No abstract

Teleost sex change is an important model to understand general principles of sexual differentiation and plasticity in the adult brain. The present study is the first to examine the proliferation zones in the adult brain of males, females and sex-changing individuals of a protandrous teleost species (Sparus aurata), by means of 5-bromo-2-deoxyuridine immunocytochemistry. Postnatal neurogenesis in the marine teleost brain was found in ventricular and subventricular areas of the brain that in most cases coincided with the embryonic proliferation zones. The molecular layer of corpus and valvula cerebelli exhibited the highest mitotic activity in the adult brain. High mitotic activity was observed in the hypothalamic, thalamic and telencephalic ventricular areas, as well as the dorsal and ventral rim of the optic tectum. Most of the labeled cells were elongated, indicating the initiation of migratory activity. There were no qualitative differences in the distribution of proliferation zones between the sex phases studied with the exception of the ventricular region of the dorsal hypothalamic area. Volume fraction analysis of the area occupied by the labeled cells suggested that this region included higher densities of newborn cells in the female animals. The proliferation pattern in the adult gilthead sea bream brain is in agreement with the hypothesis of the continuous generation of new cells in the teleost brain. Moreover, our data propose that cell proliferation differences possibly existing in the ventricular region of the dorsal hypothalamus between sexual phases, might be involved in central mechanisms of sexual plasticity in protandrous hermaphrodite teleosts.

It has been reported that neurons generated in the adult brain show sex-specific differences in several brain regions of lower vertebrates and mammals. The present study questioned whether cell proliferation and survival in the adult zebrafish (Danio rerio) cerebellum, the most mitotically active area of adult teleost brain, is sexually differentiated. Adult zebrafish were treated with the thymidine analogue 5'-bromo-2'-deoxyuridine (BrdU) and allowed to survive for 24 h (short-term) and for 21 days (long-term). BrdU immunohistochemistry allowed visualization of cells incorporating BrdU at the S phase of mitosis. At short-term survival, male zebrafish had a higher number of labelled cells at proliferation sites of the molecular layer of corpus cerebelli (CCe) and the granular layer of the caudal lobe of the cerebellum (LCa) than did females. In long-term survival, BrdU-positive cells were found at their final destination, but only the granular layer of the medial division of the valvula cerebelli showed sex-specific differences in the number of labelled cells. This higher mitotic activity in male cerebellum might be related to sex-specific motor behaviour observed in male zebrafish. To investigate the role of programmed cell death, the terminal deoxynucleotidyl-mediated dUTP nick-end-labelling (TUNEL) method was applied. The vast majority of apoptotic figures occurred in the granular cell layer of valvula and CCe, only in a few cases within the BrdU-retaining cells. Apoptosis was found specifically at the sites of the final destination of proliferating cells, indicating that the close relation of cell birth and death might represent a possible plasticity mechanism in the adult zebrafish cerebellum.

The post-injury responses of retinal ganglion cells elicit a number of glial reactions which have not been completely understood. The bilateral pattern of non-neuronal retinal cell proliferation was examined in association with the differential fates of unilaterally injured adult retinal ganglion cells by means of bromodeoxyuridine (BrdU) immunocytochemistry. Lateralization of the glioproliferative events was studied by analysing both the experimental and the uninjured contralateral as well as matched retinas of sham-operated animals. Control adult rat retina included very few BrdU-positive cells within the nerve fibre and ganglion cell layers; however, experimental retinas of degenerating groups exhibited statistically significantly higher densities of newborn cells in most layers. Clusters of labelled cells were found in the inner plexiform layer related to OX-42 staining, indicating their microglial nature. Indeed, double-labelling experiments, after short-term unilateral optic nerve crushing, identified proliferating retinal glial cells in vivo. Both types of glia, astroglial and microglial cells, exhibited BrdU-positive labelling in injured as well as uninjured experimental rat retinas. Moreover, microglial proliferating cells were also identified in explanted retinal pieces after 2 days in culture. Affected and contralateral retinas responded similarly to the unilateral experimental manipulations applied with respect to BrdU labelling. The acute glial responses observed suggest that bilateral glial proliferation might represent a common response related to degeneration events in both retinas, i.e. ipsi- and contralateral to the experimental injury.

The present study aimed to determine the anatomic distribution and developmental profile of alpha(2) and beta adrenoceptors (AR) in marine teleost brain. Alpha 2 and beta adrenoceptors were studied at different developmental stages by using [(3)H]clonidine and [(3)H]dihydroalprenolol, respectively, by means of in vitro quantitative autoradiography. Furthermore, immunohistochemical localization of the receptor subtypes was performed to determine their cellular distribution. Saturation studies determined a high-affinity component of [(3)H]clonidine and [(3)H]dihydroalprenolol binding sites. High levels of both receptors were found in preglomerular complex, ventral hypothalamus, and lateral torus. Dorsal hypothalamus and isthmus included high levels of alpha(2) AR, whereas pretectum and molecular and proliferative zone of cerebellum were specifically characterized by high densities of beta AR. From the first year of life, adult levels of both AR were found in most medial telencephalic, hypothalamic, and posterior tegmental areas. Decreases in both receptors densities with age were prominent in ventral and posterior telencephalic, pretectal, ventral thalamic, hypothalamic, and tegmental brain regions. Immunohistochemical data were well correlated with autoradiography and demonstrated the presence of alpha(2A), alpha(2C), beta(1), and beta(2) AR subtype-like immunoreactivity. Both the neuronal (perikaryal or dendritic) and the glial localization of receptors was revealed. The localization and age-dependent alterations in alpha(2) and beta AR were parallel to plasticity mechanisms, such as cell proliferation in periventricular thalamus, hypothalamus, and cerebellum. In addition, the biochemical characteristics, distribution pattern, and neuronal or glial specificity of the receptors in teleost brain support a similar profile of noradrenergic transmission in vertebrate brain evolution.

Early life stress (ELS) shapes the way individuals cope with future situations. Animals use cognitive flexibility to cope with their ever-changing environment and this is mainly processed in forebrain areas. We investigated the performance of juvenile gilthead seabream, previously subjected to an ELS regime. ELS fish showed overall higher brain catecholaminergic (CA) signalling and lower brain derived neurotrophic factor (bdnf) and higher cfos expression in region-specific areas. All fish showed a normal cortisol and serotonergic response to acute stress. Brain dopaminergic activity and the expression of the α2Α adrenergic receptor were overall higher in the fish homologue to the lateral septum (Vv), suggesting that the Vv is important in CA system regulation. Interestingly, ELS prevented post-acute stress downregulation of the α2Α receptor in the amygdala homologue (Dm3). There was a lack of post-stress response in the β2 adrenergic receptor expression and a downregulation in bdnf in the Dm3 of ELS fish, which together indicate an allostatic overload in their stress coping ability. ELS fish showed higher neuronal activity (cfos) post-acute stress in the hippocampus homologue (Dlv) and the Dm3. Our results show clear long-term effects on limbic systems of seabream that may compromise their future coping ability to environmental challenges.

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The alpha(2A)-adrenoceptor (AR) subtype, a G protein-coupled receptor located both pre- and postsynaptically, mediates adrenaline/noradrenaline functions. The present study aimed to determine the alpha(2A)-AR distribution in the adult zebrafish (Danio rerio) brain by means of immunocytochemistry. Detailed mapping showed labeling of alpha(2A)-ARs, in neuropil, neuronal somata and fibers, glial processes, and blood vessels. A high density of alpha(2A)-AR immunoreactivity was found in the ventral telencephalic area, preoptic, pretectal, hypothalamic areas, torus semicircularis, oculomotor nucleus (NIII), locus coreruleus (LC), medial raphe, medial octavolateralis nucleus (MON), magnocellular octaval nucleus (MaON), reticular formation (SRF, IMRF, IRF), rhombencephalic nerves and roots (DV, V, VII, VIII, X), and cerebellar Purkinje cell layer. Moderate levels of alpha(2A)-ARs were observed in the medial and central zone nuclei of dorsal telencephalic area, in the periventricular gray zone of optic tectum, in the dorsomedial part of optic tectum layers, and in the molecular and granular layers of all cerebellum subdivisions. Glial processes were found to express alpha(2A)-ARs in rhombencephalon, intermingled with neuronal fibers. Medium-sized neurons were labeled in telencephalic, diencephalic, and mesencephlic areas, whereas densely labeled large neurons were found in rhombencephalon, locus coeruleus, reticular formation, oculomotor area, medial octavolateralis and magnocellular octaval nuclei, and Purkinje cell somata. The functional role of alpha(2A)-ARs on neurons and glial processes is not known at present; however, their strong relation to the ventricular system, somatosensory nuclei, and nerves supports a possible regulatory role of alpha(2A)-ARs in autonomic functions, nerve output, and sensory integration in adult zebrafish brain.