis in the rabbit ( 13, 16) and in the cat (9). Neurons in the medial terminal nucleus of the rabbit also respond to slowly moving textured patterns (28). Although the primary projection from this nucleus to the flocculus is via CFs arising from the dorsal cap of the inferior olive, it is possible, but not yet confirmed, that the neurons in the medial terminal nucleus project directly to the flocculus as a MF input (4,371. Comparison between Climbing and Mossy Fiber Visual Units. A striking difference in receptive field organizations between CFs and MFs was that CFs had large and homogeneous receptive fields which always included the fovea, whereas MFs had receptive fields either in the foveal or peripheral region. When the stimulus was confined either to the fovea1 or peripheral retina, MFs tended to show more prominent responses. On the other hand, CFs received similar visual inputs from both the foveal and peripheral regions. When both regions were stimulated simultaneously, the responses tended to be larger in many CFs. Receptive fields of neurons in the flocculus-related brain stem structures have not yet been studied in the primate or in other mammals, except for the dorsal cap neurons in the rabbit (29). Depending on whether or not the animal uses binocular and frontal vision, receptive field organizations of visual neurons would be considerably different. In the present experiments, in addition to the range of stimulus velocities already discussed, there were two other limitations: one was the direction of stimulus movements that were always horizontal, and the other was the area of stimulus presentation that had been limited to only the central 1 lo”. In spite of the limitations in the experimental paradigms, there were several features in the receptive field organization in the primate: (i) both CFs and MFs, in those tested, had almost
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identical receptive fields in corresponding loci of the two eyes; (ii) they had the same preferred directions; and (iii) the two eyes yielded similar responses with respect both to the gain and to the phase-relation to the stimulus. Functional Significance of the Visual Input. The most important finding in the present study was that the flocculus received visual inputs from two functionally distinct portions of the retina via different channels of MFs. One group of MFs conveyed visual inputs only from the foveal region and the other group of MFs was responsible for the visual inputs from the periphery. The signals carried by these two groups of MFs were transmitted through different visual pathways, gave different effects on the P cells in the flocculus, and implied different functional roles. When stimulated with sinusoidally moving patterns, a large number of P cells in the flocculus were bidirectionally depressed (23). Among these cells, approximately three-quarters had two visual receptive fields, one in the foveal region and the other in the periphery 30 to 70” away from the fovea. When visual stimulation was confined to within lo” of fixation, the P cells were excited. The responses were velocity-dependent. When the moving stimulation Bkg was presented in the periphery, the discharge rates of the same P cells were decreased to 40 to 60% of the spontaneous activity. As the visual receptive fields of CFs were large and always included the fovea in the primate, it is unlikely that these fibers are responsible for the different responses in the same P cells. The excitatory and inhibitory responses in the P cells, therefore, must have been caused by the visual inputs transmitted by the MFs which had receptive fields in the foveal and peripheral regions of the retina, respectively. On the other hand, the inhibitory visual input from the periphery must have been conveyed by a group of visual MFs with peripheral receptive fields and the visuomotor MFs. All visuomotor MFs have their visual receptive fields in the periphery. The parent cells of the fibers are most likely in the vestibular nuclei (2 1). The origin of the visual MFs with peripheral receptive fields must be some brain stem nuclei such as the reticularis tegmenti pontis and the medial terminal nucleus which are known to send MFs to the flocculus. The pathway for the inhibitory visual input to the floccular P cells would be, therefore, related to the direct projection from the retina to the brain stem visual nuclei which are responsible for the generation of the optokinetic nystagmus. It is generally agreed that the flocculus serves to control both smooth pursuit and optokinetic nystagmus in the primate. The visual input arising in the foveal region, possibly transmitted through the cerebropontinecerebcllar pathway to the flocculus, may serve the smooth-pursuit function while the input from the peripheral retina, possibly taking the direct retino brain stem pathway, may be related to the optokinetic function. The visually responsive
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MFs discovered by the present study may serve to transmit these signals to the flocculus. REFERENCES 1. ALLEY, K., R. BAKER, AND J. I. SIMPSON. 1975. Afferents to the vestibulo-cerebellum and the origin of the visual climbing fibers in the rabbit. Bruin Res. 98: 582-589. 2. BAKER, R. 1977. The nucleus prepositus hypoglossi. Pages 145-178 in B. A. BROOKS AND F. J. BAJANDAS, Eds., Eye Movements. Plenum, New York/London. 3. BARMACK, N. H., AND D. T. HESS. 1980. Multiple-unit activity evoked in dorsal cap of inferior olive of the rabbit by visual stimulation. J. Neurophysiol. 43: 15 I - 164. 4. BRAUTH, S. E., AND H. J. KARTEN. 1977. Direct accessory optic projections to the vestibulo-cerebellum. A possible channel for oculomotor systems.Exp. Brain Res. 27: 73-84. 5. FRIES, W. 198 1. The projection from striate and prestriate visual cortex onto the pontine nuclei in the macaque monkey. Sot. Neurosci. Abstr. 7: 762. 6. GALLETTI, C., M. G. MAIOLI, S. !~QUATITRO,AND P. F. BATTAGLINI. 1982. Cortico-pontine projections from the visual area of the superior temporal sulcus in the macaque monkey. Arch. Ital. Biol. 120: 411-416. 7. GLICKSTEIN, M., AND J. MAY. 1982. Visual control of movement: the circuits which link visual and motor areas of the brain with special reference to the pons and cerebellum. Pages 103-145 in W. D. NEFF, Ed., Contributions to Sensory PhysioZogy. Academic Press, New York. 8. GLICKSTEIN, M., J. MAY, AND B. MERCER. 1985. Cortico-pontine projection in the macaque: the distribution oflabelled cortical cells after large injections of horseradish peroxidase in the pontine nuclei. J. Comp. Neural. 235: 343-359. 9. KAWASAKI, T., Y. SATO, AND I. KATO. 1980. The visual input pathway to the cerebellar tlocculus. Equilib. Res. 39: l-6. IO. LANGER, T., A. F. FUCHS, C. A. SCUDDER, AND M. C. CHUBB. 1985. Afferents to the flocculus of the cerebellum in the rhesus macaque as revealed by retrograde transport of horseradish peroxidase. J. Comp. Neural. 235: I-25. 11. LISBERGER, S. G., AND A. F. FUCHS. 1978. Role of primate flocculus during rapid behavioral modification of vestibulo-ocular reflex. I. Purkinje cell activity during visually guided horizontal smooth pursuit eye movements and passive head rotation. J. Neurophysiol. 41: 733-763. 12. MACPHERSON, J. M., AND J. W. ALDRIDGE. 1979. A quantitative method of computer analysis of spike train data collected from behaving animals. Brain Res. 175: 183- 187. 13. MAEKAWA, K., M. KIMURA, AND T. TAKEDA. 198 1. Mossy fiber activation ofthe cerebellar flocculus from the visual system.Ann. N. Y. Acad. Sci. 374: 476-490. 14. MAEKAWA, K., AND J. I. SIMPSON. 1973. Climbing fiber response evoked in vestibulocerebellum of rabbit from visual system.J. Neurophysiol. 36: 649-666. 15. MAEKAWA, K., AND T. TAKEDA. 1976. Electrophysiological identification of the climbing and mossy fiber pathways from the rabbit’s retina to the contralateral cerebellar flocculus. Brain Res. 109: 169- 174. 16. MAEKAWA, K., AND T. TAKEDA. 1977. Afferent pathways from the visual system to the cerebellar flocculus of the rabbit. Pages 187-196, in R. BAKER AND A. BERTHOZ, Eds., Control of Gaze by Brain Stem Neurons, Elsevier/North-Holland, Amsterdam/New York. 17. MILES, F. A., AND J. H. FULLER. 1975. Visual tracking and the primate flocculus. Science 189: 1000-1002.
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