Heterogeneity of parvalbumin expression in the avian cerebellar cortex and comparisons with zebrin II.

The cerebellar cortex has a fundamental parasagittal organization that is reflected in the physiological responses of Purkinje cells, afferent and efferent connections, and the expression of several molecular markers. The most thoroughly studied of these molecular markers is zebrin II (ZII; a.k.a. aldolase C). ZII is differentially expressed in Purkinje cells, resulting in a pattern of sagittal stripes of high expression interdigitated with stripes of little or no expression. In this study, we examined the expression of the calcium binding protein parvalbumin (PV) in the cerebellum of several avian species (pigeons, hummingbirds, zebra finches) and compared it to the expression of ZII. We found that PV immunoreactivity was distributed across the cerebellar cortex such that there were sagittal stripes of PV immunopositive (PV+) Purkinje cells alternating with PV immunonegative (PV-) Purkinje cells. Although most Purkinje cells in the anterior lobe were PV+, there were several thin (i.e. only a few Purkinje cells wide) PV- stripes spanning the folia. In the posterior lobe, PV+ and PV- stripes were also apparent, but the PV- stripes were much wider than in the anterior lobe. In sections processed for both ZII and PV, the expression was generally complementary: PV+ stripes were ZII-, and vice-versa. This complementary expression was most apparent in folia II-IV and VIII-IXcd. The complementary expression was not, however, absolute; some Purkinje cells co-expressed PV and ZII whereas others lacked both. These novel findings relate to the complex neurochemical organization of the cerebellum, and are likely important to issues regarding cerebellar plasticity.

Congruence of zebrin II expression and functional zones defined by climbing fiber topography in the flocculus.

The cerebellum is organized into parasagittal zones with respect to the topography of climbing fiber (CF) afferents and the expression of molecular markers such as zebrin II. Zebrin is expressed by a subset of Purkinje cells that are distributed as a parasagittal array of immunopositive and immunonegative stripes. Several studies in rodents suggest that, in general, CFs to the zebrin negative stripes convey somatosensory information, whereas CFs to the zebrin positive stripes convey information from visual and other sensory systems. The pigeon flocculus consists of four pairs of zebrin+/- stripes (P4 +/- through P7 +/-), however the CF input consists entirely of visual inputs. Thus, because the correspondence of zebrin expression and CF information must be different from that proposed for rodents, we investigated this relationship in the pigeon flocculus. Floccular Purkinje cells respond to patterns of optic flow resulting from self-rotation about one of two axes: either the vertical axis (zones 0 and 2), or a horizontal axis (zones 1 and 3). Visual CF afferents projecting to the flocculus arise from the medial column of the inferior olive (mcIO). Zones 0 and 2 receive input from the caudal mcIO, whereas zones 1 and 3 receive input from the rostral mcIO. We injected a fluorescent anterograde tracer into the rostral and/or caudal mcIO and visualized zebrin expression. There was a strict concordance between CF organization and zebrin labeling: caudal mcIO injections resulted in CFs in zebrin bands P4 +/- and P6 +/-, whereas rostral mcIO injections resulted in CFs in zebrin bands P5 +/- and P7 +/-. Thus, zebrin stripes P4 +/- and P6 +/- correspond to the vertical axis zones 0 and 2, whereas P5 +/- and P7 +/- correspond to the horizontal axis zones 1 and 3. This is the first explicit demonstration that a series of zebrin stripes corresponds with functional zones in the cerebellum.

A comparison of ventral tegmental neurons projecting to optic flow regions of the inferior olive vs. the hippocampal formation.

The ventral tegmental area (catecholaminergic group A10) is a midbrain region characterized by concentrated dopaminergic immunoreactivity. Previous studies in pigeons show that the ventral tegmental area provides a robust projection to the hippocampal formation and to the medial column of the inferior olive. However, the distribution, morphology, and neurochemical content of the neurons that constitute these projections have not been resolved. In this study, we used a combination of retrograde tracing techniques and immunofluorohistochemistry to address these issues. Retrograde tracers were used to demonstrate that the distribution of ventral tegmental area neurons projecting to the hippocampus and the inferior olive overlap in the caudo-ventral ventral tegmental area. The hippocampus- and inferior olive-projecting ventral tegmental area neurons could not be distinguished based on morphology: most neurons had small- to medium-sized multipolar or fusiform soma. Double-labeling with fluorescent retrograde tracers revealed that the hippocampus- and medial column of the inferior olive-projecting neurons were found intermingled in the ventral tegmental area, but no cells were double labeled; i.e. individual ventral tegmental area neurons do not project to both the hippocampal formation and medial column of the inferior olive. Finally, we found that a minority (8.2%) of ventral tegmental area neurons providing input to the hippocampus were tyrosine hydroxylase-immunoreactive, whereas none of the inferior olive-projecting neurons were tyrosine hydroxylase positive. Combined, our findings show that the projections to the hippocampus and olivocerebellar pathway arise from intermixed subpopulations of ventral tegmental area neurons with indistinguishable morphology but only the hippocampal projection involves dopaminergic neurons. We suggest that equivalent projections from the ventral tegmental area to the hippocampal formation and inferior olive exist in mammals and discuss their potential role in the processing of optic flow and the analysis of self-motion.

Responses of neurons in the medial column of the inferior olive in pigeons to translational and rotational optic flowfields.

The responses of neurons in the medial column of the inferior olive to translational and rotational optic flow were recorded from anaesthetized pigeons. Panoramic translational or rotational flowfields were produced by mechanical devices that projected optic flow patterns onto the walls, ceiling and floor of the room. The axis of rotation/translation could be positioned to any orientation in three-dimensional space such that axis tuning could be determined. Each neuron was assigned a vector representing the axis about/along which the animal would rotate/translate to produce the flowfield that elicited maximal modulation. Both translation-sensitive and rotation-sensitive neurons were found. For neurons responsive to translational optic flow, the preferred axis is described with reference to a standard right-handed coordinate system, where +x, +y and +z represent rightward, upward and forward translation of the animal, respectively (assuming that all recordings were from the right side of the brain). t(+y) neurons were maximally excited in response to a translational optic flowfield that results from self-translation upward along the vertical (y) axis. t(-y) neurons also responded best to translational optic flow along the vertical axis but showed the opposite direction preference. The two remaining groups, t(-x+z) and t(-x-z) neurons, responded best to translational optic flow along horizontal axes that were oriented 45 degrees to the midline. There were two types of neurons responsive to rotational optic flow: rVA neurons preferred rotation about the vertical axis, and rH135c neurons preferred rotation about a horizontal axis at 135 degrees contralateral azimuth. The locations of marking lesions indicated a clear topographical organization of the six response types. In summary, our results reinforce that the olivo-cerebellar system dedicated to the analysis of optic flow is organized according to a reference frame consisting of three approximately orthogonal axes: the vertical axis, and two horizontal axes oriented 45 degrees to either side the midline. Previous research has shown that the eye muscles, vestibular semicircular canals and postural control system all share a similar spatial frame of reference.

Detection of glass patterns by pigeons and humans: implications for differences in higher-level processing.

Glass patterns have been used to examine mechanisms underlying form perception. The current investigation compared detection of Glass patterns by pigeons and humans and provides evidence for substantial species differences in global form perception. Subjects were required to discriminate, on a simultaneous display, a random dot pattern from a Glass pattern. Four different randomly presented Glass patterns were used (concentric, radial, parallel-vertical, and parallel-horizontal). Detection thresholds were measured by degrading the Glass patterns through the addition of random noise. For both humans and pigeons, discrimination decreased systematically with the addition of noise. Humans showed detection differences among the four patterns, with lowest thresholds to radial and concentric patterns and highest thresholds to the parallel-horiZontal pattern. Pigeons did not show a detection difference across the four patterns. Implications for differences in neural processing of complex forms are discussed.

Fast and slow neurons in the nucleus of the basal optic root in pigeons.

The nucleus of the basal optic root (nBOR) is involved in the generation of the optokinetic response. Previous studies showed that most nBOR neurons exhibit direction-selectivity in response to largefield motion. We investigated the responses of pigeon nBOR neurons to drifting sine wave gratings of varying spatial and temporal frequency (SF,TF). Two groups of neurons were revealed. The first group preferred gratings of low SF (mean, 0.07 cycles per degree (cpd)) and high TF (mean, 0.76 Hz) ('fast' stimuli). The second group preferred gratings of high SF (mean, 0.56 cpd) and lower TF (mean, 0.33 Hz) ('slow' stimuli). Previous studies have demonstrated fast and slow neurons in pretectal nucleus lentiformis mesencephali, which is also involved in the generation of the optokinetic response.

Projections from the nucleus of the basal optic root and nucleus lentiformis mesencephali to the inferior olive in pigeons (Columba livia).

The nucleus of the basal optic root (nBOR) of the accessory optic system (AOS) and the pretectal nucleus lentiformis mesencephali (LM) are involved in the analysis of optic flow and the generation of the optokinetic response. Previous studies have shown that the nBOR projects bilaterally to the medial column (mc) of the inferior olive (IO) and the LM projects to the ipsilateral mc. In the present study the retrograde tracer cholera toxin subunit B was injected into either the caudal or rostral mc. From all injections, retrogradely labeled cells were seen in the ipsilateral pretectum along the border of the medial and lateral subnuclei of the LM. Cells were also seen in bilaterally in the nBOR. On the contralateral side, a discrete group of cells was labeled in the rostral margin of the nBOR. These cells were localized in the dorsal portion of the nBOR proper and some were found in the adjacent nBOR dorsalis. On the ipsilateral side, a diffuse group of cells was seen in the caudal nBOR. Most of these cells were in the nBOR dorsalis and outside the nBOR complex in the area ventralis of Tsai and the reticular formation. From the injections into the caudal mc, a greater proportion of labeled cells was found in the LM, whereas a greater proportion of cells was found in the nBOR from the injections into the rostral mc. This differential projection from LM and nBOR to the caudal and rostral mc is consistent with the optic flow preferences of neurons in the mc, and a similar pattern of connectivity has been found in mammalian species.

Spatiotemporal properties of fast and slow neurons in the pretectal nucleus lentiformis mesencephali in pigeons.

Neurons in the pretectal nucleus lentiformis mesencephali (LM) are involved in the analysis of optic flow that results from self-motion. Previous studies have shown that LM neurons have large receptive fields in the contralateral eye, are excited in response to largefield stimuli moving in a particular (preferred) direction, and are inhibited in response to motion in the opposite (anti-preferred) direction. We investigated the responses of LM neurons to sine wave gratings of varying spatial and temporal frequency drifting in the preferred and anti-preferred directions. The LM neurons fell into two categories. "Fast" neurons were maximally excited by gratings of low spatial [0.03-0.25 cycles/ degrees (cpd)] and mid-high temporal frequencies (0.5-16 Hz). "Slow" neurons were maximally excited by gratings of high spatial (0.35-2 cpd) and low-mid temporal frequencies (0.125-2 Hz). Of the slow neurons, all but one preferred forward (temporal to nasal) motion. The fast group included neurons that preferred forward, backward, upward, and downward motion. For most cells (81%), the spatial and temporal frequency that elicited maximal excitation to motion in the preferred direction did not coincide with the spatial and temporal frequency that elicited maximal inhibition to gratings moving in the anti-preferred direction. With respect to motion in the anti-preferred direction, a substantial proportion of the LM neurons (32%) showed bi-directional responses. That is, the spatiotemporal plots contained domains of excitation in addition to the region of inhibition. Neurons tuned to stimulus velocity across different spatial frequency were rare (5%), but some neurons (39%) were tuned to temporal frequency. These results are discussed in relation to previous studies of the responses of neurons in the accessory optic system and pretectum to drifting gratings and other largefield stimuli.

Binocular neurons in the nucleus lentiformis mesencephali in pigeons: responses to translational and rotational optic flowfields.

The pretectal nucleus lentiformis mesencephali (LM) receives direct input from the contralateral retina and is dedicated to the analysis of optic flowfields resulting from self-motion. The activity of 126 LM neurons in response to optic flow stimuli was recorded. As with previous studies, it was found that most neurons (approximately 90%) exhibited direction-selectivity to large-field stimuli moving in the contralateral hemifield. However, some neurons (approximately 10%) responded to stimulation of both eyes and had receptive field structures conducive for detection of particular patterns of optic flow resulting from either self-translation or self-rotation. These binocular neurons were maximally responsive to panoramic optic flowfields simulating either translational or rotational optic flow.

Topographic organization of inferior olive cells projecting to translational zones in the vestibulocerebellum of pigeons.

In the nodulus and ventral uvula of pigeons, there are four parasagittal zones containing Purkinje cells responsive to patterns of optic flow that results from self-translation along a particular axis in three-dimensional space. By using a three-axis system to describe the preferred direction of translational optic flow, where +X, +Y, and +Z represent rightward, upward, and forward self-motion, respectively, the four cell types are: +Y, -Y, -X-Z, and -X+Z (assuming recording from the left side of the head). The -X-Z zone is the most medial, followed in sequence by the -X+Z, -Y zone, and the +Y zones. In this study, we injected the retrograde tracer cholera toxin subunit B into each of the four translational zones to determine the origin of the climbing fiber inputs in the inferior olive. Retrograde labeling in the inferior olive was found in the ventrolateral margin of the medial column from injections into all four translational zones; however, there was a clear functional topography. Retrograde labeling from -Y zone injections was found most rostrally in the medial column, whereas retrogradely labeled cells from -X-Z zone injections were found most caudally in the medial column. Labeling from +Y and -X+Z zone injections were found between the labeling from -Y zones and -X-Z zones, with +Y labeling located slightly caudal to -X+Z labeling.

Perception of coherent motion in random dot displays by pigeons and humans.

Pigeons and humans were required to discriminate coherent from random motion in dynamic random dot displays. Coherence and velocity thresholds were determined for both species, and both thresholds were found to be substantially higher for pigeons than for humans. The results are discussed with reference to differences in motion processing in mammals and birds. It is suggested that the inferior motion sensitivity of pigeons can be attributed to poorer spatiotemporal motion integration.

Projections of purkinje cells in the translation and rotation zones of the vestibulocerebellum in pigeon (Columba livia).

Previous electrophysiological studies have shown that the pigeon vestibulocerebellum (ventral uvula, nodulus, and flocculus) can be divided into two parasagittal zones based on responses to optic flow stimuli. The medial zone (ventral uvula and nodulus) responds best to optic flow resulting from self-translation, whereas the lateral zone (flocculus) responds best to optic flow resulting from self-rotation. In this study we investigated the projections of the Purkinje cells in the translation and rotation zones of the vestibulocerebellum by using the anterograde tracer biotinylated dextran amine. Extracellular recording of Purkinje cell activity (complex spikes) in response to large-field visual stimuli were used to identify the injection sites. Injections into the translation zone resulted in extremely heavy terminal labeling in the cerebellovestibular process adjacent to the medial cerebellar nucleus. A moderate amount of terminal labeling was found in the medial cerebellar nucleus, the superior vestibular nucleus (laterally, dorsally, and medially), and the descending vestibular nucleus, particularly in the lateral half. Light terminal labeling was observed in the dorsolateral vestibular nucleus, the medial vestibular nucleus, the tangential nucleus, and the lateral vestibular nucleus pars ventralis. Injections into the rotation zone resulted in heavy terminal labeling in the superior vestibular nucleus (particularly dorsally and medially), the descending vestibular nucleus (particularly medially), and the medial vestibular nucleus. A moderate amount of terminal labeling was seen in the cerebellovestibular process adjacent to the lateral cerebellar nucleus, and the dorsolateral vestibular nucleus. A small amount of terminal labeling was found in the lateral cerebellar nucleus, the tangential nucleus, the prepositus hypoglossi, and the lateral vestibular nucleus pars ventralis.

Projections from the medial column of the inferior olive to different classes of rotation-sensitive Purkinje cells in the flocculus of pigeons.

In the flocculus of pigeons, as in other species, there are two major types of Purkinje cell responses to rotational optokinetic stimuli. One type prefers rotation about the vertical axis (VA neurons) whereas the other prefers rotation about an horizontal axis oriented at 135 degrees ipsilateral azimuth (H-135 neurons). In this study, we injected the retrograde tracer cholera toxin subunit B into the VA and H-135 zones in attempt to determine the origin of inferior olive inputs. We found that VA and H-135 zones received input from the caudal and rostral margins of the medial column of the inferior olive, respectively. There is a similar pattern of connectivity in mammalian species.

Optic flow input to the hippocampal formation from the accessory optic system.

Recent studies in rodents have implicated the hippocampal formation in "path integration": the ability to use self-motion cues (ideothesis) to guide spatial behavior. Such models of hippocampal function assume that self-motion information arises from the vestibular system. In the present study we used the retrograde tracer cholera toxin subunit B, the anterograde tracer biotinylated dextran amine, and standard extracellular recording techniques to investigate whether the hippocampal formation [which consists of the hippocampus proper and the area parahippocampalis (Hp/APH) in pigeons] receives information from the accessory optic system (AOS). The AOS is a visual pathway dedicated to the analysis of the "optic flow fields" that result from self-motion. Optic flow constitutes a rich source of ideothetic information that could be used for navigation. Both the nucleus of the basal optic root (nBOR) and nucleus lentiformis mesencephali of the AOS were shown to project to the area ventralis of Tsai (AVT), which in turn was shown to project to the Hp/APH. A smaller direct projection from the nBOR pars dorsalis to the hippocampus was also revealed. During extracellular recording experiments, about half of the cells within the AVT responded to optic flow stimuli. Together these results illustrate that the Hp/APH receives information about self-motion from the AOS. We postulate that this optic flow information is used for path integration. A review of the current literature suggests that an analogous neuronal circuit exists in mammals, but it has simply been overlooked.

Complex spike activity of Purkinje cells in the ventral uvula and nodulus of pigeons in response to translational optic flow.

The complex spike (CS) activity of Purkinje cells in the ventral uvula and nodulus of the vestibulocerebellum was recorded from anesthetized pigeons in response to translational optic flow. Translational optic flow was produced using a "translator" projector: a mechanical device that projected a translational optic flowfield onto the walls, ceiling, and floor of the room and encompassed the entire binocular visual field. CS activity was broadly tuned but maximally modulated in response to translational optic flow along a "best" axis. Each neuron was assigned a vector representing the direction in which the animal would need to translate to produce the optic flowfield that resulted in maximal excitation. The vector is described with reference to a standard right-handed coordinate system, where the vectors, +x, +y, and +z represent, rightward, upward, and forward translation of the animal, respectively. Neurons could be grouped into four response types based on the vector of maximal excitation. +y neurons were modulated maximally in response to a translational optic flowfield that results from self-motion upward along the vertical (y) axis. -y neurons also responded best to translational optic flow along the vertical axis but showed the opposite direction preference. The two remaining groups responded best to translational optic flow along horizontal axes: -x + z neurons and -x-z neurons. In summary, our results suggest that the olivocerebellar system dedicated to the analysis of translational optic flow is organized according to a reference frame consisting of three approximately orthogonal axes: the vertical axis, and two horizontal axes oriented 45 degrees to either side the midline. Previous research has shown that the rotational optic flow system, the eye muscles, the vestibular semicircular canals and the postural control system all share a similar spatial frame of reference.

Responses of neurons in the nucleus of the basal optic root to translational and rotational flowfields.

The nucleus of the basal optic root (nBOR) receives direct input from the contralateral retina and is the first step in a pathway dedicated to the analysis of optic flowfields resulting from self-motion. Previous studies have shown that most nBOR neurons exhibit direction selectivity in response to large-field stimuli moving in the contralateral hemifield, but a subpopulation of nBOR neurons has binocular receptive fields. In this study, the activity of binocular nBOR neurons was recorded in anesthetized pigeons in response to panoramic translational and rotational optic flow. Translational optic flow was produced by the "translator" projector described in the companion paper, and rotational optic flow was produced by a "planetarium projector" described by Wylie and Frost. The axis of rotation or translation could be positioned to any orientation in three-dimensional space. We recorded from 37 cells, most of which exhibited a strong contralateral dominance. Most of these cells were located in the caudal and dorsal aspects of the nBOR complex and many were localized to the subnucleus nBOR dorsalis. Other units were located outside the boundaries of the nBOR complex in the adjacent area ventralis of Tsai or mesencephalic reticular formation. Six cells responded best to rotational flowfields, whereas 31 responded best to translational flowfields. Of the rotation cells, three preferred rotation about the vertical axis and three preferred horizontal axes. Of the translation cells, 3 responded best to a flowfield simulating downward translation of the bird along a vertical axis, whereas the remaining 28 responded best to flowfields resulting from translation along axes in the horizontal plane. Seventeen of these cells preferred a flowfield resulting from the animal translating backward along an axis oriented approximately 45 degrees to the midline, but the best axes of the remaining eleven cells were distributed throughout the horizontal plane with no definitive clustering. These data are compared with the responses of vestibulocerebellar Purkinje cells.

Topographical organization of inferior olive cells projecting to translation and rotation zones in the vestibulocerebellum of pigeons.

Previous electrophysiological studies in pigeons have shown that the vestibulocerebellum can be divided into two parasagittal zones based on responses to optic flow stimuli. The medial zone responds best to optic flow resulting from self-translation, whereas the lateral zone responds best to optic flow resulting from self-rotation. This information arrives from the retina via a projection from the accessory optic system to the medial column of the inferior olive. In this study we investigated inferior olive projections to translational and rotational zones of the vestibulocerebellum using the retrograde tracer cholera toxin subunit B. Extracellular recordings of Purkinje cell activity (complex spikes) in response to large-field visual stimuli were used to identify the injection sites. We found a distinct segregation of inferior olive cells projecting to translational and rotational zones of the vestibulocerebellum. Translation zone injections resulted in retrogradely labeled cells in the ventrolateral area of the medial column, whereas rotation zone injections resulted in retrogradely labeled cells in the dorsomedial region of the medial column. Motion of any object through space, including self-motion of organisms, can be described with reference to translation and rotation in three-dimensional space. Our results show that, in pigeons, the brainstem visual systems responsible for detecting optic flow are segregated into channels responsible for the analysis of translational and rotational optic flow in the inferior olive, which is only two synapses from the retina.

Common reference frame for neural coding of translational and rotational optic flow.

Self-movement of an organism through the environment is guided jointly by information provided by the vestibular system and by visual pathways that are specialized for detecting 'optic flow'. Motion of any object through space, including the self-motion of organisms, can be described with reference to six degrees of freedom: rotation about three orthogonal axes, and translation along these axes. Here we describe neurons in the pigeon brain that respond best to optic flow resulting from translation along one of the three orthogonal axes. We show that these translational optic flow neurons, like rotational optic flow neurons, share a common spatial frame of reference with the semicircular canals of the vestibular system. The three axes to which these neurons respond best are the vertical axis and two horizontal axes orientated at 45 degrees to either side of the body midline.

Projections from the accessory optic system and pretectum to the dorsolateral thalamus in the pigeon (Columbia livia): a study using both anteretrograde and retrograde tracers.

In birds, optic flow is analyzed by two retinal-recipient nuclei: the nucleus of the basal optic root (nBOR) of the accessory optic system (AOS), and the pretectal nucleus, lentiformis mesencephali (LM). Previous anatomical studies have shown that both of these nuclei have descending projections to structures involved in oculomotor, head movement, and postural control. In this report, using biotinylated dextran amine (BDA) and cholera toxin subunit B (CTB) for anterograde and retrograde labelling, respectively, we investigated projections from the nBOR and LM to the dorsal thalamus. After injections of BDA into the nBOR and LM, terminals were consistently found in the nucleus dorsolateralis anterior pars lateralis and pars medialis, and the nucleus dorsalis intermedius ventralis anterior of the thalamus. Some terminals were also found in the nucleus dorsolateralis anterior, nucleus dorsomedialis anterior pars magnocellularis, nucleus dorsolateralis posterior, nucleus superficialis parvocellularis, and the ventrointermediate area. Injections of CTB into the dorsal thalamus resulted in retrogradely labelled cells in the pretectal region, including LM. Numerous cells were also seen in the nBOR pars lateralis and pars dorsalis, but fewer were seen in the nBOR proper. We suggest that the AOS is providing input to a thalamotelencephalic system that may be involved in several functions including: (1) multi-sensory analysis of self-motion, (2) perception of self-motion, (3) perception of the three-dimensional layout of the environment, (4) distinguishing object-motion from self-motion, and (5) spatial cognition.