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Guinea pig Anti-VGLUT2, Unconjugated antibody

RRID:AB_1587626

Antibody ID

AB_1587626

Target Antigen

VGLUT2 mouse, rat, mouse, rat

Proper Citation

(Millipore Cat# AB2251, RRID:AB_1587626)

Clonality

polyclonal antibody

Comments

Discontinued; seller recommendations: Immunohistochemistry; Western Blot; Western Blotting, Immunohistochemistry

Host Organism

guinea pig

Vendor

Millipore

Neurotransmitter diversity in pre-synaptic terminals located in the parvicellular neuroendocrine paraventricular nucleus of the rat and mouse hypothalamus.

  • Johnson CS
  • J. Comp. Neurol.
  • 2018 Jun 1

Literature context:


Abstract:

Virtually all rodent neuroendocrine corticotropin-releasing-hormone (CRH) neurons are in the dorsal medial parvicellular (mpd) part of the paraventricular nucleus of the hypothalamus (PVH). They form the final common pathway for adrenocortical stress responses. Their activity is controlled by sets of GABA-, glutamate-, and catecholamine-containing inputs arranged in an interactive pre-motor network. Defining the nature and arrangement of these inputs can help clarify how stressor type and intensity information is conveyed to neuroendocrine neurons. Here we use immunohistochemistry with high-resolution 3-dimensional image analyses to examine the arrangement of single- and co-occurring GABA, glutamate, and catecholamine markers in synaptophysin-defined pre-synaptic terminals in the PVHmpd of unstressed rats and Crh-IRES-Cre;Ai14 transgenic mice: respectively, vesicular glutamate transporter 2 (VGluT2), vesicular GABA transporter (VGAT), dopamine β-hydroxylase (DBH), and phenylethanolamine n-methyltransferase (PNMT). Just over half of all PVHmpd pre-synaptic terminals contain VGAT, with slightly less containing VGluT2. The vast majority of terminal appositions with mouse CRH neurons occur non-somatically. However, there are significantly more somatic VGAT than VGluT2 appositions. In the rat PVHmpd, about five times as many pre-synaptic terminals contain PNMT than DBH only. However, because epinephrine release has never been detected in the PVH, PNMT terminals may functionally be noradrenergic not adrenergic. PNMT and VGluT2 co-occur in some pre-synaptic terminals indicating the potential for co-transmission of glutamate and norepinephrine. Collectively, these results provide a structural basis for how GABA/glutamate/catecholamine interactions enable adrenocortical responses to fast-onset interosensory stimuli, and more broadly, how combinations of PVH neurotransmitters and neuromodulators interact dynamically to control adrenocortical activity.

Funding information:
  • NINDS NIH HHS - R01 NS029728()
  • NINDS NIH HHS - U54 NS048843(United States)

A Novel Role for Lymphotactin (XCL1) Signaling in the Nervous System: XCL1 Acts via its Receptor XCR1 to Increase Trigeminal Neuronal Excitability.

  • Bird EV
  • Neuroscience
  • 2018 May 21

Literature context:


Abstract:

Chemokines are known to have a role in the nervous system, influencing a range of processes including the development of chronic pain. To date there are very few studies describing the functions of the chemokine lymphotactin (XCL1) or its receptor (XCR1) in the nervous system. We investigated the role of the XCL1-XCR1 axis in nociceptive processing, using a combination of immunohistochemical, pharmacological and electrophysiological techniques. Expression of XCR1 in the rat mental nerve was elevated 3 days following chronic constriction injury (CCI), compared with 11 days post-CCI and sham controls. XCR1 co-existed with neuronal marker PGP9.5, leukocyte common antigen CD45 and Schwann cell marker S-100. In the trigeminal root and white matter of the brainstem, XCR1-positive cells co-expressed the oligodendrocyte marker Olig2. In trigeminal subnucleus caudalis (Vc), XCR1 immunoreactivity was present in the outer laminae and was colocalized with vesicular glutamate transporter 2 (VGlut2), but not calcitonin gene-related peptide (CGRP) or isolectin B4 (IB4). Incubation of brainstem slices with XCL1 induced activation of c-Fos, ERK and p38 in the superficial layers of Vc, and enhanced levels of intrinsic excitability. These effects were blocked by the XCR1 antagonist viral CC chemokine macrophage inhibitory protein-II (vMIP-II). This study has identified for the first time a role for XCL1-XCR1 in nociceptive processing, demonstrating upregulation of XCR1 at nerve injury sites and identifying XCL1 as a modulator of central excitability and signaling via XCR1 in Vc, a key area for modulation of orofacial pain, thus indicating XCR1 as a potential target for novel analgesics.

Funding information:
  • Medical Research Council - 87834(United Kingdom)

Genetic Ablation of All Cerebellins Reveals Synapse Organizer Functions in Multiple Regions Throughout the Brain.

  • Seigneur E
  • J. Neurosci.
  • 2018 May 16

Literature context:


Abstract:

Cerebellins are synaptic organizer molecules that bind to presynaptic neurexins and postsynaptic receptors. They are well studied in the cerebellum, but three of the four cerebellins (Cbln1, Cbln2, and Cbln4) are also broadly expressed outside of the cerebellum, suggesting that they perform general functions throughout the brain. Here, we generated male and female constitutive single (KO), double KO (dKO), and triple KO (tKO) mice of Cbln1, Cbln2, and Cbln4. We found that all constitutive cerebellin-deficient mice were viable and fertile, suggesting that cerebellins are not essential for survival. Cbln1/2 dKO mice exhibited salience-induced seizures that were aggravated in Cbln1/2/4 tKO mice, suggesting that all cerebellins contribute to brain function. As described previously, Cbln1 KO mice displayed major motor impairments that were aggravated by additional KO of Cbln2. Strikingly, the Cbln1/2 dKO did not cause alterations in synapse density in the hippocampus of young adult (1- and 2-month-old) mice, but produced a selective ∼50% decrease in hippocampal synapse density in the stratum lacunosum moleculare of the CA1 region and in the dentate gyrus of aging, 6-month-old mice. A similar decrease in excitatory synapse density was observed in the striatum and retrosplenial cortex. Behaviorally, the Cbln1 KO produced dramatic changes in motor behaviors that were partly aggravated by additional deletion of Cbln2 and/or Cbln4. Our results show that cerebellins are not essential for survival and do not contribute to initial synapse formation, but perform multiple functions throughout the brain; as a consequence, their ablation results in a delayed loss of synapses and in behavioral impairments.SIGNIFICANCE STATEMENT Cerebellins (Cbln1-4) are trans-synaptic cell adhesion molecules. In the cerebellum, Cbln1 functions as a bidirectional organizer of parallel fiber-Purkinje cell synapses by binding to presynaptic neurexins and postsynaptic GluRδ2. Little is known about the function of cerebellins outside of the cerebellum; therefore, the present study used single, double, and triple constitutive KO mice of Cbln1, Cbln2, and Cbln4 to analyze the overall function of cerebellins. We show that cerebellins act as important synaptic organizers in specific subsets of neurons and likely contribute to many different brain functions. We also show that cerebellins are not initially required for synapse formation, but rather for specification and long-term synapse maintenance and demonstrate that all cerebellins, not just Cbln1, contribute to brain function.

Funding information:
  • NICHD NIH HHS - T32 HD07190(United States)
  • NIMH NIH HHS - R37 MH052804()

An Optical Neuron-Astrocyte Proximity Assay at Synaptic Distance Scales.

  • Octeau JC
  • Neuron
  • 2018 Apr 4

Literature context:


Abstract:

Astrocytes are complex bushy cells that serve important functions through close contacts between their processes and synapses. However, the spatial interactions and dynamics of astrocyte processes relative to synapses have proven problematic to study in adult living brain tissue. Here, we report a genetically targeted neuron-astrocyte proximity assay (NAPA) to measure astrocyte-synapse spatial interactions within intact brain preparations and at synaptic distance scales. The method exploits resonance energy transfer between extracellularly displayed fluorescent proteins targeted to synapses and astrocyte processes. We validated the method in the striatal microcircuitry following in vivo expression. We determined the proximity of striatal astrocyte processes to distinct neuronal input pathways, to D1 and D2 medium spiny neuron synapses, and we evaluated how astrocyte-to-excitatory synapse proximity changed following cortical afferent stimulation, during ischemia and in a model of Huntington's disease. NAPA provides a simple approach to measure astrocyte-synapse spatial interactions in a variety of experimental scenarios. VIDEO ABSTRACT.

Funding information:
  • NCI NIH HHS - R01 CA104926(United States)

Reducing Pericyte-Derived Scarring Promotes Recovery after Spinal Cord Injury.

  • Dias DO
  • Cell
  • 2018 Mar 22

Literature context:


Abstract:

CNS injury often severs axons. Scar tissue that forms locally at the lesion site is thought to block axonal regeneration, resulting in permanent functional deficits. We report that inhibiting the generation of progeny by a subclass of pericytes led to decreased fibrosis and extracellular matrix deposition after spinal cord injury in mice. Regeneration of raphespinal and corticospinal tract axons was enhanced and sensorimotor function recovery improved following spinal cord injury in animals with attenuated pericyte-derived scarring. Using optogenetic stimulation, we demonstrate that regenerated corticospinal tract axons integrated into the local spinal cord circuitry below the lesion site. The number of regenerated axons correlated with improved sensorimotor function recovery. In conclusion, attenuation of pericyte-derived fibrosis represents a promising therapeutic approach to facilitate recovery following CNS injury.

Funding information:
  • Intramural NIH HHS - Z01 DE000698-10(United States)

Morphology of P2X3-immunoreactive nerve endings in the rat tracheal mucosa.

  • Yamamoto Y
  • J. Comp. Neurol.
  • 2018 Feb 15

Literature context:


Abstract:

Nerve endings with immunoreactivity for the P2X3 purinoreceptor (P2X3) in the rat tracheal mucosa were examined by immunohistochemistry of whole-mount preparations with confocal scanning laser microscopy. P2X3 immunoreactivity was observed in ramified endings distributed in the whole length of the trachea. The myelinated parent axons of P2X3-immunoreactive nerve endings ramified into several branches that extended two-dimensionally in every direction at the interface between the epithelial layer and lamina propria. The axonal branches of P2X3-immunoreactive endings branched off many twigs located just beneath the epithelium, and continued to intraepithelial axon terminals. The axon terminals of P2X3-immunoreactive endings were beaded, rounded, or club-like in shape and terminated between tracheal epithelial cells. Flat axon terminals sometimes partly ensheathed neuroendocrine cells with immunoreactivity for SNAP25 or CGRP. Some axons and axon terminals with P2X3 immunoreactivity were immunoreactive for P2X2, while some terminals were immunoreactive for vGLUT2. Furthermore, a retrograde tracing method using fast blue (FB) revealed that 88.4% of FB-labeled cells with P2X3 immunoreactivity originated from the nodose ganglion. In conclusion, P2X3-immunoreactive nerve endings in the rat tracheal mucosa have unique morphological characteristics, and these endings may be rapidly adapting receptors and/or irritant receptors that are activated by mucosal irritant stimuli.

Tonotopic alterations in inhibitory input to the medial nucleus of the trapezoid body in a mouse model of Fragile X syndrome.

  • McCullagh EA
  • J. Comp. Neurol.
  • 2017 Nov 1

Literature context:


Abstract:

Hyperexcitability and the imbalance of excitation/inhibition are one of the leading causes of abnormal sensory processing in Fragile X syndrome (FXS). The precise timing and distribution of excitation and inhibition is crucial for auditory processing at the level of the auditory brainstem, which is responsible for sound localization ability. Sound localization is one of the sensory abilities disrupted by loss of the Fragile X Mental Retardation 1 (Fmr1) gene. Using triple immunofluorescence staining we tested whether there were alterations in the number and size of presynaptic structures for the three primary neurotransmitters (glutamate, glycine, and GABA) in the auditory brainstem of Fmr1 knockout mice. We found decreases in either glycinergic or GABAergic inhibition to the medial nucleus of the trapezoid body (MNTB) specific to the tonotopic location within the nucleus. MNTB is one of the primary inhibitory nuclei in the auditory brainstem and participates in the sound localization process with fast and well-timed inhibition. Thus, a decrease in inhibitory afferents to MNTB neurons should lead to greater inhibitory output to the projections from this nucleus. In contrast, we did not see any other significant alterations in balance of excitation/inhibition in any of the other auditory brainstem nuclei measured, suggesting that the alterations observed in the MNTB are both nucleus and frequency specific. We furthermore show that glycinergic inhibition may be an important contributor to imbalances in excitation and inhibition in FXS and that the auditory brainstem is a useful circuit for testing these imbalances.

Experience-Dependent Synaptic Plasticity in V1 Occurs without Microglial CX3CR1.

  • Schecter RW
  • J. Neurosci.
  • 2017 Nov 1

Literature context:


Abstract:

Brief monocular deprivation (MD) shifts ocular dominance and reduces the density of thalamic synapses in layer 4 of the mouse primary visual cortex (V1). We found that microglial lysosome content is also increased as a result of MD. Previous studies have shown that the microglial fractalkine receptor CX3CR1 is involved in synaptic development and hippocampal plasticity. We therefore tested the hypothesis that neuron-to-microglial communication via CX3CR1 is an essential component of visual cortical development and plasticity in male mice. Our data show that CX3CR1 is not required for normal development of V1 responses to visual stimulation, multiple forms of experience-dependent plasticity, or the synapse loss that accompanies MD in layer 4. By ruling out an essential role for fractalkine signaling, our study narrows the search for understanding how microglia respond to active synapse modification in the visual cortex.SIGNIFICANCE STATEMENT Microglia in the visual cortex respond to monocular deprivation with increased lysosome content, but signaling through the fractalkine receptor CX3CR1 is not an essential component in the mechanisms of visual cortical development or experience-dependent synaptic plasticity.

Funding information:
  • NEI NIH HHS - R01 EY012309()

Functional Convergence at the Retinogeniculate Synapse.

  • Litvina EY
  • Neuron
  • 2017 Oct 11

Literature context:


Abstract:

Precise connectivity between retinal ganglion cells (RGCs) and thalamocortical (TC) relay neurons is thought to be essential for the transmission of visual information. Consistent with this view, electrophysiological measurements have previously estimated that 1-3 RGCs converge onto a mouse geniculate TC neuron. Recent advances in connectomics and rabies tracing have yielded much higher estimates of retinogeniculate convergence, although not all identified contacts may be functional. Here we use optogenetics and a computational simulation to determine the number of functionally relevant retinogeniculate inputs onto TC neurons in mice. We find an average of ten RGCs converging onto a mature TC neuron, in contrast to >30 inputs before developmental refinement. However, only 30% of retinogeniculate inputs exceed the threshold for dominating postsynaptic activity. These results signify a greater role for the thalamus in visual processing and provide a functional perspective of anatomical connectivity data.

Light reintroduction after dark exposure reactivates plasticity in adults via perisynaptic activation of MMP-9.

  • Murase S
  • Elife
  • 2017 Sep 6

Literature context:


Abstract:

The sensitivity of ocular dominance to regulation by monocular deprivation is the canonical model of plasticity confined to a critical period. However, we have previously shown that visual deprivation through dark exposure (DE) reactivates critical period plasticity in adults. Previous work assumed that the elimination of visual input was sufficient to enhance plasticity in the adult mouse visual cortex. In contrast, here we show that light reintroduction (LRx) after DE is responsible for the reactivation of plasticity. LRx triggers degradation of the ECM, which is blocked by pharmacological inhibition or genetic ablation of matrix metalloproteinase-9 (MMP-9). LRx induces an increase in MMP-9 activity that is perisynaptic and enriched at thalamo-cortical synapses. The reactivation of plasticity by LRx is absent in Mmp9-/- mice, and is rescued by hyaluronidase, an enzyme that degrades core ECM components. Thus, the LRx-induced increase in MMP-9 removes constraints on structural and functional plasticity in the mature cortex.

Funding information:
  • Canadian Institutes of Health Research - (Canada)

Distinct Ventral Pallidal Neural Populations Mediate Separate Symptoms of Depression.

  • Knowland D
  • Cell
  • 2017 Jul 13

Literature context:


Abstract:

Major depressive disorder (MDD) patients display a common but often variable set of symptoms making successful, sustained treatment difficult to achieve. Separate depressive symptoms may be encoded by differential changes in distinct circuits in the brain, yet how discrete circuits underlie behavioral subsets of depression and how they adapt in response to stress has not been addressed. We identify two discrete circuits of parvalbumin-positive (PV) neurons in the ventral pallidum (VP) projecting to either the lateral habenula or ventral tegmental area contributing to depression. We find that these populations undergo different electrophysiological adaptations in response to social defeat stress, which are normalized by antidepressant treatment. Furthermore, manipulation of each population mediates either social withdrawal or behavioral despair, but not both. We propose that distinct components of the VP PV circuit can subserve related, yet separate depressive-like phenotypes in mice, which could ultimately provide a platform for symptom-specific treatments of depression.

Funding information:
  • NIMH NIH HHS - R01 MH107742()
  • NIMH NIH HHS - R01 MH108594()

Peripheral and central anatomical organization of cutaneous afferent subtypes in a rat nociceptive intersegmental spinal reflex.

  • Lee HJ
  • J. Comp. Neurol.
  • 2017 Jun 15

Literature context:


Abstract:

Stimulation of rat segmental dorsal cutaneous nerves (DCNs) evokes the nociceptive intersegmental cutaneus trunci muscle (CTM) reflex. The reflex consists of early and late responses, mediated by Aδ and C fibers, respectively, based on required stimulation strengths, and shows segmental differences in terms of amplitude and duration. We have now investigated whether the peripheral or central anatomy of nociceptive afferent subtypes in different DCNs also vary in a segmental manner. The numbers of different axon subtypes, determined by axon diameter, were analyzed across peripheral DCNs from T6 to L1. The central projections of T7 and T13 DCN afferents were traced using DCN injections of cholera toxin subunit B (CTB) for myelinated A fibers and isolectin B4 (IB4) for unmyelinated C fibers and both labels were quantified in the dorsal horns. Peripheral axon subtype numbers did not differ significantly across DCNs. Centrally, IB4+ , but not CTB+ , projection areas were different between T7 and T13, consistent with different segmental CTM neurogram responses. At both levels, A fibers projected to deeper layers of the dorsal horn than did C fibers. These termination sites are consistent with both mono- and polysynaptic connections between these afferents and the ascending propriospinal interneurons of the reflex. Also analyzed were the spatial distribution, the synaptic termination, and the glutamatergic transporter profiles of DCN A and C fibers and their relationship to calcitonin gene-related peptide (CGRP) staining in the dorsal horn.

Conditional Deletion of All Neurexins Defines Diversity of Essential Synaptic Organizer Functions for Neurexins.

  • Chen LY
  • Neuron
  • 2017 May 3

Literature context:


Abstract:

Neurexins are recognized as key organizers of synapses that are essential for normal brain function. However, it is unclear whether neurexins are fundamental building blocks of all synapses with similar overall functions or context-dependent specifiers of synapse properties. To address this question, we produced triple cKO (conditional knockout) mice that allow ablating all neurexin expression in mice. Using neuron-specific manipulations combined with immunocytochemistry, paired recordings, and two-photon Ca2+ imaging, we analyzed excitatory synapses formed by climbing fibers on Purkinje cells in cerebellum and inhibitory synapses formed by parvalbumin- or somatostatin-positive interneurons on pyramidal layer 5 neurons in the medial prefrontal cortex. After pan-neurexin deletions, we observed in these synapses severe but dramatically different synaptic phenotypes that ranged from major impairments in their distribution and function (climbing-fiber synapses) to large decreases in synapse numbers (parvalbumin-positive synapses) and severe alterations in action potential-induced presynaptic Ca2+ transients (somatostatin-positive synapses). Thus, neurexins function primarily as context-dependent specifiers of synapses.

Funding information:
  • NIMH NIH HHS - F32 MH100745()
  • NIMH NIH HHS - R37 MH052804()

Probabilistic fluorescence-based synapse detection.

  • Simhal AK
  • PLoS Comput. Biol.
  • 2017 Apr 17

Literature context:


Abstract:

Deeper exploration of the brain's vast synaptic networks will require new tools for high-throughput structural and molecular profiling of the diverse populations of synapses that compose those networks. Fluorescence microscopy (FM) and electron microscopy (EM) offer complementary advantages and disadvantages for single-synapse analysis. FM combines exquisite molecular discrimination capacities with high speed and low cost, but rigorous discrimination between synaptic and non-synaptic fluorescence signals is challenging. In contrast, EM remains the gold standard for reliable identification of a synapse, but offers only limited molecular discrimination and is slow and costly. To develop and test single-synapse image analysis methods, we have used datasets from conjugate array tomography (cAT), which provides voxel-conjugate FM and EM (annotated) images of the same individual synapses. We report a novel unsupervised probabilistic method for detection of synapses from multiplex FM (muxFM) image data, and evaluate this method both by comparison to EM gold standard annotated data and by examining its capacity to reproduce known important features of cortical synapse distributions. The proposed probabilistic model-based synapse detector accepts molecular-morphological synapse models as user queries, and delivers a volumetric map of the probability that each voxel represents part of a synapse. Taking human annotation of cAT EM data as ground truth, we show that our algorithm detects synapses from muxFM data alone as successfully as human annotators seeing only the muxFM data, and accurately reproduces known architectural features of cortical synapse distributions. This approach opens the door to data-driven discovery of new synapse types and their density. We suggest that our probabilistic synapse detector will also be useful for analysis of standard confocal and super-resolution FM images, where EM cross-validation is not practical.

Speed and segmentation control mechanisms characterized in rhythmically-active circuits created from spinal neurons produced from genetically-tagged embryonic stem cells.

  • Sternfeld MJ
  • Elife
  • 2017 Feb 14

Literature context:


Abstract:

Flexible neural networks, such as the interconnected spinal neurons that control distinct motor actions, can switch their activity to produce different behaviors. Both excitatory (E) and inhibitory (I) spinal neurons are necessary for motor behavior, but the influence of recruiting different ratios of E-to-I cells remains unclear. We constructed synthetic microphysical neural networks, called circuitoids, using precise combinations of spinal neuron subtypes derived from mouse stem cells. Circuitoids of purified excitatory interneurons were sufficient to generate oscillatory bursts with properties similar to in vivo central pattern generators. Inhibitory V1 neurons provided dual layers of regulation within excitatory rhythmogenic networks - they increased the rhythmic burst frequency of excitatory V3 neurons, and segmented excitatory motor neuron activity into sub-networks. Accordingly, the speed and pattern of spinal circuits that underlie complex motor behaviors may be regulated by quantitatively gating the intra-network cellular activity ratio of E-to-I neurons.

Funding information:
  • NINDS NIH HHS - F31 NS080340()
  • NINDS NIH HHS - R01 NS090919()

Excitatory Hindbrain-Forebrain Communication Is Required for Cisplatin-Induced Anorexia and Weight Loss.

  • Alhadeff AL
  • J. Neurosci.
  • 2017 Jan 11

Literature context:


Abstract:

Cisplatin chemotherapy is commonly used to treat cancer despite severe energy balance side effects. In rats, cisplatin activates nucleus tractus solitarius (NTS) projections to the lateral parabrachial nucleus (lPBN) and calcitonin-gene related peptide (CGRP) projections from the lPBN to the central nucleus of the amygdala (CeA). We demonstrated previously that CeA glutamate receptor signaling mediates cisplatin-induced anorexia and body weight loss. Here, we used neuroanatomical tracing, immunofluorescence, and confocal imaging to demonstrate that virtually all NTS→lPBN and lPBN→CeA CGRP projections coexpress vesicular glutamate transporter 2 (VGLUT2), providing evidence that excitatory projections mediate cisplatin-induced energy balance dysregulation. To test whether lPBN→CeA projection neurons are required for cisplatin-induced anorexia and weight loss, we inhibited these neurons chemogenetically using a retrograde Cre-recombinase-expressing canine adenovirus-2 in combination with Cre-dependent inhibitory Designer Receptors Exclusive Activated by Designer Drugs (DREADDs) before cisplatin treatment. Inhibition of lPBN→CeA neurons attenuated cisplatin-induced anorexia and body weight loss significantly. Using a similar approach, we additionally demonstrated that inhibition of NTS→lPBN neurons attenuated cisplatin-induced anorexia and body weight loss significantly. Together, our data support the view that excitatory hindbrain-forebrain projections are necessary for cisplatin's untoward effects on energy intake, elucidating a key neuroanatomical circuit driving pathological anorexia and weight loss that accompanies chemotherapy treatment. SIGNIFICANCE STATEMENT: Chemotherapy treatments are commonly used to treat cancers despite accompanying anorexia and weight loss that may limit treatment adherence and reduce patient quality of life. Strikingly, we lack a neural understanding of, and effective treatments for, chemotherapy-induced anorexia and weight loss. The current data characterize the excitatory nature of neural projections activated by cisplatin in rats and reveal the necessity of specific hindbrain-forebrain projections for cisplatin-induced anorexia and weight loss. Together, these findings help to characterize the neural mechanisms mediating cisplatin-induced anorexia, advancing opportunities to develop better-tolerated chemotherapies and adjuvant therapies to prevent anorexia and concurrent nutritional deficiencies during cancer treatment.

Funding information:
  • NIDDK NIH HHS - R01 DK021397()
  • NIDDK NIH HHS - R01 DK100685()
  • NIDDK NIH HHS - R56 DK021397()
  • NIMH NIH HHS - R01 MH059911()

Spatial organization of astrocytes in ferret visual cortex.

  • López-Hidalgo M
  • J. Comp. Neurol.
  • 2016 Dec 1

Literature context:


Abstract:

Astrocytes form an intricate partnership with neural circuits to influence numerous cellular and synaptic processes. One prominent organizational feature of astrocytes is the "tiling" of the brain with non-overlapping territories. There are some documented species and brain region-specific astrocyte specializations, but the extent of astrocyte diversity and circuit specificity are still unknown. We quantitatively defined the rules that govern the spatial arrangement of astrocyte somata and territory overlap in ferret visual cortex using a combination of in vivo two-photon imaging, morphological reconstruction, immunostaining, and model simulations. We found that ferret astrocytes share, on average, half of their territory with other astrocytes. However, a specific class of astrocytes, abundant in thalamo-recipient cortical layers ("kissing" astrocytes), overlap markedly less. Together, these results demonstrate novel features of astrocyte organization indicating that different classes of astrocytes are arranged in a circuit-specific manner and that tiling does not apply universally across brain regions and species. J. Comp. Neurol. 524:3561-3576, 2016. © 2016 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc.

Somatodendritic Expression of JAM2 Inhibits Oligodendrocyte Myelination.

  • Redmond SA
  • Neuron
  • 2016 Aug 17

Literature context:


Abstract:

Myelination occurs selectively around neuronal axons to increase the efficiency and velocity of action potentials. While oligodendrocytes are capable of myelinating permissive structures in the absence of molecular cues, structurally permissive neuronal somata and dendrites remain unmyelinated. Utilizing a purified spinal cord neuron-oligodendrocyte myelinating co-culture system, we demonstrate that disruption of dynamic neuron-oligodendrocyte signaling by chemical cross-linking results in aberrant myelination of the somatodendritic compartment of neurons. We hypothesize that an inhibitory somatodendritic cue is necessary to prevent non-axonal myelination. Using next-generation sequencing and candidate profiling, we identify neuronal junction adhesion molecule 2 (JAM2) as an inhibitory myelin-guidance molecule. Taken together, our results demonstrate that the somatodendritic compartment directly inhibits myelination and suggest a model in which broadly indiscriminate myelination is tailored by inhibitory signaling to meet local myelination requirements.

Proteomic Analysis of Unbounded Cellular Compartments: Synaptic Clefts.

  • Loh KH
  • Cell
  • 2016 Aug 25

Literature context:


Abstract:

Cellular compartments that cannot be biochemically isolated are challenging to characterize. Here we demonstrate the proteomic characterization of the synaptic clefts that exist at both excitatory and inhibitory synapses. Normal brain function relies on the careful balance of these opposing neural connections, and understanding how this balance is achieved relies on knowledge of their protein compositions. Using a spatially restricted enzymatic tagging strategy, we mapped the proteomes of two of the most common excitatory and inhibitory synaptic clefts in living neurons. These proteomes reveal dozens of synaptic candidates and assign numerous known synaptic proteins to a specific cleft type. The molecular differentiation of each cleft allowed us to identify Mdga2 as a potential specificity factor influencing Neuroligin-2's recruitment of presynaptic neurotransmitters at inhibitory synapses.

Ultrastructure of geniculocortical synaptic connections in the tree shrew striate cortex.

  • Familtsev D
  • J. Comp. Neurol.
  • 2016 Apr 15

Literature context:


Abstract:

To determine whether thalamocortical synaptic circuits differ across cortical areas, we examined the ultrastructure of geniculocortical terminals in the tree shrew striate cortex to compare directly the characteristics of these terminals with those of pulvinocortical terminals (examined previously in the temporal cortex of the same species; Chomsung et al. [] Cereb Cortex 20:997-1011). Tree shrews are considered to represent a prototype of early prosimian primates but are unique in that sublaminae of striate cortex layer IV respond preferentially to light onset (IVa) or offset (IVb). We examined geniculocortical inputs to these two sublayers labeled by tracer or virus injections or an antibody against the type 2 vesicular glutamate antibody (vGLUT2). We found that layer IV geniculocortical terminals, as well as their postsynaptic targets, were significantly larger than pulvinocortical terminals and their postsynaptic targets. In addition, we found that 9-10% of geniculocortical terminals in each sublamina contacted GABAergic interneurons, whereas pulvinocortical terminals were not found to contact any interneurons. Moreover, we found that the majority of geniculocortical terminals in both IVa and IVb contained dendritic protrusions, whereas pulvinocortical terminals do not contain these structures. Finally, we found that synaptopodin, a protein uniquely associated with the spine apparatus, and telencephalin (TLCN, or intercellular adhesion molecule type 5), a protein associated with maturation of dendritic spines, are largely excluded from geniculocortical recipient layers of the striate cortex. Together our results suggest major differences in the synaptic organization of thalamocortical pathways in striate and extrastriate areas.

Differential expression patterns of K(+) /Cl(-) cotransporter 2 in neurons within the superficial spinal dorsal horn of rats.

  • Javdani F
  • J. Comp. Neurol.
  • 2015 Sep 1

Literature context:


Abstract:

γ-Aminobutyric acid (GABA)- and glycine-mediated hyperpolarizing inhibition is associated with a chloride influx that depends on the inwardly directed chloride electrochemical gradient. In neurons, the extrusion of chloride from the cytosol primarily depends on the expression of an isoform of potassium-chloride cotransporters (KCC2s). KCC2 is crucial in the regulation of the inhibitory tone of neural circuits, including pain processing neural assemblies. Thus we investigated the cellular distribution of KCC2 in neurons underlying pain processing in the superficial spinal dorsal horn of rats by using high-resolution immunocytochemical methods. We demonstrated that perikarya and dendrites widely expressed KCC2, but axon terminals proved to be negative for KCC2. In single ultrathin sections, silver deposits labeling KCC2 molecules showed different densities on the surface of dendritic profiles, some of which were negative for KCC2. In freeze fracture replicas and tissue sections double stained for the β3-subunit of GABAA receptors and KCC2, GABAA receptors were revealed on dendritic segments with high and also with low KCC2 densities. By measuring the distances between spots immunoreactive for gephyrin (a scaffolding protein of GABAA and glycine receptors) and KCC2 on the surface of neurokinin 1 (NK1) receptor-immunoreactive dendrites, we found that gephyrin-immunoreactive spots were located at various distances from KCC2 cotransporters; 5.7 % of them were recovered in the middle of 4-10-µm-long dendritic segments that were free of KCC2 immunostaining. The variable local densities of KCC2 may result in variable postsynaptic potentials evoked by the activation of GABAA and glycine receptors along the dendrites of spinal neurons.

Connectivity of pacemaker neurons in the neonatal rat superficial dorsal horn.

  • Li J
  • J. Comp. Neurol.
  • 2015 May 1

Literature context:


Abstract:

Pacemaker neurons with an intrinsic ability to generate rhythmic burst-firing have been characterized in lamina I of the neonatal spinal cord, where they are innervated by high-threshold sensory afferents. However, little is known about the output of these pacemakers, as the neuronal populations that are targeted by pacemaker axons have yet to be identified. The present study combines patch-clamp recordings in the intact neonatal rat spinal cord with tract-tracing to demonstrate that lamina I pacemaker neurons contact multiple spinal motor pathways during early life. Retrograde labeling of premotor interneurons with the trans-synaptic pseudorabies virus PRV-152 revealed the presence of burst-firing in PRV-infected lamina I neurons, thereby confirming that pacemakers are synaptically coupled to motor networks in the spinal ventral horn. Notably, two classes of pacemakers could be distinguished in lamina I based on cell size and the pattern of their axonal projections. Whereas small pacemaker neurons possessed ramified axons that contacted ipsilateral motor circuits, large pacemaker neurons had unbranched axons that crossed the midline and ascended rostrally in the contralateral white matter. Recordings from identified spino-parabrachial and spino-periaqueductal gray neurons indicated the presence of pacemaker activity within neonatal lamina I projection neurons. Overall, these results show that lamina I pacemakers are positioned to regulate both the level of activity in developing motor circuits and the ascending flow of nociceptive information to the brain, thus highlighting a potential role for pacemaker activity in the maturation of pain and sensorimotor networks in the central nervous system.

Funding information:
  • NIDCD NIH HHS - T32-DC00046(United States)

Cadherin-8 expression, synaptic localization, and molecular control of neuronal form in prefrontal corticostriatal circuits.

  • Friedman LG
  • J. Comp. Neurol.
  • 2015 Jan 1

Literature context:


Abstract:

Neocortical interactions with the dorsal striatum support many motor and executive functions, and such underlying functional networks are particularly vulnerable to a variety of developmental, neurological, and psychiatric brain disorders, including autism spectrum disorders, Parkinson's disease, and Huntington's disease. Relatively little is known about the development of functional corticostriatal interactions, and in particular, virtually nothing is known of the molecular mechanisms that control generation of prefrontal cortex-striatal circuits. Here, we used regional and cellular in situ hybridization techniques coupled with neuronal tract tracing to show that Cadherin-8 (Cdh8), a homophilic adhesion protein encoded by a gene associated with autism spectrum disorders and learning disability susceptibility, is enriched within striatal projection neurons in the medial prefrontal cortex and in striatal medium spiny neurons forming the direct or indirect pathways. Developmental analysis of quantitative real-time polymerase chain reaction and western blot data show that Cdh8 expression peaks in the prefrontal cortex and striatum at P10, when cortical projections start to form synapses in the striatum. High-resolution immunoelectron microscopy shows that Cdh8 is concentrated at excitatory synapses in the dorsal striatum, and Cdh8 knockdown in cortical neurons impairs dendritic arborization and dendrite self-avoidance. Taken together, our findings indicate that Cdh8 delineates developing corticostriatal circuits where it is a strong candidate for regulating the generation of normal cortical projections, neuronal morphology, and corticostriatal synapses.

Huntingtin is required for normal excitatory synapse development in cortical and striatal circuits.

  • McKinstry SU
  • J. Neurosci.
  • 2014 Jul 9

Literature context:


Abstract:

Huntington's disease (HD) is a neurodegenerative disease caused by the expansion of a poly-glutamine (poly-Q) stretch in the huntingtin (Htt) protein. Gain-of-function effects of mutant Htt have been extensively investigated as the major driver of neurodegeneration in HD. However, loss-of-function effects of poly-Q mutations recently emerged as potential drivers of disease pathophysiology. Early synaptic problems in the excitatory cortical and striatal connections have been reported in HD, but the role of Htt protein in synaptic connectivity was unknown. Therefore, we investigated the role of Htt in synaptic connectivity in vivo by conditionally silencing Htt in the developing mouse cortex. When cortical Htt function was silenced, cortical and striatal excitatory synapses formed and matured at an accelerated pace through postnatal day 21 (P21). This exuberant synaptic connectivity was lost over time in the cortex, resulting in the deterioration of synapses by 5 weeks. Synaptic decline in the cortex was accompanied with layer- and region-specific reactive gliosis without cell loss. To determine whether the disease-causing poly-Q mutation in Htt affects synapse development, we next investigated the synaptic connectivity in a full-length knock-in mouse model of HD, the zQ175 mouse. Similar to the cortical conditional knock-outs, we found excessive excitatory synapse formation and maturation in the cortices of P21 zQ175, which was lost by 5 weeks. Together, our findings reveal that cortical Htt is required for the correct establishment of cortical and striatal excitatory circuits, and this function of Htt is lost when the mutant Htt is present.

Funding information:
  • NIGMS NIH HHS - R01 GM072881(United States)

Conditional viral tracing reveals that steroidogenic factor 1-positive neurons of the dorsomedial subdivision of the ventromedial hypothalamus project to autonomic centers of the hypothalamus and hindbrain.

  • Lindberg D
  • J. Comp. Neurol.
  • 2013 Oct 1

Literature context:


Abstract:

Excitation of neurons in the ventromedial hypothalamus (VMH), especially those residing in the dorsomedial part of the nucleus (VMHdm), evokes sympathetic nervous system (SNS) outflow, modulating a number of physiological functions including feeding and blood glucose homeostasis. However, the anatomical basis of VMH-mediated SNS activation has thus far proved elusive. To understand how VMH neurons exercise output functions and describe an anatomical link between these neurons and the SNS, we identified downstream neural targets of the VMHdm by injecting an adenoviral vector encoding Cre recombinase (Cre)-regulated farnesylated green fluorescent protein (GFPf ) into the VMHdm of mice that express Cre in neurons expressing the VMH-specific transcription factor steroidogenic factor 1 (SF1). We confirm previously described projection patterns of the VMHdm and report the existence of a formerly unidentified projection pathway to a number of autonomic centers in the brainstem. These VMH efferents travel caudally through the periaqueductal gray (PAG) and then ventrally through the lateral lemniscus to the ventral surface of the brain, where they eventually reach caudal autonomic centers including the C1 catecholamine cell group of the rostral ventrolateral medulla (RVLM) and the nucleus of the solitary tract (NTS), where VMH efferents make close contacts with catecholaminergic neurons. We also found that VMHdm fibers reach a number of brainstem areas, including the retrotrapezoid nucleus (RTN), which are important in regulating respiration. Thus, the present study indicates that the VMH may modulate sympathetic and autonomic activity via synaptic contacts in the RTN, NTS, and RVLM and provides significant anatomical evidence to support a role of the VMH in respiratory regulation.

Funding information:
  • NHGRI NIH HHS - U54 HG004576(United States)

Axon diversity of lamina I local-circuit neurons in the lumbar spinal cord.

  • Szucs P
  • J. Comp. Neurol.
  • 2013 Aug 15

Literature context:


Abstract:

Spinal lamina I is a key area for relaying and integrating information from nociceptive primary afferents with various other sources of inputs. Although lamina I projection neurons have been intensively studied, much less attention has been given to local-circuit neurons (LCNs), which form the majority of the lamina I neuronal population. In this work the infrared light-emitting diode oblique illumination technique was used to visualize and label LCNs, allowing reconstruction and analysis of their dendritic and extensive axonal trees. We show that the majority of lamina I neurons with locally branching axons fall into the multipolar (with ventrally protruding dendrites) and flattened (dendrites limited to lamina I) somatodendritic categories. Analysis of their axons revealed that the initial myelinated part gives rise to several unmyelinated small-diameter branches that have a high number of densely packed, large varicosities and an extensive rostrocaudal (two or three segments), mediolateral, and dorsoventral (reaching laminae III-IV) distribution. The extent of the axon and the occasional presence of long, solitary branches suggest that LCNs may also form short and long propriospinal connections. We also found that the distribution of axon varicosities and terminal field locations show substantial heterogeneity and that a substantial portion of LCNs is inhibitory. Our observations indicate that LCNs of lamina I form intersegmental as well as interlaminar connections and may govern large numbers of neurons, providing anatomical substrate for rostrocaudal "processing units" in the dorsal horn.

Funding information:
  • NIGMS NIH HHS - F32GM095213-01(United States)

Development of the principal nucleus trigeminal lemniscal projections in the mouse.

  • Kivrak BG
  • J. Comp. Neurol.
  • 2013 Feb 1

Literature context:


Abstract:

The principal sensory (PrV) nucleus-based trigeminal lemniscus conveys whisker-specific neural patterns to the ventroposteromedial (VPM) nucleus of the thalamus and subsequently to the primary somatosensory cortex. Here we examined the perinatal development of this pathway with carbocyanine dye labeling in embryonic and early postnatal mouse brains. We developed a novel preparation in which the embryonic hindbrain and the diencephalon are flattened out, allowing a birds-eye view of the PrV lemniscus in its entirety. For postnatal brains we used another novel approach by sectioning the brain along an empirically determined oblique horizontal angle, again preserving the trigeminothalamic pathway. PrV neurons are born along the hindbrain ventricular zone and migrate radially for a short distance to coalesce into a nucleus adjacent to the ascending trigeminal tract. During migration of the spindle-shaped cell bodies, slender axonal processes grow along the opposite direction towards the floor plate. As early as embryonic day (E) 11, pioneering axons tipped with large growth cones cross the ventral midline and immediately make a right angle turn. By E13 many PrV axons form fascicles crossing the midline and follow a rostral course. PrV axons reach the midbrain by E15 and the thalamus by E17. While the target recognition and invasion occurs prenatally, organization of PrV axon terminals into whisker-specific rows and patches takes place during the first 4 postnatal (P) days. Initially diffuse and exuberant projections in the VPM at P1 coalesce into row and whisker specific terminal zones by P4.

Funding information:
  • Canadian Institutes of Health Research - MOP-94856(Canada)

Large GABAergic neurons form a distinct subclass within the mouse dorsal cortex of the inferior colliculus with respect to intrinsic properties, synaptic inputs, sound responses, and projections.

  • Geis HR
  • J. Comp. Neurol.
  • 2013 Jan 1

Literature context:


Abstract:

Neurons in the inferior colliculus (IC) show a remarkable diversity in their responses to sound, but it has been difficult to relate these responses to their morphology. Large cells, which are found in all subdivisions of the IC, may form an exception. We found that large neurons of the mouse dorsal cortex of the IC were GABAergic and were contacted by vesicular glutamate transporter 2-containing somatic terminals, as previously observed for the rat IC. Large cells, which were targeted under two-photon guidance, typically had a low input resistance in comparison with the other cells in the dorsal cortex of the IC. Large cells received short-latency excitatory inputs and had short first-spike latencies. These excitatory inputs were often followed by long-latency inhibitory postsynaptic potentials. In four cells, it was possible to reconstruct the ascending axon following labeling with biocytin. We found evidence that they projected to both the ventral and the dorsal divisions of the medial geniculate body of the thalamus, but they also branched off large collaterals while passing through the brachium of the IC. Our data indicate that, owing to their somatic glutamatergic inputs, large GABAergic tectothalamic projection neurons can generate short-latency, well-timed, feed-forward inhibition, which affects not only the thalamus, but also other ascending nuclei. Their remarkably homogeneous properties, which generally differed from those of the other cells in the dorsal cortex of the IC, suggest that large neurons form a distinct subclass within the dorsal cortex of the IC.

Funding information:
  • Biotechnology and Biological Sciences Research Council - (United Kingdom)
  • NEI NIH HHS - R01 EY018005(United States)

Pre-Bötzinger complex receives glutamatergic innervation from galaninergic and other retrotrapezoid nucleus neurons.

  • Bochorishvili G
  • J. Comp. Neurol.
  • 2012 Apr 1

Literature context:


Abstract:

The retrotrapezoid nucleus (RTN) contains CO(2) -responsive neurons that regulate breathing frequency and amplitude. These neurons (RTN-Phox2b neurons) contain the transcription factor Phox2b, vesicular glutamate transporter 2 (VGLUT2) mRNA, and a subset contains preprogalanin mRNA. We wished to determine whether the terminals of RTN-Phox2b neurons contain galanin and VGLUT2 proteins, to identify the specific projections of the galaninergic subset, to test whether RTN-Phox2b neurons contact neurons in the pre-Bötzinger complex, and to identify the ultrastructure of these synapses. The axonal projections of RTN-Phox2b neurons were traced by using biotinylated dextran amine (BDA), and many BDA-ir boutons were found to contain galanin immunoreactivity. RTN galaninergic neurons had ipsilateral projections that were identical with those of this nucleus at large: the ventral respiratory column, the caudolateral nucleus of the solitary tract, and the pontine Kölliker-Fuse, intertrigeminal region, and lateral parabrachial nucleus. For ultrastructural studies, RTN-Phox2b neurons (galaninergic and others) were transfected with a lentiviral vector that expresses mCherry almost exclusively in Phox2b-ir neurons. After spinal cord injections of a catecholamine neuron-selective toxin, there was a depletion of C1 neurons in the RTN area; thus it was determined that the mCherry-positive terminals located in the pre-Bötzinger complex originated almost exclusively from the RTN-Phox2b (non-C1) neurons. These terminals were generally VGLUT2-immunoreactive and formed numerous close appositions with neurokinin-1 receptor-ir pre-Bötzinger complex neurons. Their boutons (n = 48) formed asymmetric synapses filled with small clear vesicles. In summary, RTN-Phox2b neurons, including the galaninergic subset, selectively innervate the respiratory pattern generator plus a portion of the dorsolateral pons. RTN-Phox2b neurons establish classic excitatory glutamatergic synapses with pre-Bötzinger complex neurons presumed to generate the respiratory rhythm.

Funding information:
  • NHLBI NIH HHS - R01HL110737-01(United States)

Synaptotagmins I and II in the developing rat auditory brainstem: Synaptotagmin I is transiently expressed in glutamate-releasing immature inhibitory terminals.

  • Cooper AP
  • J. Comp. Neurol.
  • 2011 Aug 15

Literature context:


Abstract:

The lateral superior olive (LSO), a nucleus in the auditory brainstem, computes interaural intensity differences for sound localization by comparing converging excitatory and inhibitory inputs that carry tonotopically matched information from the two ears. Tonotopic refinement in the inhibitory projection pathway from the medial nucleus of the trapezoid body (MNTB) is known to be established during the first postnatal week in rats. During this period, immature MNTB terminals in the LSO contain vesicular transporters for both inhibitory and excitatory amino acids and release glutamate. The primary Ca(2+) sensors for vesicular release in the CNS are understood to be synaptotagmins, and in adult auditory brainstem synaptotagmin 2 is the predominant synaptotagmin. We asked here whether a different Ca(2+) sensor might be expressed in the immature auditory brainstem. We have found that synaptotagmin 1 is indeed expressed transiently in the immature auditory brainstem, most highly in those areas that receive glutamate-releasing immature inhibitory inputs from the MNTB, and that during the first postnatal week synaptotagmin 1 co-localizes with the vesicular glutamate transporter VGLUT3, a marker of glutamate-releasing immature inhibitory terminals from the MNTB. We suggest that immature MNTB terminals may contain two populations of synaptic vesicles, one expressing the vesicular inhibitory amino acid transporter together with synaptotagmin 2 and another expressing VGLUT3 together with synaptotagmin 1. Because Ca(2+) sensing is an important determinant of release properties for the presynaptic terminal, differential expression of the synaptotagmins might allow the differential release of excitatory and inhibitory neurotransmitters in response to differing patterns of neural activity.

Funding information:
  • Biotechnology and Biological Sciences Research Council - BB/C507253/1(United Kingdom)

SynCAM1 expression correlates with restoration of central synapses on spinal motoneurons after two different models of peripheral nerve injury.

  • Zelano J
  • J. Comp. Neurol.
  • 2009 Dec 10

Literature context:


Abstract:

SynCAM1 and neuroligins (NLGs) are adhesion molecules that govern synapse formation in vitro. In vivo, the molecules are expressed during synaptogenesis, and altered NLG function is linked to synapse dysfunction in autism. Less is known about SynCAM1 and NLGs in adult synapse remodeling. CNS synapse elimination occurs after peripheral nerve injury, which causes a transient decrease in synapse number on spinal motoneurons. Here we have studied the expression of SynCAM1 and NLGs in relation to changes in synaptic covering on spinal motoneurons. We performed sciatic nerve transection (SNT) or crush (SNC), axotomy models that result in poor or good conditions for axon regeneration, respectively. The two lesions resulted in similar synapse elimination and in poor (SNT) and good (SNC) return of synapses after 70 days. Functional recovery was good after SNC but absent after SNT. SynCAM1 mRNA decreased after 14 days in both models and was restored 70 days after SNC, but not after SNT. NLG2 and -3 mRNAs decreased to a smaller degree after SNC than after SNT. Synaptophysin immunoreactivity correlated with SynCAM1 mRNA 70 days after SNT and NLG2 mRNA 70 days after SNC. Surprisingly, an inverse correlation was seen between NLG3 mRNA and Vglut2, a marker for excitatory synapses, 70 days after SNT. We conclude that 1) SynCAM1 mRNA levels seem to reflect the loss and restoration of synapses on motoneurons, 2) down-regulation of NLGs is not a prerequisite for synapse elimination, and 3) expression of SynCAM1 and NLGs is regulated by different mechanisms during regeneration.