bluuebunny/crossref_metadata_2025 huggingface dataset:
Experimental Study on Impact Crushing of Granite Particles
Authors: Yang Jian-hong , Chen Qi , Zhou Jian-hua , Fang Huai-ying
Published: April 2018

To study the specific influences of production parameters and structural parameters on a vertical shaft impact crusher (VSI), a pneumatic impact crushing device was designed and used to study the influence of different impact velocities, impact angles, and initial particle sizes on the crushing effect for granite particles. The resulting grains size distribution (GSD) and the actual crushing ratio (ACR) were determined via mechanical sieving and image processing, where the experimental results showed that with increasing initial impact velocity the degree of fragmentation of the granite particles increased. However, when the impact velocity increased beyond a certain amount, the trend became more moderate. Further, there existed an optimum original particle size that allowed the degree of fragmentation of the particle to obtain a maximum value. Further, as the initial impact angle of the particles increased, the degree of fragmentation of granite particles exhibited a downward trend. Through the experimental study herein, it is found that the actual production parameters and structural parameters have significant effects on the impact crushing. According to these results, we can obtain valuable routines to guide the optimal design of the VSI.
Neprilysin Gene Transfer Reduces Human Amyloid Pathology in Transgenic Mice
Authors: Marr Robert A. , Rockenstein Edward , Mukherjee Atish , Kindy Mark S. , Hersh Louis B. , Gage Fred H. , Verma Inder M. , Masliah Eliezer
Published: April 2018

The degenerative process of Alzheimer's disease is linked to a shift in the balance between amyloid-β (Aβ) production, clearance, and degradation. Neprilysin has recently been implicated as a major extracellular Aβ degrading enzyme in the brain. However, there has been no direct demonstration that neprilysin antagonizes the deposition of amyloid-βin vivo. To address this issue, a lentiviral vector expressing human neprilysin (Lenti-Nep) was tested in transgenic mouse models of amyloidosis. We show that unilateral intracerebral injection of Lenti-Nep reduced amyloid-β deposits by half relative to the untreated side. Furthermore, Lenti-Nep ameliorated neurodegenerative alterations in the frontal cortex and hippocampus of these transgenic mice. These data further support a role for neprilysin in regulating cerebral amyloid deposition and suggest that gene transfer approaches might have potential for the development of alternative therapies for Alzheimer's disease.
Stochastic Resonance within the Somatosensory System: Effects of Noise on Evoked Field Potentials Elicited by Tactile Stimuli
Authors: Manjarrez Elı́as , Rojas-Piloni Gerardo , Méndez Ignacio , Flores Amira
Published: April 2018

Stochastic resonance (SR) is commonly understood to be the enhancement, by noise, of the response of a system to a weak input signal. The aim of this study was to demonstrate the occurrence of SR in spinal and cortical evoked field potentials (EFPs) elicited by periodic tactile stimuli in the anesthetized cat. The electrodes were positioned in spinal and cortical somatosensory regions in which the largest negative EFPs were detected. The periodic tactile stimuli consisted of local skin displacements on the central pad of the hindpaw. Two series of experiments were performed. First, periodic tactile stimuli and the noisy tactile stimuli were applied with the same indenter. Second, noisy tactile stimuli were applied with an additional indenter placed on the glabrous skin of the third hindpaw digit. This last protocol ensured that the signal and noise were mixed not in the skin but in the somatosensory regions of the CNS. All cats showed distinct SR behavior at the spinal and cortical stages of the sensory encoding. Such SR was abolished in the cortical but not in the spinal recording after sectioning of the dorsal columns and the ipsilateral dorsolateral funiculus. This suggests that the spinal neurons may also contribute to the SR observed at the cortical level. To the best of our knowledge, this is the first documented evidence that such a remarkable phenomenon embodies electrical processes of the spinocortical somatosensory system itself.
GABABReceptors Inhibit Dendrodendritic Transmission in the Rat Olfactory Bulb
Authors: Isaacson Jeffry S. , Vitten Harald
Published: April 2018

In the mammalian olfactory bulb, mitral cell dendrites release glutamate onto the dendritic spines of granule cells, which in turn release GABA back onto mitral dendrites. This local synaptic circuit forms the basis for reciprocal dendrodendritic inhibition mediated by ionotropic GABAAreceptors in mitral cells. Surprisingly little is known about neurotransmitter modulation of dendrodendritic signaling in the olfactory bulb. In this study, we examine whether metabotropic GABABreceptors modulate dendrodendritic signaling between mitral and granule cells. We find that the selective GABABagonist baclofen reduces mitral cell recurrent inhibition mediated by dendrodendritic synapses. GABABreceptor activation causes only a weak inhibition of field EPSCs in the external plexiform layer and only slightly reduces glutamate-mediated mitral cell self-excitation. Although GABABreceptors depress mitral cell glutamate release only weakly, baclofen causes a marked reduction in the amplitude of granule-cell-evoked, GABAA-mediated IPSCs in mitral cells. In addition to reducing the amplitude of granule-cell-evoked IPSCs, baclofen causes a change from paired-pulse depression to paired-pulse facilitation, suggesting that GABABreceptors modulate GABA release from granule cells. To explore the mechanism of action of GABABreceptors further, we show that baclofen inhibits high-voltage-activated calcium currents in granule cells. Together, these findings suggest that GABABreceptors modulate dendrodendritic inhibition primarily by inhibiting granule cell calcium channels and reducing the release of GABA. Furthermore, we show that endogenous GABA regulates the strength of dendrodendritic inhibition via the activation of GABABautoreceptors.
Neuronal Glutamate Uptake Contributes to GABA Synthesis and Inhibitory Synaptic Strength
Authors: Mathews Gregory C. , Diamond Jeffrey S.
Published: April 2018

Neurons must maintain a supply of neurotransmitter in their presynaptic terminals to fill synaptic vesicles. GABA is taken up into inhibitory terminals by transporters or is synthesized from glutamate by glutamic acid decarboxylase. Here we report that glutamate transporters supply GABAergic terminals in the hippocampus with glutamate, which is then used to synthesize GABA for filling synaptic vesicles. Glutamate transporter antagonists reduced IPSC and miniature IPSC (mIPSC) amplitudes, consistent with a reduction in the amount of GABA packaged into each synaptic vesicle. This reduction occurred rapidly and independently of synaptic activity, suggesting that modulation of vesicular GABA content does not require vesicle release and refilling. Raising extracellular glutamate levels increased mIPSC amplitudes by enhancing glutamate uptake and, consequently, GABA synthesis. These results indicate that neuronal glutamate transporters strengthen inhibitory synapses in response to extracellular glutamate. This modulation appears to occur under normal conditions and may constitute a negative feedback mechanism to combat hyperexcitability.
Kv3 Potassium Conductance is Necessary and Kinetically Optimized for High-Frequency Action Potential Generation in Hippocampal Interneurons
Authors: Lien Cheng-Chang , Jonas Peter
Published: April 2018

Kv3 channels are thought to be essential for the fast-spiking (FS) phenotype in GABAergic interneurons, but how these channels confer the ability to generate action potentials (APs) at high frequency is unknown. To address this question, we developed a fast dynamic-clamp system (≈50 kHz) that allowed us to add a Kv3 model conductance to CA1 oriens alveus (OA) interneurons in hippocampal slices. Selective pharmacological block of Kv3 channels by 0.3 mm4-aminopyridine or 1 mmtetraethylammonium ions led to a marked broadening of APs during trains of short stimuli and a reduction in AP frequency during 1 sec stimuli. The addition of artificial Kv3 conductance restored the original AP pattern. Subtraction of Kv3 conductance by dynamic clamp mimicked the effects of the blockers. Application of artificial Kv3 conductance also led to FS in OA interneurons after complete K+channel block and even induced FS in hippocampal pyramidal neurons in the absence of blockers. Adding artificial Kv3 conductance with altered deactivation kinetics revealed a nonmonotonic relationship between mean AP frequency and deactivation rate, with a maximum slightly above the original value. Insertion of artificial Kv3 conductance with either lowered activation threshold or inactivation also led to a reduction in the mean AP frequency. However, the mechanisms were distinct. Shifting the activation threshold induced adaptation, whereas adding inactivation caused frequency-dependent AP broadening. In conclusion, Kv3 channels are necessary for the FS phenotype of OA interneurons, and several of their gating properties appear to be optimized for high-frequency repetitive activity.
The EGL-21 Carboxypeptidase E Facilitates Acetylcholine Release atCaenorhabditis elegansNeuromuscular Junctions
Authors: Jacob Tija C. , Kaplan Joshua M .
Published: April 2018

Proneuropeptides are packaged into dense-core vesicles in which they are processed into active peptides by copackaged enzymes. Proprotein convertases (PCs) cleave precursors after dibasic residues, and carboxypeptidases remove basic residues from the C terminals. We show here that theCaenorhabditis elegans egl-21gene encodes a protein that is very similar to carboxypeptidase E (CPE) and is broadly expressed in the nervous system. Mutants lacking eitheregl-21CPE oregl-3, which encodes theC. elegansortholog of PC type 2 (PC2), were defective for processing endogenously expressed FMRFamide (Phe-Met-Arg-Phe-NH2)-related peptides (FaRPs). Mutants lacking theunc-104kinesin motor protein were defective for anterograde movement of dense-core vesicle components, includingegl-3PC2,egl-21CPE, and FaRPs. We provide evidence thategl-3PC2 andegl-21CPE mutants have diminished acetylcholine release at neuromuscular junctions (NMJs). Taken together, these results suggest thategl-21CPE andegl-3PC2 process endogenous neuropeptides that facilitate acetylcholine release atC. elegansNMJs.
Resting Potential and Submembrane Calcium Concentration of Inner Hair Cells in the Isolated Mouse Cochlea Are Set by KCNQ-Type Potassium Channels
Authors: Oliver Dominik , Knipper Marlies , Derst Christian , Fakler Bernd
Published: April 2018

Cochlear inner hair cells (IHCs) transduce sound-induced vibrations into a receptor potential (RP) that controls afferent synaptic activity and, consequently, frequency and timing of action potentials in the postsynaptic auditory neurons. The RP is thought to be shaped by the two voltage-dependent K+conductances,IK,fandIK,s, that are carried by large-conductance Ca2+- and voltage-dependent K+(BK)- and KV-type K+channels. Using whole-cell voltage-clamp recordings in the acutely isolated mouse cochlea, we show that IHCs display an additional K+current that is active at the resting membrane potential (−72 mV) and deactivates on hyperpolarization. It is potently blocked by the KCNQ-channel blockers linopirdine and XE991 but is insensitive to tetraethylammonium and 4-aminopyridine, which inhibitIK,fandIK,s, respectively. Single-cell PCR and immunocytochemistry showed expression of the KCNQ4 subunit in IHCs. In current-clamp experiments, block of the KCNQ current shifted the resting membrane potential by ∼7 to −65 mV and led to a significant activation of BK channels. Using BK channels as an indicator for submembrane intracellular Ca2+concentration ([Ca2+]i), it is shown that the shift in IHC resting potential observed after block of the KCNQ channels leads to an increase in [Ca2+]ito values ≥1 μm. In conclusion, KCNQ channels set the resting membrane potential of IHCs in the isolated organ of Corti and thus maintain [Ca2+]iat low levels. Destabilization of the resting potential and increase in [Ca2+]i, as may result from impaired KCNQ4 function in IHCs, provide a novel explanation for the progressive hearing loss (DFNA2) observed in patients with defective KCNQ4 genes.
Mutant Huntingtin Causes Context-Dependent Neurodegeneration in Mice with Huntington's Disease
Authors: Yu Zhao-Xue , Li Shi-Hua , Evans Joy , Pillarisetti Ajay , Li He , Li Xiao-Jiang
Published: April 2018

Huntington's disease (HD) mouse models that express N-terminal huntingtin fragments show rapid disease progression and have been used for developing therapeutics. However, light microscopy reveals no significant neurodegeneration in these mice. It remains unclear how mutant huntingtin induces neurodegeneration. Using caspase staining, terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling, and electron microscopy, we observed that N171-82Q mice, which express the first 171 aa of mutant huntingtin, displayed more degenerated neurons than did other HD mouse models. The neurodegeneration was also evidenced by increased immunostaining for glial fibrillary acidic protein and ultrastructural features of apoptosis. R6/2 mice, which express exon 1 of mutant huntingtin, showed dark, nonapoptotic neurons and degenerated mitochondria associated with mutant huntingtin. In HD repeat knock-in mice (HdhCAG150), which express full-length mutant huntingtin, degenerated cytoplasmic organelles were found in both axons and neuronal cell bodies in association with mutant huntingtin that was not labeled by an antibody to huntingtin amino acids 342–456. Transfection of cultured cells with mutant huntingtin revealed that an N-terminal huntingtin fragment (amino acids 1–208 plus a 120 glutamine repeat) caused a greater increase in caspase activity than did exon 1 huntingtin and longer huntingtin fragments. These results suggest that context-dependent neurodegeneration in HD may be mediated by different N-terminal huntingtin fragments. In addition, this study has identified neurodegenerative markers for the evaluation of therapeutic treatments in HD mouse models.
Responses of Neurons in the Middle Temporal Visual Area After Long-Standing Lesions of the Primary Visual Cortex in Adult New World Monkeys
Authors: Collins Christine E. , Lyon David C. , Kaas Jon H.
Published: April 2018

The retinotopic organization of the middle temporal visual area (MT) was determined in six adult owl monkeys and one adult marmoset 69 d to 10 months after lesions of the dorsolateral primary visual cortex (V1). The lesions removed were limited to extensive parts of the representation of the lower visual quadrant in V1. Microelectrodes were used to record from neurons at numerous sites in MT to determine whether parts of MT normally devoted to the lower visual quadrant (1) were unresponsive to visual stimuli, (2) acquired responsiveness to inputs from intact portions of V1, or (3) became responsive to some other visually driven input such as a relay from the superior colliculus via the pulvinar to MT. All monkeys (n = 6) with moderate to moderately large lesions had unresponsive portions of MT even after 10 months of recovery. These unresponsive regions were retinotopically equivalent to the removed parts of V1 in normal animals. Thus, there was no evidence for an alternative source of activation. In addition, these results indicate that any retinotopic reorganization of MT based on inputs from intact portions of V1 was not extensive, yet neurons near the margins of responsive cortex may have acquired new receptive fields, and the smallest 5° lesion of V1 failed to produce an unresponsive zone. Deprived portions of MT were not remarkably changed in histological appearance in cytochrome oxidase, Nissl, andWisteria floribundaagglutinin preparations. Nevertheless, some reduction in myelin staining and other histological changes were suggested. We conclude that MT is highly dependent on V1 for activation in these monkeys, and alternative sources do not become effective over months when normal activation is absent. Additionally, remaining V1 inputs have only a limited capacity to expand their activation territory into deprived portions of MT.
Direct cAMP Signaling through G-Protein-Coupled Receptors Mediates Growth Cone Attraction Induced by Pituitary Adenylate Cyclase-Activating Polypeptide
Authors: Guirland Carmine , Buck Kenneth B. , Gibney Jean A. , DiCicco-Bloom Emanuel , Zheng James Q.
Published: April 2018

Developing axons are guided to their appropriate targets by environmental cues through the activation of specific receptors and intracellular signaling pathways. Here we report that gradients of pituitary adenylate cyclase-activating polypeptide (PACAP), a neuropeptide widely expressed in the developing nervous system, induce marked attraction ofXenopusgrowth conesin vitro. PACAP exerted its chemoattractive effects through PAC1, a PACAP-selective G-protein-coupled receptor (GPRC) expressed at the growth cone. Furthermore, the attraction depended on localized cAMP signaling because it was completely blocked either by global elevation of intracellular cAMP levels using forskolin or by inhibition of protein kinase A using specific inhibitors. Moreover, local direct elevation of intracellular cAMP by focal photolysis of caged cAMP compounds was sufficient to induce growth cone attraction. On the other hand, blockade of Ca2+, phospholipase C, or phosphatidyl inositol-3 kinase signaling pathways did not affect PACAP-induced growth cone attraction. Finally, PACAP-induced attraction also involved the Rho family of small GTPases and required local protein synthesis. Taken together, our results establish cAMP signaling as an independent pathway capable of mediating growth cone attraction induced by a physiologically relevant peptide acting through GPCRs. Such a direct cAMP pathway could potentially operate in other guidance systems for the accurate wiring of the nervous system.
Overexpression of Bcl-2 Reduces Sex Differences in Neuron Number in the Brain and Spinal Cord
Authors: Zup Susan L. , Carrier Heather , Waters Elizabeth M. , Tabor Abigail , Bengston Lynn , Rosen Greta J. , Simerly Richard B. , Forger Nancy G.
Published: April 2018

Several sex differences in the nervous system depend on differential cell death during development in males and females. The anti-apoptotic protein, Bcl-2, promotes the survival of many types of neurons during development and in response to injury. To determine whether Bcl-2 might similarly control cell death in sexually dimorphic regions, we compared neuron number in wild-type mice and transgenic mice overexpressing Bcl-2 under the control of a neuron-specific promoter. Three neural areas were examined: the spinal nucleus of the bulbocavernosus (SNB), in which neuron number is greater in males; the retrodorsolateral nucleus (RDLN) of the spinal cord, which exhibits no sex difference in neuron number; and the anteroventral periventricular nucleus (AVPV) of the hypothalamus, in which both overall cell density and the number of tyrosine hydroxylase immunoreactive (TH-ir) neurons are greater in females. Bcl-2 overexpression significantly increased SNB cell number in females, overall cell density of AVPV in males, and RDLN cell number in both sexes. Bcl-2 overexpression did not alter the number of TH-ir neurons in AVPV of males or females. These findings indicate that Bcl-2 can regulate sexually dimorphic cell number in the brain and spinal cord and suggest that Bcl-2 may mediate effects of testosterone on cell survival during neural development. In contrast to the regulation of overall cell density in AVPV, the sex difference in TH cell number apparently is not caused by a Bcl-2-dependent mechanism.
Glutamate Receptor δ2 Subunit in Activity-Dependent Heterologous Synaptic Competition
Authors: Cesa Roberta , Morando Laura , Strata Piergiorgio
Published: April 2018

In the adult cerebellum, the glutamate receptor δ2 subunit (GluRδ2) is selectively targeted to the spines of the distal Purkinje cell dendrites, the spiny branchlets, that are innervated by the parallel fibers. Although GluRδ2 has no known channel function, it is presumed to be involved in the formation and stabilization of these synapses. After block of electrical activity by tetrodotoxin, GluRδ2s appear in the postsynaptic densities of the proximal dendritic spines, which then lose their contact with climbing fibers and become ectopically innervated by parallel fibers. This phenomenon suggests that climbing fiber activity prevents GluRδ2 targeting to proximal dendrites and that GluRδ2s admitted to the postsynaptic density of the spine cause withdrawal of the silent climbing fiber. To test this hypothesis, we studied the distribution of GluRδ2s in the rat cerebellum by immunoelectron microscopy during the recovery period that follows removal of the electrical block, and during the sprouting of climbing fibers that follows subtotal deletion of the parent inferior olivary neurons by administration of the drug 3-acetylpyridine. We found that after removal of the electrical block, the climbing fibers reinnervate proximal spines that bear GluRδ2s and these subunits are successively repressed. Similarly, after subtotal lesion of the inferior olive, reinnervation of denervated Purkinje cells occurs on spines bearing GluRδ2s. Thus, GluRδ2s are not responsible for displacing silent climbing fibers. We propose instead that GluRδ2s are associated with climbing fiber-to-Purkinje cell synapses, during development or at early stages of climbing fiber regeneration or sprouting, and are downregulated during the process of synapse maturation.
Activity-Dependent Autocrine-Paracrine Activation of Neuronal P2Y Receptors
Authors: Moskvina Eugenia , Unterberger Ursula , Boehm Stefan
Published: April 2018

Activation of P2Y receptors by released nucleotides subserves important autocrine-paracrine functions in various non-neural tissues. To investigate how P2Y receptors are activated in a neuronal environment, we used PC12 cells in which nucleotides were found to elicit increases in inositol phosphates via P2Y2and decreases in cAMP via P2Y12receptors. Depolarization of PC12 cells raised inositol phosphates, and blockade of voltage-gated Ca2+channels by Cd2+or degradation of extracellular nucleotides by apyrase prevented this effect. In nondepolarized cells, apyrase did not affect inositol phosphates. Depolarization of PC12 cells also reduced the A2Areceptor-mediated synthesis of cAMP. This effect was again prevented by Cd2+or apyrase, but apyrase enhanced the synthesis of cAMP even in nondepolarized cells. Overexpression of rat P2Y2receptors increased the nucleotide-dependent inositol phosphate accumulation and enhanced the effect of K+depolarization. Nevertheless, apyrase still failed to alter spontaneous inositol phosphate accumulation. Expression of rat P2Y1receptors, in contrast, led to huge increases in spontaneous inositol phosphate accumulation, which was reduced by a receptor antagonist or by apyrase. This increased synthesis of inositol phosphates could not be further enhanced by depolarization or receptor agonists, but when endogenous nucleotides were removed by superfusion, recombinant P2Y1receptors could be activated to mediate an inhibition of M-type K+channels. These results indicate that nucleoside diphosphate-sensitive (P2Y12and P2Y1) receptors are activated by spontaneous nucleotide release, whereas triphosphate-sensitive (P2Y2) receptors require an excess of depolarization-evoked release to become activated.
Different Gating Mechanisms in Glutamate Receptor and K+Channels
Authors: Sobolevsky Alexander I. , Yelshansky Maria V. , Wollmuth Lonnie P.
Published: April 2018

The basic structural features of channel gating in glutamate receptors (GluRs) remain unknown. Here we used covalent modification of substituted cysteines and fast agonist application to study the contribution of the M3 segment in AMPA receptor GluR-A subunits to channel structure and gating. The pattern of accessibility of substituted cysteines to extracellularly applied methanethiosulfonate reagents and the rates of their modification by these reagents, measured in either the presence or absence of glutamate, indicate that M3 forms an α-helix that lines the pore of the channel and is involved in gating-related movements. The voltage dependence of modification rates places the tip of the M2 loop (the Q/R site) close to the middle of M3. All of these results are consistent with pore-forming domains in GluR and K+channels having a similar structure but inverted membrane topology. Nevertheless, GluRs lack a glycine residue at a homologous structural position as the gating hinge glycine in K+channels. Moreover, simultaneous substitution of the only two glycines in M3 of GluR-A with alanines produced channels with gating properties indistinguishable from wild type. Given the unique role of glycines in the flexibility ofα-helices, our results indicate that the M3 segment in GluR does not contain a glycine gating hinge and suggest that, in contrast to the homologous domain in K+channels, M3 is rigid during gating. The different positioning and functional significance of glycines in a key structural domain may represent the basis for the distinct features of gating in GluR and K+channels.
Intracellular Membrane Targeting and Suppression of Ser880Phosphorylation of Glutamate Receptor 2 by the Linker I-Set II Domain of AMPA Receptor-Binding Protein
Authors: Fu Jie , deSouza Sunita , Ziff Edward B.
Published: April 2018

AMPA receptor-binding protein (ABP) is a multi-postsynaptic density-95/discs large/zona occludens (PDZ) protein that binds to the glutamate receptor 2/3 (GluR2/3) subunits of the AMPA receptor and is implicated in receptor membrane anchorage. A palmitoylated form of ABP localizes to spine heads, whereas a nonpalmitoylated form is found in intracellular clusters. Here, we investigate intracellular cluster formation by ABP and the ability of ABP to associate with GluR2 while in these clusters. We show that ABP interacts with intracellular membranes via the ABP linker I (LI)-set II (SII) subdomain, a region consisting of ABP linker 1 and PDZ4, -5, and -6. This suggests that cluster formation results from LI-SII ABP association with the membrane of a vesicular structure. We present evidence that ABP can self-associate at intracellular membrane surfaces via interactions involving SII. ABP in such membrane clusters can bind and retain GluR2 that has trafficked endocytotically from the plasma membrane. Phosphorylation of GluR2 at serine 880, proximal to the ABP binding site, has been implicated by others in the release of ABP from GluR2 and the mobilization of AMPA receptors for trafficking. We show that binding of GluR2 to ABP blocks phosphorylation of serine 880. This suggests that ABP can stabilize its own association with GluR2. We discuss a model in which ABP can form a protein scaffold at a vesicular membrane that is capable of binding GluR2, leading to formation of an intracellular AMPA receptor pool. Receptors in such a pool may contribute to receptor endocytotic and exocytotic trafficking and recycling.
The Meninges Is a Source of Retinoic Acid for the Late-Developing Hindbrain
Authors: Zhang Jinghua , Smith Deborah , Yamamoto Miyuki , Ma Lanhua , McCaffery Peter
Published: April 2018

One general function for retinoic acid (RA) is pattern organization in the CNS. This regulatory factor has an essential role in spinal cord motor neuron and early posterior hindbrain development. In the anterior CNS, however, there is only a limited number of foci of RA synthesis, and less attention has been placed on regions such as the anterior hindbrain where RA synthesizing enzymes are absent. This study shows that a rich source of RA lies around the hindbrain from the RA synthetic enzyme retinaldehyde dehydrogenase-2 (RALDH2) present in the surrounding meninges and mesenchyme by embryonic day 13. RALDH2 is not distributed uniformly throughout the meninges but is restricted to territories over the developing hindbrain, suggesting that RA signaling may be localized to those regions. Further regulation of RA signaling is provided by the presence of a RA sink in the form of the CYP26B1 RA catabolic enzyme expressed in deeper regions of the brain. As a guide to the neural anatomy of hindbrain RA signaling, we used a mouse transgenic for alacZreporter gene driven by a RA response element (RAREhsplacZ) to identify regions of RA signaling. This reporter mouse provides evidence that RA signaling in the hindbrain after embryonic day 13 occurs in the regions of the cerebellum and precerebellar system adjacent to sources of RA, including the inferior olive and the pontine nuclei.
Cellular Location and Circadian Rhythm of Expression of the Biological Clock GenePeriod 1in the Mouse Retina
Authors: Witkovsky Paul , Veisenberger Eleonora , LeSauter Joseph , Yan Lily , Johnson Madeleine , Zhang Dao-Qi , McMahon Douglas , Silver Rae
Published: April 2018

The cellular location and rhythmic expression ofPeriod 1(Per1) circadian clock gene were examined in the retina of aPer1::GFP transgenic mouse. MousePer1(mPer1) RNA was localized to inner nuclear and ganglion cell layers but was absent in the outer nuclear (photoreceptor) layer. Green fluorescent protein (GFP), which was shown to colocalize with PER1 protein, was found in a few subtypes of amacrine neuron, including those containing tyrosine hydroxylase, calbindin, and calretinin, but not in cholinergic amacrine cells. A small subset of ganglion cells also contained GFP immunoreactivity (GFP-IR), but the melanopsin-containing subtype, which projects to the suprachiasmatic nuclei (SCN), lacked GFP-IR. Although the intensity of GFP-IR varied among the populations of amacrine cells at each time point that was examined, both diurnal and circadian rhythms were found for the fraction of neurons showing strong GFP-IR, with peak expression between Zeitgeber/circadian (ZT/CT) times 10 and 14. In SCNs that were examined in the same mice used for the retinal measures, the peak in GFP-IR also occurred at approximately ZT/CT 10. Our results are the first to demonstrate a circadian rhythm of a biological clock component in identified neurons of a mammalian retina.
Motor Dysfunction and Altered Synaptic Transmission at the Parallel Fiber-Purkinje Cell Synapse in Mice Lacking Potassium Channels Kv3.1 and Kv3.3
Authors: Matsukawa Hiroshi , Wolf Alexander M. , Matsushita Shinichi , Joho Rolf H. , Knöpfel Thomas
Published: April 2018

Micelacking both Kv3.1 and both Kv3.3 K+channel alleles display severe motor deficits such as tremor, myoclonus, and ataxic gait. Micelacking one to three alleles at the Kv3.1 and Kv3.3 loci exhibit in an allele dose-dependent manner a modest degree of ataxia. Cerebellar granule cells coexpress Kv3.1 and Kv3.3 K+channels and are therefore candidate neurons that might be involved in these behavioral deficits. Hence, we investigated the synaptic mechanisms of transmission in the parallel fiber-Purkinje cell system. Action potentials of parallel fibers were broader in mice lacking both Kv3.1 and both Kv3.3 alleles and in mice lacking both Kv3.1 and a single Kv3.3 allele compared with those of wild-type mice. The transmission of high-frequency trains of action potentials was only impaired at 200 Hz but not at 100 Hz in mice lacking both Kv3.1 and Kv3.3 genes. However, paired-pulse facilitation (PPF) at parallel fiber-Purkinje cell synapses was dramatically reduced in a gene dose-dependent manner in mice lacking Kv3.1 or Kv3.3 alleles. Normal PPF could be restored by reducing the extracellular Ca2+concentration indicating that increased activity-dependent presynaptic Ca2+influx, at least in part caused the altered PPF in mutant mice. Induction of metabotropic glutamate receptor-mediated EPSCs was facilitated, whereas longterm depression was not impaired but rather facilitated in Kv3.1/Kv3.3 double-knockout mice. These results demonstrate the importance of Kv3 potassium channels in regulating the dynamics of synaptic transmission at the parallel fiber-Purkinje cell synapse and suggest a correlation between short-term plasticity at the parallel fiber-Purkinje cell synapse and motor performance.
Expression of Functional Tyrosine Kinase B Receptors by Rhythmically Active Respiratory Neurons in the Pre-Bötzinger Complex of Neonatal Mice
Authors: Thoby-Brisson Muriel , Cauli Bruno , Champagnat Jean , Fortin Gilles , Katz David M.
Published: April 2018

Genetic loss of brain-derived neurotrophic factor (BDNF) severely disrupts brainstem control of respiratory rhythmogenesis in newborn mice; however, the sites at which BDNF acts to regulate respiratory rhythmogenesis are unknown. Using immunochemical and multiplex RT-PCR analysis in mouse brainstem slices, we report that the BDNF receptor, Tyrosine kinase B (TrkB), is strongly expressed in the pre-Bötzinger complex (PBC), the presumed site for rhythm generation, and colocalizes with neurokinin 1 (NK1), a marker of neurons critical for breathing. The period of the respiratory rhythm generated by PBC neuronsin vitrowas increased by 30% after BDNF treatment (100 ng/ml) and not by nerve growth factor (100 ng/ml) or BDNF (100 ng/ml) in the presence of the tyrosine kinase inhibitor K252a (200 nm). Both synaptic and voltage-dependent properties of PBC neurons were modified by BDNF. Synaptic currents underlying spontaneous rhythmic bursts and glutamate-evoked currents were enhanced by 66 and 33%, respectively. BDNF reduced theIhcurrent amplitude in rhythmic neurons by 46% and shifted its activation curve by -17 mV. All neurons expressing TrkB mRNA (n= 8) also expressed mRNAs for theIhcurrent [hyperpolarization-activated cyclic nucleotide-sensitive cation nonselective channel (HCN1)], and three of four NK1-positive neurons coexpressed TrkB and HCN mRNA. Six of 16 PBC neurons expressed BDNF mRNA, supporting the possibility of autocrine and paracrine actions of BDNF within the respiratory pattern generator. Our data demonstrate that BDNF can modulate respiratory network activity through TrkB signaling in rhythmic PBC neurons.
Insulin-Like Growth Factor (IGF) Signaling through Type 1 IGF Receptor Plays an Important Role in Remyelination
Authors: Mason Jeffrey L. , Xuan Shouhong , Dragatsis Ioannis , Efstratiadis Argiris , Goldman James E.
Published: April 2018

We examined the role of IGF signaling in the remyelination process by disrupting the gene encoding the type 1 IGF receptor (IGF1R) specifically in the mouse brain by Cre-mediated recombination and then exposing these mutants and normal siblings to cuprizone. This neurotoxicant induces a demyelinating lesion in the corpus callosum that is reversible on termination of the insult. Acute demyelination and oligodendrocyte depletion were the same in mutants and controls, but the mutants did not remyelinate adequately. We observed that oligodendrocyte progenitors did not accumulate, proliferate, or survive within the mutant mice, compared with wild type, indicating that signaling through the IGF1R plays a critical role in remyelination via effects on oligodendrocyte progenitors.
NMDA-Dependent Proteolysis of Presynaptic Adhesion Molecule L1 in the Hippocampus by Neuropsin
Authors: Matsumoto-Miyai Kazumasa , Ninomiya Ayako , Yamasaki Hironobu , Tamura Hideki , Nakamura Yukiko , Shiosaka Sadao
Published: April 2018

Synaptic plasticity requires an activity-dependent, rapid, and long-lasting modification of synaptic character, including morphology and coupling strength. Here we show that a serine protease, neuropsin, directly and specifically modifies the synaptic adhesion molecule L1, which was localized to the presynaptic site of the asymmetric synapse in the mouse hippocampus. Increased neural activity triggered the rapid, transient activation of the precursor form of neuropsin in an NMDA receptor-dependent manner. The activated neuropsin immediately cleaved L1 and released a neuropsin-specific extracellular 180 kDa fragment. This neuropsin-specific L1-cleaving system is involved in NMDA receptor-dependent synaptic plasticity, such as the Schaffer collateral long-term potentiation.
Regulatory Domains in the Intergenic Region of the Oxytocin and Vasopressin Genes that Control their Hypothalamus-Specific ExpressionIn Vitro
Authors: Fields Raymond L. , House Shirley B. , Gainer Harold
Published: April 2018

Previous studies of oxytocin (OT) and vasopressin (VP) cell-specific gene expression in the hypothalamus using transgenic mouse and rat models focused attention on the intergenic region (IGR) as the site of critical enhancer elements. In this study, we used organotypic slice-explant cultures of rat hypothalamus asin vitromodels, and particle-mediated gene transfer (biolistics) transfection methods to identify critical DNA sequences in the IGR between the OT and VP genes responsible for hypothalamic-specific gene expression. Reducing the 5′ flanking region in the mouse VP gene from 3.5 kbp to 288 bp did not alter the efficacy of its expression in hypothalamic slices. All subsequent VP constructs were based on this 288 bp VP gene construct with changes made only to the IGR. These studies, which used various constructs with OT and VP promoters driving enhanced green fluorescent protein reporter gene expression, demonstrated that the IGR is necessary for OT and VP gene expression in hypothalamic slicesin vitro. The DNA sequences in the IGR responsible for both OT and VP gene expression were located in a 178 bp domain immediately downstream of exon 3 of the VP gene. In addition, another domain in the IGR, 430 bp immediately downstream of exon 3 of the OT gene, contained a positive regulatory element for OT gene expression in the hypothalamus. Alignment of the DNA sequences in the 178 and 430 bp domains reveals four common sequences (motifs) that may be candidates for the putative enhancers in the IGR that regulate OT and VP gene hypothalamic-specific expression.
Axon Regeneration in Goldfish and Rat Retinal Ganglion Cells: Differential Responsiveness to Carbohydrates and cAMP
Authors: Li Yiming , Irwin Nina , Yin Yuqin , Lanser Marc , Benowitz Larry I.
Published: April 2018

Mammalian retinal ganglion cells (RGCs) do not normally regenerate their axons through an injured optic nerve, but can be stimulated to do so by activating macrophages intraocularly. In a cell culture model of this phenomenon, we found that a small molecule that is constitutively present in the vitreous, acting in concert with macrophage-derived proteins, stimulates mature rat RGCs to regenerate their axons if intracellular cAMP is elevated. In lower vertebrates, RGCs regenerate their axons spontaneouslyin vivo, and in culture, the most potent axon-promoting factor for these cells is a molecule that resembles the small vitreous-derived growth factor from the rat. This molecule was isolated chromatographically and was shown by mass spectrometry to be a carbohydrate. In agreement with this finding,d-mannose proved to be a potent axon-promoting factor for rat RGCs (ED50∼10 μm); this response was cAMP-dependent and was augmented further by macrophage-derived proteins. Goldfish RGCs showed far less selectivity, responding strongly to eitherd-mannose ord-glucose in a cAMP-independent manner. These findings accord well with the success or failure of optic nerves to regenerate in higher and lower vertebratesin vivo. The axon-promoting effects of mannose are highly specific and are unrelated to energy metabolism or glycoprotein synthesis.
Transition from Interictal to Ictal Activity in Limbic NetworksIn Vitro
Authors: Dzhala Volodymyr I. , Staley Kevin J.
Published: April 2018

The transition from brief bursts of synchronous population activity characteristic of interictal epileptiform discharges (IEDs) to more prolonged epochs of population activity characteristic of seizures (ictal-like activity) was recorded in juvenile rat hippocampal-entorhinal cortex slices and hippocampal slices using multiple-site extracellular electrodes. Epileptiform activity was elicited by either increased extracellular potassium or 4-AP. IEDs originated in the CA3 a-b region and spread bidirectionally into CA1 and CA3c dentate gyrus. The transition from IEDs to ictal-like sustained epileptiform activity was reliably preceded by (1) increase in IED propagation velocity, (2) increase in IED secondary afterdischarges and their reverberation between CA3a and CA3c, and (3) shift in the IED initiation area from CA3 a-b to CA3c. Ictal-like sustained network oscillations (10-20 Hz) originated in CA3c and spread to CA1. The pattern of hippocampal ictal-like activity was unaffected by removal of the entorhinal cortex. These findings indicate that interictal and ictal activity can originate in the same neural network, and that the transition from interictal to ictal-like-sustained activity is preceded by predictable alterations in the origin and spread of IEDs. These findings elucidate new targets for investigating the proximate causes, prediction, and treatment of seizures.
Glutamate Decreases Mitochondrial Size and Movement in Primary Forebrain Neurons
Authors: Rintoul Gordon L. , Filiano Anthony J. , Brocard Jacques B. , Kress Geraldine J. , Reynolds Ian J.
Published: April 2018

Mitochondria are essential to maintain neuronal viability. In addition to the generation of ATP and maintenance of calcium homeostasis, the effective delivery of mitochondria to the appropriate location within neurons is also likely to influence their function. In this study we examined mitochondrial movement and morphology in primary cultures of rat forebrain using a mitochondrially targeted enhanced yellow fluorescent protein (mt-eYFP). Mt-eYFP-labeled mitochondria display a characteristic elongated phenotype and also move extensively. Application of glutamate to cultures results in a rapid diminution of movement and also an alteration from elongated to rounded morphology. This effect required the entry of calcium and was mediated by activation of the NMDA subtype of glutamate receptor. Treatment of cultures with an uncoupler or blocking ATP synthesis with oligomycin also stopped movement but did not alter morphology. Interestingly, application of glutamate together with the uncoupler did not prevent the changes in movement or shape but facilitated recovery after washout of the stimuli. This suggests that the critical target for calcium in this paradigm is cytosolic. These studies demonstrate that in addition to altering the bioenergetic properties of mitochondria, neurotoxins can also alter mitochondrial movement and morphology. We speculate that neurotoxin-mediated impairment of mitochondrial delivery may contribute to the injurious effects of neurotoxins.
Dissociating Valence of Outcome from Behavioral Control in Human Orbital and Ventral Prefrontal Cortices
Authors: O'Doherty John , Critchley Hugo , Deichmann Ralf , Dolan Raymond J.
Published: April 2018

The precise role of orbitofrontal cortex (OFC) in affective processing is still debated. One view suggests OFC represents stimulus reward value and supports learning and relearning of stimulus-reward associations. An alternate view implicates OFC in behavioral control after rewarding or punishing feedback. To discriminate between these possibilities, we used event-related functional magnetic resonance imaging in subjects performing a reversal task in which, on each trial, selection of the correct stimulus led to a 70% probability of receiving a monetary reward and a 30% probability of obtaining a monetary punishment. The incorrect stimulus had the reverse contingency. In one condition (choice), subjects had to choose which stimulus to select and switch their response to the other stimulus once contingencies had changed. In another condition (imperative), subjects had simply to track the currently rewarded stimulus. In some regions of OFC and medial prefrontal cortex, activity was related to valence of outcome, whereas in adjacent areas activity was associated with behavioral choice, signaling maintenance of the current response strategy on a subsequent trial. Caudolateral OFC-anterior insula was activated by punishing feedback preceding a switch in stimulus in both the choice and imperative conditions, indicating a possible role for this region in signaling a change in reward contingencies. These results suggest functional heterogeneity within the OFC, with a role for this region in representing stimulus-reward values, signaling changes in reinforcement contingencies and in behavioral control.
Depression of Activity in the Corticospinal Pathway during Human Motor Behavior after Strong Voluntary Contractions
Authors: Petersen Nicolas T. , Taylor Janet L. , Butler Jane E. , Gandevia Simon C.
Published: April 2018

The corticospinal system plays an important role in control of voluntary movements in primates. Recently, we demonstrated that the effectiveness of this system is depressed after maximal exercise. Because the depression was absent after antidromic activation of the motoneurons, we argued that transmission across corticospinal synapses was involved. Here, we explore the possible functional consequences of such a depression. In humans, direct electrical stimulation of axons of corticospinal neurons at the cervicomedullary level evokes motor potentials in elbow flexor muscles. When tested during relaxation after a maximal voluntary contraction (MVC) of the elbow flexors, potentials in biceps brachii and brachioradialis were depressed for ∼90 sec. The potentials were also depressed, although less markedly, when tested during a weak elbow flexion. Brief intermittent MVCs abolished the depression transiently, but during the intervening periods of relaxation, the depression appeared similar to that during continuous relaxation. The depression was greatest during relaxation after a 10 sec MVC and smaller after submaximal contractions. To look for effects of the depression on voluntary activity, we compared bilateral matching weak elbow flexions. After a conditioning 10 sec maximal elbow flexion of one arm, the electromyographic activity produced on that side was reduced relative to the activity on the contralateral side. Our findings support the view that synapses in the corticospinal system are depressed after strong voluntary contractions during both relaxation and activity. Furthermore, this depression can affect the production of voluntary movement.
Modulation of GABAAReceptors by Hydrogen Ions Reveals Synaptic GABA Transient and a Crucial Role of the Desensitization Process
Authors: Mozrzymas Jerzy W. , Żarmowska Ewa D. , Pytel Maria , Mercik Katarzyna
Published: April 2018

Protons are the most ubiquitous and very potent modulators of the biological systems. Hydrogen ions are known to modulate GABAAreceptors (GABAARs), but the mechanism whereby these ions affect IPSCs and the gating of GABAARs is not clear. In the present study we examined the effect of protons on miniature IPSCs (mIPSCs) and found that hydrogen ions strongly affected both their amplitude and time course. To explore the underlying mechanisms with resolution adequate to the time scale of synaptic transmission, we recorded current responses to ultrafast GABA applications at various pH. These experiments revealed that the major effect of protons on GABAAR gating is a strong enhancement of desensitization and binding rates at increasing pH. This analysis also indicated that desensitization rate is the fastest ligand-independent transition in the GABAAR gating scheme. Although proton effects on the time course of mIPSCs and current responses to saturating [GABA] were similar, the pH dependencies of amplitudes were almost opposite. Our quantitative analysis, based on model simulations, indicated that this difference resulted from a much shorter receptor exposure to agonist in the case of mIPSCs. Modeling of IPSCs as current responses to brief exponentially decaying GABA applications was sufficient to reproduce correctly the pH dependence of mIPSCs, and optimal fit was obtained for peak [GABA] of 1.5-3 mmand a clearance time constant of 0.075-0.125 msec. Our analysis indicates that, for these parameters of GABA transient, in control conditions (pH 7.2) mIPSCs are not saturated.