bluuebunny/crossref_metadata_2025 huggingface dataset:
Spontaneous Network Activity Transiently Depresses Synaptic Transmission in the Embryonic Chick Spinal Cord
Authors: Fedirchuk Brent , Wenner Peter , Whelan Patrick J. , Ho Stephen , Tabak Joel , O’Donovan Michael J.
Published: April 2018

We examined the effects of spontaneous or evoked episodes of rhythmic activity on synaptic transmission in several spinal pathways of embryonic day 9–12 chick embryos. We compared the amplitude of synaptic potentials evoked by stimulation of the ventrolateral funiculus (VLF), the dorsal or ventral roots, before and after episodes of activity. With the exception of the short-latency responses evoked by dorsal root stimulation, the potentials were briefly potentiated and then reduced for several minutes after an episode of rhythmic activity. Their amplitude progressively recovered in the interval between successive episodes. The lack of post-episode depression in the short-latency component of the dorsal root evoked responses is probably attributable to the absence of firing in cut muscle afferents during an episode of activity.

The post-episode depression of VLF-evoked potentials was mimicked by prolonged stimulation of the VLF, subthreshold for an episode of activity. By contrast, antidromically induced motoneuron firing and the accompanying calcium entry did not depress VLF-evoked potentials recorded from the stimulated ventral root. In addition, post-episode depression of VLF-evoked synaptic currents was observed in voltage-clamped spinal neurons. Collectively, these findings suggest that somatic postsynaptic activity and calcium entry are not required for the depression. We propose instead that the mechanism may involve a form of long-lasting activity-induced synaptic depression, possibly a combination of transmitter depletion and ligand-induced changes in the postsynaptic current accompanying transmitter release. This activity-dependent depression appears to be an important mechanism underlying the occurrence of spontaneous activity in developing spinal networks.
Immunocytochemical and Morphological Evidence for Intracellular Self-Repair as an Important Contributor to Mammalian Hair Cell Recovery
Authors: Zheng J. Lisa , Keller Gilbert , Gao Wei-Qiang
Published: April 2018

Although recent studies have provided evidence for hair cell regeneration in mammalian inner ears, the mechanism underlying this regenerative process is still under debate. Here we report immunocytochemical, histological, electron microscopic, and autoradiographic evidence that, in cultured postnatal rat utricles, a substantial number of hair cells can survive gentamicin insult even their stereocilia are lost. These partially damaged hair cells can survive for a prolonged time and regrow the stereocilia. Although the number of stereocilia-bearing hair cells increases over time after gentamicin insult, hair cell and supporting cell numbers remain essentially unchanged. Tritiated thymidine autoradiography and bromodeoxyuridine immunocytochemistry of the cultures demonstrate that cell proliferation in the sensory epithelium is very limited and is far below the number of recovered hair cells. Furthermore, terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling analysis indicates that gentamicin-induced apoptosis in the sensory epithelium occurs mainly during a 2 d treatment period, and additional cell death is minimal 2–11 d after treatment. Considered together, intracellular repair of partially damaged hair cells can be an important contributor to spontaneous hair cell recovery in mammalian inner ears.
The Distribution of Zinc Selenite and Expression of Metallothionein-III mRNA in the Spinal Cord and Dorsal Root Ganglia of the Rat Suggest a Role for Zinc in Sensory Transmission
Authors: Velázquez Rubén A. , Cai Yongjiu , Shi Qiuying , Larson Alice A.
Published: April 2018

Zinc appears to play a role in synaptic transmission in the hippocampus. We tested the hypothesis that zinc is similarly involved in sensory transmission by determining whether vesicular zinc and metallothionein-III (MT-III), a zinc-binding protein, are localized in rat primary afferent neurons. MT-III mRNA, measured using RT-PCR, and MT-III immunoreactivity, were both present in the spinal cord as well as the thoracic and lumbar dorsal root ganglia (DRG). At a time (24 hr) that allows retrograde transport of zinc selenite to cell bodies, only small-diameter neurons and neurons scattered throughout lamina V of the spinal cord were stained by sodium selenite injected intrathecally. This stain disappeared if a ligature was placed on the dorsal root to block axonal transport, demonstrating that these cells are, in fact, zinc-containing primary afferent neurons. When assessed 1 hr after sodium selenite, stain was distributed throughout the neuropil of the spinal cord, especially in lamina III and the area surrounding the central canal. Even in rhizotomized animals, large- and small-diameter DRG neuronal cell bodies were also stained with either selenite (1 hr) or 6-methoxy 8-para-toluene sulfonamide quinoline (TSQ). Paradoxically, this unique pool of zinc was eliminated in large-diameter DRG neurons after neonatal capsaicin treatment, which had no effect on selenite stain or MT-III mRNA content in small-diameter DRG neurons. In summary, we demonstrate that there is a population of capsaicin-insensitive small-diameter primary afferent neurons that are zinc-containing. In addition, there is a unique pool of capsaicin-sensitive zinc that is associated with large-diameter cell bodies.
Presynaptic Inhibition of GABAB-Mediated Synaptic Potentials in the Ventral Tegmental Area during Morphine Withdrawal
Authors: Shoji Yoshihisa , Delfs Jill , Williams John T.
Published: April 2018

Opioids increase the firing of dopamine cells in the ventral tegmental area by presynaptic inhibition of GABA release. This report describes an acute presynaptic inhibition of GABAB-mediated IPSPs by μ- and κ-opioid receptors and the effects of withdrawal from chronic morphine treatment on the release of GABA at this synapse. In slices taken from morphine-treated guinea pigs after washing out the morphine (withdrawn slices), a low concentration of a μ receptor agonist increased, rather than decreased, the amplitude of the GABABIPSP. In withdrawn slices, after blocking A1-adenosine receptors with 8-cyclopentyl-1,3-dipropylxantine, μ-opioid receptor activation inhibited the IPSP at all concentrations and increased the maximal inhibition. In addition, during withdrawal, there was a tonic increase in adenosine tone that was further increased by forskolin or D1-dopamine receptor activation, suggesting that metabolism of cAMP was the source of adenosine. The results indicate that during acute morphine withdrawal, there was an upregulation of the basal level of an opioid-sensitive adenylyl cyclase. Inhibition of this basal activity by opioids had two effects. First, a decrease in the formation of cAMP that decreased adenosine tone. This effect predominated at low μ receptor occupancy and increased the amplitude of the IPSP. Higher agonist concentrations inhibited transmitter release by both kinase-dependent and -independent pathways. This study indicates that the consequences of the morphine-induced upregulation of the cAMP cascade on synaptic transmission are dependent on the makeup of receptors and second messenger pathways present on any given terminal.
Dissociation in Effects of Lesions of the Nucleus Accumbens Core and Shell on Appetitive Pavlovian Approach Behavior and the Potentiation of Conditioned Reinforcement and Locomotor Activity byd-Amphetamine
Authors: Parkinson John A. , Olmstead Mary C. , Burns Lindsay H. , Robbins Trevor W. , Everitt Barry J.
Published: April 2018

Dopamine release within the nucleus accumbens (NAcc) has been associated with both the rewarding and locomotor-stimulant effects of abused drugs. The functions of the NAcc core and shell were investigated in mediating amphetamine-potentiated conditioned reinforcement and locomotion. Rats were initially trained to associate a neutral stimulus (Pavlovian CS) with food reinforcement (US). After excitotoxic lesions that selectively destroyed either the NAcc core or shell, animals underwent additional CS–US training sessions and then were tested for the acquisition of a new instrumental response that produced the CS acting as a conditioned reinforcer (CR). Animals were infused intra-NAcc withd-amphetamine (0, 1, 3, 10, or 20 μg) before each session. Shell lesions affected neither Pavlovian nor instrumental conditioning but completely abolished the potentiative effect of intra-NAcc amphetamine on responding with CR. Core-lesioned animals were impaired during the Pavlovian retraining sessions but showed no deficit in the acquisition of responding with CR. However, the selectivity in stimulant-induced potentiation of the CR lever was reduced, as intra-NAcc amphetamine infusions dose-dependently increased responding on both the CR lever and a nonreinforced (control) lever. Shell lesions produced hypoactivity and attenuated amphetamine-induced activity. In contrast, core lesions resulted in hyperactivity and enhanced the locomotor-stimulating effect of amphetamine. These results indicate a functional dissociation of subregions of the NAcc; the shell is a critical site for stimulant effects underlying the enhancement of responding with CR and locomotion after intra-NAcc injections of amphetamine, whereas the core is implicated in mechanisms underlying the expression of CS–US associations.
Effects of Phosphatidylinositol 4,5-Bisphosphate and Phosphatidylinositol 4-Phosphate on a Na+-Gated Nonselective Cation Channel
Authors: Zhainazarov Aslbek B. , Ache Barry W.
Published: April 2018

Olfactory receptor neurons in the lobster express a nonselective cation channel that is activated by intracellular Na+and carries a substantial part of the depolarizing receptor current. Here, we show that phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and phosphatidylinositol 4-phosphate [PI(4)P] applied to the intracellular face of cell-free patches activate the channel in the absence of Na+and that antibodies against the respective phospholipids irreversibly inhibit the evoked activity. Further, we show that applying PI(4,5)P2or PI(4)P in the presence of Na+decreases the concentration of Na+required to activate the channel from an EC50of 74 to 22 mmfor PI(4,5)P2and to 29 mmfor PI(4)P, respectively. Na+-gated channel activity was irreversibly inhibited by monoclonal antibodies against PI(4,5)P2and PI(4)P in patches never exposed to exogenous phosphatidylinositols, suggesting that endogenous inositol phospholipids are required for the activation of the channel by intracellular Na+. Our findings suggest that PI(4,5)P2and/or PI(4)P may serve as intracellular signaling molecules in these primary sensory neurons and provide a general mechanism to explain how the sensitivity of Na+-gated channels to Na+could be much greater in intact cells than in excised membrane patches.
The Metabotropic Receptor mGluR6 May Signal Through Go, But Not Phosphodiesterase, in Retinal Bipolar Cells
Authors: Nawy Scott
Published: April 2018

Bipolar cells are retinal interneurons that receive synaptic input from photoreceptors. Glutamate, the photoreceptor transmitter, hyperpolarizes On bipolar cells by closing nonselective cation channels, an effect mediated by the metabotropic receptor mGluR6. Previous studies of mGluR6 transduction have suggested that the receptor couples to a phosphodiesterase (PDE) that preferentially hydrolyzes cGMP, and that cGMP directly gates the nonselective cation channel. This hypothesis was tested by dialyzing On bipolar cells with nonhydrolyzable analogs of cGMP. Whole-cell recordings were obtained from On bipolar cells in slices of larval tiger salamander retina. Surprisingly, On bipolar cells dialyzed with 8-(4-chlorophenylthio)-cyclic GMP (8-pCPT-cGMP), or 8-bromo-cyclic GMP (8-Br-cGMP) responded normally to glutamate orl-2-amino-4-phosphonobutyrate (l-APB). Response amplitudes and kinetics were not significantly altered compared with cells dialyzed with cGMP alone. Comparable results were obtained with the PDE inhibitor 3-isobutyl-1-methyl-xanthine (IBMX) or with 8-pCPT-cGMP and IBMX together, indicating that PDE is not required for mGluR6 signal transduction. Addition of the G-protein subunit Goα to the pipette solution suppressed the cation current and occluded the glutamate response, whereas dialysis with Giα or with transducin Gβγ had no significant effect on either the cation current or the response. Dialysis of an antibody directed against Goα also reduced the glutamate response, indicating a functional role for endogenous Goα. These results indicate that mGluR6 may signal through Go, rather than a transducin-like G-protein.
A Changing Pattern of Brain-Derived Neurotrophic Factor Expression Correlates with the Rearrangement of Fibers during Cochlear Development of Rats and Mice
Authors: Wiechers Barbara , Gestwa Glikeria , Mack Andreas , Carroll Patrick , Zenner Hans-Peter , Knipper Marlies
Published: April 2018

The reorganization of specific neuronal connections is a typical feature of the developing nervous system. It is assumed that the refinement of connections in sensory systems requires spontaneous activity before the onset of cochlear function and selective sensory experience during the ensuing period. The mechanism of refinement through sensory experience is currently postulated as being based on the selective reinforcement of active projections by neurotrophins. We studied a presumed role of neurotrophins for rearrangement of afferent and efferent fibers before the onset of sensory function in the precisely innervated auditory end organ, the cochlea. We observed a spatiotemporal change in the localization of brain-derived neurotrophic factor (BDNF) protein and mRNA, which correlated with the reorganization of fibers. Thus, BDNF decreased in target hair cells during fiber retraction and was subsequently upregulated in neurons, target hair cells, and adjacent supporting cells concomitant with the formation of new synaptic contacts. Analysis of the innervation pattern in BDNF gene-deleted mice by immunohistochemistry and confocal microscopy revealed a failure in the rearrangement of fibers and a BDNF dependency of distinct neuronal projections that reorganize in control animals. Our data suggest that, before the onset of auditory function, a spatiotemporal change in BDNF expression in sensory, epithelial, and neuronal cells may guide the initial steps of refinement of the innervation pattern.
Neurotrophin Modulation of the Monosynaptic Reflex after Peripheral Nerve Transection
Authors: Mendell Lorne M. , Johnson Richard D. , Munson John B.
Published: April 2018

The effects of neurotrophin-3 (NT-3) and NT-4/5 on the function of axotomized group Ia afferents and motoneurons comprising the monosynaptic reflex pathway were investigated. The axotomized medial gastrocnemius (MG) nerve was provided with NT-3 or NT-4/5 for 8–35 d via an osmotic minipump attached to its central end at the time of axotomy. After this treatment, monosynaptic EPSPs were recorded intracellularly from MG or lateral gastrocnemius soleus (LGS) motoneurons in response to stimulation of the heteronymous nerve under pentobarbital anesthesia. Controls were preparations with axotomized nerves treated directly with vehicle; other axotomized controls were administered subcutaneous NT-3. Direct NT-3 administration (60 μg/d) not only prevented the decline in EPSP amplitude from axotomized afferents (stimulate MG, record LGS) observed in axotomy controls but, after 5 weeks, led to EPSPs larger than those from intact afferents. These central changes were paralleled by recovery of group I afferent conduction velocity. Removal of NT-3 4–5 weeks after beginning treatment resulted in a decline of conduction velocity and EPSP amplitude within 1 week to values characteristic of axotomy. The increased synaptic efficacy after NT-3 treatment was associated with enhanced connectivity of single afferents to motoneurons. NT-4/5 induced modest recovery in group I afferent conduction velocity but not of the EPSPs they elicited. NT-3 or NT-4/5 had no effect on the properties of treated motoneurons or their monosynaptic EPSPs. We conclude that NT-3, and to a limited extent NT-4/5, promotes recovery of axotomized group Ia afferents but not axotomized motoneurons or the synapses on them.
Hormonal Regulation of Glutamate Receptor Gene Expression in the Anteroventral Periventricular Nucleus of the Hypothalamus
Authors: Gu Guibao , Varoqueaux Frederique , Simerly Richard B.
Published: April 2018

Glutamate plays an important role in mediating the positive feedback effects of ovarian steroids on gonadotropin secretion, and the preoptic region of the hypothalamus is a likely site of action of glutamate. The anteroventral periventricular nucleus (AVPV) of the preoptic region is an essential part of neural pathways mediating hormonal feedback on gonadotropin secretion, and it appears to provide direct inputs to gonadotropin releasing hormone (GnRH)-containing neurons. Immunohistochemistry andin situhybridization were used in this study to define the distribution and hormonal regulation of glutamate receptor subtypes in the AVPV of juvenile female rats. Neurons that express the NMDAR1 receptor subtype are abundant in the AVPV, as are cells that express AMPA receptor subtypes (GluR1, GluR2, and GluR3 but not GluR4), and the AVPV appears to contain a dense plexus of NMDAR1-immunoreactive presynaptic terminals. However, AVPV neurons do not seem to express detectable levels of kainate receptor (GluR5, GluR6, and GluR7) or metabotropic receptor (mGluR1–6) subtypes. Treatment of ovariectomized juvenile rats with estradiol induced expression of GluR1 mRNA but did not alter levels of GluR2 or GluR3 mRNA. Treatment of estrogen-primed ovariectomized juvenile rats with progesterone caused an initial increase in GluR1 mRNA expression, followed by a small decrease 24 hr after treatment. In contrast, estrogen appears to suppress levels of NMDAR1 mRNA in the AVPV, which remained unchanged after progesterone treatment. Thus, one mechanism whereby ovarian steroids may provide positive feedback to GnRH neurons is by altering the sensitivity of AVPV neurons to glutamatergic activation.
Vesicular Monoamine Transporter-2 and Aromaticl-Amino Acid Decarboxylase Enhance Dopamine Delivery afterl-3,4-Dihydroxyphenylalanine Administration in Parkinsonian Rats
Authors: Lee Won Yong , Chang Jin Woo , Nemeth Nicole L. , Kang Un Jung
Published: April 2018

Medical therapy in Parkinson’s disease (PD) is limited by the short-duration response and development of dyskinesia that result from chronicl-3,4-dihydroxyphenylalanine (l-DOPA) therapy. These problems occur partly because the loss of dopamine storage sites leads to erratic dopamine delivery. Vesicular monoamine transporter-2 (VMAT-2) plays a critical role in dopamine storage by packaging dopamine into synaptic vesicles and regulating sustained release of dopamine. To restore the capacity to produce and store dopamine in parkinsonian rats, primary skin fibroblast cells (PF) were genetically modified with aromaticl-amino acid decarboxylase (AADC) and VMAT-2 genes. After incubation withl-DOPA in culture, the doubly transduced fibroblast cells (PFVMAA) produced and stored dopamine at a much higher level than the cells with either gene alone. PFVMAA cells in culture released dopamine gradually in a constitutive manner. Genetically modified fibroblast cells were grafted in parkinsonian rat striata, andl-DOPA was systemically administered. Higher dopamine levels were sustained for a longer duration in rats grafted with PFVMAA cells than in those grafted with either control cells or cells with AADC alone. These findings underscore the importance of dopamine storage capacity in determining the efficacy ofl-DOPA therapy and illustrate a novel method of gene therapy combined with precursor administration to overcome the major obstacles of PD treatment.
Epidermal and Fibroblast Growth Factors Behave as Mitogenic Regulators for a Single Multipotent Stem Cell-Like Population from the Subventricular Region of the Adult Mouse Forebrain
Authors: Gritti Angela , Frölichsthal-Schoeller Paola , Galli Rossella , Parati Eugenio A. , Cova Lidia , Pagano Stefano F. , Bjornson Christopher R. , Vescovi Angelo L.
Published: April 2018

The subventricular zone (SVZ) of the adult mammalian forebrain contains kinetically distinct precursor populations that contribute new neurons to the olfactory bulb. Because among forebrain precursors there are stem-like cells that can be cultured in the presence of mitogens such as epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2), we asked whether distinct subsets of stem-like cells coexist within the SVZ or whether the proliferation of a single type of SVZ stem-like cell is controlled by several GFs. We show that the latter is the case. Thus cells isolated from the SVZ coexpress the EGF and FGF receptors; by quantitative analysis, the number of stem-like cells isolated from the SVZ by either FGF2 or EGF is the same, whereas no additive effect occurs when these factors are used together. Furthermore, short-term administration of high-dose [3H]thymidinein vivodepletes both the EGF- and FGF2-responsive stem-like cell populations equally, showing they possess closely similar proliferation kinetics and likely belong to the constitutively proliferating SVZ compartment. By subcloning and population analysis, we demonstrate that responsiveness to more than one GF endows SVZ cells with an essential stem cell feature, the ability to vary self-renewal, that was until now undocumented in CNS stem-like cells. The multipotent stem cell-like population that expands slowly in the presence of FGF2 in culture switches to a faster growth mode when exposed to EGF alone and expands even faster when exposed to both GFs together. Analogous responses are observed when the GFs are used in the reverse order, and furthermore, these growth rate modifications are fully reversible.
Transcription Factors in Neuroendocrine Regulation: Rhythmic Changes in pCREB and ICER Levels Frame Melatonin Synthesis
Authors: Maronde Erik , Pfeffer Martina , Olcese James , Molina Carlos A. , Schlotter Florence , Dehghani Faramarz , Korf Horst-Werner , Stehle Jörg H.
Published: April 2018

Neurotransmitter-driven activation of transcription factors is important for control of neuronal and neuroendocrine functions. We show with anin vivoapproach that the norepinephrine cAMP-dependent rhythmic hormone production in rat pineal gland is accompanied by a temporally regulated switch in the ratio of a transcriptional activator, phosphorylated cAMP-responsive element–binding protein (pCREB), and a transcriptional inhibitor, inducible cAMP early repressor (ICER). pCREB accumulates endogenously at the beginning of the dark period and declines during the second half of the night. Concomitant with this decline, the amount of ICER rises. The changing ratio between pCREB and ICER shapes thein vivodynamics in mRNA and, thus, protein levels of arylalkylamine-N-acetyltransferase, the rate-limiting enzyme of melatonin synthesis. Consequently, a silenced ICER expression in pinealocytes leads to a disinhibited arylalkylamine-N-acetyltransferase transcription and a primarily enhanced melatonin synthesis.
From Embryo to Adult: Persistent Neurogenesis and Apoptotic Cell Death Shape the Lobster Deutocerebrum
Authors: Harzsch Steffen , Miller Julie , Benton Jeannie , Beltz Barbara
Published: April 2018

Neuronal plasticity and synaptic remodeling play important roles during the development of the invertebrate nervous system. In addition, structural neuroplasticity as a result of long-term environmental changes, behavioral modifications, age, and experience have been demonstrated in the brains of sexually mature insects. In adult vertebrates, persistent neurogenesis is found in the granule cell layer of the mammalian hippocampus and the subventricular zone, as well as in the telencephalon of songbirds, indicating that persistent neurogenesis, which is presumably related to plasticity and learning, may be an integral part of the normal biology of the mature brain. In decapod crustaceans, persistent neurogenesis among olfactory projection neurons is a common principle that shapes the adult brain, indicating a remarkable degree of life-long structural plasticity. The present study closes a gap in our knowledge of this phenomenon by describing the continuous cell proliferation and gradual displacement of proliferation domains in the central olfactory pathway of the American lobsterHomarus americanusfrom early embryonic through larval and juvenile stages into adult life. Neurogenesis in the deutocerebrum was examined by thein vivoincorporation of bromodeoxyuridine, and development and structural maturation of the deutocerebral neuropils were studied using immunohistochemistry againstDrosophilasynapsin. The role of apoptotic cell death in shaping the developing deutocerebrum was studied using the terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling method, combined with immunolabeling using an antiphospho histone H3 mitosis marker. Our results indicate that, in juvenile and adult lobsters, birth and death of olfactory interneurons occur in parallel, suggesting a turnover of these cells. When the persistent neurogenesis and concurrent death of interneurons in the central olfactory pathway of the crustacean brain are taken into account with the life-long turnover of olfactory receptor cells in crustacean antennules, a new, highly dynamic picture of olfaction in crustaceans emerges.
Voltage-Dependent Neuromodulation of Na+Channels by D1-Like Dopamine Receptors in Rat Hippocampal Neurons
Authors: Cantrell Angela R. , Scheuer Todd , Catterall William A.
Published: April 2018

Activation of D1-like dopamine (DA) receptors reduces peak Na+current in acutely isolated hippocampal neurons through phosphorylation of the α subunit of the Na+channel by cAMP-dependent protein kinase (PKA). Here we report that neuromodulation of Na+currents by DA receptors via PKA is voltage-dependent in the range of −110 to −70 mV and is also sensitive to concurrent activation of protein kinase C (PKC). Depolarization enhanced the ability of D1-like DA receptors to reduce peak Na+currents via the PKA pathway. Similar voltage-dependent modulation was observed when PKA was activated directly with the membrane-permeant PKA activator DCl-cBIMPS (cBIMPS; 20 μm), indicating that the membrane potential dependence occurs downstream of PKA. PKA activation caused only a small (−2.9 mV) shift in the voltage dependence of steady-state inactivation and had no effect on slow inactivation or on the rates of entry into the fast or slow inactivated states, suggesting that another mechanism is responsible for coupling of membrane potential changes to PKA modulation. Activation of PKC with a low concentration of the membrane-permeant diacylglycerol analog oleylacetyl glycerol also potentiated modulation by SKF 81297 or cBIMPS, and these effects were most striking at hyperpolarized membrane potentials where PKA modulation was not stimulated by membrane depolarization. Thus, activation of D1-like DA receptors causes a strong reduction in Na+current via the PKA pathway, but it is effective primarily when it is combined with depolarization or activation of PKC. The convergence of these three distinct signaling modalities on the Na+channel provides an intriguing mechanism for integration of information from multiple signaling pathways in the hippocampus and CNS.
Selective Disruption of “Late Onset” Sagittal Banding Patterns by Ectopic Expression of Engrailed-2 in Cerebellar Purkinje Cells
Authors: Baader Stephan L. , Vogel Michael W. , Sanlioglu Salih , Zhang Xulun , Oberdick John
Published: April 2018

To explore the role of Engrailed proteins in development of the cerebellum, Engrailed-2 (En-2) was ectopically expressed in cerebellar Purkinje cells from the late embryonic stage into adulthood. The fundamental organization of Purkinje cell sagittal zones as revealed by the “early onset” markers L7-β-gal and cadherin-8 was found to be virtually identical to that in wild type. In contrast, “late onset” sagittal banding patterns revealed by Purkinje cell markers zebrin I, zebrin II, and 9-O-acetyl GD3Ganglioside (P-Path), and the granule cell marker NADPH-diaphorase, were disrupted. In general, although some evidence of banding was still detectable, boundaries defined by the latter markers were poorly defined, and the patterns overall took on a diffuse appearance. In parallel with the changes in late onset markers, anterograde tracing of spinocerebellar axons revealed a general diffusion of the mossy fiber projection pattern in lobule VIII and the anterior lobe. These observations suggest that at least two separate mediolateral boundary systems exist in the cerebellum, and these are differentially affected by ectopic En-2 expression. Alternatively, one boundary system exists that remains primarily intact in the mutant, but recognition of this system by a set of late developmental events is perturbed.
Expression of the Striatal DARPP-32/ARPP-21 Phenotype in GABAergic Neurons Requires NeurotrophinsIn VivoandIn Vitro
Authors: Ivkovic Sanja , Ehrlich Michelle E.
Published: April 2018

The medium spiny neuron (MSN) is the major output neuron of the caudate nucleus and uses GABA as its primary neurotransmitter. A majority of MSNs coexpress DARPP-32 and ARPP-21, two dopamine and cyclic AMP-regulated phosphoproteins, and most of the matrix neurons express calbindin. DARPP-32 is the most commonly used MSN marker, but previous attempts to express this genein vitrohave failed. In this study we found that DARPP-32 is expressed in <12% of E13- or E17-derived striatal neurons when they are grown in defined media at high or low density in serum, dopamine, or Neurobasal/N2 (Life Technologies), and ARPP-21 is expressed in <1%. The percentage increases to 25% for DARPP-32 and 10% for ARPP-21 when the same cells are grown in Neurobasal/B27 (Life Technologies) for 7 d. After growth in Neurobasal/B27 plus brain-derived neurotrophic factor (BDNF) for 7 d, E13-derived MSNs are 53.7% DARPP-32-positive and 29.0% ARPP-21-positive; E17-derived MSNs are 66.8% DARPP-32-positive and 51.5% ARPP-21-positive. The percentage of calbindin-positive neurons also is increased under these conditions. Finally, ARPP-21 expression is reduced in mice with a targeted deletion of the BDNF gene. We conclude that BDNF is required for the maturation of a large subset of patch and matrix MSNsin vivoandin vitro. In addition, we introduce a culture system in which highly differentiated MSNs may be generated, maintained, and studied.
Different Proctolin Neurons Elicit Distinct Motor Patterns from a Multifunctional Neuronal Network
Authors: Blitz Dawn M. , Christie Andrew E. , Coleman Melissa J. , Norris Brian J. , Marder Eve , Nusbaum Michael P.
Published: April 2018

Distinct motor patterns are selected from a multifunctional neuronal network by activation of different modulatory projection neurons. Subsets of these projection neurons can contain the same neuromodulator(s), yet little is known about the relative influence of such neurons on network activity. We have addressed this issue in the stomatogastric nervous system of the crabCancer borealis. Within this system, there is a neuronal network in the stomatogastric ganglion (STG) that produces many versions of the pyloric and gastric mill rhythms. These different rhythms result from activation of different projection neurons that innervate the STG from neighboring ganglia and modulate STG network activity. Three pairs of these projection neurons contain the neuropeptide proctolin. These include the previously identified modulatory proctolin neuron and modulatory commissural neuron 1 (MCN1) and the newly identified modulatory commissural neuron 7 (MCN7). We document here that each of these neurons contains a unique complement of cotransmitters and that each of these neurons elicits a distinct version of the pyloric motor pattern. Moreover, only one of them (MCN1) also elicits a gastric mill rhythm. The MCN7-elicited pyloric rhythm includes a pivotal switch by one STG network neuron from playing a minor to a major role in motor pattern generation. Therefore, modulatory neurons that share a peptide transmitter can elicit distinct motor patterns from a common target network.
Exacerbation of Damage and Altered NF-κB Activation in Mice Lacking Tumor Necrosis Factor Receptors after Traumatic Brain Injury
Authors: Sullivan Patrick G. , Bruce-Keller Annadora J. , Rabchevsky Alexander G. , Christakos Sylivia , Clair Daret K. St. , Mattson Mark P. , Scheff Stephen W.
Published: April 2018

Tumor necrosis factor α (TNFα) is widely expressed in both neurons and glia and has been shown to be upregulated after traumatic brain injury (TBI). TNFα receptor activation results in activation of the transcription factor nuclear factor κB (NF-κB), which may serve an antiapoptotic role via the induction of target genes manganese superoxide dismutase (MnSOD) and/or calbindin. In the present study, we used a controlled cortical impact model of TBI with pertinent lines of transgenic mice to combine both morphological characterization and molecular analysis to elucidate the role of TNFα after TBI. Measurements of both the lesion volume and the blood–brain barrier breach indicated exacerbations in mice rendered genetically deficient in both the p55 and p75 TNFα receptors (TNFR-KO) compared with wild-type animals. Additionally, animals genetically altered to overexpress MnSOD showed a significant decrease in lesion volume compared with that of control littermates, whereas no alterations were observed in mice lacking the calcium-binding protein calbindin D28k. Analysis of NF-κB activation and relative levels of MnSOD revealed delayed responses in the injured cortex of TNFR-KO animals compared with wild-type animals, implying that endogenous TNFα may be neuroprotective after TBI.
Conformational Ensembles: The Role of Neuropeptide Structures in Receptor Binding
Authors: Edison Arthur S. , Espinoza Eduardo , Zachariah Cherian
Published: April 2018

Conformational properties of several similar FMRFamide-like neuropeptides from mollusks were investigated by nuclear magnetic resonance (NMR) spectroscopy. It was found that amino acid substitutions in the N-terminal variable regions of the peptides had dramatic effects on the populations of reverse turns in solution. The populations of turns, as measured by two independent NMR parameters, were found to be highly correlated (r2= 0.93 and 0.82) with IC50values using receptor membrane preparations fromHelix aspersa(Payza, 1987; Payza et al., 1989). These results suggest that the amount of turn in the free peptide can influence the receptor binding affinities of that peptide. On the basis of these observations, a model was developed in which only a single species from a conformational ensemble of an unbound peptide will bind to a particular receptor. Thus, the conformational ensemble reduces the effective concentration of a particular peptide with respect to a particular receptor.
Association of AMPA Receptors with a Subset of Glutamate Receptor-Interacting ProteinIn Vivo
Authors: Wyszynski Michael , Valtschanoff Juli G. , Naisbitt Scott , Dunah Anthone W. , Kim Eunjoon , Standaert David G. , Weinberg Richard , Sheng Morgan
Published: April 2018

The NMDA and AMPA classes of ionotropic glutamate receptors are concentrated at postsynaptic sites in excitatory synapses. NMDA receptors interact via their NR2 subunits with PSD-95/SAP90 family proteins, whereas AMPA receptors bind via their GluR2/3 subunits to glutamate receptor-interacting protein (GRIP), AMPA receptor-binding protein (ABP), and protein interacting with C kinase 1 (PICK1). We report here a novel cDNA (termed ABP-L/GRIP2) that is virtually identical to ABP except for additional GRIP-like sequences at the N-terminal and C-terminal ends. Like GRIP (which we now term GRIP1), ABP-L/GRIP2 contains a seventh PDZ domain at its C terminus. Using antibodies that recognize both these proteins, we examined the subcellular localization of GRIP1 and ABP-L/GRIP2 (collectively termed GRIP) and their biochemical association with AMPA receptors. Immunogold electron microscopy revealed the presence of GRIP at excitatory synapses and also at nonsynaptic membranes and within intracellular compartments. The association of native GRIP and AMPA receptors was confirmed biochemically by coimmunoprecipitation from rat brain extracts. A majority of detergent-extractable GluR2/3 was complexed with GRIP in the brain. However, only approximately half of GRIP was associated with AMPA receptors. Unexpectedly, immunocytochemistry of cultured hippocampal neurons and rat brain at the light microscopic level showed enrichment of GRIP in GABAergic neurons and in GABAergic nerve terminals. Thus GRIP is associated with inhibitory as well as excitatory synapses. Collectively, these findings support a role for GRIP in the synaptic anchoring of AMPA receptors but also suggest that GRIP has additional functions unrelated to the binding of AMPA receptors.
Glutamate Inhibits Thalamic Reticular Neurons
Authors: Cox Charles L. , Sherman S. Murray
Published: April 2018

Activation of metabotropic glutamate receptors (mGluRs) can result in long-lasting modulation of neuronal excitability. Multiple mGluR subtypes are localized within the rat thalamic reticular nucleus (TRN), and we have examined the effects of activating these different receptor subtypes on the excitability of these neurons using anin vitroslice preparation. Typical of most mGluR-sensitive preparations, the general mGluR agonist, (±)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (ACPD) produced a robust, long-lasting excitatory response. Surprisingly, ACPD produced a membrane hyperpolarization in some neurons. Using selective mGluR agonists, we found that activation of group II mGluRs produces the hyperpolarization, whereas the depolarization is mediated by group I mGluRs. While the polarity of the postsynaptic response (hyperpolarization vs depolarization) was dependent on the mGluR subtype activated, both actions appear to result from modification of a linear K+conductance. The inhibitory action of Glutamate, via group II mGluRs, provides an avenue for a disinhibitory effect that could have interesting consequences upon a well-investigated, model neuronal circuit, turning its assumed functional role upside down.
Identification of Transduction Elements for Benzodiazepine Modulation of the GABAAReceptor: Three Residues Are Required for Allosteric Coupling
Authors: Boileau Andrew J. , Czajkowski Cynthia
Published: April 2018

Modulation of GABAAreceptors by benzodiazepines (BZDs) is believed to involve two distinct steps: a recognition step in which BZDs bind and a conformational transition step in which the affinity of the receptor for GABA changes. Previously, using γ2/α1chimeric subunits (χ), we demonstrated that although the N-terminal 167 γ2amino acid residues confer high-affinity BZD binding, other γ2domains couple BZD binding to potentiation of the GABA-mediated Cl−current (IGABA). To determine which γ2regions couple binding to potentiation, we generated χs with longer N-terminal γ2segments for voltage-clamp experiments inXenopusoocytes. Chimeras containing greater than the N-terminal 167 γ2residues showed incremental gains in maximal potentiation for diazepam enhancement ofIGABA. Residues in γ2199–236, γ2224–236 (pre-M1), and particularly γ2257–297 (M2 and surrounding loops) are important for BZD potentiation. For several positive BZD modulators tested, the same regions restored potentiation ofIGABA. In contrast, β-carboline inverse-agonism was unaltered in chimeric receptors, suggesting that structural determinants for positive and negative BZD allosteric modulation are different. Dissection of the γ2257–297 domain revealed that three residues in concert, γ2T281, γ2I282 (M2 channel vestibule), and γ2S291 (M2–M3 loop) are necessary to impart full BZD potentiation to chimeric receptors. Thus, these residues participate in coupling distant BZD-binding events to conformational changes in the GABAAreceptor. The location of these novel residues provides insight into the mechanisms underlying allosteric coupling for other members of the ligand-gated ion channel superfamily.
Protein Kinase C Activators Inhibit the Visual Cascade inLimulusVentral Photoreceptors at an Early Stage
Authors: Dabdoub Alain , Payne Richard
Published: April 2018

The phosphoinositide cascade mediates visual transduction in invertebrate photoreceptors. Phospholipase C (PLC) catalyzes the hydrolysis of phosphatidylinositol bisphosphate, producing inositol trisphosphate (InsP3) and diacylglycerol (DAG). Protein kinase C (PKC) is a major target of DAG in many cell types. We have used PKC activators to investigate the function of the kinase in the phototransduction cascade inLimulus polyphemusventral photoreceptors. Extracellular application of (−)-indolactam V (0.03–30 μm) or phorbol-12,13-dibutyrate (10 μm) reversibly reduced the sensitivity of the electrical response of the photoreceptors to light by up to 1000-fold. The inert stereoisomer (+)-indolactam V and 4α-phorbol had no effect. The effect of (−)-indolactam V was antagonized by the PKC inhibitors bisindolylmaleimide I and Gö 6976. Coapplication of bisindolylmaleimide V, used as a negative control compound for PKC inhibition, did not reduce the effectiveness of (−)-indolactam V. These findings are consistent with (−)-indolactam V activating PKC and desensitizing the light response. Furthermore, our pharmacological results indicate that PKC activation does not appear to play a role in light adaptation. We localized the position of the target of PKC in the visual cascade. We chemically excited the cascade at various stages to determine the kinase's target. PKC activation by (−)-indolactam V decreased the light-induced elevation of intracellular calcium but had no effect on the photoreceptor's excitatory response to intracellular injection of InsP3. However, the PKC activator greatly reduced the excitation caused by GTP-γ-S injection. We propose that PKC inhibits the visual transduction cascade at the G-protein and/or PLC stage.
Conservation of Expression of Neuropeptide Y5 Receptor between Human and Rat Hypothalamus and Limbic Regions Suggests an Integral Role in Central Neuroendocrine Control
Authors: Nichol Kerry Anne , Morey Adrienne , Couzens Michelle Heather , Shine John , Herzog Herbert , Cunningham Anne Marie
Published: April 2018

Neuropeptide Y receptors belong to the G-protein-coupled receptor superfamily and mediate a wide variety of physiological functions, including blood pressure regulation, hormone release, appetite control, seizure propensity, cognition, and emotion. The recent description of a new neuropeptide Y receptor, Y5, expressed in hypothalamic nuclei in rat brain, raised the possibility that Y5 was the receptor mediating the feeding and appetite-related functions of neuropeptide Y. This was supported by subsequent data showing a downregulation of this “feeding” receptor in the brain of the obese Zucker rat (Widdowson, 1997). We have performed a detailed analysis of Y5 expression in rat brain usingin situhybridization histochemistry with digoxygenin-labeled riboprobes and compared this to expression of Y5 in human brain regions. mRNA for the human Y5 receptor was highly expressed in human hypothalamic and thalamic nuclei. In particular, the arcuate and paraventricular nuclei of the hypothalamus, midline thalamic nuclei, and amygdala showed very high levels of expression with high levels in hippocampus. The striking conservation of expression of the rat and human Y5 receptors in relevant hypothalamic and other nuclei implies sharing of a major neuroendocrine functional role by this receptor.
Cerebellar Granule Cell-Specific and Inducible Expression of Cre Recombinase in the Mouse
Authors: Tsujita Mika , Mori Hisashi , Watanabe Masahiko , Suzuki Misao , Miyazaki Jun-ichi , Mishina Masayoshi
Published: April 2018

To develop a cell type-specific and temporal regulation system of gene targeting in the cerebellum, we used the NMDA-type glutamate receptor GluRε3 subunit gene and Cre recombinase-progesterone receptor fusion (CrePR) gene in combination. Injection of the CrePR gene placed under the control of the 10 kb 5′ region of the GluRε3 gene into C57BL/6 eggs yielded the ECP25 line that strongly expressed the CrePR mRNA selectively in the granule cells of the cerebellum. Using a transgenic mouse carrying a reporter gene for Cre-mediated recombination, we showed that antiprogestins could induce the recombinase activity of CrePR protein in the cerebellar granule cells of the ECP25 line. Thus, the established mouse line will provide a valuable tool to investigate the mechanism of cerebellar function by manipulating molecules in the temporally regulated and granule cell-specific manner.
L-Type Voltage-Gated Calcium Channels Mediate NMDA-Independent Associative Long-Term Potentiation at Thalamic Input Synapses to the Amygdala
Authors: Weisskopf Marc G. , Bauer Elizabeth P. , LeDoux Joseph E.
Published: April 2018

Long-term potentiation (LTP) in the amygdala is a leading candidate mechanism to explain fear conditioning, a prominent model of emotional memory. LTP occurs in the pathway from the auditory thalamus to the lateral amygdala, and during fear conditioning LTP-like changes occur in the synapses of this pathway. Nevertheless, LTP has not been investigated in the thalamoamygdala pathway usingin vitrorecordings; hence little is known about the underlying mechanisms. We therefore examined thalamoamygdala LTPin vitrousing visualized whole-cell patch recording. LTP at these synapses was dependent on postsynaptic calcium entry, similar to synaptic plasticity in other regions of the brain. However, unlike many forms of synaptic plasticity, thalamoamygdala LTP was independent of NMDA receptors, despite their presence at these synapses, and instead was dependent on L-type voltage-gated calcium channels. This was true when LTP was induced by pairing presynaptic activity with either action potentials or constant depolarization in the postsynaptic cell. In addition, the LTP was associative, in that it required concurrent pre- and postsynaptic activity, and it was synapse specific. Thus, although this LTP is different from that described at other synapses in the brain, it is nonetheless well suited to mediate classical fear conditioning.
Presenilin-1 Mutation Increases Neuronal Vulnerability to Focal IschemiaIn Vivoand to Hypoxia and Glucose Deprivation in Cell Culture: Involvement of Perturbed Calcium Homeostasis
Authors: Mattson Mark P. , Zhu Haiyan , Yu Jin , Kindy Mark S.
Published: April 2018

Many cases of early-onset inherited Alzheimer's disease (AD) are caused by mutations in the presenilin-1 (PS1) gene. Studies of cultured neural cells suggest that PS1 mutations result in perturbed cellular calcium homeostasis and may thereby render neurons vulnerable to apoptosis. In light of evidence that metabolic impairment plays a role in AD, that cerebral ischemia may be a risk factor for AD, and that individuals with AD have increased morbidity and mortality after stroke, we examined the impact of a PS1 mutation on neuronal vulnerability to ischemic injury. We report that the extent of brain injury after focal cerebral ischemia reperfusion is increased, and behavioral outcome is worsened, in PS1 mutant knock-in mice compared to wild-type mice. Cultured cortical neurons from PS1 mutant mice exhibit increased vulnerability to glucose deprivation and chemical hypoxia compared to their wild-type counterparts. Calcium imaging studies demonstrated enhanced elevation of intracellular calcium levels after glucose deprivation and chemical hypoxia in neurons from PS1 mutant mice. Agents that block calcium release from IP3- and ryanodine-sensitive stores (xestospongin and dantrolene, respectively) protected against the endangering action of the PS1 mutation. Our data suggest that presenilin mutations may promote neuronal degeneration in AD by increasing the sensitivity of neurons to age-related ischemia-like conditions. The data further suggest that drugs that stabilize endoplasmic reticulum calcium homeostasis may prove effective in suppressing the neurodegenerative process in AD patients.