Oressia Zalucki

Neuropilin 1-Sema Signaling Regulates Crossing of Cingulate Pioneering Axons during Development of the Corpus Callosum

Pioneer axons from the cingulate cortex initiate corpus callosum (CC) development, yet nothing is known about the molecules that regulate their guidance. We demonstrate that neuropilin 1 (Npn1) plays an integral role in the development of the CC. Npn1 is localized to axons of cingulate neurons as they cross the midline, and multiple class 3 semaphorins (Semas) are expressed around the developing CC, implicating these guidance molecules in the regulation of Npn1-expressing axons emanating from the cingulate cortex. Furthermore, axons from the cingulate cortex display guidance errors in Npn1(Sema-) mice, a knockin mouse line in which Npn1 is unable to bind Semas. Analysis of mice deficient in the transcription factor Emx2 demonstrated that the cingulate cortex of these mice was significantly reduced in comparison to wild-type controls at E17 and that the CC was absent in rostral sections. Expression of Npn1 was absent in rostral sections of Emx2 mutants, suggesting that Npn1-expressing cingulate pioneers are required for CC formation. These data highlight a central role for Npn1 in the development of projections from the cingulate cortex and further illustrate the importance of these pioneer axons in the formation of the CC.

Netrin-DCC Signaling Regulates Corpus Callosum Formation Through Attraction of Pioneering Axons and by Modulating Slit2-Mediated Repulsion

The left and right sides of the nervous system communicate via commissural axons that cross the midline during development using evolutionarily conserved molecules. These guidance cues have been particularly well studied in the mammalian spinal cord, but it remains unclear whether these guidance mechanisms for commissural axons are similar in the developing forebrain, in particular for the corpus callosum, the largest and most important commissure for cortical function. Here, we show that Netrin1 initially attracts callosal pioneering axons derived from the cingulate cortex, but surprisingly is not attractive for the neocortical callosal axons that make up the bulk of the projection. Instead, we show that Netrin-deleted in colorectal cancer signaling acts in a fundamentally different manner, to prevent the Slit2-mediated repulsion of precrossing axons thereby allowing them to approach and cross the midline. These results provide the first evidence for how callosal axons integrate multiple guidance cues to navigate the midline.

A Sleep/Wake Circuit Controls Isoflurane Sensitivity in Drosophila

General anesthesia remains a mysterious phenomenon, even though a number of compelling target proteins and processes have been proposed [1]. General anesthetics such as isoflurane abolish behavioral responsiveness in all animals, and in the mammalian brain, these diverse compounds probably achieve this in part by targeting endogenous sleep mechanisms [2, 3]. However, most animals sleep [4], and they are therefore likely to have conserved sleep processes. A decade of neurogenetic studies of arousal in Drosophila melanogaster have identified a number of different neurons and brain structures that modulate sleep duration in the fly brain [5-9], but it has remained unclear until recently whether any neurons might form part of a dedicated circuit that actively controls sleep and wake states in the fly brain, as has been proposed for the mammalian brain [10]. We studied general anesthesia in Drosophila by measuring stimulus-induced locomotion under isoflurane gas exposure. Using a syntaxin1A gain-of-function construct, we found that increasing synaptic activity in different Drosophila neurons could produce hypersensitivity or resistance to isoflurane. We uncover a common pathway in the fly brain controlling both sleep duration and isoflurane sensitivity, centered on monoaminergic modulation of sleep-promoting neurons of the fan-shaped body.

Insights into the Biology and Therapeutic Applications of Neural Stem Cells

The cerebral cortex is essential for our higher cognitive functions and emotional reasoning. Arguably, this brain structure is the distinguishing feature of our species, and yet our remarkable cognitive capacity has seemingly come at a cost to the regenerative capacity of the human brain. Indeed, the capacity for regeneration and neurogenesis of the brains of vertebrates has declined over the course of evolution, from fish to rodents to primates. Nevertheless, recent evidence supporting the existence of neural stem cells (NSCs) in the adult human brain raises new questions about the biological significance of adult neurogenesis in relation to ageing and the possibility that such endogenous sources of NSCs might provide therapeutic options for the treatment of brain injury and disease. Here, we highlight recent insights and perspectives on NSCs within both the developing and adult cerebral cortex. Our review of NSCs during development focuses upon the diversity and therapeutic potential of these cells for use in cellular transplantation and in the modeling of neurodevelopmental disorders. Finally, we describe the cellular and molecular characteristics of NSCs within the adult brain and strategies to harness the therapeutic potential of these cell populations in the treatment of brain injury and disease.

Transcriptional regulation of intermediate progenitor cell generation during hippocampal development

During forebrain development, radial glia generate neurons through the production of intermediate progenitor cells (IPCs). The production of IPCs is a central tenet underlying the generation of the appropriate number of cortical neurons, but the transcriptional logic underpinning this process remains poorly defined. Here, we examined IPC production using mice lacking the transcription factor nuclear factor I/X (Nfix). We show that Nfix deficiency delays IPC production and prolongs the neurogenic window, resulting in an increased number of neurons in the postnatal forebrain. Loss of additional Nfi alleles (Nfib) resulted in a severe delay in IPC generation while, conversely, overexpression of NFIX led to precocious IPC generation. Mechanistically, analyses of microarray and ChIP-seq datasets, coupled with the investigation of spindle orientation during radial glial cell division, revealed that NFIX promotes the generation of IPCs via the transcriptional upregulation of inscuteable (Insc). These data thereby provide novel insights into the mechanisms controlling the timely transition of radial glia into IPCs during forebrain development. Summary: The Nfix and Nfib transcription factors are required for the timely transition of radial glia into intermediate progenitor cells during mouse forebrain development.

Behavioral and electrophysiological analysis of general anesthesia in 3 background strains of Drosophila melanogaster

General anesthetics achieve behavioral unresponsiveness via a mechanism that is incompletely understood. The study of genetic model systems such as the fruit fly Drosophila melanogaster is crucial to advancing our understanding of how anesthetic drugs render animals unresponsive. Previous studies have shown that wild-type control strains differ significantly in their sensitivity to general anesthetics, which potentially introduces confounding factors for comparing genetic mutations placed on these wild-type backgrounds. Here, we examined a variety of behavioral and electrophysiological endpoints in Drosophila, in both adult and larval animals. We characterized these endpoints in 3 commonly used fly strains: wild-type Canton Special (CS), and 2 commonly used white-eyed strains, isoCJ1 and w1118. We found that CS and isoCJ1 show remarkably similar sensitivity to isoflurane across a variety of behavioral and electrophysiological endpoints. In contrast, w1118 is resistant to isoflurane compared to the other 2 strains at both the adult and larval stages. This resistance is however not reflected at the level of neurotransmitter release at the larval neuromuscular junction (NMJ). This suggests that the w1118 strain harbors another mutation that produces isoflurane resistance, by acting on an arousal pathway that is most likely preserved between larval and adult brains. This mutation probably also affects sleep, as marked differences between isoCJ1 and w1118 have also recently been found for behavioral responsiveness and sleep intensity measures.

Syntaxin1A-mediated Resistance and Hypersensitivity to Isoflurane in Drosophila melanogaster

Background: Recent evidence suggests that general anesthetics activate endogenous sleep pathways, yet this mechanism cannot explain the entirety of general anesthesia. General anesthetics could disrupt synaptic release processes, as previous work in Caenorhabditis elegans and in vitro cell preparations suggested a role for the soluble NSF attachment protein receptor protein, syntaxin1A, in mediating resistance to several general anesthetics. The authors questioned whether the syntaxin1A-mediated effects found in these reductionist systems reflected a common anesthetic mechanism distinct from sleep-related processes. Methods: Using the fruit fly model, Drosophila melanogaster, the authors investigated the relevance of syntaxin1A manipulations to general anesthesia. The authors used different behavioral and electrophysiological endpoints to test the effect of syntaxin1A mutations on sensitivity to isoflurane. Results: The authors found two syntaxin1A mutations that confer opposite general anesthesia phenotypes: syxH3-C, a 14-amino acid deletion mutant, is resistant to isoflurane (n = 40 flies), and syxKARRAA, a strain with two amino acid substitutions, is hypersensitive to the drug (n = 40 flies). Crucially, these opposing effects are maintained across different behavioral endpoints and life stages. The authors determined the isoflurane sensitivity of syxH3-C at the larval neuromuscular junction to assess effects on synaptic release. The authors find that although isoflurane slightly attenuates synaptic release in wild-type animals (n = 8), syxH3-C preserves synaptic release in the presence of isoflurane (n = 8). Conclusion: The study results are evidence that volatile general anesthetics target synaptic release mechanisms; in addition to first activating sleep pathways, a major consequence of these drugs may be to decrease the efficacy of neurotransmission.

What is unconsciousness in a fly or a worm? A review of general anesthesia in different animal models

All animals are rendered unresponsive by general anesthetics. In humans, this is observed as a succession of endpoints from memory loss to unconsciousness to immobility. Across animals, anesthesia endpoints such as loss of responsiveness or immobility appear to require significantly different drug concentrations. A closer examination in key model organisms such as the mouse, fly, or the worm, uncovers a trend: more complex behaviors, either requiring several sub-behaviors, or multiple neural circuits working together, are more sensitive to volatile general anesthetics. This trend is also evident when measuring neural correlates of general anesthesia. Here, we review this complexity hypothesis in humans and model organisms, and attempt to reconcile these findings with the more recent view that general anesthetics potentiate endogenous sleep pathways in most animals. Finally, we propose a presynaptic mechanism, and thus an explanation for how these drugs might compromise a succession of brain functions of increasing complexity.

Neurogenic differentiation by hippocampal neural stem and progenitor cells is biased by NFIX expression

ABSTRACT Our understanding of the transcriptional programme underpinning adult hippocampal neurogenesis is incomplete. In mice, under basal conditions, adult hippocampal neural stem cells (AH-NSCs) generate neurons and astrocytes, but not oligodendrocytes. The factors limiting oligodendrocyte production, however, remain unclear. Here, we reveal that the transcription factor NFIX plays a key role in this process. NFIX is expressed by AH-NSCs, and its expression is sharply upregulated in adult hippocampal neuroblasts. Conditional ablation of Nfix from AH-NSCs, coupled with lineage tracing, transcriptomic sequencing and behavioural studies collectively reveal that NFIX is cell-autonomously required for neuroblast maturation and survival. Moreover, a small number of AH-NSCs also develop into oligodendrocytes following Nfix deletion. Remarkably, when Nfix is deleted specifically from intermediate progenitor cells and neuroblasts using a Dcx-creERT2 driver, these cells also display elevated signatures of oligodendrocyte gene expression. Together, these results demonstrate the central role played by NFIX in neuroblasts within the adult hippocampal stem cell neurogenic niche in promoting the maturation and survival of these cells, while concomitantly repressing oligodendrocyte gene expression signatures. Summary: In addition to its role in neuroblast maturation and differentiation, NFIX is shown to suppress the expression of oligodendrocyte-associated genes within the adult mouse hippocampus.

Local versus global effects of isoflurane anesthesia on visual processing in the fly brain

Abstract What characteristics of neural activity distinguish the awake and anesthetized brain? Drugs such as isoflurane abolish behavioral responsiveness in all animals, implying evolutionarily conserved mechanisms. However, it is unclear whether this conservation is reflected at the level of neural activity. Studies in humans have shown that anesthesia is characterized by spatially distinct spectral and coherence signatures that have also been implicated in the global impairment of cortical communication. We questioned whether anesthesia has similar effects on global and local neural processing in one of the smallest brains, that of the fruit fly (Drosophila melanogaster). Using a recently developed multielectrode technique, we recorded local field potentials from different areas of the fly brain simultaneously, while manipulating the concentration of isoflurane. Flickering visual stimuli (‘frequency tags’) allowed us to track evoked responses in the frequency domain and measure the effects of isoflurane throughout the brain. We found that isoflurane reduced power and coherence at the tagging frequency (13 or 17 Hz) in central brain regions. Unexpectedly, isoflurane increased power and coherence at twice the tag frequency (26 or 34 Hz) in the optic lobes of the fly, but only for specific stimulus configurations. By modeling the periodic responses, we show that the increase in power in peripheral areas can be attributed to local neuroanatomy. We further show that the effects on coherence can be explained by impacted signal-to-noise ratios. Together, our results show that general anesthesia has distinct local and global effects on neuronal processing in the fruit fly brain.