bluuebunny/crossref_metadata_2025 huggingface dataset:
Chronic elevation of secreted amyloid precursor protein in subcortically lesioned rats, and its exacerbation in aged rats
Authors: Wallace WC , Lieberburg I , Schenk D , Vigo-Pelfrey C , Davis KL , Haroutunian V
Published: April 2018

Subcortically lesioned rats were used as an animal model of some of the neurochemical and behavioral deficits of Alzheimer's disease (AD) to investigate the in vivo expression and metabolism of amyloid precursor protein (APP). Previously, the rapid and persistent induction of APP was described in cerebral cortices after disruption of its cholinergic, serotonergic, or noradrenergic afferents. In the present study, this induction was found to lead to the elevated secretion of APP into the cerebrospinal fluid of lesioned animals. Lesions of the forebrain cholinergic system in aged rats caused an even greater increase in the CSF levels of secreted APP. Antibodies to the extracellular domain of APP detected the protein whereas antibodies to the cytoplasmic region did not, indicating that the APP present in CSF was of the soluble form. Immunoprecipitation with an A beta sequence-specific antibody followed by immunoblot analysis indicated that a significant portion of secreted APP was of the species that contains at least the first 28 amino acids of the A beta sequence (APP gamma or APPA beta). By contrast, very low levels of A beta peptide were detected in CSF. The secretion was accompanied by an elevation of cellular C-terminal fragments of the APP in the lesioned cortex. Consistent with our previous results, this increased APP secretion was caused by lesions of subcortical cholinergic and serotonergic systems. The postlesion time course of APP secretion showed an initial reduction of APP (1 hr postlesion) in CSF followed by an eventual twofold elevation 1–6 weeks later. These results indicate that the induction of APP in response to loss of subcortical innervation leads to elevated secretion of a soluble form of cortically derived APP that contains significant portions of the A beta sequence.
Nerve growth factor (NGF) differentially regulates the chemosensitivity of adult rat cultured sensory neurons
Authors: Bevan S , Winter J
Published: April 2018

We have studied the effects of NGF on the chemosensitivity of adult rat DRG neurons over a 1–2 week period in vitro, using voltage-clamp and radioactive ion flux methods. A sustained proton evoked current was reversibly lost in NGF-free medium after 1 week. Proton-evoked efflux of radioactive 86Rb+ ions was also depressed in NGF deprived cultures, although depolarization with 40 mM potassium still evoked a large 86Rb+ efflux. A similar reversible loss of capsaicin sensitivity was noted. The response to GABA and a second, transient proton evoked current were also regulated by NGF, but over a longer time course. In contrast, the sensitivity to ATP was not influenced by the presence or absence of NGF. These data show that NGF regulates some, but not all, chemosensitivities of DRG neurons and that loss of sensitivity occurs at different rates for different agonists. The precise co-regulation of the response to capsaicin and the sustained response to protons provides further evidence that protons activate capsaicin-operated ion channels.
Three phases of TRH-induced facilitation of exocytosis by single lactotrophs
Authors: Fomina AF , Levitan ES
Published: April 2018

Membrane capacitance measurements were used to study neuropeptide modulation of exocytosis by perforated patch clamped rat lactotrophs. We report that depolarizing voltage-clamp pulses evoke exocytosis that is steeply dependent on Ca2+ influx through voltage-gated Ca2+ channels. Furthermore, we find that the neuropeptide TRH (thyrotropin- releasing hormone) acts in three phases to promote exocytosis. First, TRH transiently (within approximately 0.5 min) triggers depolarization- and extracellular Ca(2+)-independent exocytosis. Second, within 3 min of application, TRH facilitates depolarization-evoked exocytosis while inhibiting voltage-gated Ca2+ current. Finally, after 8 min, TRH further enhances depolarization-evoked exocytosis by increasing high- voltage-activated (HVA) Ca2+ channel current. Activation of protein kinase C (PKC) with a phorbol ester also stimulates depolarization- evoked exocytosis by increasing Ca2+ current. Therefore, PKC can only account for the last effect of TRH. Thus, a single neuromodulator may employ several temporally distinct mechanisms to stimulate peptide secretion.
A presynaptic mechanism accounts for the differential block of nicotinic synapses on sympathetic B and C neurons by d-tubocurarine
Authors: Shen WX , Horn JP
Published: April 2018

The effects of d-tubocurarine (dTC) on nicotinic synapses on sympathetic B and C neurons in the bullfrog were compared by recording trains of postganglionic compound action potentials (CAPs) at 0.5–20 Hz. Block by dTC was strongly use dependent in that the IC50 shifted almost 10-fold with increasing stimulus frequency. Maximum sensitivity to dTC occurred at 5 Hz in the B system and at 20 Hz in the C system. Recovery during posttrain periods was characterized by transient reduction of CAP amplitude in the B system and by transient enhancement of CAP amplitude in the C system. Thus, dTC distinguished between nicotinic synapses on the two cell types. The cell-specific effects of dTC could arise from differences in postsynaptic or presynaptic nicotinic receptors, or from differences in acetylcholine (ACh) release. We tested these possibilities using intracellular recording. Based on comparison with iontophoretic responses to ACh, changes in EPSP amplitude during repetitive stimulation in dTC could not be explained by altered postsynaptic sensitivity. The block of nicotinic receptors was further analyzed by recording synaptic currents. In B and C cells, 3 microM dTC competitively antagonized EPSC amplitude by 54% without any sign of open-channel block. In B cells, comparison of trains in normal Ringer and dTC revealed a constant fractional reduction in EPSC amplitude, thereby indicating that presynaptic nicotinic receptors do not influence release. In the C system, dTC had no effect on peptidergic EPSPs, thereby suggesting that presynaptic nicotinic receptors also do not influence peptide release. We conclude that the differential effects of dTC on nicotinic transmission are indirect consequences of differences in ACh release by preganglionic B and C neurons.
Neurotrophin-4/5 enhances survival of cultured spiral ganglion neurons and protects them from cisplatin neurotoxicity
Authors: Zheng JL , Stewart RR , Gao WQ
Published: April 2018

Destruction of spiral ganglion neurons (SGNs) induced by injury and toxins is one of the major causes for hearing loss. Here we report that neurotrophin-4/5 (NT-4/5), a member of the nerve growth factor family, promoted survival of postnatal rat SGNs up to threefold in dissociated cell cultures. The survival-promoting potency of NT-4/5 was equivalent to that of BDNF and stronger than that of NT-3. In contrast, NGF showed no detectable effects. Immunohistochemistry, with TrkB and TrkA antisera, revealed that these neurons produced TrkB protein, the functional receptor for NT-4/5 and BDNF, but not TrkA protein, the high- affinity receptor for NGF. The survival-promoting activity of NT-4/5 was completely inhibited by TrkB-IgG fusion protein. These results suggest that NT-4/5 is a specific survival factor for SGNs. In addition, NT-4/5 protected the SGNs from neurotoxic effects of the anti- cancer drug, cisplatin. Thus, NT-4/5 may have therapeutic value in preventing hearing impairment caused by damage to primary auditory afferent neurons.
Binaural tuning of auditory units in the forebrain archistriatal gaze fields of the barn owl: local organization but no space map
Authors: Cohen YE , Knudsen EI
Published: April 2018

We identified a region in the archistriatum of the barn owl forebrain that contains neurons sensitive to auditory stimuli. Nearly all of these neurons are tuned for binaural localization cues. The archistriatum is known to be the primary source of motor-related output from the avian forebrain and, in barn owls, contributes to the control of gaze, much like the frontal eye fields in monkeys. The auditory region is located in the medial portion of the archistriatum, at the level of the anterior commissure, and is within the region of the archistriatum from which head saccades can be elicited by electrical microstimulation (see preceding companion article, Knudsen et al., 1995). Free-field measurements revealed that auditory sites have large, spatial receptive fields. However, within these large receptive fields, responses are tuned sharply for sound source location. Dichotic measurements showed that auditory sites are tuned broadly for frequency and that the majority are tuned to particular values of interaural time differences and interaural level differences, the principal cues used by barn owls for sound localization. The tuning of sites to these binaural cues is essentially independent of sound level. The auditory properties of units in the medial archistriatum are similar to those of units in the optic tectum, a structure that also contributes to gaze control. Unlike the optic tectum, however, the auditory region of the archistriatum does not contain a single, continuous auditory map of space. Instead, it is organized into dorsoventral clusters of sites with similar binaural (spatial) tuning. The different representations of auditory space in closely related structures in the forebrain (archistriatum) and midbrain (optic tectum) probably reflect the fact that the forebrain contributes to a wide variety of sensorimotor tasks more complicated than gaze control.
The major delayed rectifier in both Drosophila neurons and muscle is encoded by Shab
Authors: Tsunoda S , Salkoff L
Published: April 2018

The delayed rectifier K+ current in Drosophila is similar to the classical delayed rectifier, originally described by Hodgkin and Huxley. Drosophila provides unique tools of mutant analysis to unambiguously determine the genetic identity of this native K+ current. We identified the Shab gene as the exclusive gene underlying delayed rectifier currents in both muscle and neurons. In muscles, a genetic mutation of Shab removes virtually all the whole cell delayed rectifier current (IK), while leaving unaltered the transient A-current encoded by the Shaker gene. In neurons, the Shab mutation also removes the bulk of IK, but leaves unaltered the transient A-current encoded by the Shal gene. Although most of the delayed rectifier current is the product of the Shab gene, the Shaw gene contributes a small “leak” current to most neurons and muscle cells. Thus, in contrast to the A-currents which are encoded by different genes in muscle and neuronal cell bodies (Shaker and Shal, respectively), the predominant IK in both muscle and neurons is encoded by the same gene (Shab). With the genetic identity of IK confirmed, all of the major K+ currents in embryonic Drosophila neurons and muscle are now known.
Lesions of the fornix but not the entorhinal or perirhinal cortex interfere with contextual fear conditioning
Authors: Phillips RG , LeDoux JE
Published: April 2018

The effects of entorhinal cortex lesions, combined entorhinal and perirhinal cortex lesions, and fornix lesions on the conditioning of fear responses (freezing) to contextual stimuli were examined using a conditioning procedure known to produce hippocampal-dependent contextual conditioning. Lesions of the entorhinal and or entorhinal plus perirhinal cortex did not disrupt contextual conditioning, but lesions of the fornix did. None of the lesions affected conditioning to an explicit conditioned stimulus. Given that the entorhinal cortex is the primary linkage between the neocortex and the hippocampus and that the fornix is the primary linkage with subcortical structures, subcortical inputs to and outputs of the hippocampus appear to be sufficient to mediate contextual fear conditioning. As a result, the presumption that neocortical information is required for contextual fear conditioning, and perhaps other hippocampal-dependent functions, should be reevaluated.
Intraventricular administration of antibodies to nerve growth factor retards kindling and blocks mossy fiber sprouting in adult rats
Authors: Van der Zee CE , Rashid K , Le K , Moore KA , Stanisz J , Diamond J , Racine RJ , Fahnestock M
Published: April 2018

Repeated subconvulsive electrical stimulation of certain areas of the forebrain leads to kindling, a progressive and permanent amplification of evoked epileptiform activity, which is a model for human temporal lobe epilepsy. Recent studies have shown that kindling induces synthesis of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) but not neurotrophin-3 (NT-3) in the hippocampus and cortex. Kindling also elicits mossy fiber sprouting and functional synaptogenesis in the supragranular layer, the hilus, and the CA3 region of the hippocampus. Intraventricular administration of antibodies to NGF has been shown to effectively block septohippocampal sprouting in the adult rat, and has been reported to retard amygdaloid kindling. In the present study, we have investigated the possible role of NGF in both kindling and kindling-associated sprouting. We have confirmed a kindling-induced sprouting of the mossy fibers into the stratum oriens of the CA3 region of the hippocampus, utilizing a new semiquantitative method of analysis based on Timm staining. Previous studies found no overt signs of hippocampal damage with this kindling paradigm, indicating that the increased Timm staining likely reflects a purely activity-induced sprouting. Intraventricular infusion of affinity-purified anti-NGF IgGs (which cross-react with NT-3 but not BDNF) resulted in both significant retardation of kindling and inhibition of the kindling-induced mossy fiber sprouting. The findings suggest a role for NGF in both these phenomena.
Presynaptic differentiation induced in cultured neurons by local application of basic fibroblast growth factor
Authors: Dai Z , Peng HB
Published: April 2018

Recent studies have suggested a role for molecules residing at the muscle surface in signaling presynaptic development at the neuromuscular junction (NMJ). Since heparan sulfate-proteoglycan is a major component of the extracellular matrix of skeletal muscle, factors that are bound to this proteoglycan, such as basic fibroblast growth factor (bFGF), are in a strategic position for neuronal signaling. To test this idea, we applied bFGF to cultured Xenopus spinal cord neurons and monitored the change in intracellular Ca2+ level with fura-2 ratio imaging. In one-third of the neurons, local application of bFGF elicited a 30–140% increase in intracellular Ca2+ level. Ca(2+)-free medium or suramin abolished this change. Latex beads coated with bFGF induced clustering of synaptic vesicles at the bead-neurite contacts as evidenced by anti-synaptotagmin antibody labeling and electron microscopy. This response was also blocked by Ca(2+)-free medium and by suramin. Uncoated beads or beads coated with PDGF were ineffective. This induction was also inhibited by a tyrosine kinase inhibitor, tyrphostin RG-50864, suggesting the role of receptor tyrosine kinase activation in this process. In addition, bFGF-coated beads also induced the localization of depolarization-dependent Ca2+ influx to the bead- neurite contacts. In contrast, depolarization caused a distributed Ca2+ elevation in untreated neurites. These results suggest that local presentation of bFGF can mimic the muscle target in signaling the development of both a cytoplasmic and a membranous specialization for excitation-secretion coupling observed at the NMJ.
Two factors secreted by the goldfish optic nerve induce retinal ganglion cells to regenerate axons in culture
Authors: Schwalb JM , Boulis NM , Gu MF , Winickoff J , Jackson PS , Irwin N , Benowitz LI
Published: April 2018

Unlike mammals, lower vertebrates can regenerate an injured optic nerve and other pathways of the CNS throughout life. We report here that in dissociated cell culture, goldfish retinal ganglion cells regenerate their axons in response to two factors derived from the sheath cells of the optic nerve. Axogenesis factor 1 (AF-1) is a small peptide (700–900 Da) that is inactivated by treatment with proteinase K but heat stable. A second factor, AF-2, is a polypeptide of ca 12 kDa. In the absence of these factors, dissociated retinal cells remained viable in serum-free, defined media for at least a week but showed little outgrowth, as visualized using the vital dye 5,6-carboxyfluorescein diacetate (5,6- CFDA). The addition of AF-1 induced up to 25% of cells in culture to extend processes > 75 microns in length by 6 d; AF-2 had a lesser but highly significant effect. To verify that neurite outgrowth was from retinal ganglion cells per se, we applied the lipophilic dye 4-Di-10- ASP to the optic tectum and allowed it to diffuse up the optic nerve for several days before culturing the retina. A far greater percentage of cells containing the dye showed axonal outgrowth than was observed from the overall cell population, indicating that ganglion cells are selective targets of the factors. The effects of AF-1 or AF-2 were not secondary to enhanced viability, since neither overall cell survival nor the number of retinal ganglion cells remaining in culture after 6 d was affected by the presence of the factors. The activity of AF-1 and AF-2 was not mimicked by several defined factors tested over a broad concentration range, for example, NGF, BDNF, NT-3, CNTF, taurine, retinoic acid, acidic or basic fibroblast growth factors. The concentration of AF-1 is considerably higher in CM than in optic nerve homogenates, suggesting that it is actively secreted; AF-2 has a similar concentration intra- and extracellularly. Insofar as AF-1 and AF-2 derive from cells of the optic nerve and act upon retinal ganglion cells, they are likely to be important in inducing optic nerve regeneration in vivo.
Alterations in cerebral cortical galanin concentrations following neurotransmitter-specific subcortical lesions in the rat
Authors: Gabriel SM , Knott PJ , Haroutunian V
Published: April 2018

Galanin is associated with multiple projection neurons, and its immunoreactivity in the cerebral cortex may be derived from diverse sources. We investigated the effects of subcortical lesions on cerebral cortical galanin concentrations. Lesions of the anterior noradrenergic bundle (ANB) comparably reduced cerebral cortical galanin and norepinephrine (NE) concentrations. The effects of the ANB lesions on galanin were immediate and became most pronounced 1 week later. Extensive unilateral lesions of the nucleus basalis of Meynert (NBM) decreased galanin concentrations, although not as markedly as after ANB lesions. The NBM lesions had no additional effect in the presence of an ANB lesion. Decreases in cerebral cortical galanin concentrations depended upon the extent and the duration of the NBM lesion and were not as pronounced as the decreases in markers of cholinergic activity. Acute treatments with physostigmine, which inhibit cerebral cortical AChE, had no effect on galanin concentrations. The depletion of galanin following an NBM lesion was most pronounced within hours of the insult, while the depletion of ChAT following the same lesions required several days to develop. Cortical concentrations of galanin and 5-HT increased 1 hr after dorsal raphe nucleus (DRN) lesions and then decreased 7 d later. Six weeks later, galanin concentrations recovered in the cerebral cortex despite the continued depletion of 5-HT. These studies suggest that a substantial portion of cerebral cortical galanin may derive from noradrenergic neurons and may be modulated by cortically- projecting ACh and 5-HT neurons.
Repeated confocal imaging of individual dendritic spines in the living hippocampal slice: evidence for changes in length and orientation associated with chemically induced LTP
Authors: Hosokawa T , Rusakov DA , Bliss TV , Fine A
Published: April 2018

Using confocal microscopy in conjunction with microdrop application of Dil, we have imaged and measured individual dendritic spines of living hippocampal CA1 pyramidal neurons in acute brain slices, before and approximately 3 hr after induction of long-term potentiation by chemical means. Statistical analysis of changes in the length of individual spines, and comparison with results of Monte Carlo simulations, suggests that two forms of structural change occur in chemically induced long-term potentiation: growth of a subpopulation of small spines, and angular displacement of spines. These changes could provide a structural basis for the expression of long-term potentiation.
CNS-derived neural progenitor cells for gene transfer of nerve growth factor to the adult rat brain: complete rescue of axotomized cholinergic neurons after transplantation into the septum
Authors: Martinez-Serrano A , Lundberg C , Horellou P , Fischer W , Bentlage C , Campbell K , McKay RD , Mallet J , Bjorklund A
Published: April 2018

A CNS-derived conditionally immortalized temperature-sensitive neural progenitor (CINP) cell line was used to generate NGF-secreting cells suitable for intracerebral transplantation. The cells were transduced by repeated retroviral infection, using a vector containing the mouse NGF cDNA under the control of the LTR promoter. Subcloning at the permissive temperature (33 degrees C) identified a highly NGF-secreting clone (NGF-CINP), which contained multiple copies of the transgene and released NGF at a rate of 2 ng/hr/10(5) cells in vitro, both at 33 and 37 degrees C, which was approximately 1 order of magnitude higher than what was possible to achieve in the heterogeneously infected cell cultures. After transplantation to the brain, the NGF-CINPs differentiated into cells with a predominant glia-like morphology and migrated for a distance of 1–1.5 mm from the implantation site into the surrounding host tissue, without any signs of overgrowth and tumor formation. Grafts of NGF-CINP cells implanted into the septum of adult rats with complete fimbria-fornix lesion blocked over 90% of the cholinergic cell loss in the medial septum and grafts placed in the intact striatum induced accumulation of low-affinity NGF receptor positive fibers around the implantation site. Expression of the NGF transgene in vivo was demonstrated by RT-PCR at 2 weeks after grafting. It is concluded that the immortalized neural progenitors have a number of advantageous properties that make them highly useful experimental tools for gene transfer to the adult CNS.
Toxin-insensitive Ca current in dorsal raphe neurons
Authors: Penington NJ , Fox AP
Published: April 2018

About 54% of the whole-cell Ca current recorded in dorsal raphe neurons cannot be categorized as N-, L-, or P-type Ca current. This current, ICa-Raphe, was not blocked by a combination of nimodipine, omega-CgTx- GVIA, and omega-AGA-IVA. Differences in toxin sensitivity and voltage dependence suggest that ICa-Raphe is distinct from Q- or R-type Ca currents. In raphe neurons activation of 5-HT1A receptors by 5-HT inhibits approximately 50% of the Ca current and slows activation. 5-HT inhibits both N-type Ca channels and ICa-Raphe channels by approximately 50% and slows the activation of both currents to a similar extent. Other similarities between ICa-Raphe and N-type Ca current were observed; they are both blocked to a similar extent by Ni2+, their activation properties, their current kinetics and channel availability as a function of holding potential are almost identical. Thus, ICa-Raphe represents a current that is not sensitive to known antagonists, but which is similar to N-type Ca current. Although it is possible that ICa-Raphe belongs to a heretofore undiscovered family of Ca channels it is also possible that it represents an omega-CgTx GVIA- insensitive isoform of the N-type Ca channel family.
Characterization of a rat gene, rMAL, encoding a protein with four hydrophobic domains in central and peripheral myelin
Authors: Schaeren-Wiemers N , Valenzuela DM , Frank M , Schwab ME
Published: April 2018

Wrapping and compaction of myelin sheaths around axons require specific membrane and membrane-associated proteins. Transmembrane proteins like proteolipid protein (PLP), the peripheral myelin protein 22 (PMP-22) and P0 as well as myelin basic protein (MBP) are crucial for this process. We have isolated a rat cDNA, initially denominated NS 3, that is mainly expressed in the myelinating cells of the nervous system, the oligodendrocytes and Schwann cells. The cDNA encodes a highly hydrophobic protein of 16.8 kDa with four putative transmembrane domains. The putative NS 3 protein lacks a N-terminal hydrophobic leader sequence and has no consensus sequence for N-linked glycosylation. In contrast to PLP and PMP-22, the first and third putative transmembrane domain of the NS 3 protein contain charged amino acids, a feature which resembles the structure of gap junction proteins. Sequence analysis showed that NS 3 is the rat homolog of a human gene called MAL that was cloned from, and is expressed in various T-cell lines. Therefore, we call this gene rMAL (rat MAL). In the nervous system, the expression of rMAL, mRNA begins after birth and is highest during myelination. In situ hybridization shows that rMAL mRNA is exclusively expressed in white and gray matter oligodendrocytes in the CNS and in myelinating Schwann cells in peripheral nerves. Immunohistochemistry using a peptide-specific antibody localized the rMAL protein in the myelinated areas of the CNS and PNS. Furthermore, we demonstrate by immunoblot analysis that rMAL is a component of myelin. Its structure and distribution suggest that the rMAL protein might play an important role in compact myelin. We propose that the name rMAL protein refers to rat Myelin And Lymphocyte protein.
Retrograde synaptic communication via gap junctions coupling auditory afferents to the Mauthner cell
Authors: Pereda AE , Bell TD , Faber DS
Published: April 2018

Large myelinated club endings of the goldfish eighth nerve arise in the sacculus and establish mixed electrotonic and chemical synapses with the distal part of the Mauthner (M-) cell's lateral dendrite. We show here, using paired pre- and postsynaptic recordings, that depolarizing currents generated postsynaptically (specifically, the mixed synaptic potential produced by activation of part of the afferent population) can in some cases excite the presynaptic fibers and cause them to backfire. Strikingly, while in some systems junctional properties prevent the antidromic spread of depolarizing currents, physiological properties of these afferents and the gap junctions promote backfiring: the amplitude of the coupling potential recorded from an afferent fiber is voltage dependent, increasing with depolarization and being reduced during hyperpolarization. Two mechanisms, with different kinetics, underlie this voltage dependence. One, a nonlinear membrane property of the afferent fiber itself, enhances the coupling potential as the afferent membrane depolarizes. The second mechanism, which is less sensitive to voltage and is symmetric about the resting potential, most likely represents voltage dependence of the junctional membrane. Additionally, we also show retrograde diffusion of low molecular weight substances, as the fluorescent dye Lucifer yellow and the tracer Neurobiotin were found in the terminals of afferent fibers after being injected postsynaptically into the M-cell. These results suggest that the gap junctions in these primary afferents are not only involved in fast anterograde synaptic transmission but also provide the substrate for a retrograde intercellular communication. The electrical coupling may modify the input-output relation between eighth nerve afferents and the lateral dendrite by synchronizing the population of already active fibers and by promoting the recruitment of new fibers via backfiring, such that weaker inputs produce relatively larger responses.
Ultrastructural immunolabeling shows prominent presynaptic vesicular localization of delta-opioid receptor within both enkephalin- and nonenkephalin-containing axon terminals in the superficial layers of the rat cervical spinal cord
Authors: Cheng PY , Svingos AL , Wang H , Clarke CL , Jenab S , Beczkowska IW , Inturrisi CE , Pickel VM
Published: April 2018

Opioid peptides, Met5- and Leu5-enkephalin, are known endogenous ligands for the delta-opioid receptor (DOR) associated with opioid analgesia at the spinal level. To determine the cellular sites for DOR- mediated actions, we examined the ultrastructural localization of DOR and Met5-enkephalin (ME) in the spinal cord by combining immunoperoxidase and immunogold-silver labeling for antibodies against DOR and ME, respectively. Antibodies for DOR localization were raised in guinea pig against peptide 34–47 (p34), an amino acid sequence within the extracellular N-terminus of the DOR recently cloned from mouse neuroblastoma glioma (NG-108) cells. Selective immunoperoxidase labeling for DOR was detected by light microscopy in NG-108 cells and in the lamina I and II of the dorsal horn of the spinal cord (C2-C4). Electron microscopy of these spinal laminae revealed that the majority of the punctate varicosities seen by light microscopy were axon terminals. delta-opioid receptor-like immunoreactivity (DOR-LI) in axon terminals was most prominently associated with large dense core vesicles, and sometimes seen along the membranes of small clear vesicles and segments of the plasmalemma. A semiquantitative analysis of dually labeled sections revealed that of the terminals showing DOR- LI, 23/102 (23%) also contained Met5-enkephalin-like immunoreactivity (ME-LI). Conversely, 23/35 (66%) of the terminals showing ME-LI also showed DOR-LI. In addition to the presynaptic localization, selective postsynaptic densities within dendrites were also occasionally (9%) immunolabeled for the opioid receptor. These results provide the first ultrastructural evidence that DOR may serve autoreceptor functions on ME terminals as well as presynaptic modulation of other transmitters in the dorsal horn of the rat spinal cord. Additionally, the vesicular localization of DOR-LI in axon terminals suggests the involvement of these organelles in the transport of the receptors to the plasma membrane.
Highly restricted origin of prefrontal cortical inputs to striosomes in the macaque monkey
Authors: Eblen F , Graybiel AM
Published: April 2018

The prefrontal cortex is made up of neocortical areas thought to mediate aspects of the temporal and spatial organization of behavior. One of the prime output targets of the prefrontal cortex is the striatum, which is thought to operate in series with the prefrontal cortex in some neural computations. We have analyzed this prefronto- striatal projection in cynomolgus monkeys by combining anterograde neuronal tract tracing methods with neurochemical markers for the striosome and matrix compartments of the striatum. Our results single out two parts of the frontal cortex as projecting densely to the striosome compartment of the striatum: the posterior orbitofrontal/anterior insular cortex and the mediofrontal prelimbic/anterior cingulate cortex. These areas jointly innervated striosomes in the anterior and ventromedial striatum, mainly in the caudate nucleus. Striosomes in the dorsolateral striatum were never labeled. Thus, the anatomical subsystem defined by striosome affiliation includes three cortical and striatal regions that, in humans, have been implicated in obsessive-compulsive disorder. Nearly all of the remaining parts of the prefrontal cortex studied projected preferentially to the matrix compartment. Most of these prefrontal inputs were also patchy, and many of the patches (matrisomes) were selectively paired with nearby striosomes. The highly fractionated organization of prefrontal inputs to striosomes and matrisomes could form a template for computational networks in the striatum that redistribute prefrontal corticostriatal inputs to serve in context- dependent behavioral planning.
Maintained L-type Ca2+ channel activity in excised patches of PTX- treated granule cells of the cerebellum
Authors: Lambert RC , Feltz A
Published: April 2018

Activity of high-threshold voltage activated neuronal Ca2+ channels, including dihydropyridine-sensitive (L-type) channels, rapidly disappears during cell dialysis in whole-cell recording conditions or after excision of a patch. To date, this phenomenom has been mainly related to phosphatase or protease activity. On the other hand, it has been suggested that Ca2+ channels may be regulated by G-proteins. Therefore, disruption of this regulatory pathway may also be involved directly or indirectly in the rundown process. Here, we show that treatment of cultured cerebellar granule cells with pertussis toxin (PTX) increases to 70% the probability for excising patches that display L-type Ca2+ channels activity in the inside-out recording configuration. Quantitative study indicates that, except a half decrease in the open probability, most features of the channel activity are retained after patch excision with minor modifications. The characteristics of the channel activity did not change with time during at least the first 9 min of the inside-out configuration. In addition, comparison of unitary currents recorded in the cell-attached, configuration on treated and nontreated cells demonstrates that the PTX treatment slows the activation kinetics of the current and increases the duration of channel openings evoked at -20 mV but not at 0 mV depolarizing potential. These data suggest that L-type Ca2+ channel activity are under a tonic regulation of a PTX-sensitive mechanism, which is implied in the run-down process.
Glucocorticoids, the hippocampus, and behavioral inhibition in the preweanling rat
Authors: Takahashi LK
Published: April 2018

Endogenous corticosteroids influence brain development and behavioral expression. In rat pups, a corticosteroid-dependent developmental response is behavioral inhibition, which occurs in situations involving threat. Behavioral inhibition consists of freezing and a reduction in ongoing behavior. It is presently unknown which brain region(s) that bind corticosterone (CORT) is involved in the development of freezing. The hippocampus (HC), however, is the principal target site of CORT that regulates the postnatal development of HC dentate granule cells. Therefore, this study examined whether the HC, and in particular, the dentate granule cells, plays a major role in the early appearance of behavioral inhibition. On postnatal day 9, rat pups received bilateral HC electrolytic lesions, or bilateral HC infusions of colchicine, a neurotoxin selective for dentate granule cells, or bilateral HC infusions of kainic acid, a neurotoxin selective for pyramidal cells in the CA3 field. Control rats received sham operations. After the operations, all rats were adrenalectomized (ADX) and injected daily with 3.0 mg/kg CORT, except on the day of the behavioral test. On day 14, all pups were tested for behavioral inhibition, which consisted of removing the pup from the nest box and placing it in a temperature- controlled enclosure subdivided into two compartments by a wire-mesh partition. The pup was placed in one compartment and an unfamiliar anesthetized adult male rat was placed in the adjacent compartment. Results indicated that preweanling rats with electrolytic lesions ranging from the dorsal to the ventral HC exhibited significant deficits in freezing. Importantly, similar deficits in freezing were present in pups treated with colchicine but not KA. Hence, administration of exogenous CORT is not effective in facilitating the occurrence of freezing in preweanling pups lacking dentate granule cells. To determine whether the dorsal HC dentate gyrus is an essential target site of CORT in facilitating freezing, 9-d-old rats were implanted bilaterally with 30 gauge cannula filled with either CORT or cholesterol. After the operation, all rats were ADX and tested for behavioral inhibition on day 14. During testing, ADX pups with CORT- filled cannulae showed significantly higher levels of freezing than ADX control pups. Taken together, results suggest that during the early postnatal period, the action of endogenous CORT in the HC influences the development of dentate granule cells that play an essential role in mediating the appearance of behavioral inhibition.
The cell cycle of the pseudostratified ventricular epithelium of the embryonic murine cerebral wall
Authors: Takahashi T , Nowakowski RS , Caviness VS
Published: April 2018

Neurons destined for the cerebral neocortex are formed in the pseudostratified ventricular epithelium (PVE) lining the ventricular cavity of the developing cerebral wall. The present study, based upon cumulative S-phase labeling with bromodeoxyuridine, is an analysis of cell cycle parameters of the PVE. It is undertaken in the dorsomedial cerebral wall of mouse embryos from the eleventh to the seventeenth gestational day (E11-E17, day of conception = E0) corresponding to the complete period of neuronogenesis. The growth fraction (fraction of cells in the population which is proliferating) is virtually 1.0 from E11 through E16. The length of the cell cycle increases from 8.1 to 18.4 hr, which corresponds to a sequence of 11 integer cell cycles over the course of neuronal cytogenesis in mice. The increase in the length of the cell cycle is due essentially to a fourfold increase in the length of G1 phase which is the only phase of the cell cycle which varies systematically. Thus, the G1 phase is most likely to be the phase of the cell cycle which is modulated by extrinsically and intrinsically acting mechanisms involved in the regulation of neuronal cytogenesis.
Reversal of a muscle response to GABA during C. elegans male development
Authors: Reiner DJ , Thomas JH
Published: April 2018

In the C. elegans hermaphrodite the expulsion step of defecation depends on the coordinated contraction of three enteric muscle groups: the anal depressor muscle, the intestinal muscles, and the sphincter muscle. These muscles are activated by excitatory GABA neurotransmission. Mutations in 13 genes that affect activation of these enteric muscles have previously been identified. We show that the larval male defecates by contracting the same set of enteric muscles, and that these contractions require 12 of these 13 genes. However, near the end of the last larval stage, the male anal region undergoes a developmental change, including dramatic hypertrophy of the anal sphincter muscle and the opening of a cloacal canal. We find that this modified sphincter must now relax to permit defecation. In contrast to the larval male, we find that in the adult male only 2 of the 13 genes required for enteric muscle contraction, unc-25 and unc-47, are important for sphincter muscle relaxation. unc-25 and unc-47 are required for the synthesis and utilization of GABA. We also find that two other genes, unc-46 and unc-49, previously implicated in the inhibitory action of GABA on body-wall muscle, are also required for normal adult male sphincter relaxation. In these mutants, failure to relax the sphincter muscle results in a constipated phenotype, and killing the sphincter muscle rescues this phenotype. We also find that a GABA agonist or GABA itself can suppress the adult male sphincter relaxation defect of unc-25 mutants.
Growth factor upregulation of a phosphoinositide-coupled metabotropic glutamate receptor in cortical astrocytes
Authors: Miller S , Romano C , Cotman CW
Published: April 2018

CNS function depends on a capacity for plasticity during development, following injury, and in response to changing environmental conditions. Functional alterations in signal transduction pathways and in neurotransmitter receptor expression are possible mechanisms for the expression of such plasticity. In the present report, we demonstrate that exposure of astrocytes to specific growth factors alters both the functional activity and the protein levels of a specific glutamate receptor. Exposure of astrocytes to basic fibroblast growth factor, epidermal growth factor, or transforming growth factor-alpha produced marked increases in the ability of metabotropic glutamate receptor (mGluR) agonists to stimulate phosphoinositide hydrolysis. Using Western immunoblotting, we demonstrate that an increase in the levels of one of the phosphoinositide-coupled mGluR subtypes, mGluR5, accompanies the increased ability of mGluR agonists to stimulate phosphoinositide hydrolysis. In contrast, another phosphoinositide- coupled subtype of this receptor family, mGluR1 alpha, was not present at detectable levels in these cultures. The enhanced stimulation of phosphoinositide hydrolysis showed little sensitivity to pertussis toxin, and appeared to be selective to mGluR agonists, as there was not a similar increase in the ability of norepinephrine or carbachol to stimulate phosphoinositide hydrolysis. These findings demonstrate that expression of mGluRs in astrocytes is plastic, and indicate a novel pathway through which specific growth factors may selectively modulate neurotransmitter action.
Inactivating and noninactivating Ca(2+)- and voltage-dependent K+ current in rat adrenal chromaffin cells
Authors: Solaro CR , Prakriya M , Ding JP , Lingle CJ
Published: April 2018

The properties of Ca(2+)- and voltage-dependent K+ currents and their role in defining membrane potential were studied in cultured rat chromaffin cells. Two variants of large-conductance, Ca2+ and voltage- dependent BK channels, one noninactivating and one inactivating, were largely segregated among patches. Whole-cell noninactivating and inactivating currents resulting from each of these channels were segregated among different chromaffin cells. Cell-to-cell variation in the rate and extent of whole-cell current decay was not explained by differences in cytosolic [Ca2+] regulation among cells; rather, variation was due to differences in the intrinsic properties of the underlying BK channels. About 75% of rat chromaffin cells and patches express inactivating BK current (termed BKi) while the remainder express noninactivating BK current (termed BKs). The activation time course of both currents is similar, as is the dependence of activation on [Ca2+] and membrane potential. However, deactivation of BKi channels is slower than that of BKs channels. The functional role of these BK channel variants was studied in current-clamp recordings. Although both BKi and BKs currents contribute to action potential repolarization, cells expressing BKi current are better able to fire repetitively in response to constant current injection. Blockade of BKi current by charybdotoxin abolishes this behavior, showing that afterhyperpolarizations mediated by BKi current are permissive for repetitive firing. Thus, important properties of chromaffin cell membrane excitability are determined by the type of BK current expressed.
Corticospinal control during reach, grasp, and precision lift in man
Authors: Lemon RN , Johansson RS , Westling G
Published: April 2018

Transcranial magnetic brain stimulation (TMS) was used to assess the influence of the corticospinal system on motor output in seven human subjects during a task in which they had to reach out, grasp, and lift an object. Stimuli, directed at the hand area of the motor cortex, were delivered at eight defined points during the task: during reach, at grip closure, during object manipulation, during the parallel isometric increase in grip and load forces, during the lifting movement, and while the object was held in air. The amplitudes of short-latency EMG responses evoked by TMS in six arm and hand muscles showed a striking modulation across the different phases of the task. This modulation may well reflect phasic changes in corticospinal excitability because: (1) it did not simply reflect phasic changes in muscular activity associated with task performance, (2) it could vary inversely with the amplitude of the background EMG, and (3) it was only obtained with weak TMS intensities, below threshold for evoking responses in hand muscles of the relaxed subject. Our results suggest that the cortical representations of extrinsic hand muscles, which act to orientate the hand and finger tips, were subjected to a strong excitatory drive throughout the reach. This drive was also observed for brachioradialis and anterior deltoid, which contribute to transport of the hand. In contrast, the intrinsic hand muscles appear to receive their strongest cortical input as the digits closed around the object, and just after the subject first touched the object at the onset of manipulation. The isometric parallel increase in load and grip forces necessary to lift the object, which is normally triggered by tactile contact, was delayed by TMS delivered late during the reach. TMS at this time may disrupt processing necessary to control this critical phase of the task.
Amyloid beta-peptide impairs ion-motive ATPase activities: evidence for a role in loss of neuronal Ca2+ homeostasis and cell death
Authors: Mark RJ , Hensley K , Butterfield DA , Mattson MP
Published: April 2018

The amyloid beta-peptide (A beta) that accumulates as insoluble plaques in the brain in Alzheimer's disease can be directly neurotoxic and can increase neuronal vulnerability to excitotoxic insults. The mechanism of A beta toxicity is unclear but is believed to involve generation of reactive oxygen species (ROS) and loss of calcium homeostasis. We now report that exposure of cultured rat hippocampal neurons to A beta 1–40 or A beta 25–35 causes a selective reduction in Na+/K(+)-ATPase activity which precedes loss of calcium homeostasis and cell degeneration. Na+/K(+)-ATPase activity was reduced within 30 min of exposure to A beta 25–35 and declined to less than 40% of basal level by 3 hr. A beta did not impair other Mg(2+)-dependent ATPase activities or Na+/Ca2+ exchange. Experiments with ouabain, a specific inhibitor of the Na+/K(+)-ATPase, demonstrated that impairment of this enzyme was sufficient to induce an elevation of [Ca2+]i and neuronal injury. Impairment of Na+/K(+)-ATPase activity appeared to be causally involved in the elevation of [Ca2+]i and neurotoxicity since suppression of Na+ influx significantly reduced A beta- and ouabain-induced [Ca2+]i elevation and neuronal death. Neuronal degeneration induced by ouabain appeared to be of an apoptotic form as indicated by nuclear condensation and DNA fragmentation. The antioxidant free radical scavengers vitamin E and propylgallate significantly attenuated A beta- induced impairment of Na+/K(+)-ATPase activity, elevation of [Ca2+]i and neurotoxicity, suggesting a role for ROS. Finally, exposure of synaptosomes from postmortem human hippocampus to A beta resulted in a significant and specific reduction in Na+/K(+)-ATPase and Ca(2+)-ATPase activities, without affecting other Mg(2+)-dependent ATPase activities or Na+/Ca2+ exchange. These data suggest that impairment of ion-motive ATPases may play a role in the pathogenesis of neuronal injury in Alzheimer's disease.
Brain space for learned song in birds develops independently of song learning
Authors: Brenowitz EA , Lent K , Kroodsma DE
Published: April 2018

In numerous species of birds, individuals or species that sing larger numbers of song types have larger song control nuclei in their brains. The direction of the cause and effect relationship between the complexity of song behavior and brain space is unknown, however. The hypothesis that birds that learn large song repertoires develop large song nuclei was therefore tested with a songbird, the marsh wren (Cistothorus palustris). Males were hand-reared and tutored in the laboratory with either small (n = 8 males heard 5 song types) or large (n = 8 males heard 45 song types) repertoires. When the birds were adults, the number of song types each male sang was first determined, and then the volume and certain cellular attributes of the song nuclei HVC and RA were measured. The two groups of wrens showed large behavioral differences in the size of their learned song repertoires, but did not differ in either the volumes of HVC and RA or in neuronal size, number, or density within these nuclei. These results suggest that the relationship between behavioral song complexity and brain space found in this and other species develops largely independently of early song learning experience and the later production of those songs.