Showing papers in "The Journal of Comparative Neurology in 2015"
TL;DR: An updated map of the Drosophila central complex will facilitate a deeper behavioral and physiological dissection of this sophisticated set of structures and provides new insights into the anatomical structure of the four components of the central complex and its accessory neuropils.
Abstract: Insects exhibit an elaborate repertoire of behaviors in response to environmental stimuli. The central complex plays a key role in combining various modalities of sensory information with an insect's internal state and past experience to select appropriate responses. Progress has been made in understanding the broad spectrum of outputs from the central complex neuropils and circuits involved in numerous behaviors. Many resident neurons have also been identified. However, the specific roles of these intricate structures and the functional connections between them remain largely obscure. Significant gains rely on obtaining a comprehensive catalog of the neurons and associated GAL4 lines that arborize within these brain regions, and on mapping neuronal pathways connecting these structures. To this end, small populations of neurons in the Drosophila melanogaster central complex were stochastically labeled using the multicolor flip-out technique and a catalog was created of the neurons, their morphologies, trajectories, relative arrangements, and corresponding GAL4 lines. This report focuses on one structure of the central complex, the protocerebral bridge, and identifies just 17 morphologically distinct cell types that arborize in this structure. This work also provides new insights into the anatomical structure of the four components of the central complex and its accessory neuropils. Most strikingly, we found that the protocerebral bridge contains 18 glomeruli, not 16, as previously believed. Revised wiring diagrams that take into account this updated architectural design are presented. This updated map of the Drosophila central complex will facilitate a deeper behavioral and physiological dissection of this sophisticated set of structures. J. Comp. Neurol. 523:997–1037, 2015. © 2014 Wiley Periodicals, Inc.
262 citations
TL;DR: The anatomical basis for systematic studies of LHb function in neural circuits and behavior in mice is provided, and anterograde tracing with adeno‐associated virus–mediated expression of green fluorescent protein, combined with serial two‐photon tomography, are used to map the efferents of the LHb on a standard coordinate system for the entire mouse brain, and reconstruct the efferences in three dimensions.
Abstract: The lateral habenula (LHb) is part of the habenula complex of the dorsal thalamus Recent studies of the LHb have focused on its projections to the ventral tegmental area (VTA) and rostromedial tegmental nucleus (RMTg), which contain γ-aminobutyric acid (GABA)ergic neurons that mediate reward prediction error via inhibition of dopaminergic activity However, older studies in the rat have also identified LHb outputs to the lateral and posterior hypothalamus, median raphe, dorsal raphe, and dorsal tegmentum Although these studies have shown that the medial and lateral divisions of the LHb have somewhat distinct projections, the topographic specificity of LHb efferents is not completely understood, and the relative extent of these projections to brainstem targets is unknown Here we have used anterograde tracing with adeno-associated virus-mediated expression of green fluorescent protein, combined with serial two-photon tomography, to map the efferents of the LHb on a standard coordinate system for the entire mouse brain, and reconstruct the efferent pathways of the LHb in three dimensions Using automated quantitation of fiber density, we show that in addition to the RMTg, the median raphe, caudal dorsal raphe, and pontine central gray are major recipients of LHb efferents By using retrograde tract tracing with cholera toxin subunit B, we show that LHb neurons projecting to the hypothalamus, VTA, median raphe, caudal dorsal raphe, and pontine central gray reside in characteristic, but sometimes overlapping regions of the LHb Together these results provide the anatomical basis for systematic studies of LHb function in neural circuits and behavior in mice J Comp Neurol 523:32-60, 2015 © 2014 Wiley Periodicals, Inc
131 citations
TL;DR: Detailed organization and parcellation of all subfields of the hippocampal head and body regions are revealed on the base of a combined analysis of multiple cyto‐ and chemoarchitectural stains and dense sequential section sampling, facilitating accurate segmentation and measurement of HF subfields in the human brain on MRI scans.
Abstract: The hippocampal formation (HF) is one of the hottest regions in neuroscience because it is critical to learning, memory, and cognition, while being vulnerable to many neurological and mental disorders. With increasing high-resolution imaging techniques, many scientists have started to use distinct landmarks along the anterior-posterior axis of HF to allow segmentation into individual subfields in order to identify specific functions in both normal and diseased conditions. These studies urgently call for more reliable and accurate segmentation of the HF subfields DG, CA3, CA2, CA1, prosubiculum, subiculum, presubiculum, and parasubiculum. Unfortunately, very limited data are available on detailed parcellation of the HF subfields, especially in the complex, curved hippocampal head region. In this study we revealed detailed organization and parcellation of all subfields of the hippocampal head and body regions on the base of a combined analysis of multiple cyto- and chemoarchitectural stains and dense sequential section sampling. We also correlated these subfields to macro-anatomical landmarks, which are visible on magnetic resonance imaging (MRI) scans. Furthermore, we created three versions of the detailed anatomic atlas for the hippocampal head region to account for brains with four, three, or two hippocampal digitations. These results will provide a fundamental basis for understanding the organization, parcellation, and anterior-posterior difference of human HF, facilitating accurate segmentation and measurement of HF subfields in the human brain on MRI scans.
116 citations
TL;DR: This study shows that micro‐CT is highly suitable for analyzing arthropod neuroarchitecture in situ and allows specific neuropils to be distinguished within the brain to extract quantitative data such as neuropil volumes, and data acquisition is considerably faster compared with many classical histological techniques.
Abstract: Classical histology or immunohistochemistry combined with fluorescence or confocal laser scanning microscopy are common techniques in arthropod neuroanatomy, and these methods often require time-consuming and difficult dissections and sample preparations. Moreover, these methods are prone to artifacts due to compression and distortion of tissues, which often result in information loss and especially affect the spatial relationships of the examined parts of the nervous system in their natural anatomical context. Noninvasive approaches such as X-ray micro-computed tomography (micro-CT) can overcome such limitations and have been shown to be a valuable tool for understanding and visualizing internal anatomy and structural complexity. Nevertheless, knowledge about the potential of this method for analyzing the anatomy and organization of nervous systems, especially of taxa with smaller body size (e.g., many arthropods), is limited. This study set out to analyze the brains of selected arthropods with micro-CT, and to compare these results with available histological and immunohistochemical data. Specifically, we explored the influence of different sample preparation procedures. Our study shows that micro-CT is highly suitable for analyzing arthropod neuroarchitecture in situ and allows specific neuropils to be distinguished within the brain to extract quantitative data such as neuropil volumes. Moreover, data acquisition is considerably faster compared with many classical histological techniques. Thus, we conclude that micro-CT is highly suitable for targeting neuroanatomy, as it reduces the risk of artifacts and is faster than classical techniques. J. Comp. Neurol. 523:1281–1295, 2015. © 2015 Wiley Periodicals, Inc.
113 citations
TL;DR: A digital 3D atlas of the live Drosophila AL is generated, the first of its kind, which provides an excellent geometric match for in vivo studies and characterized the innervation patterns of the two most frequently used GAL4‐lines in olfactory research: Orco and GH146‐GAL4.
Abstract: As a model for primary olfactory perception, the antennal lobe (AL) of Drosophila melanogaster is among the most thoroughly investigated and well-understood neuronal structures. Most studies investigating the functional properties and neuronal wiring of the AL are conducted in vivo, although so far the AL morphology has been mainly analyzed in vitro. Identifying the morphological subunits of the AL—the olfactory glomeruli—is usually done using in vitro AL atlases. However, the dissection and fixation procedure causes not only strong volumetric but also geometrical modifications; the result is unpredictable dislocation and a distortion of the AL glomeruli between the in vitro and in vivo brains. Hence, to characterize these artifacts, which are caused by in vitro processing, and to reliably identify glomeruli for in vivo applications, we generated a transgenic fly that expresses the red fluorescent protein DsRed directly fused to the presynaptic protein n-synaptobrevin, under the control of the pan-neuronal promotor elav to label the neuropil in the live animal. Using this fly line, we generated a digital 3D atlas of the live Drosophila AL; this atlas, the first of its kind, provides an excellent geometric match for in vivo studies. We verified the identity of 63% of AL glomeruli by mapping the projections of 34 GAL4-lines of individual chemosensory receptor genes. Moreover, we characterized the innervation patterns of the two most frequently used GAL4-lines in olfactory research: Orco- and GH146-GAL4. The new in vivo AL atlas will be accessible online to the neuroscience community. J. Comp. Neurol. 523:530–544, 2015. © 2014 Wiley Periodicals, Inc.
87 citations
TL;DR: The organization of a subset of nucleocortical projections that constitute an experimentally accessible corollary discharge pathway within the cerebellum is clarified.
Abstract: Motor commands computed by the cerebellum are hypothesized to use corollary discharge, or copies of outgoing commands, to accelerate motor corrections. Identifying sources of corollary discharge, therefore, is critical for testing this hypothesis. Here we verified that the pathway from the cerebellar nuclei to the cerebellar cortex in mice includes collaterals of cerebellar premotor output neurons, mapped this collateral pathway, and identified its postsynaptic targets. Following bidirectional tracer injections into a distal target of the cerebellar nuclei, the ventrolateral thalamus, we observed retrogradely labeled somata in the cerebellar nuclei and mossy fiber terminals in the cerebellar granule layer, consistent with collateral branching. Corroborating these observations, bidirectional tracer injections into the cerebellar cortex retrogradely labeled somata in the cerebellar nuclei and boutons in the ventrolateral thalamus. To test whether nuclear output neurons projecting to the red nucleus also collateralize to the cerebellar cortex, we used a Cre-dependent viral approach, avoiding potential confounds of direct red nucleus-to-cerebellum projections. Injections of a Cre-dependent GFP-expressing virus into Ntsr1-Cre mice, which express Cre selectively in the cerebellar nuclei, retrogradely labeled somata in the interposed nucleus, and putative collateral branches terminating as mossy fibers in the cerebellar cortex. Postsynaptic targets of all labeled mossy fiber terminals were identified using immunohistochemical Golgi cell markers and electron microscopic profiles of granule cells, indicating that the collaterals of nuclear output neurons contact both Golgi and granule cells. These results clarify the organization of a subset of nucleocortical projections that constitute an experimentally accessible corollary discharge pathway within the cerebellum.
86 citations
TL;DR: Data indicated that, while the afferents of the RMTg, VTA, and LHb do originate within the same large pool of central nervous system structures, each is also related to structures that project more strongly to it than to the others.
Abstract: Profound inhibitory control exerted on midbrain dopaminergic neurons by the lateral habenula (LHb), which has mainly excitatory outputs, is mediated by the GABAergic rostromedial tegmental nucleus (RMTg), which strongly innervates dopaminergic neurons in the ventral midbrain. Early reports indicated that the afferent connections of the RMTg, excepting its very strong LHb inputs, do not differ appreciably from those of the ventral tegmental area (VTA). Presumably, however, the RMTg contributes more to behavioral synthesis than to simply invert the valence of the excitatory signal coming from the LHb. Therefore, the present study was done to directly compare the inputs to the RMTg and VTA and, in deference to its substantial involvement with this circuitry, the LHb was also included in the comparison. Data indicated that, while the afferents of the RMTg, VTA, and LHb do originate within the same large pool of central nervous system (CNS) structures, each is also related to structures that project more strongly to it than to the others. The VTA gets robust input from ventral striatopallidum and extended amygdala, whereas RMTg biased inputs arise in structures with a more direct impact on motor function, such as deep layers of the contralateral superior colliculus, deep cerebellar and several brainstem nuclei, and, via a relay in the LHb, the entopeduncular nucleus. Input from the ventral pallidal-lateral preoptic-lateral hypothalamus continuum is strong in the RMTg and VTA and dominant in the LHb. Axon collateralization was also investigated, providing additional insights into the organization of the circuitry of this important triad of structures.
81 citations
TL;DR: Analysis of the subparaventricular zone of the hypothalamus suggests that the circadian timing system may have several parallel neural outflow pathways that provide a road map for understanding how they subserve different functions.
Abstract: The subparaventricular zone of the hypothalamus (SPZ) is the main efferent target of neural projections from the suprachiasmatic nucleus (SCN) and an important relay for the circadian timing system. Although the SPZ is fairly homogeneous cytoarchitecturally and neurochemically, it has been divided into distinct functional and connectional subdivisions. The dorsal subdivision of the SPZ (dSPZ) plays an important role in relaying signals from the SCN controlling body temperature rhythms, while the ventral subdivision (vSPZ) is critical for rhythms of sleep and locomotor activity (Lu et al. [2001] J Neurosci 21:4864–4874). On the other hand, the medial part of the SPZ receives input mainly from the dorsomedial SCN, whereas the lateral SPZ receives input from the ventrolateral SCN and the retinohypothalamic tract (Leak and Moore [2001] J Comp Neurol 433:312–334). We therefore investigated whether there are corresponding differences in efferent outputs from these four quadrants of the SPZ (dorsolateral, ventrolateral, dorsomedial, and ventromedial) by a combination of anterograde and retrograde tracing. We found that, while all four subdivisions of the SPZ share a similar backbone of major projection pathways to the septal region, thalamus, hypothalamus, and brainstem, each segment of the SPZ has a specific set of targets where its projections dominate. Furthermore, we observed intra-SPZ projections of varying densities between the four subdivisions. Taken together, this pattern of organization suggests that the circadian timing system may have several parallel neural outflow pathways that provide a road map for understanding how they subserve different functions. J. Comp. Neurol. 523:2714–2737, 2015. © 2015 Wiley Periodicals, Inc.
75 citations
TL;DR: It is found that hypercapnia activated c‐Fos expression in neurons in the external lateral, lateral crescent, and Kölliker‐Fuse PB subnuclei and that most of these neurons were glutamatergic and virtually none γ‐aminobutyric acidergic.
Abstract: In patients with obstructive sleep apnea, airway obstruction during sleep produces hypercapnia which in turn activates respiratory muscles that pump air into the lungs (e.g. the diaphragm) and that dilate and stabilize the upper airway (e.g., the genioglossus). We hypothesized that these responses are facilitated by glutamatergic neurons in the parabrachial complex (PB) that respond to hypercapnia and project to premotor and motor neurons that innervate the diaphragm and genioglossus muscles. To test this hypothesis we combined c-Fos immunohistochemistry with in situ hybridization for vGluT2 or GAD67 or with retrograde tracing from the ventrolateral medullary region that contains phrenic premotor neurons, the phrenic motor nucleus in the C3-C5 spinal ventral horn, or the hypoglossal motor nucleus. We found that hypercapnia (10% CO2 for 2 hours) activated c-Fos expression in neurons in the external lateral, lateral crescent (PBcr), and Kolliker-Fuse (KF) PB subnuclei and that most of these neurons were glutamatergic and virtually none GABAergic. Numerous CO2− responsive neurons in the KF and PBcr were labeled after retrograde tracer injection into the ventrolateral medulla or hypoglossal motor nuclei, and in the KF after injections into the spinal cord, making them candidates for mediating respiratory-facilitatory and upper airway stabilizing effects of hypercapnia.
70 citations
TL;DR: It is shown that the forebrain part of this circuit that makes a relatively rare direct connection to brainstem vocal motor neurons in independent lineages of vocal learning birds has specialized regulation of axon guidance genes from the SLIT–ROBO molecular pathway.
Abstract: Only a few distantly related mammals and birds have the trait of complex vocal learning, which is the ability to imitate novel sounds. This ability is critical for speech acquisition and production in humans, and is attributed to specialized forebrain vocal control circuits that have several unique connections relative to adjacent brain circuits. As a result, it has been hypothesized that there could exist convergent changes in genes involved in neural connectivity of vocal learning circuits. In support of this hypothesis, expanding on our related study (Pfenning et al. [2014] Science 346: 1256846), here we show that the forebrain part of this circuit that makes a relatively rare direct connection to brainstem vocal motor neurons in independent lineages of vocal learning birds (songbird, parrot, and hummingbird) has specialized regulation of axon guidance genes from the SLIT–ROBO molecular pathway. The SLIT1 ligand was differentially downregulated in the motor song output nucleus that makes the direct projection, whereas its receptor ROBO1 was developmentally upregulated during critical periods for vocal learning. Vocal nonlearning bird species and male mice, which have much more limited vocal plasticity and associated circuits, did not show comparable specialized regulation of SLIT–ROBO genes in their nonvocal motor cortical regions. These findings are consistent with SLIT and ROBO gene dysfunctions associated with autism, dyslexia, and speech sound language disorders and suggest that convergent evolution of vocal learning was associated with convergent changes in the SLIT–ROBO axon guidance pathway. J. Comp. Neurol. 523:892–906, 2015. © 2014 Wiley Periodicals, Inc.
69 citations
TL;DR: Data show that activated LK cells stimulate diuresis in vivo, and that LK and IPC signaling affect food intake in opposite directions, and suggest that serotonin via 5‐HT1B diminishes activity in the LK neurons and thereby modulates functions regulated by LK peptide.
Abstract: Metabolic homeostasis and water balance is maintained by tight hormonal and neuronal regulation. In Drosophila, insulin-like peptides (DILPs) are key regulators of metabolism, and the neuropeptide leucokinin (LK) is a diuretic hormone that also modulates feeding. However, it is not known whether LK and DILPs act together to regulate feeding and water homeostasis. Because LK neurons express the insulin receptor (dInR), we tested functional links between DILP and LK signaling in feeding and water balance. Thus, we performed constitutive and conditional manipulations of activity in LK neurons and insulin-producing cells (IPCs) in adult flies and monitored food intake, responses to desiccation, and peptide expression levels. We also measured in vivo changes in LK and DILP levels in neurons in response to desiccation and drinking. Our data show that activated LK cells stimulate diuresis in vivo, and that LK and IPC signaling affect food intake in opposite directions. Overexpression of the dInR in LK neurons decreases the LK peptide levels, but only caused a subtle decrease in feeding, and had no effect on water balance. Next we demonstrated that LK neurons express the serotonin receptor 5-HT1B. Knockdown of this receptor in LK neurons diminished LK expression, increased desiccation resistance, and diminished food intake. Live calcium imaging indicates that serotonin inhibits spontaneous activity in abdominal LK neurons. Our results suggest that serotonin via 5-HT1B diminishes activity in the LK neurons and thereby modulates functions regulated by LK peptide, but the action of the dInR in these neurons remains less clear. J. Comp. Neurol. 523:1840–1863, 2015. © 2015 Wiley Periodicals, Inc.
TL;DR: This paper investigated the onset of phagocytosis by Olfactory ensheathing cells (OECs) in the developing mouse olfactory system by utilizing two transgenic reporter lines: OMP-ZsGreen mice which express bright green fluorescent protein in OECs.
Abstract: During development of the primary olfactory system, axon targeting is inaccurate and axons inappropriately project within the target layer or overproject into the deeper layers of the olfactory bulb. As a consequence there is considerable apoptosis of primary olfactory neurons during embryonic and postnatal development and axons of the degraded neurons need to be removed. Olfactory ensheathing cells (OECs) are the glia of the primary olfactory nerve and are known to phagocytose axon debris in the adult and postnatal animal. However, it is unclear when phagocytosis by OECs first commences. We investigated the onset of phagocytosis by OECs in the developing mouse olfactory system by utilizing two transgenic reporter lines: OMP-ZsGreen mice which express bright green fluorescent protein in primary olfactory neurons, and S100β-DsRed mice which express red fluorescent protein in OECs. In crosses of these mice, the fate of the degraded axon debris is easily visualized. We found evidence of axon degradation at embryonic day (E)13.5. Phagocytosis of the primary olfactory axon debris by OECs was first detected at E14.5. Phagocytosis of axon debris continued into the postnatal animal during the period when there was extensive mistargeting of olfactory axons. Macrophages were often present in close proximity to OECs but they contributed only a minor role to clearing the axon debris, even after widespread degeneration of olfactory neurons by unilateral bulbectomy and methimazole treatment. These results demonstrate that from early in embryonic development OECs are the primary phagocytic cells of the primary olfactory nerve.
TL;DR: It is shown that the intracardiac nervous system in the zebrafish is anatomically and neurochemically complex, providing a substrate for autonomic control of cardiac effectors in all chambers.
Abstract: In the vertebrate heart the intracardiac nervous system is the final common pathway for autonomic control of cardiac output, but the neuroanatomy of this system is not well understood. In this study we investigated the innervation of the heart in a model vertebrate, the zebrafish. We used antibodies against acetylated tubulin, human neuronal protein C/D, choline acetyltransferase, tyrosine hydroxylase, neuronal nitric oxide synthase, and vasoactive intestinal polypeptide to visualize neural elements and their neurotransmitter content. Most neurons were located at the venous pole in a plexus around the sinoatrial valve; mean total number of cells was 197 ± 23, and 92% were choline acetyltransferase positive, implying a cholinergic role. The plexus contained cholinergic, adrenergic, and nitrergic axons and vasoactive intestinal polypeptide-positive terminals, some innervating somata. Putative pacemaker cells near the plexus showed immunoreactivity for hyperpolarization-activated cyclic nucleotide-gated channel 4 (HCN4) and the transcription factor Islet-1 (Isl1). The neurotracer neurobiotin showed that extrinsic axons from the left and right vagosympathetic trunks innervated the sinoatrial plexus proximal to their entry into the heart; some extrinsic axons from each trunk also projected into the medial dorsal plexus region. Extrinsic axons also innervated the atrial and ventricular walls. An extracardiac nerve trunk innervated the bulbus arteriosus and entered the arterial pole of the heart to innervate the proximal ventricle. We have shown that the intracardiac nervous system in the zebrafish is anatomically and neurochemically complex, providing a substrate for autonomic control of cardiac effectors in all chambers. J. Comp. Neurol. 523:1683–1700, 2015. © 2015 Wiley Periodicals, Inc.
TL;DR: It is shown that Ia/II proprioceptive sensory neurons in mice harboring mutations associated with ALS are affected by ALS‐causing mutations, with pathological changes starting at their peripheral nerve endings.
Abstract: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that primarily targets the motor system. Although much is known about the effects of ALS on motor neurons and glial cells, little is known about its effect on proprioceptive sensory neurons. This study examines proprioceptive sensory neurons in mice harboring mutations associated with ALS, in SOD1(G93A) and TDP43(A315T) transgenic mice. In both transgenic lines, we found fewer proprioceptive sensory neurons containing fluorescently tagged cholera toxin in their soma five days after injecting this retrograde tracer into the tibialis anterior muscle. We asked whether this is due to neuronal loss or selective degeneration of peripheral nerve endings. We found no difference in the total number and size of proprioceptive sensory neuron soma between symptomatic SOD1(G93A) and control mice. However, analysis of proprioceptive nerve endings in muscles revealed early and significant alterations at Ia/II proprioceptive nerve endings in muscle spindles before the symptomatic phase of the disease. Although these changes occur alongside those at α-motor axons in SOD1(G93A) mice, Ia/II sensory nerve endings degenerate in the absence of obvious alterations in α-motor axons in TDP43(A315T) transgenic mice. We next asked whether proprioceptive nerve endings are similarly affected in the spinal cord and found that nerve endings terminating on α-motor neurons are affected during the symptomatic phase and after peripheral nerve endings begin to degenerate. Overall, we show that Ia/II proprioceptive sensory neurons are affected by ALS-causing mutations, with pathological changes starting at their peripheral nerve endings.
TL;DR: It is demonstrated that patterns of projections to A1 are modified following deafness, with statistically significant changes occurring within the auditory thalamus and some cortical areas.
Abstract: Following sensory deprivation, primary somatosensory and visual cortices undergo crossmodal plasticity, which subserves the remaining modalities. However, controversy remains regarding the neuroplastic potential of primary auditory cortex (A1). To examine this, we identified cortical and thalamic projections to A1 in hearing cats and those with early- and late-onset deafness. Following early deafness, inputs from second auditory cortex (A2) are amplified, whereas the number originating in the dorsal zone (DZ) decreases. In addition, inputs from the dorsal medial geniculate nucleus (dMGN) increase, whereas those from the ventral division (vMGN) are reduced. In late-deaf cats, projections from the anterior auditory field (AAF) are amplified, whereas those from the DZ decrease. Additionally, in a subset of early- and late-deaf cats, area 17 and the lateral posterior nucleus (LP) of the visual thalamus project concurrently to A1. These results demonstrate that patterns of projections to A1 are modified following deafness, with statistically significant changes occurring within the auditory thalamus and some cortical areas. Moreover, we provide anatomical evidence for small-scale crossmodal changes in projections to A1 that differ between early- and late-onset deaf animals, suggesting that potential crossmodal activation of primary auditory cortex differs depending on the age of deafness onset.
TL;DR: The results illustrate that ecological selection pressures mold the structure of invertebrate brains, and exemplify how comparative analyses across ecologically divergent species can illuminate the functional significance of variation in brain structure.
Abstract: Interspecific comparisons of brain structure can inform our functional understanding of brain regions, identify adaptations to species-specific ecologies, and explore what constrains adaptive changes in brain structure, and coevolution between functionally related structures. The value of such comparisons is enhanced when the species considered have known ecological differences. The Lepidoptera have long been a favored model in evolutionary biology, but to date descriptions of brain anatomy have largely focused on a few commonly used neurobiological model species. We describe the brain of Godyris zavaleta (Ithomiinae), a member of a subfamily of Neotropical butterflies with enhanced reliance on olfactory information. We demonstrate for the first time the presence of sexually dimorphic glomeruli within a distinct macroglomerular complex (MGC) in the antennal lobe of a diurnal butterfly. This presents a striking convergence with the well-known moth MGC, prompting a discussion of the potential mechanisms behind the independent evolution of specialized glomeruli. Interspecific analyses across four Lepidoptera further show that the relative size of sensory neuropils closely mirror interspecific variation in sensory ecology, with G. zavaleta displaying levels of sensory investment intermediate between the diurnal monarch butterfly (Danaus plexippus), which invests heavily in visual neuropil, and night-flying moths, which invest more in olfactory neuropil. We identify several traits that distinguish butterflies from moths, and several that distinguish D. plexippus and G. zavaleta. Our results illustrate that ecological selection pressures mold the structure of invertebrate brains, and exemplify how comparative analyses across ecologically divergent species can illuminate the functional significance of variation in brain structure.
TL;DR: These vectors provide a starting point for transsynaptic tracing in most vertebrates, and are also excellent candidates for gene transfer in organisms that have been refractory to other methods.
Abstract: Current limitations in technology have prevented an extensive analysis of the connections among neurons, particularly within nonmammalian organisms. We developed a transsynaptic viral tracer originally for use in mice, and then tested its utility in a broader range of organisms. By engineering the vesicular stomatitis virus (VSV) to encode a fluorophore and either the rabies virus glycoprotein (RABV-G) or its own glycoprotein (VSV-G), we created viruses that can transsynaptically label neuronal circuits in either the retrograde or anterograde direction, respectively. The vectors were investigated for their utility as polysynaptic tracers of chicken and zebrafish visual pathways. They showed patterns of connectivity consistent with previously characterized visual system connections, and revealed several potentially novel connections. Further, these vectors were shown to infect neurons in several other vertebrates, including Old and New World monkeys, seahorses, axolotls, and Xenopus. They were also shown to infect two invertebrates, Drosophila melanogaster, and the box jellyfish, Tripedalia cystophora, a species previously intractable for gene transfer, although no clear evidence of transsynaptic spread was observed in these species. These vectors provide a starting point for transsynaptic tracing in most vertebrates, and are also excellent candidates for gene transfer in organisms that have been refractory to other methods.
TL;DR: Profound changes to visual and somatosensory afferent connectivity following deafness may reflect corticocortical rewiring affording acoustically deprived AAF with cross‐modal functionality.
Abstract: Cross-modal reorganization following the loss of input from a sensory modality can recruit sensory-deprived cortical areas to process information from the remaining senses. Specifically, in early-deaf cats, the anterior auditory field (AAF) is unresponsive to auditory stimuli but can be activated by somatosensory and visual stimuli. Similarly, AAF neurons respond to tactile input in adult-deafened animals. To examine anatomical changes that may underlie this functional adaptation following early or late deafness, afferent projections to AAF were examined in hearing cats, and cats with early- or adult-onset deafness. Unilateral deposits of biotinylated dextran amine were made in AAF to retrogradely label cortical and thalamic afferents to AAF. In early-deaf cats, ipsilateral neuronal labeling in visual and somatosensory cortices increased by 329% and 101%, respectively. The largest increases arose from the anterior ectosylvian visual area and the anterolateral lateral suprasylvian visual area, as well as somatosensory areas S2 and S4. Consequently, labeling in auditory areas was reduced by 36%. The age of deafness onset appeared to influence afferent connectivity, with less marked differences observed in late-deaf cats. Profound changes to visual and somatosensory afferent connectivity following deafness may reflect corticocortical rewiring affording acoustically deprived AAF with cross-modal functionality.
TL;DR: This paper investigated whether exaggerated HPA axis responses associated with chronic variable stress (CVS; daily exposure to different stressors at unpredictable times over 14 days, followed by restraint stress on day 15) and diminished HPA output seen following repeated (14 days) restraint-stress exposure are associated with differential engagement of the limbic modulatory network.
Abstract: Emotional stressors activate a stereotyped set of limbic forebrain cell groups implicated in constraining stress-induced hypothalamic-pituitary-adrenal (HPA) axis activation by inhibiting hypophysiotropic neurons in the paraventricular hypothalamic nucleus (PVH). We previously identified a circumscribed, anterior part of the bed nuclei of the stria terminalis (aBST) that houses stress-sensitive, PVH-projecting, γ-aminobutyric acid (GABA)-ergic neurons as representing a site of convergence of stress-inhibitory influences originating from medial prefrontal and hippocampal cortices. Here we investigate whether exaggerated HPA axis responses associated with chronic variable stress (CVS; daily exposure to different stressors at unpredictable times over 14 days, followed by restraint stress on day 15) and diminished HPA output seen following repeated (14 days) restraint-stress exposure are associated with differential engagement of the limbic modulatory network. Relative to acutely restrained rats, animals subjected to CVS showed the expected increase (sensitization) in HPA responses and diminished levels of activation (Fos) of GABAergic neurons and glutamic acid decarboxylase (GAD) mRNA expression in the aBST. By contrast, repeated restraint stress produced habituation in HPA responses, maintained levels of activation of GABAergic neurons, and increased GAD expression in the aBST. aBST-projecting neurons in limbic sites implicated in HPA axis inhibition tended to show diminished activational responses in both repeated-stress paradigms, with the exception of the paraventricular thalamic nucleus, in which responsiveness was maintained in repeatedly restrained animals. The results are consistent with the view that differential engagement of HPA inhibitory mechanisms in the aBST may contribute to alterations in HPA axis responses to emotional stress in sensitization and habituation paradigms.
TL;DR: Investigation of Reelin receptors in the developing mouse cerebral cortex revealed that VLDLR is localized to the distal portion of leading processes in the marginal zone (MZ), whereas ApoER2 is mainly localized to neuronal processes and the cell membranes of multipolar cells in the multipolar cell accumulation zone (MAZ).
Abstract: In mammalian developing brain, neuronal migration is regulated by a variety of signaling cascades, including Reelin signaling. Reelin is a glycoprotein that is mainly secreted by Cajal–Retzius neurons in the marginal zone, playing essential roles in the formation of the layered neocortex via its receptors, apolipoprotein E receptor 2 (ApoER2) and very low density lipoprotein receptor (VLDLR). However, the precise mechanisms by which Reelin signaling controls the neuronal migration process remain unclear. To gain insight into how Reelin signaling controls individual migrating neurons, we generated monoclonal antibodies against ApoER2 and VLDLR and examined the localization of Reelin receptors in the developing mouse cerebral cortex. Immunohistochemical analyses revealed that VLDLR is localized to the distal portion of leading processes in the marginal zone (MZ), whereas ApoER2 is mainly localized to neuronal processes and the cell membranes of multipolar cells in the multipolar cell accumulation zone (MAZ). These different expression patterns may contribute to the distinct actions of Reelin on migrating neurons during both the early and late migratory stages in the developing cerebral cortex. J. Comp. Neurol. 523:463–478, 2015. © 2014 Wiley Periodicals, Inc.
TL;DR: It is confirmed that SP, CGRP, and IB4 can be used as reliable markers for neurons with unmyelinated fibers, and question the suitability of NF200 as a marker for neuronsWith myelinated fiber, and of Peripherin as a markers for neuron with un myelination, or fine‐caliber fibers.
Abstract: Substance P (SP), calcitonin gene-related peptide (CGRP), and isolectin B4 (IB4) are widely used as markers for peripheral neurons with unmyelinated fibers, whereas neurofilament 200 (NF200), and Peripherin are used as markers for neurons with myelinated fibers, and with unmyelinated or small-caliber fibers, respectively. To study the selectivity of these markers for specific neuronal types, we analyzed their expression in neurons in the rat trigeminal ganglion by light- and electron-microscopic immunocytochemistry. Most SP-immunopositive (+), CGRP+, and IB4+ fibers were unmyelinated, but a small fraction (∼5%) were small myelinated fibers ( 20 µm(2) in cross-sectional area, equivalent to >5 µm in diameter, Aβ fiber). Our findings confirm that SP, CGRP, and IB4 can be used as reliable markers for neurons with unmyelinated fibers, and question the suitability of NF200 as a marker for neurons with myelinated fibers, and of Peripherin as a marker for neurons with unmyelinated, or fine-caliber fibers.
TL;DR: A major break was revealed in the networks of theDR such that the caudal third of the DR was more similar in afferent innervation to the MR than it was to the rostral two‐thirds of the D; a major source of heterogeneity within the DR is inclusion of the caUDal component, which may be more accurately viewed as a dorsal extension of the MR.
Abstract: Serotonin neurons in the dorsal and median raphe nuclei (DR and MR) are clustered into heterogeneous groups that give rise to topographically organized forebrain projections. However, a compelling definition of the key subgroups of serotonin neurons within these areas has remained elusive. In order to be functionally distinct, neurons must participate in distinct networks. Therefore, we analyzed subregions of the DR and MR by their afferent input. Clustering methods and principal component analysis were applied in mouse to anterograde tract-tracing experiments available from the Allen Mouse Brain Connectivity Atlas. The results revealed a major break in the networks of the DR such that the caudal third of the DR was more similar in afferent innervation to the MR than it was to the rostral two-thirds of the DR. The rostral part of the DR is associated with networks controlling motor and motivated behavior, while the caudal DR is more closely aligned with regions that regulate rhythmic hippocampal activity. Thus, a major source of heterogeneity within the DR is inclusion of the caudal component, which may be more accurately viewed as a dorsal extension of the MR.
TL;DR: The data demonstrate that GPR151 is highly conserved, specific for a subdivision of the habenular neurocircuitry, and constitutes a promising novel target for psychiatric drug development.
Abstract: The habenula is a phylogenetically conserved brain structure in the epithalamus. It is a major node in the information flow between fronto-limbic brain regions and monoaminergic brainstem nuclei, and is thus anatomically and functionally ideally positioned to regulate emotional, motivational, and cognitive behaviors. Consequently, the habenula may be critically important in the pathophysiology of psychiatric disorders such as addiction and depression. Here we investigated the expression pattern of GPR151, a G protein-coupled receptor (GPCR), whose mRNA has been identified as highly and specifically enriched in habenular neurons by in situ hybridization and translating ribosome affinity purification (TRAP). In the present immunohistochemical study we demonstrate a pronounced and highly specific expression of the GPR151 protein in the medial and lateral habenula of rodent brain. Specific expression was also seen in efferent habenular fibers projecting to the interpeduncular nucleus, the rostromedial tegmental area, the rhabdoid nucleus, the mesencephalic raphe nuclei, and the dorsal tegmental nucleus. Using confocal microscopy and quantitative colocalization analysis, we found that GPR151-expressing axons and terminals overlap with cholinergic, substance P-ergic, and glutamatergic markers. Virtually identical expression patterns were observed in rat, mouse, and zebrafish brains. Our data demonstrate that GPR151 is highly conserved, specific for a subdivision of the habenular neurocircuitry, and constitutes a promising novel target for psychiatric drug development.
TL;DR: The mushroom body of Papilio xuthus receives prominent direct inputs from the optic lobe in addition to olfactory inputs, suggesting a role in visual target detection rather than sky compass orientation.
Abstract: Papilio butterflies depend highly on visual information in their flower-foraging behavior. The retina of Papilio xuthus has been studied well, whereas the visual system in the brain is poorly understood. By investigating outputs from the optic lobe to the central brain, we found that the mushroom body of P. xuthus receives prominent direct inputs from the optic lobe in addition to olfactory inputs. The mushroom body consists of three components: the calyx, the pedunculus, and the lobes. The calyx is further subdivided into two cup-shaped primary calyces and an accessory calyx. Each primary calyx consists of three concentric subareas, the inner zone, the outer zone, and the rim of the outer zone. Dextran injections into the optic lobe, the calyx, or the antennal lobe revealed three visual inputs and one olfactory input into the calyx. The visual inputs originate from the medulla, the lobula, and a newly identified neuropil, the ventral lobe of the lobula. All visual inputs first innervate the accessory calyx, and the two lobula inputs further spread their processes through the inner zone and the rim of the outer zone of the primary calyces. Visual inputs from the medulla and the ventral lobe of the lobula collect light information from ventral eye regions, suggesting a role in visual target detection rather than sky compass orientation. In contrast to visual inputs, olfactory inputs innervate only the calycal outer zone. The multisensory inputs to the mushroom bodies in P. xuthus are probably related to their flower-foraging behavior.
TL;DR: This study compared the expression pattern, laminar distribution, and cell specificity of several rAAV serotypes injected in the primary visual cortex of adult C57Bl/6J mice to select the most optimal vector system to deliver transgenes into specific rodent neocortical circuits.
Abstract: This study compared the expression pattern, laminar distribution, and cell specificity of several rAAV serotypes (2/1, 2/5, 2/7, 2/8, and 2/9) injected in the primary visual cortex (V1) of adult C57Bl/6J mice. In order to obtain specific expression in certain neuron subtypes, different promoter sequences were evaluated for excitatory cell specificity: a universal cytomegalovirus (CMV) promoter, and two versions of the excitatory neuron-specific Ca(2+) /calmodulin-dependent kinase subunit α (CaMKIIα) promoter, CaMKIIα 0.4 and CaMKIIα 1.3. The spatial distribution as well as the cell type specificity was immunohistochemically verified. Depending on the rAAV serotype used, the transduced volume expressing reporter protein differed substantially (rAAV2/5 ≫ 2/7 ≈ 2/9 ≈ 2/8 ≫ 2/1). Excitatory neuron-specific targeting was promoter-dependent, with a surprising difference between the 1.3 kb and 0.4 kb CaMKIIα promoters. While CaMKIIα 1.3 and CMV carrying vectors were comparable, with 78% of the transduced neurons being excitatory for CMV and 82% for CaMKIIα 1.3, the shorter CaMKIIα 0.4 version resulted in 95% excitatory specificity. This study therefore puts forward the CaMKIIα 0.4 promoter as the best choice to target excitatory neurons with rAAVs. Together, these results can be used as an aid to select the most optimal vector system to deliver transgenes into specific rodent neocortical circuits, allowing further elucidation of their functions.
TL;DR: It is shown that progenitor cells expressing c‐Kit, a receptor tyrosine kinase marking stem cells in a variety of embryonic tissues, are required for maintenance of the adult neuroepithelium, providing new insight into the mechanisms involved in adult olfactory neurogenesis.
Abstract: The olfactory epithelium houses chemosensory neurons, which transmit odor information from the nose to the brain. In adult mammals, the olfactory epithelium is a uniquely robust neuroproliferative zone, with the ability to replenish its neuronal and non-neuronal populations due to the presence of germinal basal cells. The stem and progenitor cells of these germinal layers, and their regulatory mechanisms, remain incompletely defined. Here we show that progenitor cells expressing c-Kit, a receptor tyrosine kinase marking stem cells in a variety of embryonic tissues, are required for maintenance of the adult neuroepithelium. Mouse genetic fate-mapping analyses show that embryonically, a c-Kit(+) population contributes to olfactory neurogenesis. In adults under conditions of normal turnover, there is relatively sparse c-Kit(+) progenitor cell (ckPC) activity. However, after experimentally induced neuroepithelial injury, ckPCs are activated such that they reconstitute the neuronal population. There are also occasional non-neuronal cells found to arise from ckPCs. Moreover, the selective depletion of the ckPC population, utilizing temporally controlled targeted diphtheria toxin A expression, results in failure of neurogenesis after experimental injury. Analysis of this model indicates that most ckPCs reside among the globose basal cell populations and act downstream of horizontal basal cells, which can serve as stem cells. Identification of the requirement for olfactory c-Kit-expressing progenitors in olfactory maintenance provides new insight into the mechanisms involved in adult olfactory neurogenesis. Additionally, we define an important and previously unrecognized site of adult c-Kit activity.
TL;DR: Hundreds of differentially expressed genes are identified involved in a variety of processes, including phototransduction, transcriptional regulation, cell adhesion, maintenance of intra‐ and extracellular structure, and metabolism in avian photoreceptor subtypes.
Abstract: Avian photoreceptors are a diverse class of neurons, comprised of four single cones, the two members of the double cone, and rods. The signaling events and transcriptional regulators driving the differentiation of these diverse photoreceptors are largely unknown. In addition, many distinctive features of photoreceptor subtypes, including spectral tuning, oil droplet size and pigmentation, synaptic targets, and spatial patterning, have been well characterized, but the molecular mechanisms underlying these attributes have not been explored. To identify genes specifically expressed in distinct chicken (Gallus gallus) photoreceptor subtypes, we developed fluorescent reporters that label photoreceptor subpopulations, isolated these subpopulations by using fluorescence-activated cell sorting, and subjected them to next-generation sequencing. By comparing the expression profiles of photoreceptors labeled with rhodopsin, red opsin, green opsin, and violet opsin reporters, we have identified hundreds of differentially expressed genes that may underlie the distinctive features of these photoreceptor subtypes. These genes are involved in a variety of processes, including phototransduction, transcriptional regulation, cell adhesion, maintenance of intra- and extracellular structure, and metabolism. Of particular note are a variety of differentially expressed transcription factors, which may drive and maintain photoreceptor diversity, and cell adhesion molecules, which may mediate spatial patterning of photoreceptors and act to establish retinal circuitry. These analyses provide a framework for future studies that will dissect the role of these various factors in the differentiation of avian photoreceptor subtypes.
TL;DR: It is demonstrated that a 15-week occurrence of obesity and insulin resistance during childhood/adolescence induces irreversible epigenetic modifications in the brain that persist following restoration of normal metabolic homeostasis, leading to brain synaptic dysfunction during aging.
Abstract: The prevalence of childhood/adolescent obesity and insulin resistance has reached an epidemic level. Obesity's immediate clinical impacts have been extensively studied; however, current clinical evidence underscores the long-term implications. The current study explored the impacts of brief childhood/adolescent obesity and insulin resistance on cognitive function in later life. To mimic childhood/adolescent obesity and insulin resistance, we exposed 9-week-old C57BL/6J mice to a high-fat diet for 15 weeks, after which the mice exhibited diet-induced obesity and insulin resistance. We then put these mice back on a normal low-fat diet, after which the mice exhibited normal body weight and glucose tolerance. However, a spatial memory test in the forms of the Morris water maze (MWM) and contextual fear conditioning at 85 weeks of age showed that these mice had severe deficits in learning and long-term memory consolidation. Mechanistic investigations identified increased expression of histone deacetylases 5, accompanied by reduced expression of brain-derived neurotrophic factor, in the brains 61 weeks after the mice had been off the high-fat diet. Electrophysiology studies showed that hippocampal slices isolated from these mice are more susceptible to synaptic impairments compared with slices isolated from the control mice. We demonstrated that a 15-week occurrence of obesity and insulin resistance during childhood/adolescence induces irreversible epigenetic modifications in the brain that persist following restoration of normal metabolic homeostasis, leading to brain synaptic dysfunction during aging. Our study provides experimental evidence that limited early-life exposure to obesity and insulin resistance may have long-term deleterious consequences in the brain, contributing to the onset/progression of cognitive dysfunction during aging.
TL;DR: The two raphe nuclei give dual innervation within the OB, with distinct innervation patterns, using recombinant adeno‐associated virusesmediated expression of green fluorescent protein (GFP)‐synaptophysin targeted specifically to neurons of the DRN or the MRN.
Abstract: The raphe nuclei provide serotonergic innervation widely in the brain, thought to mediate a variety of neuromodulatory effects. The mammalian olfactory bulb (OB) is a prominent recipient of serotonergic fibers, particularly in the glomerular layer (GL), where they are thought to gate incoming signals from the olfactory nerve. The dorsal raphe nucleus (DRN) and the median raphe nucleus (MRN) are known to densely innervate the OB. The majority of such projections are thought to terminate in the GL, but this has not been explicitly tested. We sought to investigate this using recombinant adeno-associated viruses (rAAV)-mediated expression of green fluorescent protein (GFP)-synaptophysin targeted specifically to neurons of the DRN or the MRN. With DRN injections, labeled fibers were found mostly in the granule cell layer (GCL), not the GL. Conversely, dense labeling in the GL was observed with MRN injections, suggesting that the source of GL innervation is the MRN, not the DRN, as previously thought. The two raphe nuclei thus give dual innervation within the OB, with distinct innervation patterns. J. Comp. Neurol. 523:805–813, 2015. © 2015 Wiley Periodicals, Inc.
TL;DR: The small telencephalon of the zebrafish shows a remarkable complexity in TR gene expression, and the neurogenic ventricular regions express overlapping but distinct sets of TR genes, suggesting regional differences in the Neurogenic niches in the telencecephalon.
Abstract: The zebrafish has become a model to study adult vertebrate neurogenesis. In particular, the adult telencephalon has been an intensely studied structure in the zebrafish brain. Differential expression of transcriptional regulators (TRs) is a key feature of development and tissue homeostasis. Here we report an expression map of 1,202 TR genes in the telencephalon of adult zebrafish. Our results are summarized in a database with search and clustering functions to identify genes expressed in particular regions of the telencephalon. We classified 562 genes into 13 distinct patterns, including genes expressed in the proliferative zone. The remaining 640 genes displayed unique and complex patterns of expression and could thus not be grouped into distinct classes. The neurogenic ventricular regions express overlapping but distinct sets of TR genes, suggesting regional differences in the neurogenic niches in the telencephalon. In summary, the small telencephalon of the zebrafish shows a remarkable complexity in TR gene expression.
The adult zebrafish telencephalon has become a model to study neurogenesis. We established the expression pattern of more than 1200 transcription regulators (TR) in the adult telencephalon. The neurogenic regions express overlapping but distinct sets of TR genes suggesting regional differences in the neurogenic potential. J. Comp. Neurol. 523:1202–1221, 2015. © 2015 Wiley Periodicals, Inc.