bluuebunny/crossref_metadata_2025 huggingface dataset:
Pioneer growth cone steering decisions mediated by single filopodial contacts in situ
Authors: O'Connor TP , Duerr JS , Bentley D
Published: March 2018

In grasshopper embryo limb buds, the sibling Ti1 pioneers are the first neurons to initiate axonogenesis. The pioneer growth cones migrate from the limb tip to the CNS along a in direction comprising discrete steering events. Filopodial exploration of the cellular terrain in the vicinity of the advancing growth cone appears to be important for steering. Some information is available on the identity of cells and cell types, on cell-surface characteristics, and on the involvement of basal lamina in these steering decisions. In the work reported here, we have used computer-enhanced fluorescence video microscopy to examine filopodial behavior and the process of growth cone migration and reorientation resulting from interactions with the normal guidance cues on the in situ substrate. We observed several different kinds of migration and steering events, which appear to be related to the absolute and relative affinities of the contacted substrates. On a relatively homogeneous substrate of intrasegmental epithelium, growth cones advance by extending veils between filopodia, as is commonly observed on uniform substrates in vitro. Where growth cones confront an orthogonal border between substrates of dissimilar affinity, they remain on the higher-affinity substrate by extending new branches along it. Subsequently, reorientation in the preferred direction on the higher-affinity substrate is accomplished by regression of branches extended in the nonselected direction. By contrast, a single filopodial contact with a very high-affinity substrate, such as a guidepost neuron, can reorient a growth cone, even when it is migrating on a favorable substrate. In this situation, the filopodium that contacts the high-affinity substrate expands in diameter until it becomes the nascent axon.
Expression of multiple neurotransmitter receptors by sympathetic preganglionic neurons in vitro
Authors: Clendening B , Hume RI
Published: March 2018

Neurons in the CNS generally receive inputs form multiple afferent sources. These afferent systems seldom all use the same neurotransmitter, so most central neurons are required to express multiple neurotransmitter receptors. This work addresses the issue of how multiple neurotransmitter receptors are regulated on the surface of individual neurons. We made whole-cell voltage-clamp recordings from identified chick sympathetic preganglionic neurons (SPNs) in dissociated cell cultures. The neurons were derived from stage 30-31 (7 d) chick embryos and were studied within the first week in vitro. We found that by 1 week in vitro, most SPNs responded to the application of GABA, glycine, and glutamate. The responses of SPNs to the amino acid neurotransmitters were similar to the responses of other CNS neurons to these 3 substances. SPNs became sensitive to these substances at different times in culture. At 1 d in vitro, most cells already responded to GABA, and about half of the cells also responded to glycine and kainate. In contrast, responses to NMDA and quisqualate were usually not seen until day 3-4 in vitro. Although there was a general trend for the amplitude of the responses of SPNs to each of the neurotransmitters to increase with time in vitro, there was an immense amount of cell-to-cell variability. By measuring the amplitudes of the responses of a series of SPNs to all 3 transmitters, we were able to test whether a common regulatory mechanism governed the level of responsiveness of SPNs to all 3 amino acid transmitters. We found no correlation between cells in the amplitudes of their responses to the 3 transmitters. Both the differences in time course of appearance of responsiveness and the lack of correlation in the amplitude of responses suggest that the multiple receptors on the surface of SPNs in vitro are independently regulated.
Calcium activates two types of potassium channels in rat hippocampal neurons in culture
Authors: Lancaster B , Nicoll RA , Perkel DJ
Published: March 2018

Several calcium-dependent potassium currents can contribute to the electrophysiological properties of neurons. In hippocampal pyramidal cells, 2 afterhyperpolarizations (AHPs) are mediated by different calcium-activated potassium currents. First, a rapidly activated current contributes to action-potential repolarization and the fast AHP following individual action potentials. In addition, a slowly developing current underlies the slow AHP, which occurs after a burst of action potentials and contributes substantially to the spike- frequency accommodation observed in these cells during a prolonged depolarizing current pulse. In order to investigate the single Ca2(+)- dependent channels that might underlie these currents, we performed patch-clamp experiments on hippocampal neurons in primary culture. When excised inside-out patches were exposed to 1 microM Ca2+, 2 types of channel activity were observed. In symmetrical bathing solutions containing 140 mM K+, the channels had conductances of 19 pS and 220 pS, and both were permeable mainly to potassium ions. The properties of these 2 channels differed in a number of ways. At negative membrane potentials, the small-conductance channels were more sensitive to Ca2+ than the large channels. At positive potentials, the small-conductance channels displayed a flickery block by Mg2+ ions on the cytoplasmic face of the membrane. Low concentrations of tetraethylammonium (TEA) on the extracellular face of the membrane specifically caused an apparent reduction of the large-channel conductance. The properties of the large- and small-conductance channels are in accord with those of the fast and slow AHP, respectively.
Modification of leech behavior patterns by reserpine-induced amine depletion
Authors: O'Gara BA , Chae H , Latham LB , Friesen WO
Published: March 2018

A single injection of 100 micrograms reserpine into the crop of the medicinal leech, Hirudo medicinalis, reduced CNS serotonin and dopamine levels to less than 1% of control values within 3 d. High-pressure liquid chromotography- (HPLC) determined CNS serotonin and dopamine levels remained maximally depressed for approximately 1 month following reserpine injection. Subsequently, amine levels recovered slowly, but remained depressed 6 months after reserpine injection. Following reserpine treatment, glyoxylic acid-induced fluorescence or neutral red staining closely mirrored the HPLC-determined time course of amine depletion and recovery. Acute exposure of isolated ganglia to 10 microM reserpine for periods up to 6 hr produced a 20–30% reduction of serotonin and dopamine content. The threshold concentration of reserpine necessary to produce amine depletion was approximately 1 microM. We found that reserpine treatment eliminated biting behavior within 4 d following injection. Biting behavior remained depressed below control levels for approximately 4 months, but returned to control values while CNS serotonin and dopamine levels remained significantly depressed at this time. Unexpectedly, reserpine treatment increased rather than reduced the duration of stimulus-evoked swimming activity. This behavioral change was evident within 3 d and persisted for approximately 3.5 months. To rapidly restore amine levels in reserpine-treated animals, we bathed intact leeches in pond water containing serotonin, dopamine, or octopamine. We found that biting behavior was restored following reserpine treatment by bathing intact leeches in pond water containing serotonin or dopamine, but not octopamine. Also contrary to expectations, the increase in swim duration was not reversed by bath exposure to serotonin, dopamine, octopamine, or histamine. However, all swimming activity in reserpine- treated leeches was eliminated by the amine antagonist cyproheptadine. We propose that the presence of low levels of amines is critical for the expression of both biting and swimming activity in leeches. However, the minimal levels of amines necessary for the expression of these behaviors are lower for swimming than for biting.
Acetylcholine release from the rabbit retina mediated by kainate receptors
Authors: Linn DM , Blazynski C , Redburn DA , Massey SC
Published: March 2018

The cholinergic amacrine cells of the rabbit retina may be labeled with 3H-choline (3H-Ch), and the activity of the cholinergic population may be monitored by following the release of 3H-ACh. Glutamate analogs caused massive ACh release, up to 50 times the basal efflux, with the following rank order of potency: alpha-amino-3-hydroxy-5-methyl-4- isoxazolepropionic acid (AMPA) greater than quisqualate (QQ) = kainate (KA) much greater than NMDA (in magnesium-free medium) much greater than glutamate greater than aspartate. In contrast, the release of 3H- Ch was unchanged. Submaximal doses of each agonist were used to establish the specifity of glutamate antagonists. Kynurenic acid was selective for KA much greater than QQ, and 6,7-dinitroquinoxaline-2,3- dione (DNQX) was selective for KA greater than QQ much greater than NMDA. At low doses, which selectively blocked the response to KA, both antagonists blocked the light-evoked release of ACh. These results suggest that ACh release may be produced via several glutamate receptors, but the physiological input to the cholinergic amacrine cells is mediated by KA receptors. Because these cells receive direct input from cone bipolar cells, this work supports previous evidence that the bipolar cell transmitter is glutamate.
Calcium, network activity, and the role of NMDA channels in synaptic plasticity in vitro
Authors: Fields RD , Yu C , Nelson PG
Published: March 2018

Functionally effective neuronal circuits are constructed through a competitive process that requires patterned neuronal activity elicited by structured input from the environment. To explore the mechanisms of this activity-dependent synaptic restructuring, we have developed an in vitro preparation of mouse spinal cord neurons maintained in a 3- chambered cell-culture system. Sensory afferents that received chronic electrical stimulation for 3–5 d developed stronger synaptic connections than unstimulated afferents converging onto the same postsynaptic spinal cord neuron. Exposure to 100 microM DL-2-amino-5- phosphonovaleric acid (APV), an antagonist of the NMDA channel, during the stimulation period prevented the competitive advantage associated with electric stimulation. However, when APV was applied with a higher concentration of calcium (3 mM), activity-dependent synaptic plasticity was no longer inhibited by the NMDA receptor antagonist. This reversal of APV block of the plasticity was not impaired by reducing transmitter release with 3 mM magnesium (in addition to 3 mM calcium and APV). A suppressant effect of APV on spontaneous activity was observed, which was attributed to loss of the NMDA component of the EPSP. Activity- dependent plasticity was also blocked if spontaneous activity was suppressed with dilute tetrodotoxin (TTX; 5–10 nM), a dosage that reduces excitability of neurons but is insufficient to block sodium- dependent action potentials. These experiments bring into question how NMDA channel activation is involved in the processes of synaptic remodeling during development. The data suggest that postsynaptic activity is required for synaptic remodeling, but this activity need not involve NMDA receptor activation specifically for activity-evoked synaptic plasticity. Instead, the mechanism for plasticity appears to operate through calcium-dependent processes in general.
Ultrastructural evidence for GABAergic brain stem projections to spinal motoneurons in the rat
Authors: Holstege JC
Published: March 2018

In the present ultrastructural study in the rat, it was determined whether GABA was present in projections descending from the ventromedial reticular formation of the lower brain stem to motoneuronal cell groups in the lumbar spinal cord. For this purpose, the anterograde transport of WGA-HRP was combined with the postembedding immunogold technique for GABA, with the advantage that both markers could be visualized simultaneously in a single terminal. In 4 rats, WGA-HRP was injected in the ventromedial part of the brain stem reticular formation at levels between the rostral inferior olive and the caudal part of the facial nucleus. Vibratome sections were cut from the lumbar spinal cord, reacted for WGA-HRP, and processed for electron microscopy. Ultrathin sections containing the lateral motoneuronal cell groups were cut and treated following the immunogold technique using a polyclonal antibody directed against GABA. It was found that nearly 40% of the terminal profiles that were labeled with WGA-HRP reaction products from the ventromedial brain stem were also labeled for GABA (double labeled). Most of the double-labeled terminals (81%) were F-type (containing many flattened vesicles), 12% were G-type (containing many granular vesicles), and 7% were S-type (containing many spherical vesicles). The majority of the double-labeled terminals contacted proximal dendrites. It is argued that the descending GABAergic projection produces a general inhibitory effect on spinal motoneurons, counteracting the general facilitation produced by the serotonergic projection derived from the same brain stem area.(ABSTRACT TRUNCATED AT 250 WORDS)
Organization of visual inputs to the inferior temporal and posterior parietal cortex in macaques
Authors: Baizer JS , Ungerleider LG , Desimone R
Published: March 2018

It has been proposed that visual information in the extrastriate cortex is conveyed along 2 major processing pathways, a “dorsal” pathway directed to the posterior parietal cortex, underlying spatial vision, and a “ventral” pathway directed to the inferior temporal cortex, underlying object vision. To determine the relative distributions of cells projecting to the 2 pathways, we injected the posterior parietal and inferior temporal cortex with different fluorescent tracers in 5 rhesus monkeys. The parietal injections included the ventral intraparietal (VIP) and lateral intraparietal (LIP) areas, and the temporal injections included the lateral portions of cytoarchitectonic areas TE and TEO. There was a remarkable segregation of cells projecting to the 2 systems. Inputs to the parietal cortex tended to arise either from areas that have been implicated in spatial or motion analysis or from peripheral field representations in the prestriate cortex. By contrast, inputs to the temporal cortex tended to arise from areas that have been implicated in form and color analysis or from central field representations. Cells projecting to the parietal cortex were found in visual area 2 (V2), but only in the far peripheral representations of both the upper and lower visual field. Likewise, labeled cells found in visual areas 3 (V3) and 4 (V4) were densest in their peripheral representations. Heavy accumulations of labeled cells were found in the dorsal parieto-occipital cortex, including the parieto-occipital (PO) area, part A of V3 (V3A), and the dorsal prelunate area (DP). In the superior temporal sulcus, cells were found within several motion-sensitive areas, including the middle temporal area (MT), the medial superior temporal area (MST), the fundus of the superior temporal area (FST), and the superior temporal polysensory area (STP), as well as within anterior portions of the sulcus whose organization is as yet poorly defined. Cells projecting to areas TE and TEO in the temporal cortex were located within cytoarchitectonic area TG at the temporal pole and cytoarchitectonic areas TF and TH on the parahippocampal gyrus, as well as in noninjected portions of area TE buried within the superior temporal sulcus. In the prestriate cortex, labeled cells were found in V2, V3, and V4, but, in contrast to the loci labeled after parietal injections, those labeled after temporal injections were concentrated in the foveal or central field representations. Although few double-labeled cells were seen, 2 regions containing intermingled parietal- and temporal-projection cells were area V4 and the cortex at the bottom of the anterior superior temporal sulcus.(ABSTRACT TRUNCATED AT 400 WORDS)
GABAB-receptor-mediated inhibition of the N-methyl-D-aspartate component of synaptic transmission in the rat hippocampus
Authors: Morrisett RA , Mott DD , Lewis DV , Swartzwelder HS , Wilson WA
Published: March 2018

GABA receptor regulation of NMDA-receptor-mediated synaptic responses was studied in area CA1 of the rat hippocampus using extracellular and intracellular recording techniques. Picrotoxin (PTX) was used to suppress GABAA inhibition and 6,7-dinitroquinoxaline-2,3-dione (DNQX) was used to suppress non-NMDA receptor-mediated responses. In this manner, we were able to avoid the complicating factors caused by potentials induced by other excitatory and inhibitory amino acid receptors. Under these conditions, large NMDA-receptor-mediated EPSPs were observed. When paired stimuli were given at interstimulus intervals from 100 to 400 msec, powerful inhibition of the second response was observed. This inhibition was reversed by the GABAB antagonists phaclofen and 2-hydroxy-saclofen; it was also depressed by removal of Mg2+ from the bath. Examination of non-NMDA receptor- mediated synaptic responses (determined in the presence of D-2-amino-5- phosphonovalerate and PTX) showed no such inhibition, thereby supporting the hypothesis that GABAB inhibition of NMDA EPSPs is postsynaptic. This difference in paired-pulse inhibition of NMDA and non-NMDA EPSPs leads us to conclude that there was no evidence of GABAB- mediated presynaptic inhibition of excitatory transmitter release. Intracellular recordings in the presence of DNQX and PTX revealed a phaclofen-sensitive late IPSP that correlated in time with the period of inhibition of NMDA responses. Taken together, these data suggest that paired-pulse-inhibition of NMDA responses is produced by a GABAB- receptor-mediated hyperpolarization of the postsynaptic membrane, causing an enhanced block of the NMDA channels by Mg2+. Regulation of NMDA-mediated synaptic responses by GABAB receptors constitutes a powerful mechanism for control of a major excitatory system in hippocampal pyramidal cells.
Temporal interactions in the cat visual system. III. Pharmacological studies of cortical suppression suggest a presynaptic mechanism
Authors: Nelson SB
Published: March 2018

When tested with pairs of brief visual stimuli, neurons of the primary visual cortex of the cat show a long-lasting, orientation-selective suppression, termed “paired-pulse suppression.” The hypothesis that this suppression is due to GABAA-mediated inhibition was tested by performing temporal interaction tests before, during, and after iontophoretic application of the selective antagonist bicuculline methiodide (BMI). In keeping with previous reports, BMI elevated the spontaneous and evoked firing rates of cortical neurons, and altered basic receptive field properties. Under the influence of BMI, most neurons showed a reduced or abolished selectivity for stimulus orientation and direction of movement. The effects on orientation selectivity required higher ejection currents than did the effects on directional selectivity. At high ejection currents, most cells did lose selectivity for the orientation of a moving stimulus, but retained some selectivity for the orientation of a stationary stimulus. BMI, even at very high ejection currents, did not abolish paired-pulse suppression. In some cells, BMI enhanced or prolonged paired-pulse suppression. In further experiments, temporal interaction tests were performed in which one or the other of the component stimuli was replaced with a pharmacological stimulus (a pulse of glutamate or potassium). A pharmacological stimulus did not produce suppression of the response to a subsequent visual stimulus, nor did a visual stimulus suppress the response to a subsequent pharmacological stimulus. Paired-pulse suppression occurred only when both stimuli were visual. Taken together with previous results, the present data indicate that paired-pulse suppression is most likely due to a presynaptic mechanism.
NGF and excitatory neurotransmitters regulate survival and morphogenesis of cultured cerebellar Purkinje cells
Authors: Cohen-Cory S , Dreyfus CF , Black IB
Published: March 2018

The development of cerebellar Purkinje cells is subject to regulation by multiple epigenetic signals. To define mechanisms by which trophic and presynaptic stimulation may potentially regulate Purkinje cell ontogeny, we studied the effects of NGF and excitatory transmitters on Purkinje cell survival and morphological maturation in dissociated cell culture. Purkinje cells were identified by expression of vitamin D- dependent calcium-binding protein and by their characteristic morphology. NGF receptors were selectively localized to Purkinje cells by both ligand and monoclonal antibody binding, suggesting responsivity to the trophic agent. Simultaneous exposure to depolarizing agents and NGF specifically enhanced Purkinje cell survival in culture. NGF, in combination with either high potassium or veratridine markedly increased survival of Purkinje cells. Furthermore, NGF together with the excitatory neurotransmitters aspartate or glutamate promoted a 2- fold increase in survival. In addition, NGF increased Purkinje cell size and promoted neurite elaboration. These effects required simultaneous exposure to NGF and either aspartate, glutamate, or pharmacologic depolarizing agents. Effects on survival or neurite elaboration were not evoked by exposure to trophic factors or transmitters alone. Our results suggest a novel mechanism for regulation of development, in which trophic factor and afferent stimulation interact to promote survival and morphogenesis of developing Purkinje cells.
The early commitment of fetal neurons to the limbic cortex
Authors: Barbe MF , Levitt P
Published: March 2018

The limbic-system-associated membrane protein (LAMP) is expressed early in cerebral cortical development by migrating and postmigratory neurons in limbic regions such as the prefrontal and perirhinal cortices (Horton and Levitt, 1988), but not by nonlimbic neurons such as in the primary sensory and motor cortices. In the present study, we used LAMP expression to evaluate the timing and potential cell-lineage and environmental determinants of the commitment of cerebral cortical neurons to limbic and nonlimbic phenotypes. The cerebral wall, containing either presumptive perirhinal or sensorimotor cortex, was removed prior to [embryonic day (E) 12, 14] or just after (E17) the onset of LAMP expression, labeled with fast blue, and placed into cavities in either perirhinal or sensorimotor regions of postnatal day (P) 1 hosts. Host animals were allowed to survive until at least P10, and surviving transplanted cells were counted and evaluated for LAMP expression. Sensorimotor cortical neurons transplanted at E14 or E17, in either homotopic or heterotopic locations, did not express LAMP. In contrast, a high percentage of perirhinal cortical neurons transplanted at either E14 or E17, whether situated in sensorimotor or perirhinal regions of the host, expressed the limbic marker protein. Surprisingly, neurons from E12 donors exhibited patterns of LAMP expression that reflected their new location in the host rather than their embryonic origin. The data suggest that there is an early pliant period in which precursor and early differentiating cells remain uncommitted to a particular cortical regional phenotype. After this period, one aspect of the molecular phenotype (determined by LAMP expression) of neurons destined for sensorimotor and allo- and mesocortical regions has become immutable by environmental factors in the host.
Making arm movements within different parts of space: the premotor and motor cortical representation of a coordinate system for reaching to visual targets
Authors: Caminiti R , Johnson PB , Galli C , Ferraina S , Burnod Y
Published: March 2018

The activity of 156 individual arm-related neurons was studied in the premotor cortex (area 6) while monkeys made arm movements of similar directions within different parts of 3-dimensional space. This study was aimed at describing the relationship between premotor cortical cell activity and direction of arm movement and assessing the coordinate system underlying this relationship. We found that the activity of 152 (97.4%) cells varied in an orderly fashion with the direction of movement, in at least some region of the work space. Premotor cortical cells fired most for a given preferred direction and less for other directions of movement. These preferred directions covered the directional continuum in a uniform fashion across the work space. It was found that, as movements of similar directions were made within different parts of the work space, the cells' preferred directions changed their orientation. Although these changes had different magnitudes for different cells, at the population level, they followed closely the changes in orientation of the arm necessary to move the hand from one to another part of the work space. This shift of cells' preferred direction with the orientation of the arm in space has been observed with similar characteristics in the motor cortex (see Caminiti et al., 1990). In both premotor and motor cortices, neuronal movement population vectors provide a good description of movement direction. Unlike the individual cell preferred directions upon which they are based, movement population vectors did not change their spatial orientation across the work space, suggesting that they remain good predictors of movement direction regardless of the region of space in which movements are made. The firing frequency of both premotor and motor cortical neurons varied significantly with the position occupied by the hand in space. These static positional effects were observed in 88.5% of premotor and 91.8% of motor cortical cells. In a second task, monkeys made movements from differing origins to a common end point. This task was performed within 3 different parts of space and was aimed at dissociating movement direction from movement end point. It was found that in both premotor and motor cortices virtually all cells were related to the direction and not to the end point of movement. These data suggest that premotor and motor cortices use common mechanisms for coding arm movement direction. They also provide a basis for understanding the coordinate transformation required to move the hand toward visual targets in space.
Antiserum from a patient with cerebellar degeneration identifies a novel protein in Purkinje cells, cortical neurons, and neuroectodermal tumors
Authors: Darnell RB , Furneaux HM , Posner JB
Published: March 2018

The serum and cerebrospinal fluid of a patient (NB) with subacute cerebellar degeneration were found to contain a novel antineuronal autoantibody (anti-Nb). Using this antibody, we have identified and characterized antigens present in a subset of neurons in the CNS and in some neuroectodermal tumor lines. Anti-Nb antibody bound to antigens of Mr 150, 120, and 65 kDa in Western blots using extracts of human cerebellar Purkinje cells or human cerebral cortical neurons. Immunohistochemistry demonstrated relatively specific binding of anti- Nb IgG to Purkinje cells in sections of human cerebellum and to some neocortical neurons, especially those in layer VI. Because of the association of cerebellar degeneration with occult malignancies, we screened a number of tumor cell lines for immunoreactivity to anti-Nb antibody; only tumor lines of neuroectodermal origin (melanoma, small- cell lung cancer, and neuroblastoma) expressed the Nb antigen. Anti-Nb antibody thus identifies neuronal and tumor cell antigens that appear to be unique in size and distribution of expression.
Synaptic actions of identified peptidergic neuron R15 in Aplysia. II. Contraction of pleuroabdominal connectives mediated by motoneuron L7
Authors: Alevizos A , Weiss KR , Koester J
Published: March 2018

The purpose of this study was to determine the synaptic actions of the bursting peptidergic neuron R15 in Aplysia. R15 is known to be excited by the neuroendocrine bag cells, which trigger egg laying. In the two companion papers, we show that R15 mediates some of the effects of the bag cells on respiratory and reproductive organs. In this paper, we demonstrate that R15 excites L7, a multimodal motoneuron located in the abdominal ganglion. Although L7 excites several types of muscle fibers as well as neurons, the excitation of L7 by R15 is probably strong enough to cause contraction only of the sheath muscle of the pleuroabdominal connectives, which has an exceptionally low threshold for activation. The excitatory actions of R15 on L7, which desensitize profoundly, appear to be mediated by R15 alpha 1 peptide. The synaptic action of R15 on L7 and on the respiratory pumping system (Alevizos et al., 1991a) can be fully expressed only if R15 is first silenced for 2 hr by injection of hyperpolarizing current. A similar protocol for eliminating desensitization may prove to be generally useful for revealing the synaptic actions of other spontaneously active neurons that have rapidly desensitizing postsynaptic actions.
Nerve-dependent modulation of acetylcholine receptor epsilon-subunit gene expression
Authors: Martinou JC , Merlie JP
Published: March 2018

The relative abundance of mRNAs encoding the gamma- and epsilon- subunits of acetylcholine receptor changes in opposite directions during mouse development. Both gamma- and epsilon-mRNAs are expressed early in muscle development in vivo, and in aneural embryonic muscle and myogenic cell lines in vitro, though gamma-mRNA is at least 20-fold more abundant than epsilon-mRNA in these circumstances. While during normal development, gamma-mRNA decreases to an undetectable level by postnatal day 12, epsilon-mRNA first increases 10-fold between day 2 and day 12–15, then decreases to the level characteristic of adult muscle. We have found that the transition form gamma- to epsilon-mRNA is influenced by the levels of thyroid hormones. Indeed, high and low levels of thyroid hormones, respectively, accelerated and delayed the switch between gamma- and epsilon-mRNAs. Neither the dramatic postnatal rise in epsilon-mRNA nor its sensitivity to thyroid hormones was observed in denervated newborn animals. By contrast, denervation was without effect on epsilon-mRNA expression in adult muscle. These results suggest that, although not required for the initial activation of the epsilon-gene nor its maintenance in adult muscle, the nerve plays a major role in the perinatal regulation of epsilon-gene transcription.
Adenosine inhibits excitatory but not inhibitory synaptic transmission in the hippocampus
Authors: Yoon KW , Rothman SM
Published: March 2018

We examined the effects of adenosine and baclofen on inhibitory (IPSC) and excitatory (EPSC) synaptic currents in dissociated rat hippocampal neurons. Adenosine dramatically reduced monosynaptic EPSCs but failed to diminish IPSCs. This selective effect on EPSCs is likely due to inhibition of excitatory transmitter release because adenosine did not directly alter any properties of postsynaptic neurons. Baclofen depressed both EPSCs and IPSCs to approximately the same extent. These experiments indicate that the presynaptic effects of adenosine and baclofen are clearly separable and that transmitter sensitivities of inhibitory and excitatory neurons can differ. These differences could be exploited in the design of antiepileptic drugs that act at adenosine receptors to limit excitatory neurotransmission without blocking tonic inhibition.
Dependence of Ca2+ and K+ current development on RNA and protein synthesis in muscle-lineage cells of the ascidian Boltenia villosa
Authors: Simoncini L , Moody WJ
Published: March 2018

The early development of excitability of muscle-lineage cells of the ascidian Boltenia villosa is characterized by the appearance, just after gastrulation, of a Ca2+ current and a delayed outward K+ current, while an inwardly rectifying K+ current, present since fertilization, disappears. The muscle-lineage cells are the first cells in which we detect tissue-specific electrical properties after gastrulation. Here, we show that the development of electrical properties in these cells involves RNA and protein synthesis. If transcription or translation is blocked, the Ca2+ and outward K+ currents fail to appear, whereas the inward K+ current disappears normally. For the Ca2+ current, the sensitive period for transcription extends until just before gastrulation, while the sensitive period for translation extends until after gastrulation. The oocyte has a Ca2+ current present at about 5– 10% the density of that in the muscle-lineage cells; this current disappears by gastrulation. A comparison of the oocyte and muscle Ca2+ currents indicates that they are similar in voltage dependence and inactivation mechanism. A small difference in permeability sequence can be attributed to different surface charge properties at the two stages of development.
Morphogenesis and territorial coverage by isolated mammalian retinal ganglion cells
Authors: Montague PR , Friedlander MJ
Published: March 2018

Identified retinal ganglion cells were isolated from postnatal cat retinas and their dendrites were removed by trituration and centrifugation. The denuded cells were placed in a cell culture system and allowed to reexpress dendritic arbors in the absence of afferent input, target tissue, and interactions with neighboring ganglion cells. The retinal ganglion cells were grown above a feeder layer of astrocytes on glass coverslips equipped with paraffin pedestals. The spatial patterns of the reexpressed neurites were quantitatively analyzed using a number of measures, including an estimate of the Hausdorff dimension, H, which was used as a scale-independent metric for how well the neurite patterns filled in a restricted spatial domain. As assessed by the estimation of the Hausdorff dimensions, the neurites from a single cell achieve uniform coverage of a restricted territory independent of the total neurite length or the total number of inter-branch-point segments. A comparison with H values of ganglion cells from the intact retina revealed a similar trend. These results suggest that these cultured ganglion cells can express an intrinsic growth strategy for the uniform coverage of a restricted territory. The arbors expressed in the culture system displayed a limited range of diameters and exhibited morphology similar to the alpha-, beta-, and gamma-ganglion cells of the intact retina in the absence of afferent input or the influences of neighboring cells and target tissue. Time- lapse video data revealed that individual cultured cells showed extensive dendritic remodeling during their growth; however, after about 3 d in culture, this remodeling did not appreciably affect the territorial coverage of a cell. In the intact retina, the existence of dendritic sheets that independently and uniformly sample visual space may result from this intrinsic ability to elaborate dendrites that uniformly cover or fill in a restricted territory.
A monoclonal antibody that distinguishes between temporal and nasal retinal axons
Authors: McLoon SC
Published: March 2018

A monoclonal antibody was developed that recognizes an antigen with an asymmetric distribution in the chick retina. Immunohistochemistry showed that this antigen, temporal retinal axon protein (TRAP), was present on most if not all axons that arose from the temporal side of the retina. Very few of the axons from the nasal side of the retina were positive for TRAP. The nasal-temporal difference appeared to be in the number of axons that stained with this antibody rather than in the intensity of staining. The transition between nasal and temporal retina based on TRAP distribution appeared to be a vertical line centered on the optic fissure. A competition-based ELISA was developed to quantify the average amount of TRAP on axons in different regions of the retina. This assay also suggested that the pattern of TRAP distribution across the retina was a step function, though the results did not completely rule out the possibility of a continuous concentration gradient oriented circumferentially around the retina. Explants of embryonic nasal and temporal retina had a similar dichotomy in TRAP expression during the first 1 or 2 d in culture. The antibody to TRAP bound to retinal neurites in culture without the cell membrane being made permeable, which suggests that TRAP is a cell-surface molecule. In culture, TRAP was also expressed on the growth cones. Immunoblots showed that TRAP is trypsin sensitive and has an approximate molecular mass of 135 kDa. This is the first molecule identified with an asymmetric distribution in the nasal-temporal axis of the retina.(ABSTRACT TRUNCATED AT 250 WORDS)
Alien intracellular calcium chelators attenuate neurotransmitter release at the squid giant synapse
Authors: Adler EM , Augustine GJ , Duffy SN , Charlton MP
Published: March 2018

A number of calcium buffers were examined for their ability to reduce evoked transmitter release when injected into the presynaptic terminal of the squid giant synapse. Injection of EGTA was virtually ineffective at reducing transmitter release, even at estimated intracellular concentrations up to 80 mM. Conversely, the buffer 1,2-bis(2- aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA), which has an equilibrium affinity for calcium similar to that of EGTA at pH 7.2, produced a substantial reduction in transmitter release when injected presynaptically. This effect of BAPTA was reversible, presumably because the buffer diffused out of the terminal and into uninjected regions of the presynaptic axon. BAPTA derivatives with estimated intracellular calcium dissociation constants (Kd) ranging from 0.18 to 4.9 microM were effective at reducing transmitter release at similar estimated concentrations. A BAPTA derivative with an estimated intracellular Kd of 31 mM was less effective. BAPTA did not affect presynaptic action potentials or calcium spikes in ways that could explain its ability to reduce transmitter release. The relative effects of presynaptic injections of BAPTA and derivatives are consistent with the calcium-buffering capability of these compounds if the presynaptic calcium transient that triggers release is hundreds of microM or larger. The superior potency of BAPTA compared to EGTA apparently results from the faster calcium-binding kinetics of BAPTA and suggests that the calcium-binding molecule that triggers release binds calcium in considerably less than 200 microsec and is located very close to calcium channels.
Differential gene expression for glutamic acid decarboxylase and type II calcium-calmodulin-dependent protein kinase in basal ganglia, thalamus, and hypothalamus of the monkey
Authors: Benson DL , Isackson PJ , Hendry SH , Jones EG
Published: March 2018

In situ hybridization histochemistry, using cRNA probes, revealed a complementarity in the distributions of cells in the basal ganglia, basal nucleus of Meynert, thalamus, hypothalamus, and rostral part of the midbrain that showed gene expression for glutamic acid decarboxylase (GAD) or the alpha-subunit of type II calcium-calmodulin- dependent protein kinase (CAM II kinase-alpha). Cells in certain nuclei such as the thalamic reticular nucleus, globus pallidus, and pars reticulata of the substantia nigra show GAD gene expression only; others in nuclei such as the basal nucleus of Meynert, medial mamillary nuclei, and ventromedial hypothalamic nuclei show CAM II kinase-alpha gene expression only. A few nuclei, for example, the pars compacta of the substantia nigra and the greater part of the subthalamic nucleus, display gene expression for neither GAD nor CAM II kinase-alpha. In other nuclei, notably those of the dorsal thalamus, and possibly in the striatum, GAD- and CAM II kinase-expressing cells appear to form two separate populations that, in most thalamic nuclei, together account for the total cell population. In situ hybridization reveals large amounts of CAM II kinase-alpha mRNA in the neuropil of most nuclei containing CAM II kinase-alpha-positive cells, suggesting its association with dendritic polyribosomes. The message may thus be translated at those sites, close to the synapses with which the protein is associated. The in situ hybridization results, coupled with those from immunocytochemical staining for CAM II kinase-alpha protein, indicate that CAM II kinase-alpha is commonly found in certain non- GABAergic afferent fiber systems but is not necessarily present in the postsynaptic cells on which they terminate. It appears to be absent from most GABAergic fiber systems but can be present in the cells on which they terminate. This suggests that the kinase may be differentially engaged in pre- and postsynaptic functions at certain synapses.
Intracellular calcium levels do not change during contact-mediated collapse of chick DRG growth cone structure
Authors: Ivins JK , Raper JA , Pittman RN
Published: March 2018

When the growth cone of a chick dorsal root ganglion (DRG) neurite contacts the neurite of a chick retinal ganglion cell in vitro, the growth cone typically responds by withdrawing its lamellipodia and filopodia and collapsing. We have used the fluorescent calcium indicator dye fura-2 and digital imaging microscopy to measure calcium levels within DRG growth cones and to determine whether changes in calcium levels are responsible for the collapse of growth cone morphology when a DRG growth cone contacts a retinal ganglion cell neurite. Calcium levels within DRG growth cones were stable during neurite outgrowth. Calcium was typically distributed homogeneously throughout the growth cone, though occasionally gradients of free calcium were present. When calcium gradients were observed, calcium levels appeared higher in the active veil regions than in the central core region. Calcium levels in DRG growth cones appeared to remain stable during the period of contact-mediated growth cone collapse. Low concentrations of the calcium ionophore ionomycin increased calcium levels two- to threefold without having any observable morphological effects on DRG growth cones. Likewise, depolarization with 15 mM KCl caused a transient two- to threefold increase in calcium levels without having any observable morphological effect. These results suggest that changes in calcium levels are not responsible for contact-mediated collapse of growth cone structure. A growth cone collapsing activity has been solubilized from embryonic chick brain (Raper and Kapfhammer, 1990). Application of this material to cultures of DRG neurons caused growth cones to collapse but had no effect on calcium levels within the growth cones. The crude growth cone collapsing activity was not blocked by the presence of cobalt, nickel, lanthanum, nifedipine, or reduced- calcium medium, suggesting that transmembrane calcium fluxes were not required for growth cone collapse. These results suggest that the morphological changes associated with the collapse of growth cone structure can be independent of changes in growth cone calcium levels, and that second messengers other than calcium are likely to be involved in the regulation of many growth cone behaviors.
Dentate hilar cells with dendrites in the molecular layer have lower thresholds for synaptic activation by perforant path than granule cells
Authors: Scharfman HE
Published: March 2018

Neurons in the dentate hilus or area CA3c of rat hippocampal slices were recorded intracellularly with electrodes containing the fluorescent dye Lucifer yellow. Stimulation of perforant path fibers in the molecular layer of the fascia dentata strongly excited most hilar neurons, with a much lower threshold for action potential generation than granule cells and area CA3c pyramidal cells that were recorded in the same area of the slice. Examination of dye-filled hilar neurons with a confocal microscope showed that hilar cells with a low threshold were morphologically heterogeneous: some were spiny “mossy” cells, and others were aspiny interneurons. However, all hilar cells with low thresholds possessed dendrites that penetrated the granule cell layer and passed into the molecular layer, often reaching the outer molecular layer. The few hilar cells that had a threshold similar to, or greater than, granule cells did not possess visible dendrites in the molecular layer. The results suggest that the circuitry of the dentate region allows for (1) excitation of both granule cells and hilar cells by perforant path stimuli, and (2) strong excitation of most hilar cells when most granule cells are subthreshold for action potential generation. Given that hilar neurons project to many different sites in the ipsilateral and contralateral fascia dentata (Blackstad, 1956; Zimmer, 1971; Swanson et al., 1978; Laurberg and Sorensen, 1981), it is quite likely that hilar neurons are involved in the processing of information passing from entorhinal cortex to hippocampus.
Phospholipid-mediated delivery of anti-GAP-43 antibodies into neuroblastoma cells prevents neuritogenesis
Authors: Shea TB , Perrone-Bizzozero NI , Beermann ML , Benowitz LI
Published: March 2018

The neuronal growth-associated protein GAP-43 is expressed during axonal outgrowth and regeneration (for review, see Benowitz and Routtenberg, 1987). In the present study, we demonstrate that GAP-43 is constitutively expressed by NB2a/d1 neuroblastoma cells. The initial, most rapid outgrowth period of neuritogenesis [0–4 hr after dibutyryl adenosine 3′,5′-cyclic monophosphate (dbcAMP) treatment] is accompanied by intense GAP-43 immunoreactivity along the entire length of most neurites. However, this immunoreactivity declined nearly to background levels within hours during continued neurite outgrowth and persisted only at varicosities and growth cones. GAP-43 was detectable by metabolic labeling and immunoblot analysis in undifferentiated cells, and synthetic rates and steady-state levels of GAP-43 underwent only a modest (approximately twofold) increase during dbcAMP-induced differentiation. Unlike levels observed in neurites, perikarya of undifferentiated and differentiated cells contained similar, intense levels of GAP-43 immunoreactivity. Neurite elaboration and GAP-43 immunoreactivity were unaffected by treatment with cycloheximide, suggesting that translocation of perikaryal GAP-43 pools, rather than de novo synthesis, contributes to the transient burst of GAP-43 observed in developing neurites. Phosphatidylcholine-mediated delivery of anti-GAP-43 antibodies (alpha GAP) into cells immediately before dbcAMP treatment arrested neuritogenesis but did not induce the retraction of existing neurites. These results indicate that, while GAP- 43 expression is insufficient to induce neuritogenesis in NB2a/d1 cells, GAP-43 is nevertheless essential for the initial, dynamic phase of neurite outgrowth.