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Based on the idea of differentiation-related changes in the glycosylation pattern of neurons, the expression of two cell surface oligosaccharide epitopes, N-acetyl-lactosamine (NALA), and its sulpho-glucuronyl derivative (HNK-1), was studied, by immunohistochemistry and Western blot experiments, in the developing chick retina beginning on day 2 of incubation (E2) until day 18 post-hatching. NALA was detectable on neuroepithelial cells as soon as the primary optic vesicles formed, and this pattern continued until E3. During subsequent retinal development NALA expression became progressively restricted in concert with the appearance of postmitotic neurons as revealed by neurite outgrowth, and with the formation of synaptic contacts until it disappeared at the end of the incubation period. The pattern of NALA expression was the inverse of HNK-1 which was detected for the first time at E3 on postmitotic ganglion cells accumulating at the vitreal surface. The number of HNK-1+ cells steadily increased until around E10, when the entire neural epithelium was labelled. Synchronously to synaptogenesis, most neurons lost their HNK-1 immunoreactivity. At the time of hatching the adult-like pattern was found, characterised by subpopulations of labelled horizontal, bipolar, amacrine, and ganglion cells. Immunoblot experiments demonstrated transient NALA glycosylation of protein bands, partially identical in their apparent molecular weight to those proteins with HNK-1 glycosylation. The observed temporospatial changes in the glycosylation patterns of distinct proteins during retinal development suggest NALA as a suitable marker for neuronal proliferation, and HNK-1 for differentiation and establishment of final synaptic configuration.

The distribution of N-acetyl-lactosamine (NALA), a cell-surface carbohydrate epitope of the lactoseries, has been studied in the retina of representative species of all vertebrate classes by light microscope immunohistochemistry. In only some species of different classes (fish, amphibia and mammals) was NALA expression detected, and in these animals the distribution showed profound interspecies variability. In fishes and amphibia in which NALA was present, patterns ranged from single immunopositive cells to homogeneous labelling of cell layers. In mammals, NALA was found only in retinas that are cone dominated (tree squirrel and primates). In the tree squirrel, there was a dense cellular staining of the photoreceptor cell layer; whereas in primates, the carbohydrate epitope occurred only on some photoreceptor cells. From these receptor cells, positive axons could be traced to the inner plexiform layer. In spite of the profound interspecies differences, NALA is not randomly expressed, as its exclusive expression in mammals with cone-dominated vision indicates. The suggestion of a functional relevance for NALA glycosylation of retinal cells is supported by the labelling pattern for HNK-1 in these species, which was different from the pattern found in rod-dominated mammalian retinas.

The distribution of the carbohydrate epitope CD15, a putative cell adhesion molecule, was studied in adult vertebrate retinas by light-microscopic immunohistochemistry. Except for Old World primates, in which no immunoreactivity was detectable, all other species expressed the epitope on retinal interneurones. Subpopulations of stratified amacrine cells were found in all species with the exception of bats and marmoset monkeys, and bipolar cells were immunoreactive in frogs and all amniotic animals. Ganglion cells were labelled in urodelian, in all sauromorphian, as well as in some mammalian species. In some species, the distribution of immunoreactive neurones was correlated to areas of retinal specialization such as the visual streak in frogs and the dorsotemporal field in birds. In these parts of the retina with enhanced visual acuity, more CD15 glycosylated bipolar cells were found than in other parts. Among mammals, labelled bipolar cells were found exclusively in species with cone-dominated retinas. This comparative study shows that CD15 expression is consistently membrane associated in morphologically defined subsets of amacrine, bipolar, and ganglion cells throughout the vertebrate phylum. Its distribution pattern was found to depend more on the visual behavior of a given species (cone-dominated or rod-dominated retina) than on phylogenetic proximity between species.

The spatiotemporal distribution pattern of the carbohydrate epitope CD15 was examined in the developing chick retina. CD15 expression appeared for the first time at E13 in the INL and GCL. The developmental profile of the INL, from E13 to E16, showed increasing numbers of stratified amacrine cells, whereas diffuse amacrine subtypes appeared later, beginning at E15. Smaller populations of bipolar cells were seen at E17. Three types of CD15-positive ganglion cells could be differentiated by E15. A gradient in the appearance of identified immunoreactive amacrine cells extended from the dorsotemporal to the ventral and nasal retina. The adult-like pattern of CD15 expression did not become established until E19. Adult-like densities of immunoreactive cells were reached toward the end of the embryonic period between E18 in the dorsotemporal and ventral retina, and E19 in the dorsonasal retina. In the adult-like retina, labelled cells became particularly numerous at its greatest circumference, and were most densely packed in the dorsotemporal retinal quadrant. From E16 to P5, three membrane-bound, CD15-positive glycoproteins of 20, 32 and 34 kDa were identified by Western blots. The time course in the appearance of the membrane-associated CD15 recognition molecule on differentiating amacrine, bipolar and ganglion cells is correlated to the establishment of synaptic contacts.

Light microscopic autoradiography performed subsequent to intraocular injection of 3H-leucine revealed silver grains (SG) above axons of the optic tract which could be followed into the ventral and caudal portion of the suprachiasmatic nuclei (SCN) and above the contralateral anterior hypothalamic nucleus (AHN). By high resolution photometric measurement and computer processing the labelled areas were analysed, thus yielding statistical data of the relative grain distribution. The highest SG density was found in the ventrolateral part of both SCN (SCvl), confirming earlier reports concerning retinohypothalamic connections. That area exhibiting a cytoarchitecture different from the remaining nucleus was traversed, however, by numerous labelled axons. In the caudal part of both SCN a specific projection field of retinal fibres could be located. Here, almost no traversing fibres contribute to the rather circumscribed marked area. In the ventral part of the contralateral AHN, diffuse labelling well above background levels could be observed. Distinction between bypassing and terminating fibres within the SCvl could not be made using light microscopy. Analysis of SG distribution of the SCvl with electron microscopic autoradiography revealed a specific localization of SG within presynaptic terminals containing clear vesicles and pale mitochondria.

Intraocular injections of tritiated leucine were performed in normal and in unilaterally enucleated adult albino rats. Light microscopical evaluation of the autoradiograms revealed evidence for retinal afferents to distribute within a rather circumscribed area of the lateral hypothalamus. The silver grains were consistently, however, observed only in the group of enucleated animals, indicating differential transport under basic and experimental conditions.

The introduction of new techniques in neuroanatomy has helped to clarify a number of important issues on the nature of the accessory optic and retino-hypothalamic system. In this study, an effort was made, to collect all data obtainable, and to arrange them on the basis of some common parameter, so that the various results may be directly compared. These data were then interpreted on specific issues, although only limited coverage has been given to functional aspects, as described in older publications. The absence of a universally accepted concept of the function of both systems to date suggests the need for a thorough analysis of both contemporary and older literature dealing with a characterization of the structural components of the accessory optic and the retino-hypothalamic systems. On the basis of these anatomical studies it is concluded that the accessory optic system (AOS) is much more complex than formerly believed. Its development in various vertebrae classes is quite different. Among the mammals, the AOS is very well established in rodents but in primates and man its anatomical and functional role seems markedly reduced. In the rodents it consists of both superficially and deeply arranged axons which are in close topical relation to the cerebral peduncles. The axons not only supply the classic accessory otpic terminal nuclei but also the subthalamic nucleus and the substantia nigra. The secondary projections have not yet been established. The retino-hypothalamic system (RHS) mainly but not exclusively supplies the suprachismatic nucleus (sc). Crossed as well as uncrossed retinal fibres project to the ventro-lateral and to the posterior part of this nucleus. The cells within the sc combine both secretory and neuronal properties. The possible functions exerted are discussed. Both, the AOS and the RHS, are so closely related within the suprachiasmatic area that most methods fail in correlating experimental results with either system. The numerous problems which still remain to be clarified are also discussed.