bluuebunny/crossref_metadata_2025 huggingface dataset:
Interaction of Muscle and Brain Sodium Channels with Multiple Members of the Syntrophin Family of Dystrophin-Associated Proteins
Authors: Gee Stephen H. , Madhavan Raghavan , Levinson S. Rock , Caldwell John H. , Sealock Robert , Froehner Stanley C.
Published: April 2018

Syntrophins are cytoplasmic peripheral membrane proteins of the dystrophin-associated protein complex (DAPC). Three syntrophin isoforms, α1, β1, and β2, are encoded by distinct genes. Each contains two pleckstrin homology (PH) domains, a syntrophin-unique (SU) domain, and a PDZ domain. The name PDZ comes from the first three proteins found to contain repeats of this domain (PSD-95,Drosophiladiscs large protein, and the zona occludens protein 1). PDZ domains in other proteins bind to the C termini of ion channels and neurotransmitter receptors containing the consensus sequence (S/T)XV-COOH and mediate the clustering or synaptic localization of these proteins. Two voltage-gated sodium channels (NaChs), SkM1 and SkM2, of skeletal and cardiac muscle, respectively, have this consensus sequence. Because NaChs are sarcolemmal components like syntrophins, we have investigated possible interactions between these proteins. NaChs copurify with syntrophin and dystrophin from extracts of skeletal and cardiac muscle. Peptides corresponding to the C-terminal 10 amino acids of SkM1 and SkM2 are sufficient to bind detergent-solubilized muscle syntrophins, to inhibit the binding of native NaChs to syntrophin PDZ domain fusion proteins, and to bind specifically to PDZ domains from α1-, β1-, and β2-syntrophin. These peptides also inhibit binding of the syntrophin PDZ domain to the PDZ domain of neuronal nitric oxide synthase, an interaction that is not mediated by C-terminal sequences. Brain NaChs, which lack the (S/T)XV consensus sequence, also copurify with syntrophin and dystrophin, an interaction that does not appear to be mediated by the PDZ domain of syntrophin. Collectively, our data suggest that syntrophins link NaChs to the actin cytoskeleton and the extracellular matrix via dystrophin and the DAPC.
Mitochondrial Susceptibility to Oxidative Stress Exacerbates Cerebral Infarction That Follows Permanent Focal Cerebral Ischemia in Mutant Mice with Manganese Superoxide Dismutase Deficiency
Authors: Murakami Kensuke , Kondo Takeo , Kawase Makoto , Li Yibing , Sato Shuzo , Chen Sylvia F. , Chan Pak H.
Published: April 2018

Mitochondrial injury has been implicated in ischemic neuronal injury. Mitochondria, producing adenosine triphosphate by virtue of electron flow, have been shown to be both the sites of superoxide anion (O2−) production and the target of free radical attacks. We evaluated these mechanisms in anin vivocerebral ischemia model, using mutant mice with a heterozygous knock-out gene (Sod2−/+) encoding mitochondrial manganese superoxide dismutase (Mn-SOD).Sod2−/+ mice demonstrated a prominent increase in O2−production under normal physiological conditions and in ischemia, as evidenced by specific oxidation of a fluorescent probe, hydroethidine, reflecting decreased activity of Mn-SOD. A mitochondrial viability assay that used rhodamine 123, which is accumulated by transmembrane potential of viable mitochondria, demonstrated accelerated development of mitochondrial injury. This rapid progress of ischemic injury resulted in exacerbation of infarct size and hemisphere enlargement, causing advanced neurological deficits but without altering DNA fragmentation induction. The present study suggests that O2−overproduced in a mitochondrial compartment, when uncoupled from antioxidant defenses, induces impairment of mitochondrial function and causes exacerbation of cerebral infarction after ischemia.
Conversion of Sensory Signals into Motor Commands in Primary Motor Cortex
Authors: Salinas Emilio , Romo Ranulfo
Published: April 2018

Movement triggered by sensory stimuli requires that the networks generating the motor commands receive an adequate driving input, which, in general, is a transformed version of the initial sensory signal. We investigated the nature of this transformation in a task in which monkeys categorize the speed of tactile stimuli as either low or high, reaching for one of two pushbuttons to indicate their choice. Extracellular recordings from primary motor cortex revealed two types of neurons selective for the speed categories: ones that fire at higher rates for low versus high speeds, and others that do the opposite. These differential responses are task-specific; no firing rate modulation was seen when identical arm movements were triggered by visual cues or when stimuli were delivered passively. Analyses using decoding and modeling techniques produced two main results. First, the neurons accurately encode the chosen category; an observer measuring their responses can exhibit a psychophysical performance during categorization identical to the monkey’s. Second, by analyzing separately the trials in which hits and errors were scored, it is possible to distinguish purely sensory activity from activity exclusively related to arm motion. The recorded responses did not match either of these alternatives but were consistent with a model in which the category-tuned neurons are the link between the output of the sensory categorization process and the motor command used to indicate the animal’s decision. Thus, the observed activity seems to encode a preprocessed version of the sensory stimulus and to participate in driving the arm motion.
Identification of Amino Acid Residues of the NR2A Subunit That Control Glutamate Potency in Recombinant NR1/NR2A NMDA Receptors
Authors: Anson Lesley C. , Chen Philip E. , Wyllie David J. A. , Colquhoun David , Schoepfer Ralf
Published: April 2018

The NMDA type of ligand-gated glutamate receptor requires the presence of both glutamate and glycine for gating. These receptors are hetero-oligomers of NR1 and NR2 subunits. Previously it was thought that the binding sites for glycine and glutamate were formed by residues on the NR1 subunit. Indeed, it has been shown that the effects of glycine are controlled by residues on the NR1 subunit, and a “Venus flytrap” model for the glycine binding site has been suggested by analogy with bacterial periplasmic amino acid binding proteins. By analysis of 10 mutant NMDA receptors, we now show that residues on the NR2A subunit control glutamate potency in recombinant NR1/NR2A receptors, without affecting glycine potency. Furthermore, we provide evidence that, at least for some mutated residues, the reduced potency of glutamate cannot be explained by alteration of gating but has to be caused primarily by impairing the binding of the agonist to the resting state of the receptor. One NR2A mutant, NR2A(T671A), had anEC50for glutamate 1000-fold greater than wild type and a 255-fold reduced affinity for APV, yet it had single-channel openings very similar to those of wild type. Therefore we propose that the glutamate binding site is located on NR2 subunits and (taking our data together with previous work) is not on the NR1 subunit. Our data further imply that each NMDA receptor subunit possesses a binding site for an agonist (glutamate or glycine).
Overloaded Endoplasmic Reticulum–Golgi Compartments, a Possible Pathomechanism of Peripheral Neuropathies Caused by Mutations of the Peripheral Myelin Protein PMP22
Authors: D’Urso Donatella , Prior Reinhard , Greiner–Petter Regine , Gabreëls–Festen Anneke A. W. M. , Müller Hans Werner
Published: April 2018

Nonconservative point mutations of the peripheral myelin protein 22 (PMP22) are associated with Charcot-Marie-Tooth type 1A disease, the most common inherited peripheral neuropathy in humans, and with the Trembler J (TrJ) and Trembler (Tr) alleles in mice. We investigated the intracellular transport of wild-type PMP22 and its TrJ and Tr mutant forms in Schwann cells and in a non-neuronal cell line. In contrast to wild type, mutant proteins were not inserted into the plasma membrane and accumulated in the endoplasmic reticulum and Golgi compartments. Coexpression of each mutant with wild-type PMP22 confirmed the different intracellular distribution of the mutant forms, indicating that neither the TrJ nor Tr protein has a dominant-negative effect on the cellular distribution of wild-type PMP22. Accumulation of PMP22 immunoreactivity in the cell body of myelinating Schwann cells was also observed in nerve biopsies obtained from CMT1A patients carrying the TrJ point mutation. We propose that impaired trafficking of mutated PMP22 affects Schwann cell physiology leading to myelin instability and loss.
Functional Analysis of the Rat I Sodium Channel inXenopusOocytes
Authors: Smith Raymond D. , Goldin Alan L.
Published: April 2018

Voltage-gated sodium channels in the mammalian CNS initiate and propagate action potentials when excitatory inputs achieve threshold membrane depolarization. There are multiple sodium channel isoforms expressed in rat brain (types I, II, III, 6, and NaG). We have constructed a full-length cDNA clone encoding type I and compared the electrophysiological properties of type I (Rat1) and II (Rat2) channels in the absence and presence of the two accessory subunits β1and β2. Injection intoXenopusoocytes of RNA encoding Rat1 resulted in functional sodium currents that were blocked by tetrodotoxin, with Kapp= 9.6 nm. Rat1 sodium channels had a slower time course of fast inactivation than Rat2. Coexpression of β1accelerated inactivation of both Rat1 and Rat2, resulting in comparable inactivation kinetics. Rat1 recovered from fast inactivation more rapidly than Rat2, regardless of whether β1or β2was present. The voltage dependence of activation was similar for Rat1 and Rat2 without the β subunits, but it was more positive for Rat1 when β1and β2were coexpressed. The voltage dependence of inactivation was more positive for Rat1 than for Rat2, and coexpression with β1and β2accentuated that difference. Finally, sodium current amplitudes were reduced by 7–9% for both Rat1 and Rat2 channels when protein kinase A phosphorylation was induced. It has been suggested previously that Rat1 and Rat6 channels mediate transient and maintained sodium conductances, respectively, in Purkinje cells, and the electrophysiological properties of Rat1 currents are consistent with a role for this channel in mediating the rapidly inactivating, transient current.
Multiple Pathways of Neuronal Death Induced by DNA-Damaging Agents, NGF Deprivation, and Oxidative Stress
Authors: Park David S. , Morris Erick J. , Stefanis Leonidas , Troy Carol M. , Shelanski Michael L. , Geller Herbert M. , Greene Lloyd A.
Published: April 2018

Here, we compare the pathways by which DNA-damaging agents, NGF deprivation, and superoxide dismutase 1 (SOD1) depletion evoke apoptosis of sympathetic neurons. Previous work raised the hypothesis that cell cycle signaling plays a required role in neuronal apoptosis elicited by NGF deprivation and the DNA-damaging agent camptothecin. To test this hypothesis, we extended our investigation of DNA-damaging agents to cytosine arabinoside (AraC) and UV irradiation. As with NGF deprivation and camptothecin treatment, the cyclin-dependent kinase inhibitors flavopiridol and olomoucine protected neurons from apoptosis induced by AraC and UV treatment. These observations support the model that camptothecin, AraC, and UV treatment cause DNA damage, which leads to apoptosis by a mechanism that, as in the case of NGF deprivation, includes activation of cell cycle components. Flavopiridol and olomoucine, however, had no effect on death induced by SOD1 depletion, suggesting that CDKs do not play a role in this paradigm of neuronal death. To compare further the mechanisms of death evoked by NGF withdrawal, SOD1 depletion, and DNA-damaging agents, we investigated their responses to inhibitors of cysteine aspartases, elements of apoptotic pathways. The V-ICEinhand BAF, two peptide inhibitors of cysteine aspartases, protected neurons in all three death paradigms. In contrast, the cysteine aspartase inhibitory peptide zVAD-fmk conferred protection from NGF withdrawal and SOD1 depletion, but not DNA-damaging agents, whereas acYVAD-cmk protected only from SOD1 depletion. Taken together, these findings indicate that three different apoptotic stimuli activate separate pathways of death in the same neuron type.
Primed Facilitation of Homosynaptic Long-Term Depression and Depotentiation in Rat Hippocampus
Authors: Holland Lorne L. , Wagner John J.
Published: April 2018

Previous studies have demonstrated that prior synaptic activity can influence the subsequent induction of synaptic plasticity in the brain. Such temporal modulation of synaptic plasticity has been called “metaplasticity.” In this report, we describe the facilitatory effects of high-frequency stimulation on the induction of homosynaptic long-term depression (LTD) in the CA1 region of the rat hippocampus. The LTD induced by low-frequency stimulation (1 Hz) protocols was found to be homosynaptic and NMDA receptor-dependent. The facilitatory effects of the high-frequency stimulation-induced priming event itself were found to be NMDA receptor-independent and to have a duration of at least 90 min. The effects of priming also were heterosynaptic, because the induction of synaptic plasticity by low-frequency stimulation was enhanced at an unprimed synaptic pathway after the priming of an independent pathway. In addition to enhancing LTD, priming also enhanced the reversal of long-term potentiation elicited by a 5 Hz depotentiation protocol. Our results provide examples of how metaplasticity may play a key role in the ongoing modulation of the induction and stabilization of alterations in synaptic strength.
Neural Substrates for Species Recognition in the Time-Coding Electrosensory Pathway of Mormyrid Electric Fish
Authors: Friedman Matthew A. , Hopkins Carl D.
Published: April 2018

Mormyrid electric fish have species- and sex-typical electric organ discharges (EODs). One class of tuberous electroreceptors, the knollenorgans, plays a critical role in electric communication; one function is species recognition of EOD waveforms. In this paper, we describe cell types in the knollenorgan central pathway, which appear responsible for analysis of the temporal patterns of spikes encoded by the knollenorgans in response to EOD stimuli. Secondary sensory neurons in the nucleus of the electrosensory lateral line lobe (NELL) act as relays of peripheral responses. They fire a single phase-locked spike to an outside positive-going voltage step. Axons from the NELL project to the toral nucleus exterolateralis pars anterior (ELa). Immediately after they enter the ELa, they send collaterals to terminate on one to three ELa large cells and then continue in a lengthy neuronal pathway that traverses the ELa several times. After a path length of up to 5 mm, the NELL axon terminates on as many as 70 ELa small cells. Thus the large cells appear to be excited first, followed by the small cells, with the intervening length of the axon serving as a delay line. The large cells also respond with phase-locked spikes to voltage steps. Large cell axons extend for ∼1 mm and terminate on several small cells within the ELa. The terminals are known to be GABAergic inputs and are presumed inhibitory. We propose that small cells receive direct inhibition from large cells and delayed excitation from NELL axons. The small cells may act as anti-co-incidence detectors to analyze the temporal structure of the EOD waveform.
Calmodulin Is Involved in Membrane Depolarization-Mediated Survival of Motoneurons by Phosphatidylinositol-3 Kinase- and MAPK-Independent Pathways
Authors: Soler Rosa M. , Egea Joaquim , Mintenig Gerard M. , Sanz-Rodriguez Cesar , Iglesias Montse , Comella Joan X.
Published: April 2018

In the present work, we find that the elevation of extracellular K+concentration promotes the survival of chick spinal cord motoneuronsin vitrodeprived of any neurotrophic support. This treatment induces chronic depolarization of the neuronal plasma membrane, which activates L-type voltage-dependent Ca2+channels, resulting in Ca2+influx and elevation of the cytosolic free Ca2+concentration. Pharmacological reduction of intracellular free Ca2+or withdrawal of extracellular Ca2+reversed the effects of depolarization on survival. The intracellular Ca2+response to membrane depolarization developed as an initial peak followed by a sustained increase in intracellular Ca2+concentration. The depolarizing treatment caused tyrosine phosphorylation of mitogen-activated protein kinase (MAPK) without involving tyrosine kinase receptor activation. The calmodulin antagonist W13 inhibited the survival-promoting effect induced by membrane depolarization but not the tyrosine phosphorylation of MAPK. Moreover, depolarization did not induce phosphatidylinositol-3 kinase (PI-3K) phosphorylation in our cells, and the PI-3K inhibitor wortmannin did not suppress the survival-promoting effect of K+treatment. These results suggest that calmodulin is involved in calcium-mediated survival of motoneurons through the activation of PI-3K- and MAPK-independent pathways.
Effect of Glutamate on Dendritic Growth in Embryonic Rat Motoneurons
Authors: Metzger Friedrich , Wiese Stefan , Sendtner Michael
Published: April 2018

Glutamate is a major excitatory neurotransmitter for spinal motoneurons. We have investigated its effect on survival and neurite formation in cultures of highly enriched motoneurons from 15-d-old rat embryos. Whereas the survival of these neurons was not reduced by this treatment, a distinct and specific effect on dendrite outgrowth could be observed. Axon outgrowth was not affected by glutamate. Our data suggest that calcium influx via ionotropic AMPA/kainate (AMPA/KA) receptors is responsible for the regulation of dendrite outgrowth by excitatory neurotransmission. This was shown by the use of specific inhibitors for the different classes of glutamate receptors. The effect was reduced by continuous depolarization at 35 mmKCl and by treatment with joro spider toxin (JSTX-3, 3 μm), a blocker of Ca2+-conducting AMPA receptors. Removal of glutamate after 5 d of culture led to increased dendrite growth during the following culture period, and delayed addition resulted in a reduction in the length of already existing dendrites. Our observation that the effect is dose-dependent and reversible reflects a potential physiological function of excitatory neurotransmission on dendrite growth and morphology during a developmental period when synaptic contacts from afferent neurons to motoneurons are made in the spinal cord.
Transition of Brain Activation from Frontal to Parietal Areas in Visuomotor Sequence Learning
Authors: Sakai Katsuyuki , Hikosaka Okihide , Miyauchi Satoru , Takino Ryousuke , Sasaki Yuka , Pütz Benno
Published: April 2018

We studied the neural correlates of visuomotor sequence learning using functional magnetic resonance imaging (fMRI). In the test condition, subjects learned, by trial and error, the correct order of pressing two buttons consecutively for 10 pairs of buttons (2 × 10 task); in the control condition, they pressed buttons in any order. Comparison between the test condition and the control condition revealed four brain areas specifically related to learning: the dorsolateral prefrontal cortex (DLPFC), the presupplementary motor area (pre-SMA), the precuneus, and the intraparietal sulcus (IPS). We found that the time course of activation during learning was different between these areas. To normalize the individual differences in the speed of learning, we classified the performance of each subject into three learning stages: early, intermediate, and advanced stages. Both the relative increase of signal intensity and the number of activated pixels within the four areas showed significant changes across the learning stages, with different time courses. The two frontal areas, DLPFC and pre-SMA, were activated in the earlier stages of learning, whereas the two parietal areas, precuneus and IPS, were activated in the later stages. Specifically, DLPFC, pre-SMA, precuneus, and IPS were most highly activated in the early stage, in both the early and intermediate stages, in the intermediate stage, and in both the intermediate and advanced stages, respectively. The results suggest that the acquisition of visuomotor sequences requires frontal activation, whereas the retrieval of visuomotor sequences requires parietal activation, which might reflect the transition from the declarative stage to the procedural stage.
Cellular Sites for Activation of δ-Opioid Receptors in the Rat Nucleus Accumbens Shell: Relationship with Met5-Enkephalin
Authors: Svingos Adena L. , Clarke Cathy L. , Pickel Virginia M.
Published: April 2018

The shell compartment of the nucleus accumbens (AcbSh) is prominently involved in the rewarding aspects of δ-opioid receptor (DOR) agonists, including one of its putative endogenous ligands, Met5-enkephalin (Enk). We examined the ultrastructural immunocytochemical localization of an antipeptide DOR antiserum and an antibody against Enk to determine the major cellular sites for DOR activation and the spatial relationship between DOR and Enk in this region. Sixty percent of DOR-immunoreactive profiles were axon terminals and small unmyelinated axons, whereas the remainder were mainly dendrites and dendritic spines. In axons and terminals, DOR labeling was distributed along plasma and vesicular membranes. DOR-containing terminals were mainly large and primarily formed symmetric synapses or occasionally asymmetric synapses. DOR immunoreactivity also was associated with terminals that were small and formed punctate symmetric or nonrecognizable synapses. Dual immunoperoxidase and immunogold labeling showed that 35% of DOR-labeled axons apposed other terminals that contained Enk. In addition, 25% of the DOR-labeled terminals contained Enk. Thirty-five percent of DOR labeling was observed within dendrites and dendritic spines. DOR-labeled spines showed intense immunoreactivity within asymmetric postsynaptic junctions, which were formed by terminals that lacked Enk immunoreactivity. DOR-labeled spines, however, were apposed to Enk-containing terminals in 13% of all associations between dually labeled profiles. These results provide ultrastructural evidence that activation of DOR in the AcbSh is primarily involved in modulating the presynaptic release of mainly inhibitory, but also excitatory, neurotransmitters. These data also suggest that DOR plays a role in determining the postsynaptic response to excitatory afferents.
Acetylcholinesterase mRNA Level and Synaptic Activity in Rat Muscles Depend on Nerve-Induced Pattern of Muscle Activation
Authors: Sketelj Janez , Crne-Finderle Neva , Strukelj Borut , Trontelj Joze V. , Pette Dirk
Published: April 2018

Acetylcholinesterase (AChE) mRNA levels are severalfold higher in fast rat muscles compared with slow. We hypothesized that AChE mRNA levels and AChE activity in the neuromuscular junction depend on a specific nerve-induced pattern of motor unit activation. Chronic low-frequency stimulation, mimicking the activation pattern in slow muscles, was applied to fast muscles in rats. Molecular forms of AChE were analyzed by velocity sedimentation, and AChE mRNA levels were analyzed by Northern blots. AChE mRNA levels in stimulated fast muscles dropped to 10–20% of control after 1 week and became comparable to those in slow soleus muscles. The activity of the junctional A12AChE form in 35 d stimulated fast muscles decreased to 56% of control value, reaching that in the soleus muscle. Therefore, synaptic AChE itself depends on the muscle activation pattern. Complete inactivity after denervation also decreased the AChE mRNA level in fast muscles to <10% in 48 hr. In contrast, profuse fibrillations observed in noninnervated immature regenerating muscles maintain AChE mRNA levels at 80% of that in the innervated fast muscles. If protein synthesis was inhibited by cycloheximide, AChE mRNA levels in 3-d-old regenerating muscle, still containing myoblasts, increased approximately twofold. No significant increase after cycloheximide application was observed either in denervated mature fast muscles or in normal slow muscles. Low AChE mRNA levels observed in those muscles are probably not caused by decreased stability of AChE mRNA as demonstrated in myoblasts.
Yotiao, a Novel Protein of Neuromuscular Junction and Brain That Interacts with Specific Splice Variants of NMDA Receptor Subunit NR1
Authors: Lin Jerry W. , Wyszynski Michael , Madhavan Raghavan , Sealock Robert , Kim Jai Up , Sheng Morgan
Published: April 2018

The molecular machinery underlying neurotransmitter receptor immobilization at postsynaptic sites is poorly understood. The NMDA receptor subunit NR1 can form clusters in heterologous cells via a mechanism dependent on the alternatively spliced C1 exon cassette in its intracellular C-terminal tail, suggesting a functional interaction between NR1 and the cytoskeleton. The yeast two-hybrid screen was used here to identify yotiao, a novel coiled coil protein that interacts with NR1 in a C1 exon-dependent manner. Yotiao mRNA (11 kb) is present modestly in brain and abundantly in skeletal muscle and pancreas. On Western blots, yotiao appears as an ∼230 kDa band that is present in cerebral cortex, hippocampus, and cerebellum. Biochemical studies reveal that yotiao fractionates with cytoskeleton-associated proteins and with the postsynaptic density. With regard to immunohistochemistry, two anti-yotiao antibodies display a somatodendritic staining pattern similar to each other and to the staining pattern of NR1. Yotiao was colocalized by double-label immunocytochemistry with NR1 in rat brain and could be coimmunoprecipitated with NR1 from heterologous cells. Thus yotiao is an NR1-binding protein potentially involved in cytoskeletal attachment of NMDA receptors. Consistent with a general involvement in postsynaptic structure, yotiao was also found to be specifically concentrated at the neuromuscular junction in skeletal muscle.
BDNF Injected into the Superior Colliculus Reduces Developmental Retinal Ganglion Cell Death
Authors: Ma Yun-Tao , Hsieh Ted , Forbes M. Elizabeth , Johnson James E. , Frost Douglas O.
Published: April 2018

The role of neurotrophins as survival factors for developing CNS neurons, including retinal ganglion cells (RGCs), is uncertain. Null mutations for brain-derived neurotrophic factor (BDNF) or neurotrophin 4 (NT4), individually or together, are without apparent effect on the number of RGCs that survive beyond the period of normal, developmental RGC death. This contrasts with the BDNF dependence of RGCsin vitroand the effectiveness of BDNF in reducing RGC loss after axotomy. To investigate the effect of target-derived neurotrophins on the survival of developing RGCs, we injected BDNF into the superior colliculus (SC) of neonatal hamsters. At the age when the rate of developmental RGC death is greatest, BDNF produces, 20 hr after injection, a 13–15-fold reduction in the rate of RGC pyknosis compared with the rates in vehicle-injected and untreated hamsters. There is no effect 8 hr after injection. Electrochemiluminescence immunoassay measurements of BDNF protein in the retinae and SC of normal and BDNF-treated hamsters demonstrate that the time course of BDNF transport to RGCs supports a role for target-derived BDNF in promoting RGC survival. The effectiveness of pharmacological doses of BDNF in reducing developmental RGC death may be useful in further studies of the mechanisms of stabilization and elimination of immature central neurons.
Insulin-Like Growth Factor-I Is a Differentiation Factor for Postmitotic CNS Stem Cell-Derived Neuronal Precursors: Distinct Actions from Those of Brain-Derived Neurotrophic Factor
Authors: Arsenijevic Yvan , Weiss Samuel
Published: April 2018

Insulin-like growth factor-I (IGF-I) has been reported previously to promote the proliferation, survival, and maturation of sympathetic neuroblasts, the genesis of retinal neurons, and the survival of CNS projection and motor neurons. Here we asked whether IGF-I could promote thein vitrodifferentiation of postmitotic mammalian CNS neuronal precursors derived from multipotent epidermal growth factor (EGF)-responsive stem cells. In the absence of IGF-I, virtually no neurons were present in cultured stem cell progeny, whereas IGF-I increased neuron number by eight- to 40-fold. Brief exposures (2 hr) to IGF-I were sufficient to allow for neuronal differentiation without affecting proliferation or survival. IGF-I actions could be mimicked by insulin and IGF-II at concentrations that correspond to the pharmacology of the IGF-I receptor, the latter for which the mRNA was detected in undifferentiated stem cell progeny. Although ineffectual alone at low concentrations (10 nm) that would activate its own receptor, insulin was able to potentiate the actions of IGF-I by acting on mitotically active neural precursors. When neuronal precursor differentiation by IGF-I was examined in relation to brain-derived neurotrophic factor (BDNF), two important observations were made: (1) BDNF could potentiate the differentiating actions of IGF-I plus insulin, and (2) BDNF could act on a separate population of precursors that did not require IGF-I plus insulin for differentiation. Taken together, these results suggest that IGF-I and BDNF may act together or sequentially to promote neuronal precursor differentiation.
Vagotomy Blocks the Induction of Interleukin-1β (IL-1β) mRNA in the Brain of Rats in Response to Systemic IL-1β
Authors: Hansen Michael K. , Taishi Ping , Chen Zutang , Krueger James M.
Published: April 2018

There is considerable interest in the mechanisms by which systemic cytokines signal the CNS to elicit centrally controlled biological actions. This study determined the effects of intraperitoneal injections of interleukin-1β (IL-1β) on IL-1β mRNA and IL-1 receptor accessory protein (IL-1RAP) mRNA production in rat liver and brain using the reverse transcription-PCR. Saline or IL-1β (0.5 μg/kg) was injected intraperitoneally in subdiaphragmatically vagotomized and sham-operated (SHAM) rats. All injections were performed at dark onset, and rats were killed 2 hr after the injection. In SHAM rats, IL-1β increased IL-1β mRNA levels in the liver, hypothalamus, hippocampus, and brainstem. Subdiaphragmatic vagotomy blocked the IL-1β-induced increase in IL-1β mRNA in the brainstem and hippocampus and significantly attenuated the increase in the hypothalamus. Vagotomy did not affect IL-1β-induced IL-1β mRNA production in the liver. IL-1RAP mRNA was highly expressed in each region examined; however, no significant differences in IL-1RAP mRNA production were found in any region after IL-1β injection. The current results indicate that the vagus nerve is involved in transmitting cytokine signals to the brain and suggest that the induction of brain cytokines is a critical step in the pathway by which vagal-mediated signals result in centrally controlled symptoms of the acute phase response.
Two Receptor Tyrosine Phosphatases of the LAR Family Are Expressed in the Developing Leech by Specific Central Neurons as well as Select Peripheral Neurons, Muscles, and Other Cells
Authors: Gershon Timothy R. , Baker Michael W. , Nitabach Michael , Wu Peter , Macagno Eduardo R.
Published: April 2018

Receptor protein tyrosine phosphatases (rPTPs) are thought to play a crucial role in neuronal development, particularly in pathfinding by growing processes. We have cloned and sequenced twoHirudo medicinalisrPTPs that are homologous to theDrosophilaand vertebrate rPTPs of the Leukocyte common antigen-related (LAR) subfamily. TheseHirudorPTPs, HmLAR1 and HmLAR2, are products of different, homologous genes, both containing two tandem intracellular phosphatase domains and ectodomains with three tandem Ig domains and different numbers of tandem fibronectin type III (FIII) domains. They are expressed in distinct patterns during embryogenesis. HmLAR1 mRNA is expressed by a subset of central and peripheral neurons and by several peripheral muscular structures, whereas HmLAR2 mRNA is expressed by a different subset of central neurons and by the peripheral, neuron-like Comb cells. HmLAR1 and HmLAR2 proteins are located on the neurites of central neurons. In addition, HmLAR2 is expressed on the cell body, processes, and growth cones of the Comb cells. Because of their CAM-like ectodomains and homology to proteins known to be involved in pathfinding and because they are expressed by different subsets of neurons, we hypothesize that HmLAR1 and HmLAR2 participate in navigational decisions that distinguish the sets of neurons that express them. Furthermore, we hypothesize that HmLAR2 is also involved in setting up the highly regular array of parallel processes established by the Comb cells. Lastly, we propose that the HmLAR1 ectodomain on peripheral muscle cells plays a role in target recognition via interactions with neuronal receptors, which might include HmLAR1 or HmLAR2.
Adrenergic α2C-Receptors Modulate the Acoustic Startle Reflex, Prepulse Inhibition, and Aggression in Mice
Authors: Sallinen Jukka , Haapalinna Antti , Viitamaa Timo , Kobilka Brian K. , Scheinin Mika
Published: April 2018

Studies on animal models of stress, anxiety, aggression, and sensorimotor gating have linked specific monoamine neurotransmitter abnormalities to the cognitive and behavioral disturbances associated with many affective neuropsychiatric disorders. Although α2-adrenoceptors (α2-ARs) have been suggested to have a modulatory role in these disorders, the specific roles of each α2-AR subtype (α2A, α2B, and α2C) are largely unknown. The restricted availability of relevant animal models and the lack of subtype-selective α2-AR drugs have precluded detailed studies in this area. Therefore, transgenic mice were used to study the possible role of the α2C-AR subtype in two well established behavioral paradigms: prepulse inhibition (PPI) of the startle reflex and isolation-induced aggression. The α2C-AR-altered mice appear grossly normal, but subtle changes have been observed in their brain dopamine (DA) and serotonin (5-HT) metabolism. In this study, the mice with targeted inactivation of the gene encoding α2C-ARs (α2C-KO) had enhanced startle responses, diminished PPI, and shortened attack latency in the isolation–aggression test, whereas tissue-specific overexpression of α2C-ARs (α2C-OE) was associated with opposite effects. Correlation analyses suggested that both the magnitude of the startle response and its relative PPI (PPI%) were modulated by the mutations. In addition, the differences in PPI, observed between drug-naive α2C-OE mice and their wild-type controls, were abolished by treatment with a subtype nonselective α2-agonist and antagonist. Thus, drugs acting via α2C-ARs might have therapeutic value in disorders associated with enhanced startle responses and sensorimotor gating deficits, such as schizophrenia, attention deficit disorder, post-traumatic stress disorder, and drug withdrawal.
Overtraining Does Not Mitigate Contextual Fear Conditioning Deficits Produced by Neurotoxic Lesions of the Basolateral Amygdala
Authors: Maren Stephen
Published: April 2018

The influence of overtraining on the magnitude of fear-conditioning deficits produced by neurotoxic lesions of the basolateral amygdala (BLA) was examined. Either 1 d before or 1 week after the administration of neurotoxic BLA lesions, rats received either 1 or 25 conditioning trials consisting of the delivery of unsignaled foot shock in a novel observation chamber; freezing served as the measure of conditional fear. In this conditioning paradigm, asymptotic performance is reached in five conditioning trials, and 25 conditioning trials constitutes an overtraining procedure. The results revealed that overtraining does not affect the magnitude of the contextual freezing deficits produced by post-training BLA lesions. Similarly, overtraining did not influence the level of reacquisition obtained by rats with post-training BLA lesions after 10 reacquisition trials. A similar pattern of results was observed in rats with pretraining BLA lesions. Neurotoxic BLA lesions did not alter either motor activity or shock reactivity. These results indicate that overtraining does not limit the important role of the BLA in the acquisition and expression of contextual fear conditioning.
Selective Fasciculation and Divergent Pathfinding Decisions of Embryonic Chick Motor Axons Projecting to Fast and Slow Muscle Regions
Authors: Milner Louise D. , Rafuse Victor F. , Landmesser Lynn T.
Published: April 2018

Proper motor function requires the precise matching of motoneuron and muscle fiber properties. The lack of distinguishing markers for early motoneurons has made it difficult to determine whether this matching is established by selective innervation during development or later via motoneuron–muscle fiber interactions. To examine whether chick motoneurons selectively innervate regions of their target containing either fast or slow muscle fibers, we backlabeled neurons from each of these regions with lipophilic dyes. We found that motor axons projecting to fast and slow muscle regions sorted into separate but adjacent fascicles proximally in the limb, long before they reached the muscle. More distally, these fascicles made divergent pathfinding decisions to course directly to the appropriate muscle fiber region. In contrast, axons projecting to different areas of an all-fast muscle did not fasciculate separately and became more intermingled as they coursed through the limb. Selective fasciculation of fast- and slow-projecting motoneurons was similar both before and after motoneuron cell death, suggesting that motoneurons specifically recognized and fasciculated with axons growing to muscle regions containing the appropriate muscle fiber type. Taken together, these results strongly support the hypothesis that “fast” and “slow” motoneurons are molecularly distinct before target innervation and that they use these differences to selectively fasciculate, pathfind to, and branch within the correct muscle fiber region from the outset of neuromuscular development.
Microcircuitry and Mosaic of a Blue–Yellow Ganglion Cell in the Primate Retina
Authors: Calkins David J. , Tsukamoto Yoshihiko , Sterling Peter
Published: April 2018

Perception of hue is opponent, involving the antagonistic comparison of signals from different cone types. For blue versus yellow opponency, the antagonism is first evident at a ganglion cell with firing that increases to stimulation of short wavelength-sensitive (S) cones and decreases to stimulation of middle wavelength-sensitive (M) and long wavelength-sensitive (L) cones. This ganglion cell, termed blue–yellow (B–Y), has a distinctive morphology with dendrites in both ON and OFF strata of the inner plexiform layer (Dacey and Lee, 1994). Here we report the synaptic circuitry of the cell and its spatial density. Reconstructing neurons in macaque fovea from electron micrographs of serial sections, we identified six ganglion cells that branch in both strata and have similar circuitry. In the ON stratum each cell collects ∼33 synapses from bipolar cells traced back exclusively to invaginating contacts from S cones, and in the OFF stratum each cell collects ∼14 synapses from bipolar cells (types DB2 and DB3) traced to basal synapses from ∼20 M and L cones. This circuitry predicts that spatially coincident blue–yellow opponency arises at the level of the cone output via expression of different glutamate receptors. S cone stimuli suppress glutamate release onto metabotropic receptors of the S cone bipolar cell dendrite, thereby opening cation channels, whereas M and L cone stimuli suppress glutamate release onto ionotropic glutamate receptors of DB2 and DB3 cell dendrites, thereby closing cation channels. Although the B–Y cell is relatively rare (3% of foveal ganglion cells), its spatial density equals that of the S cone; thus it could support psychophysical discrimination of a blue–yellow grating down to the spatial cutoff of the S cone mosaic.
Calcium Waves Precede Electrophysiological Changes of Spreading Depression in Hippocampal Organ Cultures
Authors: Kunkler Phillip E. , Kraig Richard P.
Published: April 2018

Although intercellular Ca2+waves resemble spreading depression (SD) and occur in hippocampal organ cultures (HOTCs), SD has not been reported in these cultures. Accordingly, electrophysiological and Ca2+imaging techniques were used to examine potential interrelations between Ca2+waves and electrophysiological changes of SD. Our results show, for the first time, that HOTCs can support SD. Furthermore, two distinct Ca2+waves were found to precede SD. The first traveled >100 μm/sec along the pyramidal cell dendritic layer. The second subsequently traveled mostly perpendicular to the pyramidal cell layer from CA3 (or CA1) but also in all directions from its area of initiation. This second, slower wave spread with the interstitial DC change of SD at millimeters per minute but always ahead of it by 6–16 sec. Heptanol, which uncouples gap junctions, blocked both of these Ca2+waves and SD. Thus, two types of Ca2+waves occur with the initiation and propagation of SD. The first might reflect interneuronal changes linked by gap junctions, whereas the second might stem from interastrocyte changes linked via similar connections. Because individual cells can be followed in space and time for protracted periods in HOTCs, this preparation may be ideal for studies designed to explore not only the mechanisms of SD but also the long-term consequences of SD, such as ischemic tolerance.
Granule Neuron Regulation of Purkinje Cell Development: Striking a Balance Between Neurotrophin and Glutamate Signaling
Authors: Morrison Mary E. , Mason Carol A.
Published: April 2018

Granule neurons, presynaptic afferents of Purkinje cells, are potent regulators of Purkinje cell development. Purified Purkinje cells survive and differentiate poorly, whereas coculture with granule neurons enhances their survival and dendritic development. Here we investigate the role of neurotrophins in granule–Purkinje cell interactions. BDNF or NT-4 improves, but NT-3 or CNTF reduces, survival of isolated Purkinje cells. When granule neurons are present, however, BDNF or NT-4 treatment leads to Purkinje cell loss. This decrease is overcome by anti-BDNF or TrkB-IgG-blocking reagents or by CNQX, a non-NMDA glutamate receptor antagonist. Furthermore, BDNF increases the spine density on the surviving Purkinje cells. These results suggest that Purkinje cell survival and differentiation are context-dependent and require a balance between neurotrophin- and activity-dependent signaling.
Evidence for Involvement of the cGMP–Protein Kinase G Signaling System in the Induction of Long-Term Depression, But Not Long-Term Potentiation, in the Dentate GyrusIn Vitro
Authors: Wu Jianqun , Wang Yue , Rowan Michael J. , Anwyl Roger
Published: April 2018

The involvement of the cGMP–protein kinase G (PKG) signaling pathway in the induction of long-term depression (LTD) and long-term potentiation (LTP) was investigated in the medial perforant path of the dentate gyrusin vitro. Low-frequency stimulation (LFS)-induced LTD of field EPSPs was inhibited by bath perfusion of the selective soluble guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ). LFS-induced LTD of EPSPs and whole-cell patch-clamped EPSCs was also blocked by bath perfusion and postsynaptic intracellular injection, respectively, of the selective PKG inhibitor KT5823. Elevation of intracellular cGMP by perfusion of the cGMP phosphodiesterase inhibitor zaprinast resulted in induction of LTD of field EPSPs and EPSCs. Occlusion experiments showed mutual inhibition between LFS-induced LTD and zaprinast-induced LTD. The zaprinast-induced LTD of field EPSPs was inhibited by perfusion of ODQ and KT5823. In addition, zaprinast-induced LTD of EPSCs was inhibited by postsynaptic application of KT5823. Glutamate receptor stimulation, especially that of metabotropic glutamate receptors (mGluRs), was required for zaprinast-induced LTD, because cessation of test stimulation or perfusion with the mGluR antagonist (+)-α-methyl-4-carboxyphenylglycine (MCPG) inhibited zaprinast-induced LTD. No inhibitory effect of ODQ or KT5823 on the induction of LTP of EPSPs or EPSCs was found. These data indicate that the cGMP–guanyly cyclase–PKG signaling pathway in the dentate gyrus is essential for induction of LTD, although not of LTP, in the dentate gyrus.
Structural Features Determining Differential Receptor Regulation of Neuronal Ca Channels
Authors: Simen Arthur A. , Miller Richard J.
Published: April 2018

Dihydropyridine-insensitive Ca channels are subject to direct receptor G-protein-mediated inhibition to differing extents. α1Bchannels are much more strongly modulated than α1Echannels. To understand the structural basis for this difference, we have constructed and expressed various α1Band α1Echimeric Ca channels and examined their regulation by κ-opioid receptors. Replacement of the first membrane-spanning domain of α1Ewith the corresponding region of α1Bresulted in a chimeric Ca channel that was modulated by κ-opioid receptors to a significantly greater extent than α1E. Transfer of the N terminus and I/II loop from α1Bin addition to domain I resulted in a chimeric channel that was modulated to the same extent as α1B. Other regions of the molecule do not appear to contribute significantly to the degree of inhibition obtained, although the C terminus may contribute to facilitation.
Sensitive Periods for Visual Calibration of the Auditory Space Map in the Barn Owl Optic Tectum
Authors: Brainard Michael S. , Knudsen Eric I.
Published: April 2018

Previous studies have identified sensitive periods for the developing barn owl during which visual experience has a powerful influence on the calibration of sound localization behavior. Here we investigated neural correlates of these sensitive periods by assessing developmental changes in the capacity of visual experience to alter the map of auditory space in the optic tectum of the barn owl. We used two manipulations. (1) We equipped owls with prismatic spectacles that optically displaced the visual field by 23° to the left or right, and (2) we restored normal vision to prism-reared owls that had been raised wearing prisms. In agreement with previous behavioral experiments, we found that the capacity of abnormal visual experience to shift the tectal auditory space map was restricted to an early sensitive period. However, this period extended until later in life (∼200 d) than described previously in behavioral studies (∼70 d). Furthermore, unlike the previous behavioral studies that found that the capacity to recover normal sound localization after restoration of normal vision was lost at ∼200 d of age, we found that the capacity to recover a normal auditory space map was never lost. Finally, we were able to reconcile the behaviorally and neurophysiologically defined sensitive periods by taking into account differences in the richness of the environment in the two sets of experiments. We repeated the behavioral experiments and found that when owls were housed in a rich environment, the capacity to adjust sound localization away from normal extended to later in life, whereas the capacity to recover to normal was never lost. Conversely, when owls were housed in an impoverished environment, the capacity to recover a normal auditory space map was restricted to a period ending at ∼200 d of age. The results demonstrate that the timing and even the existence of sensitive periods for plasticity of a neural circuit and associated behavior can depend on multiple factors, including (1) the nature of the adjustment demanded of the system and (2) the richness of the sensory and social environment in which the plasticity is studied.