Metabolic changes in the nucleus of the optic tract after monocular enucleation as revealed by cytochrome oxidase histochemistry.

The histochemistry for the mitochondrial enzyme cytochrome oxidase (CO) was used to evaluate the levels of metabolic activity in neurons of the nucleus of the optic tract (NOT) and dorsal terminal nucleus (DTN) in the opossum (Didelphis aurita). The observations were performed in four groups: normal juveniles (4 months old), monocularly enucleated juveniles analysed when adults, normal adults (8 to 18 months old) and monocularly enucleated adults. CO labeled cells were observed to have a similar distribution along the NOT-DTN anteroposterior axis in both juvenile and adult normal animals. Monocular enucleation performed in adults produced a significant reduction of the reactive neuropil but not of the number of CO labeled cells in the deafferented NOT-DTN: the number of labeled neurons per section in the deafferented side matched those of the ipsilateral complex. In juveniles, however, this procedure caused a systematic reduction of the number of CO labeled cells in the contralateral NOT-DTN in comparison to the spared complex. The lack of reduction in the number of neurons found on the deafferented side of the NOT-DTN of monocularly enucleated adult opossums compared with the ipsilateral side might result from the presence of compensatory inputs to maintain their metabolic equivalence. However, when the monocular enucleation was performed in juvenile opossums, a statistically significant asymmetry of CO neurons in the NOT-DTN was observed. In other words, the compensatory mechanisms proposed for the adults were either absent or insufficient to achieve symmetry in juveniles, suggesting a more heavily reliance in the retinal input.

Distribution of NADPH-diaphorase cells in visual and somatosensory cortex in four mammalian species.

The distribution of the well-labeled nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd) Type I neurons was evaluated in the isocortex of four mammalian species: the Didelphis opossum, the Monodelphis opossum, the rat and the marmoset. In Didelphis opossum, laminar distribution was examined in tangential and non-tangential sections. The density increases from superficial to deep layers of the gray matter. In rats' tangential sections, infragranular and supragranular layers have higher density than layer IV. Cell density measurements in the visual and the somatosensory cortices were compared in tangential sections from flattened hemispheres of the four species. Somatosensory areas were identified histochemically in rat (barrel fields) and marmoset (S1 and S2/PV). In the opossums, areas S1 and S2/PV were identified by multiunit recording. Except in the rat, primary visual cortex (V1) was labeled histochemically by NADPHd and/or cytochrome oxidase. In the four species, cell density in somatosensory cortex was significantly higher than in visual cortex. Taken together these results demonstrate that NADPHd Type I neurons are not homogeneously distributed in the isocortex of these mammals. In conclusion, the tangential distribution of Type I neurons in the sensory areas examined, but not its laminar distribution, was similar in the four species. Given that rats, marmosets and opossums are distantly related species, and that the latter are considered to have more 'generalized' brains, it is conceivable that this pattern of tangential distribution of Type I neurons is a general feature of mammalian isocortex.

Cortical and subcortical influences on the nucleus of the optic tract of the opossum.

In the present work we propose a new phylogenetic hypothesis for the role played by cortical and subcortical afferents to the nucleus of the optical tract, the main visual relay station of the horizontal optokinetic reflex in mammals. The hypothesis is supported by anatomical and physiological data obtained in the South American opossum (Didelphis aurita) using the following experimental approaches: (i) single-unit recordings in the nucleus of the optic tract and simultaneous electrical stimulation of the contralateral nucleus of the optic tract; (ii) single-unit recordings in the nucleus of the optic tract and simultaneous electrical stimulation of the ipsilateral striate cortex; (iii) injection of cholera toxin subunit B into the striate cortex and subsequent immunohistochemical reaction to reveal the presence of the marker in the thalamus and mesencephalon; and (iv) single-unit recordings in the nucleus of the optic tract both before and after ablation of the ipsilateral visual cortex. The main results are: (i) there is a strong inhibitory reciprocal effect upon the nucleus of the optic tract following stimulation of its contralateral counterpart; (ii) electrophysiological and anatomical data imply that the visual cortex does not project directly to the nucleus of the optic tract. Rather, cortical terminals seem to target the nearby anterior and posterior pretectal nuclei and orthodromic latencies in the nucleus of the optic tract following stimulation of the visual cortex were twice as large as in the superior colicullus; and (iii) ablation of the entire visual cortex did not have any effect upon binocularity of cells in the nucleus of the optic tract. These results strengthen the model proposed here for the role of the interactions between the nuclei of the optic tract under optokinetic stimulation. The hypothesis in the present work is that the cortical influences upon the nucleus of the optical tract, in addition to the subcortical ones, appeared only recently in phylogenesis. In more primitive mammals, such as the opossum, subcortical interactions are thought to play a relatively important role. With the emergence of retinal specializations, such as the fovea, one might suppose that there followed the appearance of new ocular movements, such as the smooth pursuit and certain types of saccades, that came to join the pre-existent optokinetic reflex.

Patterns of corticocortical, corticotectal, and commissural connections in the opossum visual cortex.

Patterns of connections of the visual cortex of the South American opossum, Didelphis aurita, were revealed by using neuronal tracers to identify and characterize visual specializations of the peristriate cortex (PS). The visuotopy of corticotectal connections of the anterolateral portion of PS (PSal) is symmetrical to that of the striate cortex (ST or primary visual area [V1]). Three consecutive bands of commissural connections coincide, respectively, with the ST-PS border, the limit between the caudal and rostral PSal halves (PSc and PSr), and the border of PS with the parietal and temporal cortices. PSc and PSr contain regular commissural rings similar to those present in the peristriate cortex of eutherian mammals. ST projections define in PSc two strings of periodical foci consecutively concentric to V1 and a single focus in PSr. Although they were organized topographically, ascending, descending, and commissural connections between ST and PSal showed a high degree of convergence and divergence. These results conform to the model of a single area homologous to the second visual area (V2) bordering V1. Moreover, they suggest the possibility that PSal includes either one or two additional belt-like areas successively anterior to V2. Along with the finding of alternating bands of high and low cytochrome oxidase activity in PSal, the data further suggest that this region contains modular specializations similar to those of the peristriate cortex of primates and other eutherian mammals. The posterolateral peristriate cortex (PSpl) constitutes another visual area, since it consists of a distinct focus of reciprocal corticocortical and interhemispheric connections and a separate source of corticotectal projections. Finally, a visuomotor function for the orbital cortex is proposed based on its direct projections to optical tectal layers. The close cladistic relationship of opossums to mammalian ancestral forms suggests that the PSal parcelation into belt-like areas that contain modules reflects the primitive organization of the visual cortex. Moreover, a highly diffuse pattern of corticocortical connections may represent a requirement for a brain with few visual areas to perform global processing.

Cytochrome oxidase and NADPH-diaphorase on the afferent relay branch of the optokinetic reflex in the opossum.

In the present study, histochemical techniques combined with more conventional anatomical methods were used to refine the identification of the nucleus of the optic tract and the nuclei of the accessory optic system in the opossum. The distribution of the enzyme cytochrome oxidase (CO) was examined in the cells and the neuropil of the opossum's mesodiencephalic region. Strong CO labeling was present in the nucleus of the optic tract (NOT)-dorsal terminal nucleus (DTN). Alternate sections, taken from animals that had received bilateral injections of horseradish peroxidase centered in the region of the inferior olive, were subjected to assays for CO and horseradish peroxidase. The region occupied by CO-labeled cells in the NOT-DTN superimposed with the one defined by retrogradely labeled cells. Cell counts along the NOT-DTN anteroposterior axis revealed that although the olivary and CO-positive cells were confined within similar boundaries, the latter are up to twofold more numerous than the former. As revealed by cytochrome oxidase histochemistry, the outlines of the NOT-DTN, the other pretectal nuclei and the nuclei belonging to the accessory optic system coincided with those revealed by the histochemistry for nicotinamide dinucleotide phosphate diaphorase (NADPH-d). After an intraocular injection of cholera toxin beta subunit and alternate sections processing for NADPH-d and CO, the distribution of labeled retinal terminal fields in the mesodiencephalic region was shown to be coincident with regions of high levels of histochemical labeling. These results are discussed in the light of previous anatomofunctional assessments of the pretectum and accessory optic system.

The horizontal optokinetic reflex of the opossum (Didelphis marsupialis aurita): physiological and anatomical studies in normal and early monoenucleated specimens.

In the opossum the symmetrical binocular horizontal optokinetic nystagmus gives way to an asymmetrical monocular reflex: the nasotemporal (NT) stimulation yielding lower gain than the temporonasal (TN). In adults, monocularly enucleated at postnatal days 21-25 (pnd21-25), the gain of NT responses is markedly increased, approaching that of TN. Severe cell loss was detected in the nucleus of the optic tract (NOT) on the deafferented side in early monoenucleated specimens. In normal animals retinal afferents to the NOT are all crossed, while in animals enucleated at pnd21-25 sparse uncrossed retinal elements were observed. Although this abnormal projection might influence the increased NT response in this subgroup, it is argued that the increased symmetry in monoenucleated opossums may be the result of changes mediated by the commissural connection between both NOTs.

On the functional anatomy of the nucleus of the optic tract-dorsal terminal nucleus commissural connection in the opossum (Didelphis marsupialis aurita).

Immunocytochemical methods revealed the presence of GABA in cell bodies and terminals in the nucleus of the optic tract-dorsal terminal nucleus, the medial terminal nucleus, the lateral terminal nucleus and the interstitial nucleus of the superior fasciculus of the opossum (Didelphis marsupialis aurita). Moreover, after unilateral injections of rhodamine beads in the nucleus of the optic tract-dorsal terminal nucleus complex and processing for GABA, double-labelled cells were detected in the ipsilateral complex, up to 400 microns from the injected site, but not in the opposite. Analysis of the distributions of GABAergic and retrogradely-labelled cells throughout the contralateral nucleus of the optic tract-dorsal terminal nucleus showed that the highest density of GABAergic and rhodamine-labelled cells overlapped at the middle third of the complex. Previous electrophysiological data obtained in the opossum had suggested the existence, under certain conditions, of an inhibitory action between the nucleus of the optic tract-dorsal terminal nucleus of one side over the other. The absence of GABAergic commissural neurons may imply that this inhibition is mediated by an excitatory commissural pathway that activates GABAergic interneurons.

The nucleus of the optic tract of the opossum.

This paper reviews anatomical and electrophysiological data on the nucleus of the optic tract (NOT) of the opossum, a nucleus in the afferent branch of the horizontal optokinetic reflex. It is proposed that subcortical routes are essential for responses from the two eyes: a direct retinal projection from the contralateral eye and a commissural pathway between the two NOTs for the ipsolateral eye. In the latter case there's evidence that the commisural axons have a relay on inhibitory neurones. This circuit accounts for the differences in response pattern under monocular condition: temporo-nasal motion of the visual stimulus elicits excitation in the contralateral NOT, resulting in inhibition of the ipsolateral nucleus, while naso-temporal motion promotes inhibition in the contralateral nucleus, releasing the ipsolateral nucleus from the commissural input.

The nucleus of the optic tract (NOT) and the dorsal terminal nucleus (DTN) of opossums (Didelphis marsupialis aurita).

Wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) was injected unilaterally into the pretectocollicular region of opossums (Didelphis marsupialis aurita), primarily to investigate the existence of a commissural subcortical pathway but also to reveal afferents and efferents of the nucleus of the optic tract (NOT) and dorsal terminal nucleus (DTN) in this species. Labelled cells and terminals were observed in the contralateral NOT-DTN. Furthermore, HRP was injected bilaterally in the region of the inferior olive (IO) to verify if the distribution of labelled cells in the NOT-DTN overlapped the region of commissural labelled cells. The two subpopulations of retrogradely labelled cells coincided, being distributed within the retinal terminal field attributed to the NOT-DTN, as revealed by contralateral eye injections of HRP. The commissural cells were located slightly more ventral than the olivary cells in the optic tract. The pretectocollicular WGA-HRP injections also labelled cells and terminals bilaterally in the lateral terminal nucleus (LTN), interstitial nucleus of the superior fasciculus, posterior fibers (INSFp), ventral lateral geniculate nucleus (vLGN), and superior colliculus (SC) and ipsilaterally in the medial terminal nucleus (MTN). In addition, further caudally, labelled cells and terminals were observed bilaterally in the nuclei prepositus hypoglossi (PH) and in the medial (MVN) and lateral (LVN) vestibular nuclei. Labelled terminals were found in the ipsilateral nucleus reticularis tegmenti pontis (NRTP) and in the IO with ipsilateral predominance. This study allowed an anatomical delimitation of the NOT-DTN in this opossum species, as defined by the olivary and commissural subpopulations, as well as a hodological evaluation of this region. The existence of some common anatomical aspects with other mammalian species is discussed.

Binocularity in the nucleus of the optic tract of the opossum.

In the present work, we characterize electrophysiologically a commissural subcortical pathway which is related to binocular interactions in the nucleus of the optic tract (NOT) of the opossum. The main role played by the circuit comprising this pathway seems to be in relaying information coming from the ipsilateral eye to the NOT. The strongest evidence comes from experiments in which lidocaine was injected into the NOT and the ensuing effects in the opposite nucleus were monitored under ipsilateral monocular stimulation. It was consistently observed that during action of lidocaine the directional response normally elicited by stimulation of the ipsilateral eye did not take place in the NOT opposite to the silenced nucleus. This effect was reverted in a few minutes after recovery of the injected NOT. The response to stimulation of the contralateral eye, though, was not affected by this procedure.

The retinal distribution of ganglion cells with crossed and uncrossed projections and the visual field representation in the opossum.

The retinal distribution of ganglion cells with crossed and uncrossed projections in the South American opossum, Didelphis marsupialis, was revealed by delivering HRP to one optic tract or to retinal targets of one hemisphere. The cells with uncrossed projections are restricted to the temporal retina, comprising 1/3 of the total retinal area, with a sharp transition at the naso-temporal boundary. Besides being distributed over the nasal 2/3 of the retina, cells with crossed projections are intermingled with those with uncrossed projections over the entire temporal retina. Quantitative analysis about the representation of the horizontal meridian on four specimens revealed that the maximum density of cells with uncrossed projections is on the average located at 3.2 mm (SD = 0.21), i.e. 34.8 deg, temporal to the optic disk, falling to 10% at 2.1 mm (SD = 0.14) or 22.8 deg. On the other hand, the peak for cells with crossed projections is more nasally placed at 1.8 mm (SD = 0.18), i.e. 19.6 deg. Between these two maxima, the site wherein the densities of cells with crossed and uncrossed projections are about equal is on the average about 2.7 mm (SD = 0.25) form the optic disk, i.e. 29.3 deg. This estimate supports the hypothesis that the retinal intersection of the vertical meridian lies within the region of split representation of crossed and uncrossed ganglion cells. In addition, it was observed that the opossum's retina has a large contingent of cells with uncrossed projections temporal to an eccentricity of 2.7 mm from the optic disk, where it represents roughly 2/3 of the ganglion cells. These data corroborate the relevance of the opossum as a non-primate model for visual work.

Dynamic surrounds of receptive fields in primate striate cortex: a physiological basis for perceptual completion?

Visual receptive fields (RFs) were mapped inside and outside the cortical representation of the optic disk in the striate cortex (area V1) of anesthetized and paralyzed Cebus monkeys. Unexpectedly, most cells were found to be binocularly driven, and the RFs mapped with contralateral-eye stimulation progressed in a topographically appropriate fashion as the optic disk sector was crossed. Activation of these neurons by the contralateral eye was shown to depend on stimulation of the parts of the retina around the optic disk. Outside the optic disk representation, a similar effect was demonstrated by obstructing the "classical" RF with masks 5-10 times larger in size. In all cases, visual stimuli presented around the mask could be used to accurately interpolate the position of the hidden RF. These properties reflect, at a cellular level, the process of "filling in" that allows for completion of the visual image across natural and artificially induced scotomas.

Disparity selective units in the superior colliculus of the opossum.

Binocular visual responses can be recorded in two regions of the superficial layers of the superior colliculus of the opossum. The direct binocular region (DBR) represents the binocular portion of the contralateral hemifield whereas the rostral pole (RP) represents the binocular portion of the ipsilateral hemifield. In the present study single units from both of these regions were tested with binocular and monocular stimulation. Most cells in both regions showed response facilitation when both eyes were simultaneously stimulated and, when tested with different binocular disparities, most cells showed broadly-tuned disparity selectivity. DBR units usually preferred disparities near zero whereas RP units had a wider range of preferred disparities, with a tendency toward positive (crossed) values. This data indicates that the superior colliculus of the opossum could provide a neural substrate for a coarse analysis of depth and also might help control vergence eye movements. The different ranges of disparity selectivity of DBR and RP are consistent with the previously reported monocular receptive-field data and suggest that DBR and RP analyze different depths of the 3-dimensional visual scene.

Evidence for recrossing of the pathway from the retina to the ipsilateral nucleus of the optic tract.

Single-unit recordings of the nucleus of the optic tract (NOT) under visual stimulation were performed in 5 opossums. Most of the units were directionally selective. Receptive fields for the contralateral eye lie mainly in the contralateral field while those for the ipsilateral eye were mainly in the ipsilateral field. As the nasal retina does not project ipsilaterally, recrossing must occur in the pathway from the retina to the ipsilateral NOT. Possible sites for this recrossing are discussed.

Patterns of cytochrome oxidase activity in the visual cortex of a South American opossum (Didelphis marsupialis aurita).

The normal pattern of cytochrome oxidase (CO) activity in the posterior cortical areas of the South American opossum (Didelphis marsupialis aurita) was assessed both in horizontal sections of flattened cortices and in transversal cortical sections. The tangential distribution of CO activity was uniformly high in the striate cortex. In the peristriate region alternating bands of dense and weak staining occupied all the cortical layers with the exception of layer I. This observation suggests the existence of a functional segregation of visual processing in the peristriate cortex of the opossum similar to that present in phylogenetically more recent groups.

Visual response properties of pretectal units in the nucleus of the optic tract of the opossum.

Single-units were recorded from the nucleus of the optic tract. Most of the units showed excitation in response to random check patterns presented on a tangent screen to the contralateral eye, moving in a temporal to nasal direction and/or inhibition in the opposite direction. The excitatory response to the temporal to nasal movement, observed in most units, was unchanged throughout the range of speeds tested, except for a decrease at the slowest (0.6 deg/s) and fastest (150 deg/s) speeds. On the other hand, the inhibitory responses evoked by a nasal to temporal movement, had a peak between 3 and 16 deg/s which decreased towards both extremes. An average of 45% of the units were influenced by the stimulation of the ipsilateral eye. In one third of them the response was very weak. In the remainder, the mean frequency of spikes in one direction of the horizontal movement was more than twice that in the opposite stimulus direction. In the great majority of these units, stimulation of each eye yielded the same overall pattern of directionality, that is, movement of the stimulus towards the recording side led to excitation and/or movement in the reverse direction led to inhibition. Inhibition was stronger than excitation in most ipsilaterally responding units. Excitatory responses elicited by the ipsilateral eye were always weaker than those by the contralateral but in a few cases the ipsilateral inhibitory component was more prominent than the contralateral one.

Horizontal optokinetic reflex in the opossum Didelphis marsupialis aurita.

Electro-oculographic recordings were performed in 10 opossums. The optokinetic reflex was elicited by projecting a random dot stimulus on a cylindrical screen moving horizontally from left to right or right to left at various constant speeds. Binocular stimulation yielded the same response as the temporal to nasal monocular condition. The nasal to temporal monocular response was always less than that to the opposite direction: 50% at 3 degrees/s and 15% at 18 degrees/s. These results are discussed in a comparative context.

The ipsilateral field representation in the striate cortex of the opossum.

Reference axes for the visuotopic study of the opossum's striate cortex were estimated from corresponding binocular response fields using multi-unit recording. These central binocular axes (CBA) were derived from experimental data based on the concept that corresponding receptive fields for each eye should be mostly in register under natural conditions. Vertical reference meridians, orthogonal to these axes, define a contralateral and an ipsilateral field for each eye with respect to the recording site. An ipsilateral field representation was observed for both eyes in the striate cortex at the transition zone with peristriate. Maximal values for the center and border of ipsilateral receptive fields were, respectively, 8 and 20 degrees for the contralateral eye and 6 and 14 degrees for the ipsilateral eye. An equivalent ipsilateral field representation was found in animals that had the anterior commissure cut prior to the recording session. This suggests that the ipsilateral field of both eyes may be represented in the striate cortex via the ipsilateral optic tract. Additionally, it was observed that the region of higher ganglion cell density in the retina shows a flattened distribution and that the CBA intersects the retina at the temporal aspect of this region.

Organization of the visual thalamus: corticothalamic projections from the primary visual area in the opossum.

The projections from the visual cortex to thalamic nuclei in the opossum were investigated by degeneration and radioautographic methods. Efferent axons from the striate cortex course from the internal capsule to the thalamus in two bundles, one of which innervates the reticular and dorsal lateral geniculate nuclei, while the other bundle innervates the external layer of the ventral lateral geniculate nucleus. Both bundles course along and through terminal fields found in the lateral posterior nucleus. The visuotopic organization inferred from the corticofugal pathway shows that projection lines in the dorsal lateral geniculate nucleus (DLGN) run along medio-lateral strips including both the beta and alpha segments, across the quasi-laminae of retino-geniculate projection, and that naso-temporal axes in the visual field run in a dorso-ventral direction through the DLGN. The external layer of the ventral lateral geniculate nucleus receives a projection from the striate cortex and bilateral projections from the retina, and naso-temporal axes in the visual field are represented along a dorso-ventral direction in this layer. The striate cortical projections cover approximately the lateral two-thirds of the lateral posterior nucleus, overlapping a small retinal terminal field, and naso-temporal axes in the visual field are represented onto the cortico-recipient zone in a mirror-symmetric direction to that of the adjoining DLGN. In all these three cortico-recipient zones, dorso-ventral axes in the visual field are represented in a rostro-caudal direction.