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.

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.

Growth and restriction of the ipsilateral retinocollicular projection in the opossum.

The distribution of optic nerve fibers and terminals in the superior colliculus (SC) was followed throughout its development in pouch young opossums in order to establish the normal sequence of events leading to the formation of mature patterns. Up to 7 days of life in the pouch, labeled fibers can be followed only as far as the rostral aspect of the optic tract. The earliest evidence for crossed retinal projections in the SC is found at 10 days of age. In parasagittal sections, the label extends along the rostrocaudal tectal axis from the rostral border to the presumptive caudal pole of the SC. Unequivocal evidence for ipsilateral retinocollicular projection is found at 15 days extending to all but the caudal 5th of the rostrocaudal extent of the SC. The projections from both eyes overlap extensively in the SC at 22 days and after this age significant changes occur, mostly at the ipsilateral side: a sub-pial tier of fine label develops excluding both rostral and caudal collicular poles; a deeper tier of coarse label extends from the rostral to the caudal pole and a third, patchy tier of label is found at the prospective strata griseum superficiale and griseum intermediate. By 47 and 60 days the tangential distribution of the projections is virtually indistinguishable from the adult pattern although laminar segregation does not seem as sharp as in the adult. Comparisons of the changeable patterns of ipsilateral retinocollicular projections from 22 to 34 days with the invariant, aberrant pattern in adult animals submitted to uniocular enucleation at either age suggests that the preservation of a juvenile pattern does not provide a comprehensive explanation for the formation of aberrant projections.

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.

Genesis of neurons of the retinal ganglion cell layer in the opossum.

In this study, we have examined the genesis of neurons of the retinal ganglion cell layer of the opossum Didelphis marsupialis by [3H]-thymidine autoradiography. Our results suggest that most neurons surviving to adulthood are generated in postnatal life from day 1 to day 23. Cells are generated according to a coarse gradient from the retinal geometric center to the periphery. Regional analysis of soma size distributions in different cohorts suggest that this gradient is actually formed by two partially-overlapping, concentric waves of cell proliferation. Most medium and large ganglion cells are formed during the early wave, whereas most displaced amacrine cells and small ganglion cells are formed during the late wave. Our results confirm the appropriateness of the opossum as a model for studies of development of the mammalian visual system.

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.

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.

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.

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.

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.

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.

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.

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.

Horizontal optokinetic reflex in the opussum 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 yelded the same response as the temporal to nasal monocular condition. The nasal to temporal monocular response was always less than to the opposite direction: 50% at 3 degrees/s and 15% at 18 degress/s. These results are discussed in a comparative context

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

Single-unit recordings of the nucleous 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 does not project ipsilaterally, recrossing must occur in the pathway from the retina to the ipsilateral NOT. Possible sites for this recrossing are discussed