bluuebunny/crossref_metadata_2025 huggingface dataset:
Spectral Integration in the Inferior Colliculus of the Mustached Bat
Authors: Leroy Scott A. , Wenstrup Jeffrey J.
Published: April 2018

Acoustic behaviors including orientation and social communication depend on neural integration of information across the sound spectrum. In many species, spectral integration is performed by combination-sensitive neurons, responding best when distinct spectral elements in sounds are combined. These are generally considered a feature of information processing in the auditory forebrain. In the mustached bat's inferior colliculus (IC), they are common in frequency representations associated with sonar signals but have not been reported elsewhere in this bat's IC or the IC of other species. We examined the presence of combination-sensitive neurons in frequency representations of the mustached bat's IC not associated with biosonar. Seventy-five single-unit responses were recorded with the best frequencies in 10–23 or 32–47 kHz bands. Twenty-six displayed single excitatory tuning curves in one band with no additional responsiveness to a second signal in another band. The remaining 49 responded to sounds in both 10–23 and 32–47 kHz bands, but response types varied. Sounds in the higher band were usually excitatory, whereas sounds in the lower band either facilitated or inhibited responses to the higher frequency signal. Interactions were usually strongest when the higher and lower frequency stimuli were presented simultaneously, but the strength of interactions varied. Over one-third of the neurons formed a distinct subset; they responded most sensitively to bandpass noise, and all were combination sensitive. We suggest that these combination-sensitive interactions are activated by elements of mustached bat social vocalizations. If so, neuronal integration characterizing analysis of social vocalizations in many species occurs in the IC.
Nitric Oxide Modulation of Interleukin-1β-Evoked Intracellular Ca2+Release in Human Astrocytoma U-373 MG Cells and Brain Striatal Slices
Authors: Meini Antonella , Benocci Alberto , Frosini Maria , Sgaragli Gianpietro , Pessina Gianpaolo , Aldinucci Carlo , Youmbi Gisèle Tchuisseu , Palmi Mitri
Published: April 2018

Intracellular Ca2+mobilization and release into mammal CSF plays a fundamental role in the etiogenesis of fever induced by the proinflammatory cytokine interleukin-1β (IL-1β) and other pyrogens. The source and mechanism of IL-1β-induced intracellular Ca2+mobilization was investigated using two experimental models. IL-1β (10 ng/ml) treatment of rat striatal slices preloaded with45Ca2+elicited a delayed (30 min) and sustained increase (125–150%) in spontaneous45Ca2+release that was potentiated byl-arginine (300 μm) and counteracted byN-ω-nitro-l-arginine methyl ester (l-NAME) (1 and 3 mm). The nitric oxide (NO) donors diethylamine/NO complex (sodium salt) (0.3 and 1 mm) and spermine/NO (0.1 and 0.3 mm) mimicked the effect of IL-1β on Ca2+release. IL-1β stimulated tissue cGMP concentration, and dibutyryl cGMP enhanced Ca2+release. The guanyl cyclase inhibitors 1H-[1,2,4]oxadiazole[4,3-a] quinoxalin-1-one (100 μm) and 6-[phenylamino]-5,8 quinolinedione (50 μm) counteracted Ca2+release induced by 2.5 but not 10 ng/ml IL-1β. Ruthenium red (50 μm) and, to a lesser extent, heparin (3 mg/ml) antagonized IL-1β-induced Ca2+release, and both compounds administered together completely abolished this response. Similar results were obtained in human astrocytoma cells in which IL-1β elicited a delayed (30 min) increase in intracellular Ca2+concentration ([Ca2+]i) (402 ± 71.2% of baseline), which was abolished by 1 mml-NAME. These data indicate that the NO/cGMP-signaling pathway is part of the intracellular mechanism transducing IL-1β-evoked Ca2+mobilization in glial and striatal cells and that the ryanodine and the inositol-(1,4,5)-trisphosphate-sensitive Ca2+stores are involved.
Activation of Group II Metabotropic Glutamate Receptors Induces Long-Term Depression of Synaptic Transmission in the Rat Amygdala
Authors: Lin Hui-Ching , Wang Su-Jane , Luo Ming-Zen , Gean Po-Wu
Published: April 2018

An animal model most sensitive for measuring anticipatory anxiety is fear conditioning, which is expressed by an enduring increase in synaptic strength in the amygdala. A converse view predicts that agents that induce long-term depression (LTD) of synaptic efficacy in the amygdala may be useful in the amelioration of stress disorders. In the present study, we show that activation of group II metabotropic glutamate receptor (mGluR II) by (2S,3S,4S)-2-(carboxycyclopropyl) glycine (l-ccg) induces an LTD in the basolateral amygdala neurons. The effect was concentration-dependent with a maximal inhibition of ∼30%. The induction ofl-CCG LTD required concurrent synaptic activity, required presynaptic but not postsynaptic Ca2+increases, and was independent of NMDA receptors.l-CCG LTD was associated with an increase in the ratio of paired-pulse facilitation and was not occluded by low-frequency stimulation-induced LTD, suggesting that these two forms of LTD did not share a common underlying mechanism.

After eliciting LTD withl-CCG, application of isoproterenol increased the synaptic responses back to its original baseline, demonstrating that chemically depressed synapses could be potentiated by another chemical. A selective PKA inhibitor, KT 5720, by its own caused a depression of synaptic transmission and blockedl-CCG LTD, presumably by mimicking and thereby occluding any further depression. Together, these results suggest thatl-CCG LTD is induced by presynaptically mGluR II-mediated inhibition of Ca2+-sensitive adenylyl cyclase, resulting in a decrease in cAMP formation and PKA activation, which leads to a long-lasting decrease in transmitter release.
Unequal Expression of Allelic Kainate Receptor GluR7 mRNAs in Human Brains
Authors: Schiffer Hans H. , Swanson Geoffrey T. , Masliah Elizier , Heinemann Stephen F.
Published: April 2018

We describe here the first example of an exonic polymorphism that affects the primary structure of a human ionotropic glutamate receptor. The human kainate receptor GluR7 gene contains a thymine (T)/guanine (G) nucleotide variation that determines a serine or alanine at position 310 in the extracellular region of GluR7 receptor subunits. Our finding contrasts with a previous report that suggested that GluR7 transcripts were RNA-edited at this site. Whole-cell patch-clamp recordings did not detect differences in receptor activation and desensitization between the human GluR7 receptor isoforms expressed in HEK-293 cells. Analysis of 41 tissue samples obtained from 30 human brains revealed expression level differences between GluR7 alleles expressed in the same brain. The expression level of the allelic GluR7 mRNAs differed in 27 samples from 1.2- to 12.7-fold. Unequal expression level of allelic mRNAs is characteristic for genes that are affected by genomic imprinting or that contain mutations. Genomic imprinting in most cases is conserved between human and mice. However, we did not detect unequal expression of allelic GluR7 mRNAs in mice. Our results are important for future studies that explore a potential role or roles for GluR7 receptors in the brain and for neurological disorders.
The β2aSubunit Is a Molecular Groom for the Ca2+Channel Inactivation Gate
Authors: Restituito S. , Cens T. , Barrere C. , Geib S. , Galas S. , De Waard M. , Charnet P.
Published: April 2018

Ca2+channel inactivation is a key element in controlling the level of Ca2+entry through voltage-gated Ca2+channels. Interaction between the pore-forming α1subunit and the auxiliary β subunit is known to be a strong modulator of voltage-dependent inactivation. Here, we demonstrate that an N-terminal membrane anchoring site (MAS) of the β2asubunit strongly reduces α1A(CaV2.1) Ca2+channel inactivation. This effect can be mimicked by the addition of a transmembrane segment to the N terminus of the β2asubunit. Inhibition of inactivation by β2aalso requires a link between MAS and another important molecular determinant, the β interaction domain (BID). Our data suggest that mobility of the Ca2+channel I–II loop is necessary for channel inactivation. Interaction of this loop with other identified intracellular channel domains may constitute the basis of voltage-dependent inactivation. We thus propose a conceptually novel mechanism for slowing of inactivation by the β2asubunit, in which the immobilization of the channel inactivation gate occurs by means of MAS and BID.
Schwann Cells Are Removed from the Spinal Cord after Effecting Recovery from Paraplegia
Authors: Jasmin Luc , Janni Gabriella , Moallem Theodore M. , Lappi Douglas A. , Ohara Peter T.
Published: April 2018

Remyelination of the CNS is necessary to restore neural function in a number of demyelinating conditions. Schwann cells, the myelinating cells of the periphery, are candidates for this purpose because they have more robust regenerative properties than their central homologs, the oligodendrocytes. Although the ability of Schwann cells to remyelinate the CNS has been demonstrated, their capacity to enter the adult spinal cord in large numbers and effect functional recovery remains uncertain. We used cholera toxin B-subunit conjugated to saporin to demyelinate the rat lumbar spinal cord, remove macroglia, and produce paraplegia. After the removal of oligodendrocyte and astrocyte debris by invading macrophages, there was a spontaneous entry of Schwann cells into the spinal cord, along with axonal remyelination and concomitant functional recovery from paraplegia occurring within 75 d. The Schwann cells appeared to enter the dorsal funiculi via the dorsal root entry zone and the lateral funiculi via rootlets that had become adherent to the lateral spinal cord after the inflammation. In the following weeks, Schwann cell myelin surrounding central axons was progressively replaced by oligodendrocyte myelin without lapse in motor function. Our results show that endogenous Schwann cells can reverse a severe neurological deficit caused by CNS demyelination and enable later oligodendrocyte remyelination.
NBQX Attenuates Excitotoxic Injury in Developing White Matter
Authors: Follett Pamela L. , Rosenberg Paul A. , Volpe Joseph J. , Jensen Frances E.
Published: April 2018

The excitatory neurotransmitter glutamate is released from axons and glia under hypoxic/ischemic conditions.In vitro, oligodendrocytes (OLs) express non-NMDA glutamate receptors (GluRs) and are susceptible to GluR-mediated excitotoxicity. We evaluated the role of GluR-mediated OL excitotoxicity in hypoxic/ischemic white matter injury in the developing brain. Hypoxic/ischemic white matter injury is thought to mediate periventricular leukomalacia, an age-dependent white matter lesion seen in preterm infants and a common antecedent to cerebral palsy. Hypoxia/ischemia in rat pups at postnatal day 7 (P7) produced selective white matter lesions and OL death. Furthermore, OLs in pericallosal white matter express non-NMDA GluRs at P7. Unilateral carotid ligation in combination with hypoxia (6% O2for 1 hr) resulted in selective, subcortical white matter injury with a marked ipsilateral decrease in immature and myelin basic protein-expressing OLs that was also significantly attenuated by 6-nitro-7-sulfamoylbenzo(f)quinoxaline-2,3-dione (NBQX). Intracerebral AMPA demonstrated greater susceptibility to OL injury at P7 than in younger or older pups, and this was attenuated by systemic pretreatment with the AMPA antagonist NBQX. These results indicate a parallel, maturation-dependent susceptibility of immature OLs to AMPA and hypoxia/ischemia. The protective efficacy of NBQX suggests a role for glutamate receptor-mediated excitotoxic OL injury in immature white matterin vivo.
The Surface Accessibility of the Glycine Receptor M2–M3 Loop Is Increased in the Channel Open State
Authors: Lynch Joseph W. , Han Nian-Lin Reena , Haddrill Justine , Pierce Kerrie D. , Schofield Peter R.
Published: April 2018

Mutations in the extracellular M2–M3 loop of the glycine receptor (GlyR) α1 subunit have been shown previously to affect channel gating. In this study, the substituted cysteine accessibility method was used to investigate whether a structural rearrangement of the M2–M3 loop accompanies GlyR activation. All residues from R271C to V277C were covalently modified by both positively charged methanethiosulfonate ethyltrimethylammonium (MTSET) and negatively charged methanethiosulfonate ethylsulfonate (MTSES), implying that these residues form an irregular surface loop. The MTSET modification rate of all residues from R271C to K276C was faster in the glycine-bound state than in the unliganded state. MTSES modification of A272C, L274C, and V277C was also faster in the glycine-bound state. These results demonstrate that the surface accessibility of the M2–M3 loop is increased as the channel transitions from the closed to the open state, implying that either the loop itself or an overlying domain moves during channel activation.
Transport and Localization of the DEG/ENaC Ion Channel BNaC1α to Peripheral Mechanosensory Terminals of Dorsal Root Ganglia Neurons
Authors: Garcı́a-Añoveros Jaime , Samad Tarek A. , Žuvela-Jelaska Ljiljana , Woolf Clifford J. , Corey David P.
Published: April 2018

Mammalian brain sodium channel (BNaC, also known as BNC/ASIC) proteins form acid-sensitive and amiloride-blockable sodium channels that are related to putative mechanosensory channels. Certain BNaC isoforms are expressed exclusively in dorsal root ganglia (DRG) and have been proposed to form the ion channels mediating tissue acidosis-induced pain. With antibody labeling, we find that the BNaC1α isoform is expressed by most large DRG neurons (low-threshold mechanosensors not involved in acid-induced nociception) and few small nociceptor neurons (which include high-threshold mechanoreceptors). BNaC1α is transported from DRG cell bodies to sensory terminals in the periphery, but not to the spinal cord, and is located specifically at specialized cutaneous mechanosensory terminals, including Meissner, Merkel, penicillate, reticular, lanceolate, and hair follicle palisades as well as some intraepidermal and free myelinated nerve endings. Accordingly, BNaC1α channels might participate in the transduction of touch and painful mechanical stimuli.
Spatiotemporal Dynamics of Brain-Derived Neurotrophic Factor mRNA Induction in the Vestibulo-Olivary Network during Vestibular Compensation
Authors: Li Yue Xin , Hashimoto Takanori , Tokuyama Wataru , Miyashita Yasushi , Okuno Hiroyuki
Published: April 2018

Vestibular compensation, which is the behavioral recovery from vestibular dysfunction produced by unilateral labyrinthectomy (UL), is attributed to functional and structural reorganization of neural networks in the central vestibular system. To assess the possible contribution of brain-derived neurotrophic factor (BDNF) to this recovery process, we investigated changes in mRNA expression levels in the central vestibular system after UL. We evaluated BDNF mRNA expression levels by quantitative reverse transcription-PCR andin situhybridization. We found that BDNF mRNA is differentially induced in the medial vestibular nucleus ipsilateral to UL and in the prepositus hypoglossi and inferior olive on the contralateral side. The BDNF mRNA induction lasted for at least 24 hr and returned to the basal expression level within 72 hr after UL. In contrast to BDNF mRNA induction, the expression of an immediate-early gene, c-fos, quickly reached the maximum level at 3 hr and decreased to the basal level within 24 hr after UL. Neither BDNF or c-fosinduction was observed in sham-operated animals. The persistent induction of BDNF after UL temporally corresponded to early behavioral manifestations of vestibular compensation. We further found thattrkB mRNA was expressed in the central vestibular network at high levels, although its expression levels did not change over time after UL. Because BDNF is implicated in regulating synaptic structure and function, these results provide support for the hypothesis that BDNF is involved in neuronal reorganization that allows vestibular compensation.
Progression of Changes in Dopamine Transporter Binding Site Density as a Result of Cocaine Self-Administration in Rhesus Monkeys
Authors: Letchworth Sharon R. , Nader Michael A. , Smith Hilary R. , Friedman David P. , Porrino Linda J.
Published: April 2018

The present study examined the time course of alterations in levels of dopamine transporter (DAT) binding sites that accompany cocaine self-administration using quantitativein vitroreceptor autoradiography with [3H]WIN 35,428. The density of dopamine transporter binding sites in the striatum of rhesus monkeys with 5 d, 3.3 months, or 1.5 years of cocaine self-administration experience was compared with DAT levels in cocaine-naı̈ve control monkeys. Animals in the long-term (1.5 years) exposure group self-administered cocaine at 0.03 mg/kg per injection, whereas the initial (5 d) and chronic (3.3 months) treatment groups were each divided into lower dose (0.03 mg/kg per injection) and higher dose (0.3 mg/kg per injection) groups. Initial cocaine exposure led to moderate decreases in [3H]WIN 35,428 binding sites, with significant changes in the dorsolateral caudate (−25%) and central putamen (−19%) at the lower dose. Longer exposure, in contrast, resulted in elevated levels of striatal binding sites. The increases were most pronounced in the ventral striatum at the level of the nucleus accumbens shell. At the lower dose of the chronic phase, for example, significant increases of 21–42% were measured at the caudal level of the ventral caudate, ventral putamen, olfactory tubercle, and accumbens core and shell. Systematic variation of cocaine dose and drug exposure time demonstrated the importance of these factors in determining the intensity of increased DAT levels. With self-administration of higher doses especially, increases were more intense and included dorsal portions of the striatum so that every region at the caudal level exhibited a significant increase in DAT binding sites (20–54%). The similarity of these findings to previous studies in human cocaine addicts strongly suggest that the increased density of dopamine transporters observed in studies of human drug abusers are the result of the neurobiological effects of cocaine, ruling out confounds such as polydrug abuse, preexisting differences in DAT levels, or comorbid psychiatric conditions.
Deficiency of Growth Hormone-Releasing Hormone Signaling Is Associated with Sleep Alterations in the Dwarf Rat
Authors: Obál Ferenc , Fang Jidong , Taishi Ping , Kacsóh Balint , Gardi Janos , Krueger James M.
Published: April 2018

The somatotropic axis, and particularly growth hormone-releasing hormone (GHRH), is implicated in the regulation of sleep–wake activity. To evaluate sleep in chronic somatotropic deficiency, sleep–wake activity was studied in dwarf (dw/dw) rats that are known to have a defective GHRH signaling mechanism in the pituitary and in normal Lewis rats, the parental strain of thedw/dwrats. In addition, expression of GHRH receptor (GHRH-R) mRNA in the hypothalamus/preoptic region and in the pituitary was also determined by means of reverse transcription-PCR, and GHRH content of the hypothalamus was measured. Hypothalamic/preoptic and pituitary GHRH-R mRNA levels were decreased in thedw/dwrats, indicating deficits in the central GHRHergic transmission. Hypothalamic GHRH content indw/dwrats was also less than that found in Lewis rats. Thedw/dwrats had less spontaneous nonrapid eye movement sleep (NREMS) (light and dark period) and rapid eye movement sleep (REMS) (light period) than did the control Lewis rats. After 4 hr of sleep deprivation, rebound increases in NREMS and REMS were normal in thedw/dwrat. As determined by fast Fourier analysis of the electroencephalogram (EEG), the sleep deprivation-induced enhancements in EEG slow-wave activity in thedw/dwrats were only one-half of the response in the Lewis rats. The results are compared with sleep findings previously obtained in GHRH-deficient transgenic mice. The alterations in NREMS are attributed to the defect in GHRH signaling, whereas the decreases in REMS might result from the growth hormone deficiency in thedw/dwrat.
Ionotropic Histamine Receptors and H2Receptors Modulate Supraoptic Oxytocin Neuronal Excitability and Dye Coupling
Authors: Hatton Glenn I. , Yang Qin Zhao
Published: April 2018

Histaminergic neurons of the tuberomammillary nucleus (TM) project monosynaptically to the supraoptic nucleus (SON). This projection remains intact in our hypothalamic slices and permits investigation of both brief synaptic responses and the effects of repetitively activating this pathway. SON oxytocin (OX) neurons respond to single TM stimuli with fast IPSPs, whose kinetics resemble those of GABAAor glycine receptors. IPSPs were blocked by the Cl−channel blocker picrotoxin, but not by bicuculline or strychnine, and by histamine H2, but not by H1or H3receptor antagonists, suggesting the presence of an ionotropic histamine receptor and the possible nonspecificity of currently used H2antagonists. G-protein mediation of the IPSPs was ruled out using guanosine 5′-O-(2-thiodiphosphate) (GDP-βS), pertussis toxin, and Rp-adenosine 3′,5′-cyclic monophosphothioate triethylamine (Rp-cAMPs), none of which blocked evoked IPSPs. We also investigated the effects of synaptically released histamine on dye coupling and neuronal excitability. One hundred seventy-three OX neurons were Lucifer yellow-injected in horizontal slices. Repetitive TM stimulation (10 Hz, 5–10 min) reduced coupling, an effect blocked by H2, but not by H1or H3, receptor antagonists. Because H2receptors are linked to activation of adenylyl cyclase, TM-stimulated reduction in coupling was blocked by GDP-βS, pertussis toxin, and Rp-cAMPs and was mimicked by 8-bromo-cAMP, 3-isobutyl-1-methylxanthine, and Sp-cAMP. Membrane potentials of OX and vasopressin neurons were hyperpolarized, accompanied by decreased conductances, in response to bath application of 8-bromo-cAMP but not the membrane-impermeable cAMP. These results suggest that synaptically released histamine, in addition to evoking fast IPSPs in OX cells, mediates a prolonged decrease in excitability and uncoupling of the neurons.
Substance P Abolishes the Facilitatory Effect of ATP on Spontaneous Glycine Release in Neurons of the Trigeminal Nucleus Pars Caudalis
Authors: Wang Zhi-Ming , Katsurabayashi Shutaro , Rhee Jeong-Seop , Brodwick Malcolm , Akaike Norio
Published: April 2018

Glycine release was facilitated by the activation of presynaptic ATP receptors (P2X-type) in a preparation of dissociated trigeminal nucleus pars caudalis neurons in which the native synaptic boutons were preserved. The action of ATP was completely blocked by substance P (SP) without alteration of the miniature IPSC (mIPSC) amplitude distribution. SP itself had no effect on mIPSC frequency or amplitude. The inhibitory effect of SP on ATP action was blocked by CP99994, indicating that the SP receptors are of the neurokinin-1 type. The ATP-induced facilitation of the mIPSC frequency was unaffected by Cd2+. Moreover, SP did not inhibit the increase in mIPSC frequency induced high K+application, suggesting that SP did not modulate voltage-dependent calcium channels or subsequent steps in the release process.

KT5720 and phorbol 12-myristate 13-acetate did not block SP action, indicating that neither the cAMP–protein kinase A nor the protein kinase C pathway mediates the SP effects. However, in the presence ofN-(6-aminohexyl)-5-chloro-1-naphthalene sulphonamide (W-7), SP was no longer able to inhibit the ATP-induced stimulation of mIPSC frequency. 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-l-tyrosyl]-4-phenylpiperazine also suppressed the SP action, suggesting that SP modulates P2Xreceptors via a Ca2+/calmodulin-dependent protein kinase II-mediated pathway. In conventional whole-cell mode, the presence of W-7 in the patch pipette did not affect the SP inhibitory action. Thus, SP is not likely to be generating its modulation through the production of a retrograde signal (involving calmodulin) from the postsynaptic cell to the presynaptic boutons.

These results are the first demonstration of the modulation of one presynaptic receptor by another. Because SP inhibits the ATP stimulation of glycine release, SP may play a significant role in hyperalgesia or chronic pain.
Rearrangement of Nicotinic Receptor α Subunits during Formation of the Ligand Binding Sites
Authors: Mitra Mirna , Wanamaker Christian P. , Green William N.
Published: April 2018

Muscle nicotinic acetylcholine receptors (AChRs) are pentamers that contain two α subunits a β, γ (or ε), and δ subunit. In this paper, we have characterized subunit processing and folding events leading to formation of the two AChR ligand binding sites. α subunit residues, 187–199, which are part of overlapping ACh and α-bungarotoxin (Bgt) binding sites on AChRs, were assayed using a monoclonal antibody (mAb) specific for these residues. We found that this region was inaccessible to the mAb early during AChR assembly but became accessible as the first of two Bgt binding sites formed later during assembly, indicating that the region changes conformation as the Bgt binding site appears. Without previous reduction, 20% of the α subunits could be alkylated by bromoacetylcholine bromide as the first ACh binding site formed, which further indicated that the disulfide bond between cysteines 192 and 193 does not form until the first ACh binding site appears soon after Bgt binding site formation. When α subunits were mutated to add a glycosylation site at residue 187, the number of Bgt binding sites increased threefold, AChRs assembled more efficiently, and 2.5-fold more AChRs reached the cell surface. Our results indicate that binding site formation involves a rate-limiting rearrangement of the α subunit that exposes the 187–199 region to the endoplasmic reticulum lumen and determines when cysteines 192 and 193 disulfide bond.
Identification of Amino Acid Residues in GluR1 Responsible for Ligand Binding and Desensitization
Authors: Banke T. G. , Greenwood J. R. , Christensen J. K. , Liljefors T. , Traynelis S. F. , Schousboe A. , Pickering Darryl S.
Published: April 2018

Although GluR1oand GluR3oare homologous at the amino acid level, GluR3odesensitizes approximately threefold faster than GluR1o. By creating chimeras of GluR1oand GluR3oand point amino acid exchanges in their S2 regions, two residues were identified to be critical for GluR1odesensitization: Y716 and the R/G RNA-edited site, R757. With creation of the double-point mutant (Y716F, R757G)GluR1o, complete exchange of the desensitization rate of GluR1oto that of GluR3owas obtained. In addition, both the potency and affinity of the subtype-selective agonist bromohomoibotenic acid were exchanged by the Y716F mutation. A model is proposed of the AMPA receptor binding site whereby a hydrogen-bonding matrix of water molecules plays an important role in determining both ligand affinity and receptor desensitization properties. Residues Y716 in GluR1 and F728 in GluR3 differentially interact with this matrix to affect the binding affinity of some ligands, providing the possibility of developing subtype-selective compounds.
Interleukin-10 Prevents Glutamate-Mediated Cerebellar Granule Cell Death by Blocking Caspase-3-Like Activity
Authors: Bachis Alessia , Colangelo Anna M. , Vicini Stefano , Doe Pylord P. , De Bernardi Maria A. , Brooker Gary , Mocchetti Italo
Published: April 2018

Interleukin-10 (IL-10) has been shown to reduce neuronal degeneration after CNS injury. However, the molecular mechanisms underlying the neuroprotective properties of this cytokine are still under investigation. Glutamate exacerbates secondary injury caused by trauma. Thus, we examined whether IL-10 prevents glutamate-mediated cell death. We used rat cerebellar granule cells in culture because these neurons undergo apoptosis upon exposure to toxic concentrations of glutamate (100–500 μm) or NMDA (300 μm). Pretreatment of cerebellar granule cells with IL-10 (1–50 ng/ml) elicited a dose- and time-dependent reduction of glutamate-induced excitotoxicity. Most importantly, IL-10 reduced the number of apoptotic cells when added to the cultures together or 1 hr after glutamate. Using patch-clamping and fluorescence Ca2+imaging techniques, we examined whether IL-10 prevents glutamate toxicity by blocking the function of NMDA channel. IL-10 failed to affect NMDA channel properties and to reduce NMDA-mediated rise in intracellular Ca2+. Thus, this cytokine appears to prevent glutamate toxicity by a mechanism unrelated to a blockade of NMDA receptor function. Various proteases, such as caspase-3, and transcription factors, such as nuclear factor κB (NF-κB), have been proposed to participate in glutamate-mediated apoptosis. Thus, we examined whether IL-10 modulates the activity of these apoptotic markers. IL-10 blocked both the glutamate-mediated induction of caspase-3 as well as NF-κB DNA binding activity, suggesting that the neuroprotective properties of IL-10 may rely on its ability to block the activity of proapoptotic proteins.
Fate of Midbrain Dopaminergic Neurons Controlled by the Engrailed Genes
Authors: Simon Horst H. , Saueressig Harald , Wurst Wolfgang , Goulding Martyn D. , O'Leary Dennis D. M.
Published: April 2018

Deficiencies in neurotransmitter-specific cell groups in the midbrain result in prominent neural disorders, including Parkinson's disease, which is caused by the loss of dopaminergic neurons of the substantia nigra. We have investigated in mice the role of the engrailed homeodomain transcription factors, En-1 and En-2, in controlling the developmental fate of midbrain dopaminergic neurons.En-1is highly expressed by essentially all dopaminergic neurons in the substantia nigra and ventral tegmentum, whereasEn-2is highly expressed by a subset of them. These neurons are generated and differentiate their dopaminergic phenotype inEn-1/En-2double null mutants, but disappear soon thereafter. Use of anEn-1/tau-LacZknock-in mouse as an autonomous marker for these neurons indicates that they are lost, rather than that they change their neurotransmitter phenotype. A single allele ofEn-1on anEn-2null background is sufficient to produce a wild type-like substantia nigra and ventral tegmentum, whereas in contrast a single allele ofEn-2on anEn-1null background results in the survival of only a small proportion of these dopaminergic neurons, a finding that relates to the differential expression ofEn-1andEn-2. Additional findings indicate that En-1 and En-2 regulate expression of α-synuclein, a gene that is genetically linked to Parkinson's disease. These findings show that the engrailed genes are expressed by midbrain dopaminergic neurons from their generation to adulthood but are not required for their specification. However, the engrailed genes control the survival of midbrain dopaminergic neurons in a gene dose-dependent manner. Our findings also suggest a link between engrailed and Parkinson's disease.
Adaptive Axonal Remodeling in the Midbrain Auditory Space Map
Authors: DeBello William M. , Feldman Daniel E. , Knudsen Eric I.
Published: April 2018

The auditory space map in the external nucleus of the inferior colliculus (ICX) of barn owls is highly plastic, especially during early life. When juvenile owls are reared with prismatic spectacles (prisms) that displace the visual field laterally, the auditory spatial tuning of neurons in the ICX adjusts adaptively to match the visual displacement. In the present study, we show that this functional plasticity is accompanied by axonal remodeling.

The ICX receives auditory input from the central nucleus of the inferior colliculus (ICC) via topographic axonal projections. We used the anterograde tracer biocytin to study experience-dependent changes in the spatial pattern of axons projecting from the ICC to the ICX. The projection fields in normal adults were sparser and more restricted than those in normal juveniles. The projection fields in prism-reared adults were denser and broader than those in normal adults and contained substantially more bouton-laden axons that were appropriately positioned in the ICX to convey adaptive auditory spatial information. Quantitative comparison of results from juvenile and prism-reared owls indicated that prism experience led to topographically appropriate axonal sprouting and synaptogenesis. We conclude that this elaboration of axons represents the formation of an adaptive neuronal circuit.

The density of axons and boutons in the normal projection zone was preserved in prism-reared owls. The coexistence of two different circuits encoding alternative maps of space may underlie the ability of prism-reared owls to readapt to normal conditions as adults.
Gating of Human Theta Oscillations by a Working Memory Task
Authors: Raghavachari Sridhar , Kahana Michael J. , Rizzuto Daniel S. , Caplan Jeremy B. , Kirschen Matthew P. , Bourgeois Blaise , Madsen Joseph R. , Lisman John E.
Published: April 2018

Electrode grids on the cortical surface of epileptic patients provide a unique opportunity to observe brain activity with high temporal–spatial resolution and high signal-to-noise ratio during a cognitive task. Previous work showed that large-amplitude theta frequency oscillations occurred intermittently during a maze navigation task, but it was unclear whether theta related to the spatial or working memory components of the task. To determine whether theta occurs during a nonspatial task, we made recordings while subjects performed the Sternberg working memory task. Our results show event-related theta and reveal a new phenomenon, the cognitive “gating” of a brain oscillation: at many cortical sites, the amplitude of theta oscillations increased dramatically at the start of the trial, continued through all phases of the trial, including the delay period, and decreased sharply at the end. Gating could be seen in individual trials and varying the duration of the trial systematically varied the period of gating. These results suggest that theta oscillations could have an important role in organizing multi-item working memory.
Anxiolytic and Anti-Stress Effects of Brain Prolactin: Improved Efficacy of Antisense Targeting of the Prolactin Receptor by Molecular Modeling
Authors: Torner Luz , Toschi Nicola , Pohlinger Agnes , Landgraf Rainer , Neumann Inga D.
Published: April 2018

We provide the first evidence that prolactin is a neuromodulator of behavioral and neuroendocrine stress coping in the rat. In virgin female and male rats, intracerebral infusion of ovine prolactin (oPRL) into the lateral cerebral ventricle (intracerebroventricular) exerted an anxiolytic effect on the elevated plus-maze in a dose-dependent manner (0.1 and 1.0 μg/5 μl;p< 0.01). In contrast, downregulation of the expression of the long form of brain prolactin receptors by chronic intracerebroventricular infusion of an antisense oligodeoxynucleotide (ODN) (osmotic minipump, 0.5 μg · 0.5 μl−1 · hr−1; 5 d) increased anxiety-related behavior on the plus-maze compared with mixed bases-treated and vehicle-treated rats (p< 0.01), again demonstrating an anxiolytic effect of PRL acting at brain level. Furthermore, in jugular vein-catheterized female rats, the stress-induced increase of corticotropin secretion was decreased after chronic intracerebroventricular infusion of oPRL (osmotic minipump, 1.0 μg · 0.5 μl−1 · hr−1;p< 0.05) and, in contrast, was further elevated by antisense targeting of the brain prolactin receptors (p< 0.01). This provides evidence for a receptor-mediated attenuation of the responsiveness of the hypothalamo-pituitary-adrenal (HPA) axis by prolactin. The antisense ODN sequence was selected on the basis of secondary structure molecular modeling of the target mRNA to improve antisense ODN–mRNA hybridization. Receptor autoradiography confirmed the expected improvement in the efficacy of downregulation of prolactin receptor expression [empirically designed antisense, 30%;p> 0.05, not significant; adjustment of target position after mRNA modeling, 72%;p< 0.05). Taken together, prolactin acting at brain level has to be considered as a novel regulator of both emotionality and HPA axis reactivity.
Medial Geniculate, Amygdalar and Cingulate Cortical Training-Induced Neuronal Activity during Discriminative Avoidance Learning in Rabbits with Auditory Cortical Lesions
Authors: Duvel Adam D. , Smith David M. , Talk Andrew , Gabriel Michael
Published: April 2018

This study addressed the neural mediation of discriminative avoidance learning, wherein rabbits step in a wheel apparatus in response to an acoustic conditional stimulus, the CS+, to avoid a foot shock, and they learn to ignore a different stimulus, the CS−, not followed by foot shock. Previously, muscimol-induced inactivation of the amygdala in the first session of training prevented learning during the inactivation and permanently blocked the development of discriminative training-induced neuronal activity (TIA) in the medial division of the medial geniculate nucleus (MGm). These results suggested that amygdalar neurons induce discriminative TIA in the MGm via basolateral (BL) amygdalar axonal projections to the auditory cortex. To test this hypothesis, the activity of neurons in the MGm was recorded during learning in rabbits with lesions of the auditory cortex. Recordings were also made in the lateral and BL amygdalar nuclei and in the cingulate cortex. In support of the hypothesis, discriminative learning in rabbits with lesions was impaired significantly during early training sessions 1–4; in these same sessions, discriminative TIA was abolished in the MGm, the BL nucleus, and the anterior cingulate cortex. The lesions also blocked posterior cingulate cortical discriminative TIA in training sessions 1–2 but spared TIA in sessions 3–7. Lateral amygdalar neurons showed gradual development of discrimination that was not significantly affected by the lesions. The results demonstrate a critical role of auditory cortex in early discriminative learning and in the production of early discriminative TIA in multiple areas.
Evidence for a Role of Mixed Lineage Kinases in Neuronal Apoptosis
Authors: Mota Mónica , Reeder Melissa , Chernoff Jonathan , Bazenet Chantal E.
Published: April 2018

Superior cervical ganglion (SCG) sympathetic neurons die by apoptosis when deprived of nerve growth factor (NGF). It has been shown previously that the induction of apoptosis in these neurons at NGF withdrawal requires both the activity of the small GTP-binding protein Cdc42 and the activation of the c-Jun N-terminal kinase (JNK) pathway. The mixed lineage kinase 3 (MLK3) belongs to a family of mitogen-activated protein (MAP) kinase kinase kinases. MLK3 contains a Cdc42/Rac interactive-binding (CRIB) domain and activates both the JNK and the p38 MAP kinase pathways. In this study the role of MLK3 in the induction of apoptosis in sympathetic neurons has been investigated. Overexpression of an active MLK3 induces activation of the JNK pathway and apoptosis in SCG neurons. In addition, overexpression of kinase dead mutants of MLK3 blocks apoptosis as well as c-Jun phosphorylation induced by NGF deprivation. More importantly, MLK3 activity seems to increase by 5 hr after NGF withdrawal in both differentiated PC12 cells and SCG neurons. We also show that MLK3 lies downstream of Cdc42 in the neuronal death pathway. Regulation of MLK3 in neurons seems to be dependent on MLK3 activity and possibly on an additional cellular component, but not on its binding to Cdc42. These results suggest that MLK3, or a closely related kinase, is a physiological element of NGF withdrawal-induced activation of the Cdc42–c-Jun pathway and neuronal death. MLK3 therefore could be an interesting therapeutic target in a number of neurodegenerative diseases involving neuronal apoptosis.
A β-Strand in the γ2Subunit Lines the Benzodiazepine Binding Site of the GABAAReceptor: Structural Rearrangements Detected during Channel Gating
Authors: Teissére Jeremy A. , Czajkowski Cynthia
Published: April 2018

Benzodiazepines (BZDs) exert their effects in the CNS by binding to a modulatory site on GABAAreceptors. Individual amino acids have been implicated in BZD recognition and modulation of the GABAAreceptor, but the secondary structure of the amino acids contributing to the BZD binding site has not been elucidated. In this report we used the substituted cysteine accessibility method to understand the structural dynamics of a region of the GABAAreceptor implicated in BZD binding, γ2Y72–γ2Y83. Each residue within this region was mutated to cysteine and expressed with wild-type α1and β2subunits inXenopusoocytes. Methanethiosulfonate (MTS) reagents were used to modify covalently the engineered cysteines, and the subsequent effects on BZD modulation of the receptor were monitored functionally by two-electrode voltage clamp. We identified an alternating pattern of accessibility to sulfhydryl modification, indicating that the region γ2T73–γ2T81 adopts a β-strand conformation. By monitoring the ability of BZD ligands to impede the covalent modification of accessible cysteines, we also identified two residues within this region, γ2A79 and γ2T81, that line the BZD binding site. Sulfhydryl modification of γ2A79C or γ2T81C allosterically shifts the GABA EC50of the receptor, suggesting that certain MTS compounds may act as tethered agonists at the BZD binding site. Last, we present structural evidence that a portion of the BZD binding site undergoes a conformational change in response to GABA binding and channel gating (opening and desensitization). These data represent an important step in understanding allosteric communication in ligand-gated ion channels.
FGF Induces a Switch in Death Receptor Pathways in Neuronal Cells
Authors: Eves Eva M. , Skoczylas Christine , Yoshida Keiko , Alnemri Emad S. , Rosner Marsha R.
Published: April 2018

Basic fibroblast growth factor (FGF2) has many roles in neuronal development and maintenance including effects on mitogenesis, survival, fate determination, differentiation, and migration. Using a conditionally immortalized rat hippocampal cell line, H19-7, and primary hippocampal cultures, we now demonstrate that FGF2 treatment differentially regulates members of the tumor necrosis factor (TNF) superfamily of death domain receptors and their ligands. H19-7 cells transferred from serum to defined (N2) medium undergo apoptosis by a Fas-dependent mechanism similar to primary neurons. In contrast, H19-7 cells treated with FGF undergo apoptosis by a Fas-independent mechanism. FGF suppresses the Fas death pathway but also induces apoptosis by activation of a TNFα death pathway in both H19-7 and hippocampal progenitor cells. Expression of the TNF receptor 1 (TNFR1) or TNFR2 in H19-7 cells is sufficient to sensitize the cells to TNFα, similar to the effects of FGF. Because TNFα can be either proapoptotic or antiapoptotic, these results provide an explanation for the divergent trophic effects of FGF2 treatment and the observation that multiple trophic inputs are required for the survival of specific neurons.
Death in the Balance: Alternative Participation of the Caspase-2 and -9 Pathways in Neuronal Death Induced by Nerve Growth Factor Deprivation
Authors: Troy Carol M. , Rabacchi Sylvia A. , Hohl Justin B. , Angelastro James M. , Greene Lloyd A. , Shelanski Michael L.
Published: April 2018

The data presented here demonstrate that sympathetic neurons have the potential to activate two alternative caspase-dependent pathways either of which is capable of mediating death induced by NGF deprivation and that these neurons have the potential to switch from one pathway to the other. The presence of these two alternative pathways to trophic factor deprivation-induced death may have implications for ensuring the correct development of the nervous system. In wild-type neurons, a caspase-2-dependent pathway is required for death, and a caspase-9-dependent pathway appears to be suppressed by endogenous inhibitors of apoptosis proteins (IAPs). In contrast, for caspase-2-null neurons, death is dependent on the caspase-9 pathway. The mechanism underlying the shift is the result of a threefold compensatory elevation of caspase-9 expression and a doubling of levels of direct IAP binding protein with low pI (DIABLO)/second mitochondria-derived activator of caspase (Smac), an IAP inhibitor, both at the mRNA and protein levels. These findings resolve seemingly discrepant findings regarding the roles of various caspases after NGF deprivation and raise a cautionary note regarding the interpretation of findings with caspase-null animals. The choice of the death-mediating caspase pathway in the sympathetic neurons is thus dependent on the regulated relative expression of components of the pathways including those of caspases, IAPs, and IAP inhibitors.
Spontaneous Changes in Mitochondrial Membrane Potential in Cultured Neurons
Authors: Buckman Jennifer F. , Reynolds Ian J.
Published: April 2018

Using the mitochondrial membrane potential (ΔΨm)-sensitive fluorescent dyes 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolocarbocyanine iodide (JC-1) and tetramethylrhodamine methyl ester (TMRM), we have observed spontaneous changes in the ΔΨmof cultured forebrain neurons. These fluctuations in ΔΨmappear to represent partial, transient depolarizations of individual mitochondria. The frequency of these ΔΨmfluctuations can be significantly lowered by exposure to a photo-induced oxidant burden, an ATP synthase inhibitor, or a glutamate-induced sodium load, without changing overall JC-1 fluorescence intensity. These spontaneous fluctuations in JC-1 signal were not inhibited by altering plasma membrane activity with tetrodotoxin or MK-801 or by blocking the mitochondrial permeability transition pore (PTP) with cyclosporin A. Neurons loaded with TMRM showed similar, low-amplitude, spontaneous fluctuations in ΔΨm. We hypothesize that these ΔΨmfluctuations are dependent on the proper functioning of the mitochondria and reflect mitochondria alternating between the active and inactive states of oxidative phosphorylation.
The Leukocyte Common Antigen-Related Protein Tyrosine Phosphatase Receptor Regulates Regenerative Neurite OutgrowthIn Vivo
Authors: Xie Youmei , Yeo Tracy T. , Zhang Cheng , Yang Tao , Tisi Michelle A. , Massa Stephen M. , Longo Frank M.
Published: April 2018

Drosophilaand leech models of nervous system development demonstrate that protein tyrosine phosphatase (PTP) receptors regulate developmental neurite outgrowth. Whether PTP receptors regulate neurite outgrowth in adult systems or in regenerative states remains unknown. The leukocyte common antigen-related (LAR) receptor is known to be present in rodent dorsal root ganglion (DRG) neurons; therefore, the well established model of postcrush sciatic nerve regeneration was used to test the hypothesis that LAR is required for neurite outgrowth in the adult mammalian nervous system. In uninjured sciatic nerves, no differences in nerve morphology and sensory function were detected between wild-type and LAR-deficient littermate transgenic mice. Sciatic nerve crush resulted in increased LAR protein expression in DRG neurons. In addition, nerve injury led to an increase in the proportion of LAR protein isoforms known to have increased binding affinity to neurite-promoting laminin–nidogen complexes. Two weeks after nerve crush, morphological analysis of distal nerve segments in LAR-deficient transgenic mice demonstrated significantly decreased densities of myelinated fibers, decreased axonal areas, and increased myelin/axon area ratios compared with littermate controls. Electron microscopy analysis revealed a significant twofold reduction in the density of regenerating unmyelinated fibers in LAR−/− nerves distal to the crush site. Sensory testing at the 2 week time point revealed a corresponding 3 mm lag in the proximal-to-distal progression of functioning sensory fibers along the distal nerve segment. These studies introduce PTP receptors as a major new gene family regulating regenerative neurite outgrowthin vivoin the adult mammalian system.