bluuebunny/crossref_metadata_2025 huggingface dataset:
C-Terminal Alternative Splicing Changes the Gating Properties of a Human Spinal Cord Calcium Channel α1A Subunit
Authors: Krovetz Howard S. , Helton Thomas D. , Crews Anne L. , Horne William A.
Published: April 2018

The calcium channel α1Asubunit gene codes for proteins with diverse structure and function. This diversity may be important for fine tuning neurotransmitter release at central and peripheral synapses. The α1AC terminus, which serves a critical role in processing information from intracellular signaling molecules, is capable of undergoing extensive alternative splicing. The purpose of this study was to determine the extent to which C-terminal alternative splicing affects some of the fundamental biophysical properties of α1Asubunits. Specifically, the biophysical properties of two alternatively spliced α1Asubunits were compared. One variant was identical to an isoform identified previously in human brain, and the other was a novel isoform isolated from human spinal cord. The variants differed by two amino acids (NP) in the extracellular linker between transmembrane segments IVS3 and IVS4 and in two C-terminal regions encoded by exons 37 and 44. Expression inXenopusoocytes demonstrated that the two variants were similar with respect to current–voltage relationships and the voltage dependence of steady-state activation and inactivation. However, the rates of activation, inactivation, deactivation, and recovery from inactivation were all significantly slower for the spinal cord variant. A chimeric strategy demonstrated that the inclusion of the sequence encoded by exon 44 specifically affects the rate of inactivation. These findings demonstrate that C-terminal structural changes alone can influence the way in which α1Asubunits respond to a depolarizing stimulus and add to the developing picture of the C terminus as a critical domain in the regulation of Ca2+channel function.
Cocaine and Antidepressant-Sensitive Biogenic Amine Transporters Exist in Regulated Complexes with Protein Phosphatase 2A
Authors: Bauman Andrea L. , Apparsundaram Subbu , Ramamoorthy Sammanda , Wadzinski Brian E. , Vaughan Roxanne A. , Blakely Randy D.
Published: April 2018

Presynaptic transporter proteins regulate the clearance of extracellular biogenic amines after release and are important targets for multiple psychoactive agents, including amphetamines, cocaine, and antidepressant drugs. Recent studies reveal that dopamine (DA), norepinephrine (NE), and serotonin (5-HT) transporters (DAT, NET, and SERT, respectively) are rapidly regulated by direct or receptor-mediated activation of cellular kinases, particularly protein kinase C (PKC). With SERTs, PKC activation results in activity-dependent transporter phosphorylation and sequestration. Protein phosphatase 1/2A (PP1/PP2A) inhibitors, such as okadaic acid (OA) and calyculin A, also promote SERT phosphorylation and functional downregulation. How kinase, phosphatase, and transporter activities are linked mechanistically is unclear. In the present study, we found that okadaic acid-sensitive phosphatase activity is enriched in SERT immunoprecipitates from human SERT stably transfected cells. Moreover, blots of these immunoprecipitates reveal the presence of PP2A catalytic subunit (PP2Ac), findings replicated using brain preparations. Whole-cell treatments with okadaic acid or calyculin A diminished SERT/PP2Ac associations. Phorbol esters, which trigger SERT phosphorylation, also diminish SERT/PP2Ac associations, effects that can be blocked by PKC antagonists as well as the SERT substrate 5-HT. Similar transporter/PP2Ac complexes were also observed in coimmunoprecipitation studies with NETs and DATs. Our findings provide evidence for the existence of regulated heteromeric assemblies involving biogenic amine transporters and PP2A and suggest that the dynamic stability of these complexes may govern transporter phosphorylation and sequestration.
Evidence That Helix-Loop-Helix Proteins Collaborate with Retinoblastoma Tumor Suppressor Protein to Regulate Cortical Neurogenesis
Authors: Toma Jean G. , El-Bizri Hiba , Barnabé-Heider Fanie , Aloyz Raquel , Miller Freda D.
Published: April 2018

The retinoblastoma tumor suppressor protein (pRb) family is essential for cortical progenitors to exit the cell cycle and survive. In this report, we test the hypothesis that pRb collaborates with basic helix-loop-helix (bHLH) transcription factors to regulate cortical neurogenesis, taking advantage of the naturally occurring dominant-inhibitory HLH protein Id2. Overexpression of Id2 in cortical progenitors completely inhibited the induction of neuron-specific genes and led to apoptosis, presumably as a consequence of conflicting differentiation signals. Both of these phenotypes were rescued by coexpression of a constitutively activated pRb mutant. In contrast, Id2 overexpression in postmitotic cortical neurons affected neither neuronal gene expression nor survival. Thus, pRb collaborates with HLHs to ensure the coordinate induction of terminal mitosis and neuronal gene expression as cortical progenitors become neurons.
Evidence That Different Cation Chloride Cotransporters in Retinal Neurons Allow Opposite Responses to GABA
Authors: Vardi Noga , Zhang Ling-Li , Payne John A. , Sterling Peter
Published: April 2018

GABA gating an anion channel primarily permeable to chloride can hyperpolarize or depolarize, depending on whether the chloride equilibrium potential (ECl) is negative or positive, respectively, to the resting membrane potential (Erest). If the transmembrane Cl−gradient is set by active transport, those neurons or neuronal regions that exhibit opposite responses to GABA should express different chloride transporters. To test this, we immunostained retina for the K-Cl cotransporter (KCC2) that normally extrudes chloride and for the Na-K-Cl cotransporter (NKCC) that normally accumulates chloride. KCC2 was expressed wherever EClis either known or predicted to be negative to Erest(ganglion cells, bipolar axons, and OFF bipolar dendrites), whereas NKCC was expressed wherever EClis either known or predicted to be positive to Erest(horizontal cells and ON bipolar dendrites). Thus, in the retina, the opposite effects of GABA on different cell types and on different cellular regions are probably primarily determined by the differential targeting of these two chloride transporters.
Mice Lacking Specific Nicotinic Acetylcholine Receptor Subunits Exhibit Dramatically Altered Spontaneous Activity Patterns and Reveal a Limited Role for Retinal Waves in Forming ON and OFF Circuits in the Inner Retina
Authors: Bansal Anu , Singer Joshua H. , Hwang Bryan J. , Xu Wei , Beaudet Art , Feller Marla B.
Published: April 2018

Before phototransduction, spontaneous activity in the developing mammalian retina is required for the appropriate patterning of retinothalamic connections, and there is growing evidence that this activity influences the development of circuits within the retina itself. We demonstrate here that the neural substrate that generates waves in the mouse retina develops through three distinct stages. First, between embryonic day 16 and birth [postnatal day 0 (P0)], we observed both large, propagating waves inhibited by nicotinic acetylcholine receptor (nAChR) antagonists and small clusters of cells displaying nonpropagating, correlated calcium increases that were independent of nAChR activation. Second, between P0 and P11, we observed only larger propagating waves that were abolished by toxins specific to α3 and β2 subunit-containing nAChRs. Third, between P11 and P14 (eye opening) we observed propagating activity that was abolished by ionotropic glutamate receptor antagonists. The time course of this developmental shift was dramatically altered in retinas from mice lacking the β2 nAChR subunit or the β2 and β4 subunits. These retinas exhibited a novel circuit at P0, no spontaneous correlated activity between P1 and P8, and the premature induction at P8 of an ionotropic glutamate receptor-based circuit. Retinas from postnatal mice lacking the α3 nAChR subunit exhibited spontaneous, correlated activity patterns that were similar to those observed in embryonic wild-type mice. In α3−/− and β2−/− mice, the development and distribution of cholinergic neurons and processes and the density of retinal ganglion cells (RGCs) and the gross segregation of their dendrites into ON and OFF sublaminae were normal. However, the refinement of individual RGC dendrites is delayed. These results indicate that retinal waves mediated by nAChRs are involved in, but not required for, the development of neural circuits that define the ON and OFF sublamina of the inner plexiform layer.
Differential Expression ofCOUP-TFI, CHL1,and Two Novel Genes in Developing Neocortex Identified by Differential Display PCR
Authors: Liu Qing , Dwyer Noelle D. , O'Leary Dennis D. M.
Published: April 2018

Genes that control the specification and differentiation of the functionally specialized areas of the mammalian neocortex are likely expressed across the developing neocortex in graded or restricted patterns. To search for such genes we have performed a PCR-based differential display screen using RNAs from rostral neocortex, which included the primary motor area, and caudal neocortex, which included the primary visual area, of embryonic day 16 rats. We identified 82 differentially expressed gene fragments. Secondary screening byin situhybridization confirmed that five fragments, representing four genes, are differentially expressed across developing rat neocortex. Two of the genes,chick ovalbumin upstream transcription factor I(COUP-TFI) andclose homolog of L1(CHL1), have been cloned previously, but their differential expression in cortex has not been reported. Sequences from the other two fragments suggest that they represent novel genes. The expression patterns include graded, restricted, and discontinuous expression with abrupt borders that might correlate with those of areas. The differential expression patterns of all four genes are established before the arrival of thalamocortical afferents, suggesting that they are independent of thalamic influence, and could direct or reflect arealization. In addition,COUP-TFIandCHL1exhibit dynamic expression patterns that undergo substantial changes after thalamocortical afferents invade the cortical plate, suggesting that thalamic axons may influence their later expression. Postnatally,COUP-TFIis most prominently expressed in layer 4, in both rats and mice, andCHL1is expressed in layer 5.COUP-TFIexpression in cortex, and in ventral telencephalon and dorsal thalamus, suggests several possible causes for the loss of layer 4 neurons and the reduced thalamocortical projection reported inCOUP-TFIknock-out mice.
Semaphorin 3A Is Required for Guidance of Olfactory Axons in Mice
Authors: Schwarting Gerald A. , Kostek Christine , Ahmad Naira , Dibble Christian , Pays Laurent , Püschel Andreas W.
Published: April 2018

Semaphorin 3A (Sema3A) is a membrane-associated secreted protein that has chemorepulsive properties for neuropilin-1 (npn-1)- expressing axons. Although mice lacking the Sema3A protein display skeletal abnormalities and heart defects, most axonal projections in the CNS develop normally. We show here that Sema3A is expressed in the lamina propria surrounding the olfactory epithelium (OE) and by ensheathing cells in the nerve layer of the ventral olfactory bulb (OB) throughout development. Subsets of sensory neurons expressing npn-1 are distributed throughout the OE and extend fibers to the developing OB. In wild-type mice, npn-1-positive (npn-1+) axons extend to lateral targets in the rostral OB and medial targets in the caudal OB, avoiding regions expressing Sema3A. In Sema3A homozygous mutant mice, many npn-1+axons are misrouted into and through the ventral nerve layer, beginning as early as embryonic day 13 and continuing at least until birth. At postnatal day 0, npn-1+glomeruli are atypically located in the ventral OB of Sema3A−/−mice, indicating that aberrant axon trajectories are not corrected during development and that connections are made in inappropriate target regions. In addition, subsets of OCAM+axons that normally project to the ventrolateral OB and some lactosamine-containing glycan+axons that normally target the ventral OB are also misrouted in Sema3A mutants. These observations indicate that Sema3A expression by ensheathing cells plays an important role in guiding olfactory axons into specific compartments of the OB.
Serotonin DepletionIn VivoInhibits the Branching of Olfactory Projection Neurons in the Lobster Deutocerebrum
Authors: Sullivan Jeremy M. , Benton Jeanne L. , Beltz Barbara S.
Published: April 2018

Serotonin depletion during embryogenesis has been shown previously to retard the growth of the olfactory and accessory lobes of the lobster deutocerebrum (Benton et al., 1997). The present study was undertaken to determine whether morphological changes in the interneurons innervating these lobes contribute to this growth retardation. We examined the effects ofin vivoserotonin depletion using 5,7-dihydroxytryptamine (5,7-DHT) on the morphology of the olfactory projection neurons, one of two major classes of interneurons that innervate both lobes. Intracellular dye fills of olfactory projection neurons in normal embryos showed that each neuron extensively innervates either the olfactory or accessory lobe before projecting to neuropil regions in the protocerebrum. In embryos injected with 5,7-DHT, however, the deutocerebral arbors of 13.5% of the olfactory projection neurons examined were either markedly reduced compared with normal neurons or absent. Affected neurons also exhibited a number of additional aberrant morphological features suggesting that these neurons represent cells that were affected during their initial morphogenesis. Olfactory projection neurons with aberrant morphologies were also encountered, although less frequently (7.5% of the neurons examined), in control (sham-injected) embryos indicating that the sham injections can affect the development of the brain. This observation provides insights into the nature of effects seen in control embryos in previous experiments (Benton et al., 1997). The results of the present study indicate thatin vivoserotonin depletion inhibits the branching of olfactory projection neurons and suggest, therefore, that one of the functions of serotonin during normal development is to promote the ingrowth of these neurons into the deutocerebral neuropils.
Estrous Changes in Responses of Rat Gracile Nucleus Neurons to Stimulation of Skin and Pelvic Viscera
Authors: Bradshaw Heather B. , Berkley Karen J.
Published: April 2018

Multi- and single-unit recording was performed in the gracile nucleus in urethane-anesthetized rats to examine estrous variations in responses of its neurons to brushing the hindquarters and mechanical stimulation of the uterus, vaginal canal, cervix, and colon. Six rats each were studied in each of the four estrous stages: proestrus (P), estrus (E), metestrus (M), and diestrus (D). The magnitude of multi-unit responses to gentle brushing of the perineum, hip, and tail, but not the foot and leg, was significantly greater during proestrus than during other stages. Of 70 single units responsive to brush, 56 (80%) responded to stimulation of at least one viscus. Although this percentage did not change with estrous stage, the direction and latency of some responses did. Pressure on the cervix evoked significantly more inhibitory (vs excitatory) responses in P than in E and M, and the response latency was significantly longer in D and P than in E and M. The direction of response to vaginal distention did not change with estrous stage, but response latency was significantly longer in D than in P and E. Uterine distention evoked significantly more inhibitory responses in D than in P, with no estrous changes in latency. Responses to colon distention did not change. These variations in both magnitude of response to tactile stimulation and characteristics of response to stimulation of reproductive organs, but not the colon, correlate with changes in mating behaviors of the female rat, suggesting that the gracile nucleus is a component of neural systems that control reproductive behaviors.
Effects of Irradiance and Stimulus Duration on Early Gene Expression (Fos) in the Suprachiasmatic Nucleus: Temporal Summation and Reciprocity
Authors: Dkhissi-Benyahya O. , Sicard B. , Cooper H. M.
Published: April 2018

The daily behavioral, physiological, and hormonal rhythms in mammals are regulated by an endogenous circadian clock located in the suprachiasmatic nucleus (SCN) and are synchronized by the natural 24 hr light/dark cycle. We studied the response properties (threshold, saturation, and linearity) of the photic system to irradiance by assaying light induction of Fos, the protein product of the immediate early genec-fos. Fos expression was quantified by image analysis in the SCN and in the retina. Fos expression in the SCN and retina are unrelated because the response differs in terms of threshold, saturation, and range. In the SCN, Fos expression increases proportionately to increases in both irradiance and duration of light exposure. The photic system shows a linear temporal integration of photons for durations ranging from 3 sec to 47.5 min. The principal result of this study shows that in the SCN, Fos expression is directly proportional to the total number of photons rather than to irradiance or duration alone (reciprocity), and that integration occurs over a range of 5 log units of photon number. This report provides the first demonstration that the mechanism of photon integration by the circadian system is expressed at a cellular level in the SCN.
Stress-Level Cortisol Treatment Impairs Inhibitory Control of Behavior in Monkeys
Authors: Lyons David M. , Lopez Jacqueline M. , Yang Chou , Schatzberg Alan F.
Published: April 2018

Most studies of cortisol-induced cognitive impairments have focused on hippocampal-dependent memory. This study investigates a different aspect of cognition in a randomized placebo-controlled experiment with monkeys that were treated with cortisol according to a protocol that simulates a prolonged stress response. Young adult and older adult monkeys were assigned randomly to placebo or chronic treatment with cortisol in a 2 × 2 factorial design (n= 8 monkeys per condition). Inhibitory control of behavior was assessed with a test shown previously in primates to reflect prefrontal cortical dysfunction. Failure to inhibit a specific goal-directed response was evident more often in older adults. Treatment with cortisol increased this propensity in both older and young adult monkeys. Age-related differences in response inhibition were consistent across blocks of repeated test trials, but the treatment effects were clearly expressed only after prolonged exposure to cortisol. Aspects of performance that did not require inhibition were not altered by age or treatment with cortisol, which concurs with effects on response inhibition rather than nonspecific changes in behavior. These findings lend support to related reports that cortisol-induced disruptions in prefrontal dopamine neurotransmission may contribute to deficits in response inhibition and play a role in cognitive impairments associated with endogenous hypercortisolism in humans.
Effect of Temperature on Dopamine Transporter Function and Intracellular Accumulation of Methamphetamine: Implications for Methamphetamine-Induced Dopaminergic Neurotoxicity
Authors: Xie Tao , McCann Una D. , Kim Saejeong , Yuan Jie , Ricaurte George A.
Published: April 2018

Hyperthermia exacerbates and hypothermia attenuates methamphetamine (METH)-induced dopamine (DA) neurotoxicity. The mechanisms underlying these temperature effects are unknown. Given the essential role of the DA transporter (DAT) in the expression of METH-induced DA neurotoxicity, we hypothesized that the effect of temperature on METH-induced DA neurotoxicity is mediated, at least in part, at the level of the DAT. To test this hypothesis, the effects of small, physiologically relevant temperature changes on DAT function were evaluated in two types of cultured neuronal cells: (1) a neuroblastoma cell line stably transfected with human DAT cDNA and (2) rat embryonic mesencephalic primary cells that naturally express the DAT. Temperatures for studies of DAT function were selected based on core temperature measurements in animals exposed to METH under usual ambient (22°C) and hypothermic (6°C) temperature conditions, where METH neurotoxicity was fully expressed and blocked, respectively. DAT function, determined by measuring accumulation of radiolabeled DA and 1-methyl-4-phenylpyridinium (MPP+), was found to directly correlate with temperature, with higher levels of substrate uptake at 40°C, intermediate levels at 37°C, and lower levels at 34°C. DAT-mediated accumulation of METH also directly correlated with temperature, with greater accumulation at higher temperatures. These findings indicate that relatively small, physiologically relevant changes in temperature significantly alter DAT function and intracellular METH accumulation, and suggest that the effect of temperature on METH-induced DA neurotoxicity is mediated, at least in part, at the level of the DAT.
Evidence for Spatial Modules Mediated by Temporal Synchronization of Carbachol-Induced Gamma Rhythm in Medial Entorhinal Cortex
Authors: Dickson Clayton T. , Biella Gerardo , de Curtis Marco
Published: April 2018

Fast (γ) oscillations in the cortex underlie the rapid temporal coordination of large-scale neuronal assemblies in the processing of sensory stimuli. Cortical γ rhythm is modulatedin vivoby cholinergic innervation from the basal forebrain and can be generatedin vitroafter exogenous cholinergic stimulation. Using the isolated guinea pig brain, anin vitropreparation that allows for the study of an intact cerebrum, we studied the spatial features of γ activity evoked by the cholinomimetic carbachol (CCh) in the medial entorhinal cortex (mEC). γ activity induced by either arterial perfusion or intraparenchymal application of CCh showed a phase reversal across mEC layer II and was reduced or abolished in a spatially localized region by focal infusions of atropine, bicuculline, and CNQX. In addition, a spatially restricted zone of γ activity could be induced by passive diffusion of CCh from a recording pipette. Finally, γ oscillations recorded at multiple sites across the surface of the mEC using array electrodes during arterial perfusion of CCh demonstrated a decline in synchronization (coherence) as the interelectrode distance increased. This effect was independent of the signal amplitude and was specific for γ as opposed to theta-like activity induced by CCh in the same experiments.

These results suggest that CCh-induced γ oscillations in the mEC are mediated through direct muscarinic excitation of a highly localized reciprocal inhibitory–excitatory network located in superficial layers. We propose that functional cortical modules of highly synchronous γ oscillations may organize incoming (cortical) and outgoing (hippocampal) information in the mEC.
Cholinergic Inhibition of Ventral Midbrain Dopamine Neurons
Authors: Fiorillo Christopher D. , Williams John T.
Published: April 2018

Muscarinic acetylcholine receptors are common throughout the CNS. The predominant subtypes in the brain are positively coupled to phosphoinositide hydrolysis and have been found to modulate multiple conductances. Muscarinic receptor activation is most often observed to be excitatory because of suppression of various potassium conductances. Here it is reported that three distinct effects of muscarinic receptor activation can be observed in isolation from one another, depending on the duration of receptor activation and the concentration of agonist. Brief activation of muscarinic receptors, as is likely to occur with normal synaptic transmission, hyperpolarized dopamine neurons of the ventral midbrain through a calcium-activated potassium conductance. With repeated or persistent activation of muscarinic receptors, the hyperpolarizing response was entirely desensitized in the absence of any change in resting membrane potential. With sustained activation by higher concentrations of agonist, dopamine neurons were depolarized. This demonstrates that muscarinic receptors can mediate very diverse, and even opposing, postsynaptic effects on neurons depending on the pattern of acetylcholine release.
Long-Term Potentiation Induced by θ Frequency Stimulation Is Regulated by a Protein Phosphatase-1-Operated Gate
Authors: Brown George P. , Blitzer Robert D. , Connor John H. , Wong Tony , Shenolikar Shirish , Iyengar Ravi , Landau Emmanuel M.
Published: April 2018

Long-term potentiation (LTP) can be induced in the Schaffer collateral→CA1 synapse of hippocampus by stimulation in the θ frequency range (5–12 Hz), an effect that depends on activation of the cAMP pathway. We investigated the mechanisms of the cAMP contribution to this form of LTP in the rat hippocampal slice preparation. θ pulse stimulation (TPS; 150 stimuli at 10 Hz) by itself did not induce LTP, but the addition of either the β-adrenergic agonist isoproterenol or the cAMP analog 8-bromo-cAMP (8-Br-cAMP) enabled TPS-induced LTP. The isoproterenol effect was blocked by postsynaptic inhibition of cAMP-dependent protein kinase. Several lines of evidence indicated that cAMP enabled LTP by blocking postsynaptic protein phosphatase-1 (PP1). Activators of the cAMP pathway reduced PP1 activity in the CA1 region and increased the active form of inhibitor-1, an endogenous inhibitor of PP1. Postsynaptic injection of activated inhibitor-1 mimicked the LTP-enabling effect of cAMP pathway stimulation. TPS evoked complex spiking when isoproterenol was present. However, complex spiking was not sufficient to enable TPS-induced LTP, which additionally required the inhibition of postsynaptic PP1. PP1 inhibition seems to promote the activation of Ca2+/calmodulin-dependent protein kinase (CaMKII), because (1) a CaMKII inhibitor blocked the induction of LTP by TPS paired with either isoproterenol or activated inhibitor-1 and (2) CaMKII in area CA1 was activated by the combination of TPS and 8-Br-cAMP but not by either stimulus alone. These results indicate that the cAMP pathway enables TPS-induced LTP by inhibiting PP1, thereby enhancing Ca2+-independent CaMKII activity.
Role of Bicarbonate and Chloride in GABA- and Glycine-Induced Depolarization and [Ca2+]iRise in Fetal Rat MotoneuronsIn Situ
Authors: Kulik Anna , Nishimaru Hiroshi , Ballanyi Klaus
Published: April 2018

Ca2+imaging and (perforated) patch recording were used to analyze the mechanism of GABA- and glycine-induced depolarizations in lumbar motoneurons of spinal cord slices from fetal rats. In fura-2 ester-loaded cells, the agonist-induced depolarizations increased [Ca2+]iby up to 100 nm. The GABA- and glycine-evoked [Ca2+]itransients were suppressed by bicuculline and strychnine, respectively. Their magnitude decreased by ∼50% between embryonic days 15.5 and 19.5. The [Ca2+]iincreases were abolished by Ca2+-free superfusate and attenuated by ∼65% by nifedipine, showing that the responses were mediated by voltage-activated Ca2+channels. The [Ca2+]irises were potentiated by >300% immediately after removal of Cl−from the superfusate but recovered to values of 50–200% of control during repeated agonist administration in Cl−-free saline. Bumetanide gradually suppressed the [Ca2+]iincreases by >75%. Subsequent removal of Cl−reconstituted the responses and increased, upon repeated agonist application, the peak [Ca2+]irises to values above control. Removal of HCO3−from the Cl−-free (bumetanide-containing) superfusate reversibly abolished both the agonist-induced [Ca2+]irises and depolarizations that were reestablished by formate anions. In Cl−-containing superfusate, removal of HCO3−decreased both the peak and duration of the agonist-evoked membrane depolarization and [Ca2+]iresponse. Our findings show that HCO3−efflux has a major contribution to depolarizations mediated by GABAAand glycine receptor-coupled anion channels in prenatal neurons. We hypothesize that the HCO3−-dependent depolarizing component, which is likely to produce an intracellular acidosis, might play an important role during the early postnatal period when the Cl−-dependent component gradually shifts to hyperpolarization.
Slow Desensitization Regulates the Availability of Synaptic GABAAReceptors
Authors: Overstreet Linda S. , Jones Mathew V. , Westbrook Gary L.
Published: April 2018

At central synapses, a large and fast spike of neurotransmitter efficiently activates postsynaptic receptors. However, low concentrations of transmitter can escape the cleft and activate presynaptic and postsynaptic receptors. We report here that low concentrations of GABA reduce IPSCs in hippocampal neurons by preferentially desensitizing rather than opening GABAAchannels. GABA transporter blockade also caused desensitization by locally elevating GABA to ∼1 μm. Recovery of the IPSC required several seconds, mimicking recovery of the channel from slow desensitization. These results indicate that low levels of GABA can regulate the amplitude of IPSCs by producing a slow form of receptor desensitization. Accumulation of channels in this absorbing state allows GABAAreceptors to detect even a few molecules of GABA in the synaptic cleft.
Tumor Necrosis Factor α Induces a Metalloprotease-Disintegrin, ADAM8 (CD 156): Implications for Neuron–Glia Interactions during Neurodegeneration
Authors: Schlomann Uwe , Rathke-Hartlieb Silvia , Yamamoto Shunsuke , Jockusch Harald , Bartsch Jörg W.
Published: April 2018

ADAM proteases, defined by extracellular disintegrin and metalloprotease domains, are involved in protein processing and cell–cell interactions. Using wobbler (WR) mutant mice, we investigated the role of ADAMs in neurodegeneration and reactive glia activation in the CNS. We found that ADAM8 (CD 156), a suspected leukocyte adhesion molecule, is expressed in the CNS and highly induced in affected CNS areas of WR mice, in brainstem and spinal cord. ADAM8 mRNA and protein are found at low levels throughout the normal mouse CNS, in neurons and oligodendrocytes. In the WR CNS regions in which neurodegeneration occurs, ADAM8 is induced in neurons, reactive astrocytes, and activated microglia. Similarly, the proinflammatory cytokine tumor necrosis factor α (TNF-α) is upregulated and shows the same cellular distribution. In primary astrocytes from wild-type and WR mice, in primary cerebellar neurons, and in mouse motoneuron-like NSC19 cells, ADAM8 expression was induced up to 15-fold by mouse TNF-α, in a dose-dependent manner. In both cell types, ADAM8 was also induced by human TNF-α, indicating that TNF receptor type I (p55) is involved. Induction of ADAM8 mRNA was suppressed by treatment with an interferon-regulating factor 1 (IRF-1) antisense oligonucleotide. We conclude that IRF-1-mediated induction of ADAM8 by TNF-α is a signaling pathway relevant for neurodegenerative disorders with glia activation, proposing a role for ADAM8 in cell adhesion during neurodegeneration.
Clathrin-Mediated Endocytosis near Active Zones in Snake Motor Boutons
Authors: Teng Haibing , Wilkinson Robert S.
Published: April 2018

We have used the activity-dependent probe FM1-43 with electron microscopy (EM) to examine endocytosis at the vertebrate nerve–muscle synapse. Preparations were fixed after very brief neural stimulation at reduced temperature, and internalized FM1-43 was photoconverted into an electron-dense reaction product. To locate the reaction product, we reconstructed computer renderings of individual terminal boutons from serial EM sections. Most of the reaction product was seen in 40–60 nm vesicles. All of the labeled vesicles were clathrin-coated, and 92% of them were located within 300 nm of the plasma membrane, suggesting that they had undergone little processing after retrieval from their endocytic sites. The vesicles (and by inference the sites) were not dispersed randomly near the plane of the membrane but instead were clustered significantly near active zones. Additional reaction product was found within putative macropinosomes; these appeared to form from deep membrane invaginations near active zones. Thus two mechanisms of endocytosis were evident after brief stimulation. Endocytosis near active zones is consistent with the existence of local exo/endocytic cycling pools. This mechanism also might serve to maintain alignment of active zones with postsynaptic folds during periods of activity when vesicular and plasma membranes are interchanged.
Cell Migration and Aggregation in the Developing Telencephalon: Pulse-Labeling Chick Embryos with Bromodeoxyuridine
Authors: Striedter Georg F. , Keefer Brian P.
Published: April 2018

Previous studies had concluded that the avian telencephalon develops according to an outside-in schedule of neurogenesis, with relatively little migration of young neuroblasts past older cells. These previous studies had, however, been based on the “cumulative labeling” method, which is less accurate than the “pulse-labeling” method typically used in mammals. In the present study, we pulse-labeled chick embryos by injecting low doses of the thymidine analog bromodeoxyuridine (BrdU) directly into the circulatory system of chick embryos at 6 d of incubation. The brains of these embryos were then examined for anti-BrdU-labeled cells at postinjection survival times from 30 min to 10 d. Comparisons across different survival times, as well as with cases in which BrdU was injected on day 7, suggested that our effective pulse duration is <24 hr. This was confirmed by injecting tritiated thymidine 24 hr after the BrdU and seeing no double-labeled cells. Several deviations from the previously reported pattern of telencephalic neurogenesis were also noted. Most importantly, the cells born on day 6 in the avian Wulst, the likely homolog of mammalian neocortex, end up homogeneously distributed throughout the Wulst, which suggests that many of them are migrating past older cells. Furthermore, the cells born on day 6 in the ventral hyperstriatum and dorsal neostriatum gradually (over the course of 2–3 d) aggregate into distinct multicellular clusters, which suggests that isochronic cells in these regions adhere preferentially to one another. Finally, the data reveal a proliferative subventricular zone similar to that observed in the ganglionic eminences of mammalian embryos.
Pax6Modulates the Dorsoventral Patterning of the Mammalian Telencephalon
Authors: Stoykova Anastassia , Treichel Dieter , Hallonet Marc , Gruss Peter
Published: April 2018

ThePax6gene encodes a transcription factor with a restricted expression in the ventricular zone of the pallium and subpallium. We tested whether the function ofPax6is necessary for the correct patterning and morphogenesis of the vertebrate telencephalon. Homozygous embryos of thePax6/Small eyemutant lack functional PAX6 protein because of a point mutation of the gene. In the mutantSmall eyeembryos we detected a ventralization of the molecular patterning of the telencephalon at two borders, the pallium/subpallium and the lateral/medial ganglionic eminence. The results indicate thatPax6controls the lateral limit of the expression ofNkx2.1,Shh,andLhx6in the prechordal neural tube, the telencephalon. This finding is in agreement with previous studies and supports a model for a common genetic mechanism for modulation of the dorsoventral patterning of the prechordal and epichordal CNS. The pattern defects caused by the loss of Pax6 function result in multiple morphological abnormalities in theSmall eyebrain: dysgenesis of the piriform, insular, and lateral cortices, the claustrum–endopiriform nucleus, and a failure in the differentiation of a subpopulation of the cortical precursors. Together the results demonstrate that Pax6 has an essential role for the modulation of the dorsoventral patterning of the embryonic telencephalon, influencing thereby the forebrain morphogenesis.
Neuronal Activity and Brain-Derived Neurotrophic Factor Regulate the Density of Inhibitory Synapses in Organotypic Slice Cultures of Postnatal Hippocampus
Authors: Marty Serge , Wehrlé Rosine , Sotelo Constantino
Published: April 2018

Hippocampal interneurons inhibit pyramidal neurons through the release of the neurotransmitter GABA. Given the importance of this inhibition for the proper functioning of the hippocampus, the development of inhibitory synapses must be tightly regulated. In this study, the possibility that neuronal activity and neurotrophins regulate the density of GABAergic inhibitory synapses was investigated in organotypic slice cultures taken from postnatal day 7 rats.

In hippocampal slices cultured for 13 d in the presence of the GABAAreceptor antagonist bicuculline, the density of glutamic acid decarboxylase (GAD) 65-immunoreactive terminals was increased in the CA1 area when compared with control slices. Treatment with the glutamate receptor antagonist 6,7-dinitroquinoxaline-2,3-dione decreased the density of GAD65-immunoreactive terminals in the stratum oriens of CA1. These treatments had parallel effects on the density of GABA-immunoreactive processes. Electron microscopic analysis after postembedding immunogold labeling with antibodies against GABA indicated that bicuculline treatment increased the density of inhibitory but not excitatory synapses. Application of exogenous BDNF partly mimicked the stimulatory effect of bicuculline on GAD65-immunoreactive terminals. Finally, antibodies against BDNF, but not antibodies against nerve growth factor, decrease the density of GAD65-immunoreactive terminals in bicuculline-treated slices.

Thus, neuronal activity regulates the density of inhibitory synapses made by postnatal hippocampal interneurons, and BDNF could mediate part of this regulation. This regulation of the density of inhibitory synapses could represent a feedback mechanism aimed at maintaining an appropriate level of activity in the developing hippocampal networks.
Muscarinic Tone Sustains Impulse Flow in the Septohippocampal GABA But Not Cholinergic Pathway: Implications for Learning and Memory
Authors: Alreja Meenakshi , Wu Min , Liu Weimin , Atkins Joshua B. , Leranth Csaba , Shanabrough Marya
Published: April 2018

Systemic infusions of the muscarinic cholinergic receptor antagonists atropine and scopolamine (atr/scop) produce an amnesic syndrome in humans, subhuman primates, and rodents. In humans, this syndrome may resemble early symptoms of Alzheimer's disease. Behavioral studies in rats have demonstrated that the medial septum/diagonal band of Broca (MSDB), which sends cholinergic and GABAergic projections to the hippocampus, is a critical locus in mediating the amnesic effects of atr/scop. The amnesic effects of atr/scop in the MSDB have been presumed but not proven to be caused by a decrease in hippocampal acetylcholine (ACh) release after blockade of a muscarinic tone in the MSDB. Using electrophysiological recordings and fluorescent-labeling techniques to identify living septohippocampal neurons in rat brain slices, we now report that, contrary to current belief, a blockade of the muscarinic tone in the MSDB does not decrease impulse flow in the septohippocampal cholinergic pathway; instead, it decreases impulse flow in the septohippocampal GABAergic pathway via M3muscarinic receptors. We also report that the muscarinic tone in the MSDB is maintained by ACh that is released locally, presumably via axon collaterals of septohippocampal cholinergic neurons. As such, cognitive deficits that occur in various neurodegenerative disorders that are associated with a loss or atrophy of septohippocampal cholinergic neurons cannot be attributed solely to a decrease in hippocampal acetylcholine release. An additional, possibly more important mechanism may be the concomitant decrease in septohippocampal GABA release and a subsequent disruption in disinhibitory mechanisms in the hippocampus. Restoration of impulse flow in the septohippocampal GABA pathway, possibly via M3receptor agonists, may, therefore, be critical for successful treatment of cognitive deficits associated with neurodegenerative disorders such as Alzheimer's and Parkinson's disease.
The CircadianClockMutation Alters Sleep Homeostasis in the Mouse
Authors: Naylor Erik , Bergmann Bernard M. , Krauski Kristyn , Zee Phyllis C. , Takahashi Joseph S. , Vitaterna Martha Hotz , Turek Fred W.
Published: April 2018

The onset and duration of sleep are thought to be primarily under the control of a homeostatic mechanism affected by previous periods of wake and sleep and a circadian timing mechanism that partitions wake and sleep into different portions of the day and night. The mouseClockmutation induces pronounced changes in overall circadian organization. We sought to determine whether this genetic disruption of circadian timing would affect sleep homeostasis. TheClockmutation affected a number of sleep parameters during entrainment to a 12 hr light/dark (LD 12:12) cycle, when animals were free-running in constant darkness (DD), and during recovery from 6 hr of sleep deprivation in LD 12:12. In particular, in LD 12:12, heterozygous and homozygousClockmutants slept, respectively, ∼1 and ∼2 hr less than wild-type mice, and they had 25 and 51% smaller increases in rapid eye movement (REM) sleep during 24 hr recovery, respectively, than wild-type mice. The effects of the mutation on sleep are not readily attributable to differential entrainment to LD 12:12 because the baseline sleep differences between genotypes were also present when animals were free-running in DD. These results indicate that genetic alterations of the circadian clock system and/or its regulatory genes are likely to have widespread effects on a variety of sleep and wake parameters, including the homeostatic regulation of sleep.
Interleukin-1β-Induced Changes in Blood–Brain Barrier Permeability, Apparent Diffusion Coefficient, and Cerebral Blood Volume in the Rat Brain: A Magnetic Resonance Study
Authors: Blamire A. M. , Anthony D. C. , Rajagopalan B. , Sibson N. R. , Perry V. H. , Styles P.
Published: April 2018

The cytokine interleukin-1β (IL-1β) is implicated in a broad spectrum of CNS pathologies, in which it is thought to exacerbate neuronal loss. Here, the effects of injecting recombinant rat IL-1β into the striatum of 3-week-old rats were followed noninvasively from 2 to 123 hr using magnetic resonance imaging and spectroscopy. Four hours after injection of IL-1β (1 ng in 1 μl), cerebral blood volume was significantly increased, the blood–brain barrier (BBB) became permeable to intravenously administered contrast agent between 4.5 and 5 hr, and the apparent diffusion coefficient (ADC) of brain water fell by 6 hr (5.42 ± 0.35 × 10−4mm2/sec treated, 7.35 ± 0.77 × 10−4mm2/sec control;p< 0.001). At 24 hr the BBB was again intact, but the ADC, although partially recovered, remained depressed at both 24 and 123 hr (p< 0.03). Depleting the animals of neutrophils before IL-1β injection prevented the BBB permeability at all time points, but the ADC was still depressed at 6 hr (6.64 ± 0.34 × 10−4mm2/sec treated, 7.49 ± 0.38 × 10−4mm2/sec control;p< 0.005). No changes were seen in brain metabolites using proton spectroscopy at 6 hr after IL-1β.

Intraparenchymal injection of IL-1β caused a neutrophil-dependent transient increase in BBB permeability. The presence of neutrophils within the brain parenchyma significantly contributed to the IL-1β-induced changes in cerebral blood volume and the ADC of brain water. However, IL-1β apparently had a direct effect on the resident cell populations, which persisted well after all recruited leukocytes had disappeared. Thus the action of IL-1β alone can give rise to magnetic resonance imaging-visible changes that are normally attributed to alterations to cellular homeostasis.
Reduction of Pentylenetetrazole-Induced Seizure Activity in Awake Rats by Seizure-Triggered Trigeminal Nerve Stimulation
Authors: Fanselow Erika E. , Reid Ashlan P. , Nicolelis Miguel A. L.
Published: April 2018

Stimulation of the vagus nerve has become an effective method for desynchronizing the highly coherent neural activity typically associated with epileptic seizures. This technique has been used in several animal models of seizures as well as in humans suffering from epilepsy. However, application of this technique has been limited to unilateral stimulation of the vagus nerve, typically delivered according to a fixed duty cycle, independently of whether ongoing seizure activity is present. Here, we report that stimulation of another cranial nerve, the trigeminal nerve, can also cause cortical and thalamic desynchronization, resulting in a reduction of seizure activity in awake rats. Furthermore, we demonstrate that providing this stimulation only when seizure activity begins results in more effective and safer seizure reduction per second of stimulation than with previous methods. Seizure activity induced by intraperitoneal injection of pentylenetetrazole was recorded from microwire electrodes in the thalamus and cortex of awake rats while the infraorbital branch of the trigeminal nerve was stimulated via a chronically implanted nerve cuff electrode. Continuous unilateral stimulation of the trigeminal nerve reduced electrographic seizure activity by up to 78%, and bilateral trigeminal stimulation was even more effective. Using a device that automatically detects seizure activity in real time on the basis of multichannel field potential signals, we demonstrated that seizure-triggered stimulation was more effective than the stimulation protocol involving a fixed duty cycle, in terms of the percent seizure reduction per second of stimulation. In contrast to vagus nerve stimulation studies, no substantial cardiovascular side effects were observed by unilateral or bilateral stimulation of the trigeminal nerve. These findings suggest that trigeminal nerve stimulation is safe in awake rats and should be evaluated as a therapy for human seizures. Furthermore, the results demonstrate that seizure-triggered trigeminal nerve stimulation is technically feasible and could be further developed, in conjunction with real-time seizure-predicting paradigms, to prevent seizures and reduce exposure to nerve stimulation.
Independence of and Interactions between GABA-, Glutamate-, and Acetylcholine-Activated Cl Conductances inAplysiaNeurons
Authors: Kehoe JacSue , Vulfius Catherine
Published: April 2018

In certainAplysianeurons, glutamate, GABA, and acetylcholine (ACh) all elicit desensitizing Cl-dependent responses. This fact and the finding that the glutamate and GABA responses “cross-desensitize” led to the suggestion (Swann and Carpenter, 1975; King and Carpenter, 1987) that the responses to these transmitters were mediated by the same receptor–channel complex.

This hypothesis is incompatible with the demonstration given here that the GABA- and glutamate-gated channels are clearly distinct; the GABA channel, but not the glutamate channel, shows outward rectification (Matsumoto, 1982; King and Carpenter, 1987, 1989) and is selectively blocked by intracellular sulfate. Exploiting these distinctive characteristics and the independent expression of the receptors in some cells, we have been able to reevaluate the so-called cross-desensitization by analyzing the ability of GABA, glutamate, and other agonists to interact with each of the receptor molecules.

The cross-desensitization was found to be exclusively attributable to the ability of GABA to interact with the glutamate receptor (Oyama et al., 1990). The GABA receptor is unaffected by glutamate. Nevertheless, in cells expressing both receptors, glutamate can reduce the GABA response by auto-desensitizing the part of the response that is mediated by the glutamate receptor. No interactions were observed between ACh-induced responses and either of the responses elicited by the amino acids.

The invertebrate glutamate-gated Cl channels that have been cloned resemble the vertebrate glycine receptor (Vassilatis et al., 1997). Our pharmacological evaluation of the molluscan glutamate receptor points in the same direction.