bluuebunny/crossref_metadata_2025 huggingface dataset:
Estrogen Levels Regulate the Subcellular Distribution of Phosphorylated Akt in Hippocampal CA1 Dendrites
Authors: Znamensky Vladimir , Akama Keith T. , McEwen Bruce S. , Milner Teresa A.
Published: April 2018

In addition to genomic pathways, estrogens may regulate gene expression by activating specific signal transduction pathways, such as that involving phosphatidylinositol 3-kinase (PI3-K) and the subsequent phosphorylation of Akt (protein kinase B). The Akt pathway regulates various cellular events, including the initiation of protein synthesis. Our previous studies showed that synaptogenesis in hippocampal CA1 pyramidal cell dendritic spines is highest when brain estrogen levels are highest. To address the role of Akt in this process, the subcellular distribution of phosphorylated Akt immunoreactivity (pAkt-I) in the hippocampus of female rats across the estrous cycle and male rats was analyzed by light microscopy (LM) and electron microscopy (EM). By LM, the density of pAkt-I in stratum radiatum of CA1 was significantly higher in proestrus rats (or in estrogen-supplemented ovariectomized females) compared with diestrus, estrus, or male rats. By EM, pAkt-I was found throughout the shafts and in select spines of stratum radiatum dendrites. Quantitative ultrastructural analysis identifying pAkt-I with immunogold particles revealed that proestrus rats compared with diestrus, estrus, and male rats contained significantly higher pAkt-I associated with (1) dendritic spines (both cytoplasm and plasmalemma), (2) spine apparati located within 0.1 μm of dendritic spine bases, (3) endoplasmic reticula and polyribosomes in the cytoplasm of dendritic shafts, and (4) the plasmalemma of dendritic shafts. These findings suggest that estrogens may regulate spine formation in CA1 pyramidal neurons via Akt-mediated signaling events.
Decoding Spike Trains Instant by Instant Using Order Statistics and the Mixture-of-Poissons Model
Authors: Wiener Matthew C. , Richmond Barry J.
Published: April 2018

In the brain, spike trains are generated in time and presumably also interpreted as they unfold in time. Recent work (Oram et al., 1999; Baker and Lemon, 2000) suggests that in several areas of the monkey brain, individual spike times carry information because they reflect an underlying rate variation. Constructing a model based on this stochastic structure allows us to apply order statistics to decode spike trains instant by instant as spikes arrive or do not. Order statistics are time-consuming to compute in the general case. We demonstrate that data from neurons in primary visual cortex are well fit by a mixture of Poisson processes; in this special case, our computations are substantially faster. In these data, spike timing contributed information beyond that available from the spike count throughout the trial. At the end of the trial, a decoder based on the mixture-of-Poissons model correctly decoded about three times as many trials as expected by chance, compared with approximately twice as many as expected by chance using the spike count only. If our model perfectly described the spike trains, and enough data were available to estimate model parameters, then our Bayesian decoder would be optimal. For four-fifths of the sets of stimulus-elicited responses, the observed spike trains were consistent with the mixture-of-Poissons model. Most of the error in estimating stimulus probabilities is attributable to not having enough data to specify the parameters of the model rather than to misspecification of the model itself.
Bimodal Locomotion Elicited by Electrical Stimulation of the Midbrain in the SalamanderNotophthalmus viridescens
Authors: Cabelguen Jean-Marie , Bourcier-Lucas Céline , Dubuc Réjean
Published: April 2018

The present experiments were designed to identify the mesencephalic locomotor region (MLR) in the salamander. Anin vitrosemi-intact preparation from a decerebrate adult salamander (Notophthalmusviridescens) was developed in which the locomotor activities were monitored from electromyographic and video recordings. The results show that the two locomotor modes exhibited by salamanders (i.e., stepping and swimming) were evoked by electrical microstimulation (5–15 Hz; 0.1–10 μA; 2 msec pulses) of a circumscribed region in the caudal mesencephalon. At threshold current strength (0.5–3.5 μA at 15 Hz), rhythmic limb movements and intersegmental coordination, such as during stepping, were induced. As the stimulation strength was subsequently increased, the frequency of stepping became more rapid, and, at 2.0–5.5 μA, the limbs were held back against the body wall and swimming movements of the trunk were induced. An additional increase of the stimulation strength induced an increase of the frequency and amplitude of the swimming movements. Anatomical studies conducted in parallel revealed the presence of choline acetyltransferase immunoreactive cells in the functionally identified MLR region. Together, the present results indicate that the MLR is present in salamanders and that its level of activation determines the mode of locomotion. Walking is induced at low activation levels, and swimming, which constitutes a faster mode of locomotion, requires stronger stimulation of the MLR. Furthermore, as in other vertebrates, the MLR contains cholinergic cells.
Submillisecond Synchronization of Fast Electrical Oscillations in Neocortex
Authors: Barth Daniel S.
Published: April 2018

Fast electrical oscillations (FOs; >200 Hz) in the sensory neocortex can be recorded in a variety of species, including humans, and may reflect extremely fast integration of sensory information. This report demonstrates that, in the whisker representation of rat cortex, multivibrissa stimulation produces propagating FO field potential patterns and time-locked unit activity that are sensitive to submillisecond delays in interstimulus intervals. We propose that FOs may be produced by synchronized population spikes and their subthreshold sequelas in cortical pyramidal cells. FOs serve to accurately mark stimulus onset as a phase-encoded excitatory signal, producing phase-sensitive interactions that, in the context of exploratory whisking, may extract features of an object under exploration.
Imaging Calcium Entry Sites and Ribbon Structures in Two Presynaptic Cells
Authors: Zenisek David , Davila Viviana , Wan Lei , Almers Wolfhard
Published: April 2018

We investigated the location of calcium entry sites and synaptic ribbons in the type-Mb goldfish bipolar neuron and the bullfrog saccular hair cell. Cells were loaded with a fast calcium indicator (Fluo-3 or Fluo-5F) and an excess of a high-affinity but slow Ca buffer (EGTA). The cell surface was imaged by evanescent field microscopy. Small fluorescent “hot spots” representing calcium entry sites appeared abruptly when a voltage step opened Ca channels and disappeared or dimmed abruptly when Ca channels closed. In bipolar cells, the fluorescence of hot spots tracked the calcium influx. Hair cells showed similar Ca hot spots. Synaptic ribbons or dense bodies were labeled by immunofluorescence with an antibody that recognizes the ribbon protein ribeye. The antibody labeled punctate structures beneath the plasma membrane. In both bipolar neurons and hair cells, the number of Ca entry sites was similar or identical to that of ribbons or dense bodies, consistent with the idea that calcium-channel clusters reside near ribbons, and that both mark active zones. In bipolar cells, the number of Ca entry sites and ribeye-positive fluorescent spots is also strikingly similar to that of exocytic active zones but significantly less than the number of total exocytic sites including solitary fusion events outside active zones. We suggest that in bipolar terminals, active zones, Ca entry sites, and synaptic ribbons all colocalize, but also that a significant number of vesicles can fuse outside active zones and, hence, independently of synaptic ribbons.
Endocytosis and Vesicle Recycling at a Ribbon Synapse
Authors: Paillart Christophe , Li Jian , Matthews Gary , Sterling Peter
Published: April 2018

At ribbon synapses, where exocytosis is regulated by graded depolarization, vesicles can fuse very rapidly with the plasma membrane (complete discharge of the releasable pool in ∼200 msec). Vesicles are also retrieved very rapidly (time constant of ∼1 sec), leading us to wonder whether their retrieval uses an unusual mechanism. To study this, we exposed isolated bipolar neurons from goldfish retina to cationized ferritin. This electron-dense marker uniformly decorated the cell membrane and was carried into the cell during membrane retrieval. Endocytosis was activity-dependent and restricted to the synaptic terminal. The labeling pattern was consistent with direct retrieval from the plasma membrane of large, uncoated endosomes 60–200 nm in diameter. Even after extensive synaptic activity lasting several minutes, most of the ferritin remained in large endosomes and was present in only ∼10% of the small vesicles that constitute the reserve pool. By contrast, after brief stimulation at a conventional terminal, ferritin did not reside in endosomes but was present in ∼63% of the small vesicles. We suggest that the bipolar ribbon synapse sustains its rapid exocytosis by retrieving membrane in larger “bites” than the clathrin-dependent mechanism thought to dominate at conventional synapses. The resulting large endosomes bud off small vesicles, which reenter the reserve pool and finally the releasable pool.
Neocortical Long-Term Potentiation and Experience-Dependent Synaptic Plasticity Require α-Calcium/Calmodulin-Dependent Protein Kinase II Autophosphorylation
Authors: Hardingham Neil , Glazewski Stanislaw , Pakhotin Pavel , Mizuno Keiko , Chapman Paul F. , Giese K. Peter , Fox Kevin
Published: April 2018

Experience-dependent plasticity can be induced in the barrel cortex by removing all but one whisker, leading to potentiation of the neuronal response to the spared whisker. To determine whether this form of potentiation depends on synaptic plasticity, we studied long-term potentiation (LTP) and sensory-evoked potentials in the barrel cortex of α-calcium/calmodulin-dependent protein kinase II (αCaMKII)T286Amutant mice. We studied three different forms of LTP induction: theta-burst stimulation, spike pairing, and postsynaptic depolarization paired with low-frequency presynaptic stimulation. None of these protocols produced LTP in αCaMKIIT286Amutant mice, although all three were successful in wild-type mice. To study synaptic plasticityin vivo, we measured sensory-evoked potentials in the barrel cortex and found that single-whisker experience selectively potentiated synaptic responses evoked by sensory stimulation of the spared whisker in wild types but not in αCaMKIIT286Amice. These results demonstrate that αCaMKII autophosphorylation is required for synaptic plasticity in the neocortex, whether induced by a variety of LTP induction paradigms or by manipulation of sensory experience, thereby strengthening the case that the two forms of plasticity are related.
Degeneration of the Amygdala/Piriform Cortex and Enhanced Fear/Anxiety Behaviors in Sodium Pump α2 Subunit (Atp1a2)-Deficient Mice
Authors: Ikeda Keiko , Onaka Tatsushi , Yamakado Makoto , Nakai Junichi , Ishikawa Tomo-o , Taketo Makoto M. , Kawakami Kiyoshi
Published: April 2018

The sodium pump is the enzyme responsible for the maintenance of Na+and K+gradients across the cell membrane. Four isoforms of the catalyticα subunit have been identified, but their individual roles remain essentially unknown. To investigate the necessary functions of the α2 subunitin vivo, we generated and analyzed mice defective in the α2 subunit gene. Mice homozygous for the α2 mutation died just after birth and displayed selective neuronal apoptosis in the amygdala and piriform cortex. In these regions, high expression of c-Fos before apoptosis indicated neural hyperactivity, and re-uptake of glutamic acid and GABA into P2fraction containing crude synaptosome was impaired. These results indicate that the α2 subunit plays a critical role regulating neural activity in the developing amygdala and piriform cortex. Further supporting a role of theα2 subunit in the function of the amygdala, heterozygous adult mice showed augmented fear/anxiety behaviors and enhanced neuronal activity in the amygdala and piriform cortex after conditioned fear stimuli.
Early Maintenance of Hippocampal Mossy Fiber—Long-Term Potentiation Depends on Protein and RNA Synthesis and Presynaptic Granule Cell Integrity
Authors: Calixto Eduardo , Thiels Edda , Klann Eric , Barrionuevo Germán
Published: April 2018

The neural substrates of memory likely include long-term potentiation (LTP) of synaptic strength that results from high-frequency stimulation (HFS) of the afferent pathway. The mechanisms that underlie the maintenance of LTP include RNA and protein synthesis, although the contribution of these molecular events typically does not become essential until several hours after LTP induction. We here show that, different from this pattern, (1) LTP maintenance at the mossy fiber (MF) input to CA3 pyramidal cells in the hippocampus depends on protein and RNA synthesis soon after LTP induction, and (2) some of these molecular events are controlled by signaling from the presynaptic granule cell soma. Bath application of the protein synthesis inhibitor emetine or cycloheximide 1 hr after MF LTP induction in hippocampal slices caused loss of MF potentiation. In contrast, application of emetine 1 hr after LTP induction at the commissural—associational input to CA3 pyramidal cells had no effect on this form of LTP. Administration of emetine or the RNA synthesis inhibitor actinomycin-D before delivery of HFS to MF input also caused a rapid decay of MF potentiation, although neither drug had an effect on the amplitude or the time-constant of decay of post-tetanic potentiation (PTP). Similarly, transection of MF axons near granule cell somas had no effect on baseline or PTP parameters but caused loss of potentiation at a rate comparable with that after actinomycin-D application. These results indicate that the mechanisms that underlie MF LTP maintenance differ from those involved in LTP maintenance at other glutamatergic synapses.
Intracellular Patch Electrochemistry: Regulation of Cytosolic Catecholamines in Chromaffin Cells
Authors: Mosharov Eugene V. , Gong Liang-Wei , Khanna Bhavanna , Sulzer David , Lindau Manfred
Published: April 2018

Alterations in the cytosolic pool directly affect neurotransmitter synthesis and release and are suggested to be key factors in various neurodegenerative disorders. Although this cytosolic pool is the most metabolically active, it is miniscule compared with the amount of vesicular transmitter and has never been quantified separately. Here, we introduce intracellular patch electrochemistry (IPE), a technique that for the first time provides direct measurements of cytosolic oxidizable molecules in single mammalian cells. In amperometric mode, IPE detects total catechols, whereas in cyclic voltammetric mode, it preferentially measures catecholamines. In cultured chromaffin cells, the total cytosolic catechol concentration was 50–500 μm, of which ∼10% were catecholamines. Reserpine, a vesicular monoamine transporter inhibitor, had no effect on the catecholamine pool but increased total catechols by fourfold to fivefold. Combined with pargyline, a monoamine oxidase inhibitor, reserpine increased catecholamine levels in the cytosol by approximately sixfold. Amphetamine induced a transient approximately fivefold accumulation of cytosolic catecholamines and a slow increase of total catechols. In cells incubated with 3,4-dihydroxy-L-phenylalanine (l-DOPA), catecholamines increased by ∼2.5-fold and total catechols increased by approximately fourfold. Cytosolic catecholamines returned to control levels ≤10 min afterl-DOPA withdrawal, whereas total catechols remained approximately twofold elevated even after a 1.5 hr incubation inl-DOPA-free media. Our data indicate that cytosolic catecholamines are strictly maintained at a defined level, and drug-induced increases in their concentrations lead to the accumulation of other catecholamine derivatives, such as DOPAC and 3,4-dihydroxyphenylethyleneglycol. These derivatives reside in the cytosol for hours after treatment and may be an underlying cause of drug-related cytotoxicity.
Characterization of Depolarization-Induced Suppression of Inhibition Using Paired Interneuron–Purkinje Cell Recordings
Authors: Diana Marco A. , Marty Alain
Published: April 2018

Depolarization-induced suppression of inhibition (DSI) is a retrograde form of synaptic inhibition involving the Ca2+-dependent release of cannabinoids from the postsynaptic cell. DSI exerts multiple effects on presynaptic neurons: here, we establish the breakdown of DSI in its individual components at the synapses between basket and stellate cells and Purkinje cells. In the presence of tetrodotoxin, the change in IPSC frequency entirely accounted for the decrease of transmission during DSI; in contrast, without tetrodotoxin, the reductions of frequency and average amplitude gave equal contributions. In paired recordings, transmission displayed an irreversible rundown unless interneurons were recorded from with the perforated patch method. Under these conditions, a DSI of 68.8% was measured; the failure rate and the paired pulse ratio (at 20 msec intervals) increased from 1.2 to 20.2 and 95.6 to 132.6%, respectively, and the variance to mean ratio augmented 2.17-fold. Presynaptic dialysis with Cs+led to a major potentiation of synaptic strength and to a marked reduction of DSI with respect to control potassium conditions; DSI recovered only partially when decreasing the extracellular Ca2+concentration to match the control IPSC amplitudes. These results, combined with those of Kreitzer et al. (2002), indicate that three distinct presynaptic processes contribute to DSI: reductions of miniature frequency (13.4% of total DSI), of presynaptic action potential frequency (23.2%), and of the probability that presynaptic depolarizations elicit transmitter release (63.4%). The latter component involves a modulation of K+channels and trial-to-trial modifications of the presynaptic signal.
Enhanced Expression of a Specific Hyperpolarization-Activated Cyclic Nucleotide-Gated Cation Channel (HCN) in Surviving Dentate Gyrus Granule Cells of Human and Experimental Epileptic Hippocampus
Authors: Bender Roland A. , Soleymani Sheila V. , Brewster Amy L. , Nguyen Snow T. , Beck Heinz , Mathern Gary W. , Baram Tallie Z.
Published: April 2018

Changes in the expression of ion channels, contributing to altered neuronal excitability, are emerging as possible mechanisms in the development of certain human epilepsies. In previous immature rodent studies of experimental prolonged febrile seizures, isoform-specific changes in the expression of hyperpolarization-activated cyclic nucleotide-gated cation channels (HCNs) correlated with long-lasting hippocampal hyperexcitability and enhanced seizure susceptibility. Prolonged early-life seizures commonly precede human temporal lobe epilepsy (TLE), suggesting that transcriptional dysregulation of HCNs might contribute to the epileptogenic process. Therefore, we determined whether HCN isoform expression was modified in hippocampi of individuals with TLE. HCN1 and HCN2 expression were measured usingin situhybridization and immunocytochemistry in hippocampi from three groups: TLE with hippocampal sclerosis (HS;n= 17), epileptic hippocampi without HS, or non-HS (NHS;n= 10), and autopsy material (n= 10). The results obtained in chronic human epilepsy were validated by examining hippocampi from the pilocarpine model of chronic TLE.

In autopsy and most NHS hippocampi, HCN1 mRNA expression was substantial in pyramidal cell layers and lower in dentate gyrus granule cells (GCs). In contrast, HCN1 mRNA expression over the GC layer and in individual GCs from epileptic hippocampus was markedly increased once GC neuronal density was reduced by >50%. HCN1 mRNA changes were accompanied by enhanced immunoreactivity in the GC dendritic fields and more modest changes in HCN2 mRNA expression. Furthermore, similar robust and isoform-selective augmentation of HCN1 mRNA expression was evident also in the pilocarpine animal model of TLE. These findings indicate that the expression of HCN isoforms is dynamically regulated in human as well as in experimental hippocampal epilepsy. After experimental febrile seizures (i.e., early in the epileptogenic process), the preserved and augmented inhibition onto principal cells may lead to reduced HCN1 expression. In contrast, in chronic epileptic HS hippocampus studied here, the profound loss of interneuronal and principal cell populations and consequent reduced inhibition, coupled with increased dendritic excitation of surviving GCs, might provoke a “compensatory” enhancement of HCN1 mRNA and protein expression.
Apamin-Sensitive Small Conductance Calcium-Activated Potassium Channels, through their Selective Coupling to Voltage-Gated Calcium Channels, Are Critical Determinants of the Precision, Pace, and Pattern of Action Potential Generation in Rat Subthalamic Nucleus NeuronsIn Vitro
Authors: Hallworth Nicholas E. , Wilson Charles J. , Bevan Mark D.
Published: April 2018

Distinct activity patterns in subthalamic nucleus (STN) neurons are observed during normal voluntary movement and abnormal movement in Parkinson's disease (PD). To determine how such patterns of activity are regulated by small conductance (SK) calcium-activated potassium channels (KCa) and voltage-gated calcium (Cav) channels, STN neurons were recorded in the perforated patch configuration in slices, [which were prepared from postnatal day 16 (P16)-P30 rats and held at 37°C] and then treated with the SK KCachannel antagonist apamin or the SK KCaagonist 1-ethyl-2-benzimidazolinone or the Cavchannel antagonists ω-conotoxin GVIA (Cav2.2-selective) or nifedipine (Cav1.2-1.3-selective). In other experiments, fura-2 was introduced as an indicator of intracellular calcium dynamics.

A component of the current underlying single-spike afterhyperpolarization was sensitive to apamin, phase-locked to calcium entry via Cav2.2 channels, and necessary for precise, autonomous, single-spike oscillation. SK KCa/Cav2.2 channel coupling did not underlie spike-frequency adaptation but limited activity in response to current injection by encoding the accumulation of intracellular calcium, maintained the characteristic sigmoidal frequency-intensity relationship and generated a post-train afterhyperpolarization. In addition, SK KCachannels terminated rebound burst activity more effectively in neurons with short-duration bursts (<100 msec) than neurons with long-duration bursts (>100 msec), presumably through their activation by Cav3 channels. Cav1.2-1.3 channels were not strongly coupled to SK KCachannels and therefore supported secondary range and long-duration rebound burst firing. In summary, SK KCachannels play a fundamental role in autonomous, driven, and rebound activity and oppose the transition from autonomous, rhythmic, single-spike activity to burst firing in STN neurons.
Circadian Clock-Controlled Regulation of cGMP-Protein Kinase G in the Nocturnal Domain
Authors: Tischkau Shelley A. , Weber E. Todd , Abbott Sabra M. , Mitchell Jennifer W. , Gillette Martha U.
Published: April 2018

The suprachiasmatic nucleus (SCN) circadian clock exhibits a recurrent series of dynamic cellular states, characterized by the ability of exogenous signals to activate defined kinases that alter clock time. To explore potential relationships between kinase activation by exogenous signals and endogenous control mechanisms, we examined clock-controlled protein kinase G (PKG) regulation in the mammalian SCN. Signaling via the cGMP-PKG pathway is required for light- or glutamate (GLU)-induced phase advance in late night. Spontaneous cGMP-PKG activation occurred at the end of subjective night in free-running SCNin vitro. Phasing of the SCN rhythmin vitrowas delayed by ∼3 hr after treatment with guanylyl cyclase (GC) inhibitors, PKG inhibition, or antisense oligodeoxynucleotide (αODN) specific for PKG, but not PKA inhibitor or mismatched ODN. This sensitivity to GC-PKG inhibition was limited to the same 2 hr time window demarcated by clock-controlled activation of cGMP-PKG. Inhibition of the cGMP-PKG pathway at this time caused delays in the phasing of four endogenous rhythms: wheel-running activity, neuronal activity, cGMP, andPer1. Timing of the cGMP-PKG-necessary window in both rat and mouse depended on clock phase, established by the antecedent light/dark cycle rather than solar time. Because behavioral, neurophysiological, biochemical, and molecular rhythms showed the same temporal sensitivities and qualitative responses, we predict that clock-regulated GC-cGMP-PKG activation may provide a necessary cue as to clock state at the end of the nocturnal domain. Because sensitivity to phase advance by light-GLU-activated GC-cGMP-PKG occurs in juxtaposition, these signals may induce a premature shift to this PKG-necessary clock state.
Mechanisms of Lateral Inhibition in the Olfactory Bulb: Efficiency and Modulation of Spike-Evoked Calcium Influx into Granule Cells
Authors: Egger Veronica , Svoboda Karel , Mainen Zachary F.
Published: April 2018

Granule cells are axonless local interneurons that mediate lateral inhibitory interactions between the principal neurons of the olfactory bulb via dendrodendritic reciprocal synapses. This unusual arrangement may give rise to functional properties different from conventional lateral inhibition. Although granule cells spike, little is known about the role of the action potential with respect to their synaptic output. To investigate the signals that underlie dendritic release in these cells, two-photon microscopy in rat brain slices was used to image calcium transients in granule cell dendrites and spines. Action potentials evoked calcium transients throughout the dendrites, with amplitudes increasing with distance from soma and attaining a plateau level within the external plexiform layer, the zone of granule cell synaptic output. Transient amplitudes were, on average, equal in size in spines and adjacent dendrites. Surprisingly, both spine and dendritic amplitudes were strongly dependent on membrane potential, decreasing with depolarization and increasing with hyperpolarization from rest. Both the current-voltage relationship and the time course of inactivation were consistent with the known properties of T-type calcium channels, and the voltage dependence was blocked by application of the T-type calcium channel antagonists Ni2+and mibefradil. In addition, mibefradil reduced action potential-mediated synaptic transmission from granule to mitral cells. The implication of a transiently inactivating calcium channel in synaptic release from granule cells suggests novel mechanisms for the regulation of lateral inhibition in the olfactory bulb.
Phospholipase Cγ in Distinct Regions of the Ventral Tegmental Area Differentially Modulates Mood-Related Behaviors
Authors: Bolaños Carlos A. , Perrotti Linda I. , Edwards Scott , Eisch Amelia J. , Barrot Michel , Olson Valerie G. , Russell David S. , Neve Rachael L. , Nestler Eric J.
Published: April 2018

Neurotrophic factor signaling pathways modulate cellular and behavioral responses to drugs of abuse. In addition, chronic exposure to morphine increases expression of phospholipase Cγ1 (PLCγ1) (a protein involved in neurotrophic signaling) in the ventral tegmental area (VTA), a neural substrate for many drugs of abuse. Using viral-mediated gene transfer to locally alter the activity of PLCγ1, we show that overexpression of PLCγ1 in rostral portions of the VTA (R-VTA) results in increased morphine place preference, whereas PLCγ1 overexpression in the caudal VTA (C-VTA) results in avoidance of morphine-paired compartments. In addition, overexpression of PLCγ1 in R-VTA causes increased preference for sucrose and increased anxiety-like behavior but does not affect responses to stress or nociceptive stimuli. In contrast, overexpression of PLCγ1 in C-VTA decreases preference for sucrose and increases sensitivity to stress and nociceptive stimuli, although there was a tendency for increased anxiety-like behavior as seen for the R-VTA. These results show that levels of PLCγ1 in the VTA regulate responsiveness to drugs of abuse, natural rewards, and aversive stimuli and point to the possibility that distinct topographical regions within the VTA mediate generally positive versus negative responses to emotional stimuli. Moreover, these data also support a role for drug-induced elevations in PLCγ1 expression in the VTA in mediating long-term adaptations to drugs of abuse and aversive stimuli.
Sodium Channel β4, a New Disulfide-Linked Auxiliary Subunit with Similarity to β2
Authors: Yu Frank H. , Westenbroek Ruth E. , Silos-Santiago Inmaculada , McCormick Kimberly A. , Lawson Deborah , Ge Pei , Ferriera Holly , Lilly Jeremiah , DiStefano Peter S. , Catterall William A. , Scheuer Todd , Curtis Rory
Published: April 2018

The principal α subunit of voltage-gated sodium channels is associated with auxiliary β subunits that modify channel function and mediate protein-protein interactions. We have identified a new β subunit termed β4. Like the β1-β3 subunits, β4 contains a cleaved signal sequence, an extracellular Ig-like fold, a transmembrane segment, and a short intracellular C-terminal tail. Using TaqMan reverse transcription-PCR analysis,in situhybridization, and immunocytochemistry, we show that β4 is widely distributed in neurons in the brain, spinal cord, and some sensory neurons.β4 is most similar to theβ2 subunit (35% identity), and, like theβ2 subunit, the Ig-like fold of β4 contains an unpaired cysteine that may interact with the α subunit. Under nonreducing conditions, β4 has a molecular mass exceeding 250 kDa because of its covalent linkage to Nav1.2a, whereas on reduction, it migrates with a molecular mass of 38 kDa, similar to the mature glycosylated forms of the otherβ subunits. Coexpression ofβ4 with brain Nav1.2a and skeletal muscle Nav1.4 α subunits in tsA-201 cells resulted in a negative shift in the voltage dependence of channel activation, which overrode the opposite effects ofβ1 andβ3 subunits when they were present. This novel, disulfide-linked β subunit is likely to affect both protein-protein interactions and physiological function of multiple sodium channel α subunits.
Semaphorin 3F Antagonizes Neurotrophin-Induced Phosphatidylinositol 3-Kinase and Mitogen-Activated Protein Kinase Kinase Signaling: A Mechanism for Growth Cone Collapse
Authors: Atwal Jasvinder K. , Singh Karun K. , Tessier-Lavigne Marc , Miller Freda D. , Kaplan David R.
Published: April 2018

Peripheral nerve growth is regulated by the coordinated action of numerous external stimuli, including positively acting neurotrophin-derived growth cues and restrictive semaphorin cues. Here, we show that Semaphorin 3F (Sema 3F) can antagonize nerve growth factor (NGF)-stimulated TrkA (tyrosine receptor kinase A) signaling in sympathetic neurons, thereby apparently contributing to growth cone collapse. Sema 3F suppressed NGF-induced activation of the phosphatidylinositol 3 (PI3)-kinase-Akt and MEK (mitogen-activated protein kinase kinase)-ERK (extracellular signal-regulated kinase) pathways, both of which we show to be required to maintain growth cone structure. Sema 3F-induced growth cone collapse was partially reversed by sustained activation of the PI3-kinase and MEK pathways, which was achieved by overexpression of the Gab-1 (growth-associated binder 1) docking protein. These data indicate that a novel mechanism used by Sema 3F to collapse growth cones in sympathetic neurons is to dampen neurotrophin signaling, providing an intracellular mechanism for cross talk between positive and negative axon growth cues.
Developmental Modulation of Retinal Wave Dynamics: Shedding Light on the GABA Saga
Authors: Sernagor Evelyne , Young Carol , Eglen Stephen J.
Published: April 2018

Embryonic spontaneous activity, in the form of propagating waves, is crucial for refining visual connections. To study what aspects of this correlated activity are instructive, we must first understand how their dynamics change with development and what factors trigger their disappearance after birth. Here we report that in the turtle retina, GABA, rather than glutamate and acetylcholine, influences developmental changes in wave dynamics. Using calcium imaging of the ganglion cell layer, we report how waves switch from fast and broad, when they emerge, to slow and narrow a few days before hatching, coinciding with the emergence of excitatory GABAAreceptor-mediated activity. Around hatching, waves gradually become stationary patches, whereas GABAAshifts from excitatory to inhibitory, coinciding with the upregulation of the cotransporter KCC2, suggesting that changes in intracellular chloride underlie the shift. Dark-rearing from hatching causes correlated spontaneous activity to persist, whereas GABAAresponses remain excitatory, and KCC2 expression is weaker. We conclude that GABA plays an important regulatory role during the maturation of retinal neural activity. Using a simple and elegant mechanism, namely the switch from excitatory to inhibitory, GABAAreceptor-mediated activity is necessary and sufficient to cause retinal waves to stop propagating, ultimately leading to the disappearance of correlated spontaneous activity. Moreover, our results suggest that visual experience modulates the GABAergic switch.
Functional Synaptic Contacts by Intranuclear Axon Collaterals of Thalamic Relay Neurons
Authors: Cox Charles L. , Reichova Iva , Sherman S. Murray
Published: April 2018

Relay neurons of the lateral geniculate nucleus innervate visual cortex, but they also provide axonal collaterals to neurons in the thalamic reticular nucleus, and these thalamic reticular neurons provide feedback inhibition to relay cells. An alternative source of inhibitory inputs onto geniculate relay neurons arises from intralaminar interneurons that provide feedforward inhibition via retinogeniculate innervation, and perhaps feedback inhibition via the corticothalamic pathway, analogous to that involving thalamic reticular neurons. Several reports indicate that relay neurons may also give rise to axonal collaterals within the lateral geniculate nucleus, constituting another route for feedback or local integration. We now provide new data indicating that collaterals from geniculate relay neurons provide excitatory input to local intralaminar interneurons and that this pathway may serve as a previously unknown means of local feedback inhibition. This circuitry could prove important in such activities as surround inhibition of receptive fields or increasing signal gain over noise.
Transient Activation of Superior Prefrontal Cortex during Inhibition of Cognitive Set
Authors: Konishi Seiki , Jimura Koji , Asari Tomoki , Miyashita Yasushi
Published: April 2018

The prefrontal cortex implements a set-shifting function that includes inhibition of a previously acquired cognitive set. The impairment of the inhibitory function results in perseverative behavior that forms one characteristic feature of frontal lobe dysfunction. Previous neuroimaging studies have revealed inhibitory mechanisms in the inferior prefrontal cortex. The present functional magnetic resonance imaging study devised “dual-match” stimuli in a set-shifting paradigm that allowed us to temporally isolate the inhibitory processes recruited during exposure to a previously acquired set. Transient activation time-locked to the isolated inhibition was revealed in the left middle frontal gyrus near the superior frontal sulcus. In a control experiment conducted after subjects had been informed and made aware of the exposure, however, the superior prefrontal activation disappeared, and prominent activation was revealed in a set of brain regions that included the left posterior inferior frontal sulcus. These double dissociation results indicate inhibitory mechanisms in the superior prefrontal cortex, alternative to the inferior prefrontal ones, that are activated depending on the subjects' strategy for inhibition of cognitive set.
Novel Insights into the Regulation of the Timeless Protein
Authors: Ashmore Lesley J. , Sathyanarayanan Sriram , Silvestre David W. , Emerson Mark M. , Schotland Peter , Sehgal Amita
Published: April 2018

In theDrosophilacircadian clock,period(per) and its partner,timeless(tim), play a central role in the negative limb of an autoregulatory feedback loop. Unlikeper, the dosage of which affects the frequency (tau) of the circadian cycle, we found that increasing copies of thetimgene has no effect on clock period length. The use of thetimpromoter to expressperresults in a shortening of circadian period, also indicating that the regulation oftimis different from that ofper. DrosophilaTIM is similar to the mammalian circadian protein mPER2 in that it shuttles independently between the nucleus and cytoplasm bothin vivoandin vitro. Contrary to the current model that PER and TIM heterodimerization is a prerequisite for their nuclear entry, PER is not required to transport TIM into nuclei, although it influences TIM localization and vice versa. Blocking nuclear export led to increased nuclear expression of TIM in S2 cells and in wild-type andper01larvae, suggesting that PER may be required for nuclear retention of TIM. Unlike PER, nuclear TIM alone has no ability to repress transcription. We propose that TIM drives cycles of PER expression by regulating its stability, and in turn, PER retains TIM in the nucleus, either for the regulation of its own stability or for a novel nuclear role of TIM.
Subunit Composition of Functional Nicotinic Receptors in Dopaminergic Neurons Investigated with Knock-Out Mice
Authors: Champtiaux Nicolas , Gotti Cecilia , Cordero-Erausquin Matilde , David Denis J. , Przybylski Cédric , Léna Clément , Clementi Francesco , Moretti Milena , Rossi Francesco M. , Le Novère Nicolas , McIntosh J. Michael , Gardier Alain M. , Changeux Jean-Pierre
Published: April 2018

Nicotinic acetylcholine receptors (nAChRs) expressed by dopaminergic (DA) neurons have long been considered as potential therapeutic targets for the treatment of several neuropsychiatric diseases, including nicotine and cocaine addiction or Parkinson's disease. However, DA neurons express mRNAs coding for most, if not all, neuronal nAChR subunits, and the subunit composition of functional nAChRs has been difficult to establish. Immunoprecipitation experiments performed on mouse striatal extracts allowed us to identify three main types of heteromeric nAChRs (α4β2*, α6β2*, and α4α6β2*) in DA terminal fields. The functional relevance of these subtypes was then examined by studying nicotine-induced DA release in striatal synaptosomes and recording ACh-elicited currents in DA neurons fromα4, α6, α4α6, and β2 knock-out mice. Our results establish that α6β2*nAChRs are functional and sensitive to α-conotoxin MII inhibition. These receptors are mainly located on DA terminals and consistently do not contribute to DA release induced by systemic nicotine administration, as evidenced byin vivomicrodialysis. In contrast, (nonα6)α4β2*nAChRs represent the majority of functional heteromeric nAChRs on DA neuronal soma. Thus, whereas a combination of α6β2*and α4β2*nAChRs may mediate the endogenous cholinergic modulation of DA release at the terminal level, somato-dendritic (nonα6)α4β2*nAChRs most likely contribute to nicotine reinforcement.
Apolipoprotein E Markedly Facilitates Age-Dependent Cerebral Amyloid Angiopathy and Spontaneous Hemorrhage in Amyloid Precursor Protein Transgenic Mice
Authors: Fryer John D. , Taylor Jennie W. , DeMattos Ronald B. , Bales Kelly R. , Paul Steven M. , Parsadanian Maia , Holtzman David M.
Published: April 2018

Cerebral amyloid angiopathy (CAA) is a common cause of brain hemorrhage in the elderly. It is found in the majority of patients with Alzheimer's disease (AD). The most common form of CAA is characterized by the deposition of the amyloid-β (Aβ) peptide in the walls of cerebral vessels, and this deposition can lead to hemorrhage and infarction. As in AD, the ϵ4 allele of apolipoprotein E (APOE) is a risk factor for CAA. To determine the effect of apoE on CAA and associated hemorrhagein vivo, we used two amyloid precursor protein (APP) transgenic mouse models that develop age-dependent Aβ deposition: PDAPP and APPsw mice. We found that both models developed an age-dependent increase in CAA and associated microhemorrhage, with the APPsw model having an earlier and more severe phenotype; however, when APPsw and PDAPP mice were bred onto anApoe-/- background, no CAA was detected through 24 months of age, and there was little to no evidence of microhemorrhage. Biochemical analysis of isolated cerebral vessels from both PDAPP and APPsw mice with CAA revealed that, as in human CAA, the ratio of Aβ 40:42 was elevated relative to brain parenchyma. In contrast, the ratio of Aβ 40:42 from cerebral vessels isolated from old PDAPP,Apoe-/- mice was extremely low. These findings demonstrate that murine apoE markedly promotes the formation of CAA and associated vessel damage and that the effect of apoE combined with the level of Aβ40or the ratio of Aβ 40:42 facilitates this process.
Cerebellar Climbing Fibers Modulate Simple Spikes in Purkinje Cells
Authors: Barmack Neal H. , Yakhnitsa Vadim
Published: April 2018

Purkinje cells have two action potentials: Climbing fiber responses (CFRs) and simple spikes (SSs). CFRs reflect the discharge of a single climbing fiber at multiple synaptic sites on the proximal dendrite of the Purkinje cell. SSs reflect the summed action of a subset of parallel fiber synapses on Purkinje cell dendritic spines. Because mossy fiber afferents terminate on granule cells, the ascending axons of which bifurcate, giving rise to parallel fibers, the modulation of SSs has been attributed to mossy fiber afferent signals. This inference has never been tested. Conversely, the low discharge frequency of CFRs has led many to conclude that they have a unique and intermittent role in cerebellar signal processing. We examine the relative potency of vestibularly modulated mossy fiber and climbing fiber signals in evoking CFRs and SSs in Purkinje cells of the uvula-nodulus in chloralose-urethane-anesthetized rabbits. Vestibular primary afferents were blocked by unilateral labyrinthectomy (UL). A UL destroys the vestibular primary afferent signal to the ipsilateral uvula-nodulus, while leaving intact the vestibular climbing fiber signal from the contralateral inferior olive. After UL, vestibular stimulation modulated CFRs and SSs in ipsilateral uvula-nodular Purkinje cells, demonstrating that the primary vestibular afferent mossy fiber input to the ipsilateral uvula-nodulus was not necessary for SS modulation. Unilateral microlesions of the caudal half of the β-nucleus of the inferior olive reduced a modulated climbing fiber signal to the contralateral uvula-nodulus, causing loss of both vestibularly modulated CFRs and SSs in contralateral Purkinje cells. Vestibular climbing fibers not only evoke low-frequency CFRs, but also indirectly modulate higher-frequency SSs. This modulation must be attributed to cerebellar interneurons. Golgi cell inhibition of granule cells may provide the interneuronal mechanism for CFR-induced SS modulation.
Chemokine Expression by Glial Cells Directs Leukocytes to Sites of Axonal Injury in the CNS
Authors: Babcock Alicia A. , Kuziel William A. , Rivest Serge , Owens Trevor
Published: April 2018

Innate responses in the CNS are critical to first line defense against infection and injury. Leukocytes migrate to inflammatory sites in response to chemokines. We studied leukocyte migration and glial chemokine expression within the denervated hippocampus in response to axonal injury caused by entorhinodentate lesions. A population of Mac1/CD11b+ CD45highmacrophages (distinct from CD45lowmicroglia) was specifically detected within the lesion-reactive hippocampus by 12 hr after injury. Significant infiltration by CD3+ T cells did not occur in the denervated hippocampus until 24 hr after axotomy. A broad spectrum of chemokines [RANTES/CCL5, monocyte chemoattractant protein (MCP)-1/CCL2, interferon γ inducible protein (IP)-10/CXCL10, macrophage inflammatory protein (MIP)-1α/CCL3, MIP-1β/CCL4, and MIP-2/CXCL2] was induced at this time. RANTES/CCL5 was not significantly elevated until 24 hr after axotomy, whereas MCP-1/CCL2 was significantly induced before leukocyte infiltration occurred. Neither T cells nor macrophages infiltrated the denervated hippocampus of CCR2-deficient mice, arguing for a critical role for the CCR2 ligand MCP-1/CCL2 in leukocyte migration. Both T cells and macrophages infiltrated CCR5-deficient hippocampi, showing that CCR5 ligands (including RANTES/CCL5) are not critical to this response.In situhybridization combined with immunohistochemistry for ionized binding calcium adapter molecule (iba)1 or glial fibrillary acidic protein (GFAP) identified iba1+ microglia and GFAP+ astrocytes as major sources of MCP-1/CCL2 within the lesion-reactive hippocampus. We conclude that leukocyte responses to CNS axonal injury are directed via innate glial production of chemokines.
Binary Spiking in Auditory Cortex
Authors: DeWeese Michael R. , Wehr Michael , Zador Anthony M.
Published: April 2018

Neurons are often assumed to operate in a highly unreliable manner: a neuron can signal the same stimulus with a variable number of action potentials. However, much of the experimental evidence supporting this view was obtained in the visual cortex. We have, therefore, assessed trial-to-trial variability in the auditory cortex of the rat. To ensure single-unit isolation, we used cell-attached recording. Tone-evoked responses were usually transient, often consisting of, on average, only a single spike per stimulus. Surprisingly, the majority of responses were not just transient, but were also binary, consisting of 0 or 1 action potentials, but not more, in response to each stimulus; several dramatic examples consisted of exactly one spike on 100% of trials, with no trial-to-trial variability in spike count. The variability of such binary responses differs from comparably transient responses recorded in visual cortical areas such as area MT, and represent the lowest trial-to-trial variability mathematically possible for responses of a given firing rate. Our study thus establishes for the first time that transient responses in auditory cortex can be described as a binary process, rather than as a highly variable Poisson process. These results demonstrate that cortical architecture can support a more precise control of spike number than was previously recognized, and they suggest a re-evaluation of models of cortical processing that assume noisiness to be an inevitable feature of cortical codes.