In this study and the accompanying article (Folgueira et al., 2004a), the fluorescent carbocyanine dye 1,1'-dioctadecyl 3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) was used in fixed tissue to comprehensively analyze the connections of the different regions of the telencephalic lobes and the preoptic region of the rainbow trout. Here, we analyze the connections of the dorsal area (D; pallium) of the telencephalon, and the preoptic region, as well as the telencephalic connections of several structures in the diencephalon and brainstem of juvenile trout. The dorsal plus dorsolateral pallial zone of D (Dd+Dl-d) receives afferents from contralateral Dd+Dl-d, the ventral area of the telencephalon, preoptic nucleus, suprachiasmatic nucleus, medial thalamus, preglomerular complex, anterior and lateral tuberal nuclei, posterior tuberal nucleus, posterior hypothalamic lobe, superior raphe nucleus, and the rhombencephalic central gray and reticular formation, and projects to the central zone of D (Dc), medial thalamus, and some caudomedial hypothalamic regions. The medial zone of D (Dm) maintains reciprocal connections with the preglomerular complex and also receives afferents from the preoptic nucleus, suprachiasmatic nucleus, anterior tuberal nucleus, preglomerular tertiary gustatory nucleus, posterior tubercle, superior raphe nucleus, locus coeruleus, and the rhombencephalic central gray, and reticular formation. Dc receives fibers mainly from Dd+Dl-d, preoptic nucleus, preglomerular complex, and torus semicircularis and projects to several extratelencephalic centers, including the paracommissural nucleus, optic tectum, torus semicircularis, thalamus, preglomerular complex, posterior tubercle nuclei, and inferior hypothalamic lobes. The posterior zone of D (Dp) is mainly connected with the olfactory bulbs, the ventral and supracommissural nuclei of the ventral area (subpallium), the preoptic nucleus, and the preglomerular complex and projects to wide hypothalamic and posterior tubercular regions. The preoptic nucleus projects to the olfactory bulb, to most regions of the telencephalic lobes, and to several diencephalic and brainstem structures. These results reveal complex and specialized connectional patterns in the rainbow trout dorsal telencephalon and preoptic region. Most of these connections have not been described previously in salmonids. These connections indicate that the salmonid telencephalon is involved in multisensorial processing and modulation of brain activity.

The habenulae are bilateral nuclei located in the dorsal diencephalon that are conserved across vertebrates. Here we describe the main afferents to the habenulae in larval and adult zebrafish. We observe afferents from the subpallium, nucleus rostrolateralis, posterior tuberculum, posterior hypothalamic lobe, median raphe; we also see asymmetric afferents from olfactory bulb to the right habenula, and from the parapineal to the left habenula. In addition, we find afferents from a ventrolateral telencephalic nucleus that neurochemical and hodological data identify as the ventral entopeduncular nucleus (vENT), confirming and extending observations of Amo et al. (2014). Fate map and marker studies suggest that vENT originates from the diencephalic prethalamic eminence and extends into the lateral telencephalon from 48 to 120 hour post-fertilization (hpf). No afferents to the habenula were observed from the dorsal entopeduncular nucleus (dENT). Consequently, we confirm that the vENT (and not the dENT) should be considered as the entopeduncular nucleus “proper” in zebrafish. Furthermore, comparison with data in other vertebrates suggests that the vENT is a conserved basal ganglia nucleus, being homologous to the entopeduncular nucleus of mammals (internal segment of the globus pallidus of primates) by both embryonic origin and projections, as previously suggested by Amo et al. (2014).

BackgroundAlthough the mechanisms underlying brain patterning and regionalization are very much conserved, the morphology of different brain regions is extraordinarily variable across vertebrate phylogeny. This is especially manifest in the telencephalon, where the most dramatic variation is seen between ray-finned fish, which have an everted telencephalon, and all other vertebrates, which have an evaginated telencephalon. The mechanisms that generate these distinct morphologies are not well understood.ResultsHere we study the morphogenesis of the zebrafish telencephalon from 12 hours post fertilization (hpf) to 5 days post fertilization (dpf) by analyzing forebrain ventricle formation, evolving patterns of gene and transgene expression, neuronal organization, and fate mapping. Our results highlight two key events in telencephalon morphogenesis. First, the formation of a deep ventricular recess between telencephalon and diencephalon, the anterior intraencephalic sulcus (AIS), effectively creates a posterior ventricular wall to the telencephalic lobes. This process displaces the most posterior neuroepithelial territory of the telencephalon laterally. Second, as telencephalic growth and neurogenesis proceed between days 2 and 5 of development, the pallial region of the posterior ventricular wall of the telencephalon bulges into the dorsal aspect of the AIS. This brings the ventricular zone (VZ) into close apposition with the roof of the AIS to generate a narrow ventricular space and the thin tela choroidea (tc). As the pallial VZ expands, the tc also expands over the upper surface of the telencephalon. During this period, the major axis of growth and extension of the pallial VZ is along the anteroposterior axis. This second step effectively generates an everted telencephalon by 5 dpf.ConclusionOur description of telencephalic morphogenesis challenges the conventional model that eversion is simply due to a laterally directed outfolding of the telencephalic neuroepithelium. This may have significant bearing on understanding the eventual organization of the adult fish telencephalon.