Hardwiring of fine synaptic layers in the zebrafish visual pathway

<p>Abstract</p> <p>Background</p> <p>Neuronal connections are often arranged in layers, which are divided into sublaminae harboring synapses with similar response properties. It is still debated how fine-grained synaptic layering is established during development. Here we investigated two stratified areas of the zebrafish visual pathway, the inner plexiform layer (IPL) of the retina and the neuropil of the optic tectum, and determined if activity is required for their organization.</p> <p>Results</p> <p>The IPL of 5-day-old zebrafish larvae is composed of at least nine sublaminae, comprising the connections between different types of amacrine, bipolar, and ganglion cells (ACs, BCs, GCs). These sublaminae were distinguished by their expression of cell type-specific transgenic fluorescent reporters and immunohistochemical markers, including protein kinase Cβ (PKC), parvalbumin (Parv), zrf3, and choline acetyltransferase (ChAT). In the tectum, four retinal input layers abut a laminated array of neurites of tectal cells, which differentially express PKC and Parv. We investigated whether these patterns were affected by experimental disruptions of retinal activity in developing fish. Neither elimination of light inputs by dark rearing, nor a D, L-amino-phosphono-butyrate-induced reduction in the retinal response to light onset (but not offset) altered IPL or tectal lamination. Moreover, thorough elimination of chemical synaptic transmission with <it>Botulinum </it>toxin B left laminar synaptic arrays intact.</p> <p>Conclusion</p> <p>Our results call into question a role for activity-dependent mechanisms – instructive light signals, balanced <it>on </it>and <it>off </it>BC activity, Hebbian plasticity, or a permissive role for synaptic transmission – in the synaptic stratification we examined. We propose that genetically encoded cues are sufficient to target groups of neurites to synaptic layers in this vertebrate visual system.</p

Focusing on optic tectum circuitry through the lens of genetics

The visual pathway is tasked with processing incoming signals from the retina and converting this information into adaptive behavior. Recent studies of the larval zebrafish tectum have begun to clarify how the 'micro-circuitry' of this highly organized midbrain structure filters visual input, which arrives in the superficial layers and directs motor output through efferent projections from its deep layers. The new emphasis has been on the specific function of neuronal cell types, which can now be reproducibly labeled, imaged and manipulated using genetic and optical techniques. Here, we discuss recent advances and emerging experimental approaches for studying tectal circuits as models for visual processing and sensorimotor transformation by the vertebrate brain

Neuroendocrine transcriptional programs adapt dynamically to the supply and demand for neuropeptides as revealed in NSF mutant zebrafish

<p>Abstract</p> <p>Background</p> <p>Regulated secretion of specialized neuropeptides in the vertebrate neuroendocrine system is critical for ensuring physiological homeostasis. Expression of these cell-specific peptide markers in the differentiating hypothalamus commences prior to birth, often predating the physiological demand for secreted neuropeptides. The conserved function and spatial expression of hypothalamic peptides in vertebrates prompted us to search for critical neuroendocrine genes in newly hatched zebrafish larvae.</p> <p>Results</p> <p>We screened mutant 5 days post-fertilization zebrafish larvae that fail to undergo visually mediated background adaptation for disruption in hypothalamic <it>pomc </it>expression. To our surprise, the ATPase <it>N-ethylmaleimide sensitive factor </it>(<it>nsf</it>) was identified as an essential gene for maintenance of neuroendocrine transcriptional programs during the embryo-to-larva transition. Despite normal hypothalamic development in <it>nsf</it>^{<it>st</it>53}mutants, neuropeptidergic cells exhibited a dramatic loss of cell-specific markers by 5 days post-fertilization that is accompanied by elevated intracellular neuropeptide protein. Consistent with the role of NSF in vesicle-membrane fusion events and intracellular trafficking, cytoplasmic endoplasmic reticulum-like membranes accumulate in <it>nsf</it>^-/-hypothalamic neurons similar to that observed for <it>SEC18 </it>(<it>nsf ortholog</it>) yeast mutants. Our data support a model in which unspent neuropeptide cargo feedbacks to extinguish transcription in neuropeptidergic cells just as they become functionally required. In support of this model we found that <it>gnrh3 </it>transcripts remained unchanged in pre-migratory, non-functional gonadotropin-releasing hormone (GnRH) neurons in <it>nsf</it>^-/-zebrafish. Furthermore, <it>oxytocin-like </it>(<it>oxtl</it>, <it>intp</it>) transcripts, which are found in osmoreceptive neurons and persist in mutant zebrafish, drop precipitously after mutant zebrafish are acutely challenged with high salt.</p> <p>Conclusion</p> <p>Our analyses of <it>nsf </it>mutant zebrafish reveal an unexpected role for NSF in hypothalamic development, with mutant 5 days post-fertilization larvae exhibiting a stage-dependent loss of neuroendocrine transcripts and a corresponding accumulation of neuropeptides in the soma. Based on our collective findings, we speculate that neuroendocrine transcriptional programs adapt dynamically to both the supply and demand for neuropeptides to ensure adequate homeostatic responses.</p

Forward Genetic Analysis of Visual Behavior in Zebrafish

The visual system converts the distribution and wavelengths of photons entering the eye into patterns of neuronal activity, which then drive motor and endocrine behavioral responses. The gene products important for visual processing by a living and behaving vertebrate animal have not been identified in an unbiased fashion. Likewise, the genes that affect development of the nervous system to shape visual function later in life are largely unknown. Here we have set out to close this gap in our understanding by using a forward genetic approach in zebrafish. Moving stimuli evoke two innate reflexes in zebrafish larvae, the optomotor and the optokinetic response, providing two rapid and quantitative tests to assess visual function in wild-type (WT) and mutant animals. These behavioral assays were used in a high-throughput screen, encompassing over half a million fish. In almost 2,000 F2 families mutagenized with ethylnitrosourea, we discovered 53 recessive mutations in 41 genes. These new mutations have generated a broad spectrum of phenotypes, which vary in specificity and severity, but can be placed into only a handful of classes. Developmental phenotypes include complete absence or abnormal morphogenesis of photoreceptors, and deficits in ganglion cell differentiation or axon targeting. Other mutations evidently leave neuronal circuits intact, but disrupt phototransduction, light adaptation, or behavior-specific responses. Almost all of the mutants are morphologically indistinguishable from WT, and many survive to adulthood. Genetic linkage mapping and initial molecular analyses show that our approach was effective in identifying genes with functions specific to the visual system. This collection of zebrafish behavioral mutants provides a novel resource for the study of normal vision and its genetic disorders

Level of agreement between frequently used cardiovascular risk calculators in people living with HIV

Objectives The aim of the study was to describe agreement between the QRISK2, Framingham and Data Collection on Adverse Events of Anti‐HIV Drugs (D:A:D) cardiovascular disease (CVD) risk calculators in a large UK study of people living with HIV (PLWH). Methods PLWH enrolled in the Pharmacokinetic and Clinical Observations in People over Fifty (POPPY) study without a prior CVD event were included in this study. QRISK2, Framingham CVD and the full and reduced D:A:D CVD scores were calculated; participants were stratified into ‘low’ ( 20%) categories for each. Agreement between scores was assessed using weighted kappas and Bland–Altman plots. Results The 730 included participants were predominantly male (636; 87.1%) and of white ethnicity (645; 88.5%), with a median age of 53 [interquartile range (IQR) 49–59] years. The median calculated 10‐year CVD risk was 11.9% (IQR 6.8–18.4%), 8.9% (IQR 4.6–15.0%), 8.5% (IQR 4.8–14.6%) and 6.9% (IQR 4.1–11.1%) when using the Framingham, QRISK2, and full and reduced D:A:D scores, respectively. Agreement between the different scores was generally moderate, with the highest level of agreement being between the Framingham and QRISK2 scores (weighted kappa = 0.65) but with most other kappa coefficients in the 0.50–0.60 range. Conclusions Estimates of predicted 10‐year CVD risk obtained with commonly used CVD risk prediction tools demonstrate, in general, only moderate agreement among PLWH in the UK. While further validation with clinical endpoints is required, our findings suggest that care should be taken when interpreting any score alone

Validation of a novel multivariate method of defining HIV-associated cognitive impairment

Background. The optimum method of defining cognitive impairment in virally suppressed people living with HIV is unknown. We evaluated the relationships between cognitive impairment, including using a novel multivariate method (NMM), patient– reported outcome measures (PROMs), and neuroimaging markers of brain structure across 3 cohorts. Methods. Differences in the prevalence of cognitive impairment, PROMs, and neuroimaging data from the COBRA, CHARTER, and POPPY cohorts (total n = 908) were determined between HIV-positive participants with and without cognitive impairment defined using the HIV-associated neurocognitive disorders (HAND), global deficit score (GDS), and NMM criteria. Results. The prevalence of cognitive impairment varied by up to 27% between methods used to define impairment (eg, 48% for HAND vs 21% for NMM in the CHARTER study). Associations between objective cognitive impairment and subjective cognitive complaints generally were weak. Physical and mental health summary scores (SF-36) were lowest for NMM-defined impairment (P < .05). There were no differences in brain volumes or cortical thickness between participants with and without cognitive impairment defined using the HAND and GDS measures. In contrast, those identified with cognitive impairment by the NMM had reduced mean cortical thickness in both hemispheres (P < .05), as well as smaller brain volumes (P < .01). The associations with measures of white matter microstructure and brain-predicted age generally were weaker. Conclusion. Different methods of defining cognitive impairment identify different people with varying symptomatology and measures of brain injury. Overall, NMM-defined impairment was associated with most neuroimaging abnormalities and poorer selfreported health status. This may be due to the statistical advantage of using a multivariate approac

Depression, lifestyle factors and cognitive function in people living with HIV and comparable HIV-negative controls

We investigated whether differences in cognitive performance between people living with HIV (PLWH) and comparable HIV-negative people were mediated or moderated by depressive symptoms and lifestyle factors. METHODS: A cross-sectional study of 637 'older' PLWH aged ≥ 50 years, 340 'younger' PLWH aged < 50 years and 276 demographically matched HIV-negative controls aged ≥ 50 years enrolled in the Pharmacokinetic and Clinical Observations in People over Fifty (POPPY) study was performed. Cognitive function was assessed using a computerized battery (CogState). Scores were standardized into Z-scores [mean = 0; standard deviation (SD) = 1] and averaged to obtain a global Z-score. Depressive symptoms were evaluated via the Patient Health Questionnaire (PHQ-9). Differences between the three groups and the effects of depression, sociodemographic factors and lifestyle factors on cognitive performance were evaluated using median regression. All analyses accounted for age, gender, ethnicity and level of education. RESULTS: After adjustment for sociodemographic factors, older and younger PLWH had poorer overall cognitive scores than older HIV-negative controls (P < 0.001 and P = 0.006, respectively). Moderate or severe depressive symptoms were more prevalent in both older (27%; P < 0.001) and younger (21%; P < 0.001) PLWH compared with controls (8%). Depressive symptoms (P < 0.001) and use of hashish (P = 0.01) were associated with lower cognitive function; alcohol consumption (P = 0.02) was associated with better cognitive scores. After further adjustment for these factors, the difference between older PLWH and HIV-negative controls was no longer significant (P = 0.08), while that between younger PLWH and older HIV-negative controls remained significant (P = 0.01). CONCLUSIONS: Poorer cognitive performances in PLWH compared with HIV-negative individuals were, in part, mediated by the greater prevalence of depressive symptoms and recreational drug use reported by PLWH

A História da Alimentação: balizas historiográficas

Os M. pretenderam traçar um quadro da História da Alimentação, não como um novo ramo epistemológico da disciplina, mas como um campo em desenvolvimento de práticas e atividades especializadas, incluindo pesquisa, formação, publicações, associações, encontros acadêmicos, etc. Um breve relato das condições em que tal campo se assentou faz-se preceder de um panorama dos estudos de alimentação e temas correia tos, em geral, segundo cinco abardagens Ia biológica, a econômica, a social, a cultural e a filosófica!, assim como da identificação das contribuições mais relevantes da Antropologia, Arqueologia, Sociologia e Geografia. A fim de comentar a multiforme e volumosa bibliografia histórica, foi ela organizada segundo critérios morfológicos. A seguir, alguns tópicos importantes mereceram tratamento à parte: a fome, o alimento e o domínio religioso, as descobertas européias e a difusão mundial de alimentos, gosto e gastronomia. O artigo se encerra com um rápido balanço crítico da historiografia brasileira sobre o tema

Cellular and molecular mechanisms of neurite targeting in the zebrafish visual system

Histologically discrete, parallel layers occur frequently in the nervous system. In many cases, each lamina is a target for innervation by a subset of neurons. We are interested in how neurites select their target laminae. Young zebrafish larvae develop two laminated neuropils in the visual system: the inner plexiform layer (IPL) of the retina and the synaptic region of the optic tectum. Using cell type-specific markers, we have characterized the IPL and tectal neuropil in detail, identifying the complement of neurites that compose each lamina. Using these maps, we investigated of the role of activity in IPL sublamination, and completed a forward genetic screen to identify molecules regulating lamination in the retina and tectum. In mammals, retinal activity is important for the sublamination of ganglion cell (GC) dendrites. Using pharmacological tools and the brudas/NSF mutant, we show that the zebrafish IPL develops in an activity-independent manner, at least until 7 dpf. Hard-wiring mechanisms may be conserved across these species, but other, activity-dependent mechanisms are not. Our screen uncovered five informative mutants. We exploited the moonraker (mra) and notorious (noto) mutants to explore the importance of cell-cell interactions in IPL development. Transplantation studies with mra demonstrate that a subset of amacrine cells (ACs) rely on cell non-autonomous cues to sublaminate. The mra locus encodes DEAD-box protein 19, an RNA helicase not previously implicated in neurite targeting. In noto, GC, AC, and bipolar cell neurites appear to branch outside their target sublaminae. These IPL defects are autonomous to GCs; GC dendrites are able to instruct the sublamination of partner neurites. In the tectal neuropil, noto GC axons and some tectal dendrites are highly disorganized. While wildtype axons project to a single tectal lamina, noto GCs can form arbors in two or three. We explored the developmental relationship between GC axons tectal dendrites by characterizing the tectum of lakritz (lak) mutants, which lack GCs. Our observations uncovered only slight defects in lak mutant tectal dendritogenesis. The grossly visible aspects of lamination in the tectum are largely GC-independent