bluuebunny/crossref_metadata_2025 huggingface dataset:
Axotomy-like changes in cat motoneuron electrical properties elicited by botulinum toxin depend on the complete elimination of neuromuscular transmission
Authors: Pinter MJ , Vanden Noven S , Muccio D , Wallace N
Published: April 2018

The electrical properties of cat medial gastrocnemius (MG) spinal motoneurons were studied 14–21 d following injection of type A botulinum toxin (BTX) into the MG muscle. Treated MG muscles were atrophic, displayed pronounced fibrillation activity, and were markedly but not completely paralyzed. MG motoneuron electrical properties from animals with the highest MG muscle-twitch forces (greater than 20 gm) appeared normal, while motoneuron properties from animals with the lowest MG muscle-twitch forces (less than 10 gm) exhibited axotomy-like changes, though these changes were less pronounced than after axotomy itself. No changes in the axonal conduction velocity were observed, however. Motoneuron connectivity with MG muscle fibers was determined following intracellular stimulation of MG motoneurons by averaging EMG signals from 3 or 4 pairs of recording electrodes inserted into the BTX- treated MG muscles. Normal electrical properties were observed among motoneurons in which detectable EMG activity linked to the intracellular stimulation pulse was observed. The level of this connectivity, however, indicated that a relatively small number of muscle fibers were activated by individual motoneuron action potentials. Axotomy-like changes of electrical properties were observed in MG motoneurons that could not be associated with detectable EMG activity in the BTX-treated MG muscle following repeated trials of intracellular stimulation. These results indicate that the existence of effective neuromuscular transmission at a small number of motor terminals is sufficient to prevent the appearance of axotomy-like changes in motoneuron electrical properties, and that the absence of such transmission at all motor terminals is associated with the appearance of axotomy-like changes. The results suggest that the effects of axotomy itself on motoneuron properties may be based upon the loss or elimination of a potent interaction between muscle and motoneurons normally mediated by neuromuscular transmission.
The origin of corticospinal projections from the premotor areas in the frontal lobe
Authors: Dum RP , Strick PL
Published: April 2018

We determined the origin of corticospinal neurons in the frontal lobe. These neurons were labeled by retrograde transport of tracers after injections into either the dorsolateral funiculus at the second cervical segment or the gray matter of the spinal cord throughout the cervical enlargement. Using retrograde transport of tracer from the arm area of the primary motor cortex, we defined the arm representation in each premotor area in another set of animals. We found that corticospinal projections to cervical segments of the spinal cord originate from the primary motor cortex and from the 6 premotor areas in the frontal lobe. These are the same premotor areas that project directly to the arm area of the primary motor cortex. The premotor areas are located in parts of cytoarchitectonic area 6 on the lateral surface and medial wall of the hemisphere, as well as in subfields of areas 23 and 24 in the cingulate sulcus. The total number of corticospinal neurons in the arm representations of the premotor areas equals or exceeds the total number in the arm representation of the primary motor cortex. The premotor areas collectively comprise more than 60% of the cortical area in the frontal lobe that projects to the spinal cord. Like the primary motor cortex, each of the premotor areas contains local regions that have a high density of corticospinal neurons. These observations indicate that a substantial component of the corticospinal system originates from the premotor areas in the frontal lobe. Each of the premotor areas has direct access to the spinal cord, and as a consequence, each has the potential to influence the generation and control of movement independently of the primary motor cortex. These findings raise serious questions about the utility of viewing the primary motor cortex as the “upper motoneuron” or “final common pathway” for the central control of movement.
Differential localization of four subspecies of protein kinase C in the rat striatum and substantia nigra
Authors: Yoshihara C , Saito N , Taniyama K , Tanaka C
Published: April 2018

The distribution of protein kinase C (PKC) subspecies and their colocalization with neurotransmitters were examined in the rat striatum and substantia nigra (SN), using immunocytochemistry. The alpha- and beta I-PKC immunoreactivies were seen predominantly in the perikarya of the neurons in the striatum and SN. In contrast, the beta II- and gamma- PKC immunoreactivities were abundant in both the perikarya and the neuropils in the striatum and only in the neuropils in the SN. From electron microscopic studies, the alpha- and beta I-PKC immunoreactivities were seen adjacent to the plasma membrane, while the beta II-PKC immunoreactivity was observed in the cytoplasm around the Golgi complex. The gamma-PKC immunoreaction was dense throughout the cytoplasm. The double-staining and lesion studies revealed that the alpha-PKC-immunopositive neurons in the striatum were intrinsic cholinergic neurons, and that most of the alpha-PKC-immunoreactive neurons in the SN were dopaminergic neurons. The beta I-PKC- immunoreactive neurons were intrinsic GABAergic neurons in the striatum. Moreover, most of the beta II- and gamma-PKC-immunoreactive neurons were medium-sized neurons projecting to the SN, and over 90% of GABAergic neurons in the caudate-putamen contained beta II-PKC. The beta II-PKC-immunoreactive neurons showed no gamma-PKC immunoreactivity, and the gamma-PKC-immunoreactive neurons were not beta II-PKC immunoreactive. These findings suggest that alpha-PKC is related to the function of the nigral dopaminergic and the striatal cholinergic neurons, and that the beta I-PKC is involved in the function of the striatal intrinsic GABAergic neurons. The beta II- and gamma-PKC may also modulate a specific neuronal function in the striatonigral system.
Serotonin selectively attenuates glutamate-evoked activation of noradrenergic locus coeruleus neurons
Authors: Aston-Jones G , Akaoka H , Charlety P , Chouvet G
Published: April 2018

The effect of 5-HT on activity of noradrenergic locus coeruleus (LC) neurons was studied using microiontophoretic and micropressure drug application in anesthetized rats. 5-HT had no consistent effect on LC spontaneous discharge, eliciting a modest decrease overall. However, 5- HT reliably attenuated responses of LC neurons to excitatory amino acids (EAAs), one of the major classes of transmitters in afferents to these neurons. This effect was specific for EAA responses because it occurred for glutamate and kainate but not for ACh. In contrast, iontophoretic norepinephrine (NE) selectively attenuated spontaneous activity but not responses evoked by either glutamate or ACh. The responsiveness of LC neurons to EAAs as quantified by a response- contrast measure (evoked excitation/basal activity) was markedly reduced by 5-HT, but was increased by NE. For ACh, such responsiveness of LC cells was not changed by 5-HT, but was increased by NE. The effects of 5-HT were prevented and reversed by iontophoretically applied antagonists of 5-HT receptors, methysergide and methiothepin. Thus, 5-HT appears to selectively interact with EAA responses of LC neurons, acting as a filter to attenuate LC activity linked to its major EAA inputs while allowing other channels afferent to the LC (e.g., those utilizing ACh) to be expressed.
A coordinate system for the synthesis of visual and kinesthetic information
Authors: Tillery SI , Flanders M , Soechting JF
Published: April 2018

The results of this study suggest that information derived from kinesthetic inputs alone is not normally used to generate an estimate of the location of the hand in extrapersonal space. This finding provides support for the interpretation of previous results suggesting that a representation of a visual target in extrapersonal space must be transformed into a kinesthetic reference frame before the parameters of a targeted arm movement can be computed (Soechting and Flanders, 1989b). We asked subjects to use a pointer to indicate the spatial location of their hand following an unseen passive displacement. We found that subjects had large errors in the locations that they chose and that there was a large degree of variability for repeated trials with the same hand location. The errors were a result neither of using the pointer to indicate a spatial locus nor of an inability to make use of kinesthetic information. Instead, the errors resulted from an inability of subjects to synthesize an estimate of the hand's spatial location from only kinesthetic cues. We also asked subjects to use the pointer to indicate the location of their hand following a passive displacement when they had visual information about the passive displacement. In this case, we found that subjects performed better than when they had only kinesthetic information, but not as well as when they had only visual information about target location. This finding suggests that kinesthetic information about target location affects the processing of visual information.
Development of striatal compartmentalization following pre- or postnatal dopamine depletion
Authors: Snyder-Keller AM
Published: April 2018

Nigrostriatal dopamine (DA) projections terminate in distinct patches during the late prenatal and early postnatal period in the rat. During the first postnatal week, patches of DA fibers overlap with clusters of striatal neurons that share several identified characteristics. The early segregation of striatal cell types into either these patches or the surrounding matrix becomes a permanent organizational feature of the striatum. In order to determine whether the heterogeneous distribution of DA influences the formation of cellular patches, the developmental organization of chemically identifiable cell types was examined in normal rats and in rats DA depleted as infants (0 or 3 d) or in utero (embryonic days 17–18). During the first postnatal week, corresponding patches of DA afferents and substance P (SP)- immunoreactive neurons existed in the striatum of normal animals, and AChE-positive zones overlapped these patches in the lateral striatum. Injection of 6-hydroxydopamine into the lateral ventricles of fetal or infant rats produced a dramatic loss of striatal DA terminals. Neither the patchy distribution of SP-immunoreactive neurons nor the distinctive pattern of AChE staining present during the first 2 postnatal weeks was disrupted. During the third postnatal week, cells immunoreactive for leu-enkephalin or calbindin-D28k were confined to the matrix compartment, and this compartmentalization was also not noticeably changed by pre- or postnatal DA depletion. In adult animals, overlapping patches of leu-enkephalin- and SP-immunoreactive fibers were observed, regardless of whether any DA terminals remained. Thus, the basic organization of the striatal patch and matrix compartments develops normally in the absence of DA innervation through much of the formative period. Although these observations do not completely dismiss the possibility that the first DA afferents to appear in the striatal primordia influence contracted striatal cells to develop the patch phenotype, they suggest that the patchy distribution of DA afferents may be secondary to the early clustering of striatal neurons forming the patch compartment.
Interaction of colocalized neuropeptides: functional significance in the circadian timing system
Authors: Albers HE , Liou SY , Stopa EG , Zoeller RT
Published: April 2018

The suprachiasmatic nucleus (SCN), which appears to act as a circadian clock, contains a subpopulation of local circuit neurons in which vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI), and gastrin releasing peptide (GRP) are colocalized. To determine whether VIP, PHI, and GRP interact within the SCN to produce a signal important for circadian control, the behavioral and cellular effects of coadministration of these neuropeptides were investigated. Coadministration of VIP, PHI, and GRP within the SCN mimicked the phase- delaying effects of light on circadian control following in vivo microinjection and activated SCN single units recorded in vitro. These behavioral and cellular effects of coadministration of VIP, PHI, and GRP were significantly greater than administration of VIP, PHI, or GRP alone or coadministration of any 2 of these peptides. These data illustrate a new mechanism whereby multiple, colocalized neuropeptides interact in a functionally significant manner, and indicate that the interaction of VIP, PHI, and GRP may be involved in the regulation of circadian rhythms by the SCN.
Distributions of spinothalamic, spinohypothalamic, and spinotelencephalic fibers revealed by anterograde transport of PHA-L in rats
Authors: Cliffer KD , Burstein R , Giesler GJ
Published: April 2018

Fibers projecting from several levels of the spinal cord to the diencephalon and telencephalon were labeled anterogradely with Phaseolus vulgaris leucoagglutinin injected iontophoretically. Labeled fibers in the thalamus confirmed projections previously observed. In addition, many labeled fibers were seen in the hypothalamus and in telencephalic areas not generally recognized previously as receiving such projections. In the hypothalamus, these areas included the lateral hypothalamus (including the medial forebrain bundle), the posterior hypothalamic area, the dorsal hypothalamic area, the dorsomedial nucleus, the paraventricular nucleus, the periventricular area, the suprachiasmatic nucleus, and the lateral and medial preoptic areas. In the telencephalon, areas with labeled fibers included the ventral pallidum, the globus pallidus, the substantia innominata, the basal nucleus of Meynert, the amygdala (central nucleus), the horizontal and vertical limbs of the diagonal band of Broca, the medial and lateral septal nuclei, the bed nucleus of the stria terminalis, the nucleus accumbens, infralimbic cortex, and medial orbital cortex. These results suggest that somatosensory, possibly including visceral sensory, information is carried directly from the spinal cord to areas in the brain involved in autonomic regulation, motivation, emotion, attention, arousal, learning, memory, and sensory-motor integration. Many of these areas are associated with the limbic system.
The involvement of GABAB receptors and coupled G-proteins in spinal GABAergic presynaptic inhibition
Authors: Alford S , Grillner S
Published: April 2018

GABA acts as a presynaptic inhibitory transmitter in the spinal cord. In the lamprey, it has recently been shown that it acts in this way at both primary sensory and motor system synapses and is important in the generation of a locomotor rhythm. Both GABAA and GABAB receptors are activated at these sites by GABA released during physiological activity. In some systems, GABAB receptor activation has been shown to lead to modulation of ion channel function indirectly through the action of a pertussis toxin (PTX)-sensitive G-protein. Here we have studied the mechanism of action of the presynaptic GABAB receptor in this system. GABAB receptor activation leads to a decrease in axonal membrane impedance and also to a reduction in the axonal action potential duration. The ionic basis for this response remains unknown, though it is not, unlike the response to GABAA receptor activation, mediated by an increase in conductance to Cl-. The effects of GABAB receptor antagonism with phaclofen are mimicked by pretreatment of the spinal cord with PTX. Because this procedure inactivates certain classes of G-proteins, it seemed likely that the GABAB receptor- mediated effects are initiated via a presynaptic population of PTX- sensitive G-proteins. Experiments in which only presynaptic G-proteins were interfered with indicate that this is so. Stable analogs of GTP and GDP were used to activate permanently or to antagonize, respectively, the GTP binding site in the presynaptic component of these spinal synapses. We conclude that GABAB receptor-mediated synaptic suppression in the spinal cord is caused by GTP binding to presynaptic G-proteins linked to the GABAB receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
A computational model for the overall pattern of ocular dominance
Authors: Jones DG , Van Sluyters RC , Murphy KM
Published: April 2018

In layer IV of the primary visual cortex, in both the macaque monkey and the cat, geniculocortical terminals representing the two eyes are segregated into alternating zones known as ocular dominance bands. Viewed tangentially, in the monkey these bands take the form of a series of branching parallel stripes that run roughly perpendicular to the border of striate cortex. In the cat, the overall ocular dominance pattern consists of irregularly branching, beaded bands that exhibit no predominant orientation. If the striking differences in the appearance of these two patterns reflect important differences in the basic rules governing cortical ocular dominance, then this poses a problem for attempts to formulate general principles of visual cortical organization. However, it has been suggested that the differences in the appearance of the ocular dominance patterns in these two species could result simply from known differences in the boundary conditions of their geniculocortical pathways. This article describes the formulation and testing of a single computational model that accurately predicts the quite dissimilar ocular dominance patterns in cats and monkeys. This model also generalizes to predict the different ocular dominance patterns observed in young and old three-eyed frogs, supporting the notion that the overall pattern of ocular dominance is governed by a common set of rules. The significance of these results is discussed in terms of previous models, which have focused largely on local processes underlying the development of ocular dominance segregation. Although the present model is not a developmental one, it does shed some light on potential mechanisms for establishing retinotopy in striate cortex and on possible developmental relationships between the geniculostriate pathway and intrinsic modularity of the striate cortex.
Opiate withdrawal-induced hyperactivity of locus coeruleus neurons is substantially mediated by augmented excitatory amino acid input
Authors: Akaoka H , Aston-Jones G
Published: April 2018

Single-cell activity was recorded in the locus coeruleus (LC) of morphine-dependent, halothane-anesthetized rats. Systemic administration of the opiate antagonist naloxone (0.1 mg/kg, i.v.) robustly increased the activity of LC neurons. Local microinjection of naloxone or of its hydrophilic derivative, naloxone methiodide, into LC (10 mM, 20-40 nl) did not activate LC neurons in dependent rats. Intracerebroventricular or intracoerulear injection of kynurenate, a broad-spectrum antagonist of excitatory amino acids (EAAs), substantially but incompletely attenuated the activation of LC cells induced by intravenous naloxone-precipitated withdrawal (more than 50% blockade). Intracoerulear microinjections of the non-NMDA-receptor antagonist 6-cyano-7-dinitroquinoxaline-2,3-dione (CNQX) or the selective NMDA-receptor antagonist AP5 significantly reduced the withdrawal-induced excitation. AP5 was the least effective among all antagonists tested. Similar microinjections of kynurenate or CNQX almost completely suppressed the excitation of LC neurons induced by electrical stimulation of a rear footpad. LC responses to footpad stimulation (mediated by endogenous EAAs) or iontophoretically applied glutamate were not modified by the chronic morphine treatment. These results indicate that a substantial part of LC hyperactivity during opiate withdrawal is mediated by an augmented EAA input to LC.
The Alzheimer amyloid precursor is associated with the detergent- insoluble cytoskeleton
Authors: Refolo LM , Wittenberg IS , Friedrich VL , Robakis NK
Published: April 2018

The amyloid beta-protein (A beta P), the main component of neuritic plaques in Alzheimer's disease (AD), is derived by unknown mechanisms from a family of amyloid precursor proteins (APPs). Using a detergent extraction procedure, we have found that in brain and in neural cell lines, 50-90% of APP is bound to detergent-insoluble cytoskeleton. Labeling experiments performed in a C6 glioma cell line indicated that both cell surface and intracellular APPs are associated with the cytoskeleton. This association requires intact microtubules and is modulated by protein phosphorylation and by cell density. These findings suggest that the function of cellular APP, presently unknown, involves the cytoskeleton and particularly microtubules. The dynamic nature of the binding and its dependence on microtubules and protein phosphorylation suggest it as a possible target in AD, where abnormal cytoskeletal structures and protein phosphorylation have been reported. Altered cytoskeletal binding of APP might lead to its aberrant proteolysis and generation of the A beta P.
Testosterone differentially regulates the regenerative properties of injured hamster facial motoneurons
Authors: Kujawa KA , Emeric E , Jones KJ
Published: April 2018

We have previously demonstrated that systemic administration of testosterone accelerates functional recovery, as measured behaviorally, from facial paralysis induced by facial nerve crush axotomy in gonadectomized adult male hamsters. In this investigation, the hypothesis that testosterone enhances return of motor function by increasing the rate of axonal regeneration following injury was tested using fast axonal transport of radioactively labeled proteins to assess facial nerve regeneration. Adult castrated and intact males, and intact females, were subjected to right facial nerve crush axotomy at the stylomastoid foramen. One-half of the axotomized animals in each group received subcutaneous implants of testosterone, with the remainder of the animals sham implanted. Systemic administration of testosterone resulted in a 26-30% increase in the rate of regeneration of the fastest growing population of axons in the male experimental groups, regardless of whether the animal was castrated or not. This rate increase is similar to that observed in the conditioning lesion paradigm utilized by others. In the females, testosterone had a significant, but less pronounced, effect on the rate of axonal regeneration, which may be due in part to inherent gender-related differences in regenerative properties of facial motoneurons. A surprising finding of this study was that no shortening of the delay of sprout formation by testosterone was observed across the various experimental groups. These data suggest that the mechanism by which gonadal steroids act in the injured nervous system is partly through the differential regulation of the regenerative properties of the injured cell, presumably via hormone receptor-mediated action at the level of the neuronal genome.
Spontaneous action potential activity and synaptic currents in the embryonic turtle cerebral cortex
Authors: Blanton MG , Kriegstein AR
Published: April 2018

We used loose-patch and whole-cell recording techniques to study the development of spontaneous action potential activity and spontaneous excitatory and inhibitory synaptic currents in embryonic neurons in the cerebral hemispheres of turtles. Sporadic action potential activity appeared early in development at stage 17, soon after morphologically identifiable pyramidal and nonpyramidal neurons were first observed in the cortex. As the cortical plate matured in midembryonic stages, action potential activity became more regular and fell into one of two distinct patterns, tonic and intermittent high-frequency firing. Spontaneous excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs) appeared at developmental stages 18 and 20, respectively, after action potential activity was established. EPSCs and IPSCs exhibited characteristic ionic dependence and pharmacology throughout development. EPSCs reversed in direction at the equilibrium potential for cations and were sensitive to 6-cyano-7-nitroquinoxaline-2,3-dione, an antagonist of the non-NMDA type of glutamate receptor. IPSCs reversed at the equilibrium potential for chloride and were sensitive to bicuculline methiodide, a GABAA receptor antagonist. Spontaneous synaptic currents differed in their time course of development and in waveform parameters. Spontaneous synaptic currents differed in their time course of development and in waveform parameters. Spontaneous EPSCs appeared at stage 18 and increased progressively in frequency, from 0.2 +/- 0.1 Hz at stage 20 to 3.2 +/- 2.0 Hz at stage 26 (hatching), while spontaneous IPSCs appeared at stage 20 and surpassed EPSCs in frequency, increasing to 7.1 +/- 1.6 Hz at stage 26. EPSCs exhibited stable amplitudes during development, with a mean conductance of 126 +/- 20 pS at stage 26, while IPSCs increased in mean amplitude, from 180 +/- 12 pS at stage 18 to 260 +/- 44 pS at stage 26. The rise time to peak conductance of both types of synaptic currents increased with developmental time, for EPSCs increasing from 1.5 +/- 0.5 msec at stage 20 to 2.7 +/- 0.6 msec at stage 26 and for IPSCs increasing from 2.9 +/- 0.2 msec at stage 18 to 6.2 +/- 0.8 msec at stage 26. While the decay time constants increased for EPSCs, from 3.9 +/- 1.2 msec at stage 20 to 8.7 +/- 2.3 msec at stage 26, decay time constants for IPSCs showed a decreasing trend from 24.0 +/- 5.2 msec at stage 18 to 18.4 +/- 5.3 msec at stage 26. The excitatory and inhibitory synaptic currents were sensitive to the sodium channel blocker TTX and were thus dependent, in part, on spontaneous action potential activity.(ABSTRACT TRUNCATED AT 400 WORDS)
Sodium-bicarbonate cotransport in retinal Muller (glial) cells of the salamander
Authors: Newman EA
Published: April 2018

An electrogenic Na+/HCO3- cotransport system was studied in freshly dissociated Muller cells of the salamander retina. Cotransporter currents were recorded from isolated cells using the whole-cell, voltage-clamp technique following the block of K+ conductance with external Ba2+ and internal Cs+. At constant pHo, an outward current was evoked when extracellular HCO3- concentration was raised by pressure ejecting a HCO3(-)-buffered solution onto the surface of cells bathed in nominally HCO3(-)-free solution. The HCO3(-)-evoked outward current was reduced to 4.4% of control by 0.5 mM DIDS (4,4′- diisothiocyanatostilbene-2,2′-disulfonate), to 28.8% of control by 2 mM DNDS (4,4′-dinitrostilbene-2,2′-disulfonate), and to 28.4% of control by 2 mM harmaline. Substitution of choline for Na+ in bath and ejection solutions reduced the response to 1.3% of control. Bicarbonate-evoked currents of normal magnitude were recorded when methane sulfonate was substituted for Cl- in bath, ejection, and intracellular solutions. Similarly, an outward current was evoked when extracellular Na+ concentration was raised in the presence of HCO3-. The Na(+)-evoked response was reduced to 16.2% of control by 2 mM DNDS and was abolished by removal of HCO3- from bath and ejection solutions. Taken together, these results (block by stilbenes and harmaline, HCO3- and Na+ dependence, Cl- independence) indicate that salamander Muller cells possess an electrogenic Na+/HCO3- cotransport system. Na+/HCO3- cotransporter sites were localized primarily at the endfoot region of Muller cells. Ejection of HCO3- onto the endfoot evoked outward currents 10 times larger than currents evoked by ejections onto the opposite (distal) end of the cell. The reversal potential of the cotransporter was determined by DNDS block of cotransport current. In the absence of a transmembrane HCO3- gradient, the reversal potential varied systematically as a function of the transmembrane Na+ gradient. The reversal potential was -0.1 mV for a [Na+]o:[Na+]i ratio of 1:1 and -25.2 mV for a Na+ gradient ratio of 7.4:1. Based on these values, the estimated stoichiometry of the cotransporter was 2.80 +/- 0.13:1 (HCO3- :Na+). Possible functions of the glial cell Na+/HCO3- cotransporter, including the regulation of CO2 in the retina and the regulation of cerebral blood flow, are discussed.
Regulation of bradykinin- and ATP-activated Ca(2+)-permeable channels in rat pheochromocytoma (PC12) cells
Authors: Neuhaus R , Reber BF , Reuter H
Published: April 2018

Membrane currents activated by bradykinin (500 nM) and by extracellular ATP (50 microM) were studied in voltage-clamped, NGF-treated rat pheochromocytoma (PC12) cells. Under quasiphysiological ionic conditions, both substances caused an outward current due to opening of Ca(2+)-activated K+ channels. Bradykinin caused an additional inward current that could be studied after blockade by internal Cs+ of the initial transient outward current. The inward current became larger when the extracellular Ca2+ concentration was increased. Neither inositol-1,4,5-trisphosphate, dioctanoylglycerol, phorbol 12-myristat 13-acetate, forskolin, GTP, GTP-gamma-S, or pretreatment with pertussis toxin affected this current component. Increasing the internal Ca buffer concentration [EGTA or bis-(o-aminophenoxy)-ethane-N,N,N′,N′- tetra-acetic acid] from 1 to 10 mM had no effect on the inward current as long as the free [Ca2+]i was kept constant. However, it was modulated by the resting free [Ca2+]i. Elevation of [Ca2+]i from nominally 0 to 60 or to 180 nM increased the bradykinin-induced average peak current density from 0.14 to 1.04 or to 2.29 pA/pF, respectively. This regulation may depend on a calmodulin-dependent pathway, since CGS 9343B, a calmodulin inhibitor, blocked the effect of elevated [Ca2+]i. With ATP as an agonist, outward current was preceded by a large inward current that was partially blocked by extracellular Ca2+ in the millimolar range. Extracellular Ca2+ was also found to reduce the single-channel conductance estimated from outside-out patches treated with ATP.
The properties of intracellular calcium stores in cultured rat cerebellar neurons
Authors: Brorson JR , Bleakman D , Gibbons SJ , Miller RJ
Published: April 2018

Cerebellar Purkinje neurons contain a remarkable array of cellular components potentially concerned with regulation of the free cytoplasmic Ca2+ concentration, [Ca2+]i. These include high concentrations of Ca(2+)-binding proteins, inositol 1,4,5-triphosphate receptors (IP3R), and ryanodine receptors (RyR). The latter two molecules are thought to be associated with intracellular Ca2+ stores. We have examined the properties of such stores in cultured rat cerebellar neurons taken from 16 d rat embryos. In this system, about half of the neurons could be identified as Purkinje-like cells, as indicated by staining for the Ca(2+)-binding protein calbindin D-28k, as well as for IP3R and RyR. In double immunofluorescent staining, the IP3R and RyR immunoreactivity primarily colocalized with the staining for calbindin. The cells responded to glutamate, kainate, and quisqualate with large increases in the somatic [Ca2+]i but failed to respond directly to NMDA (10-50 microM). Furthermore, the neurons expressed active membrane conductances, repetitive action potential firing, and spontaneous firing patterns similar to those reported for cerebellar Purkinje neurons in vivo. Action potential firing produced changes in somatic [Ca2+]i that were quite small or absent in most cells. However, blocking spike repolarization with tetraethylammonium (5 mM) produced substantial transient elevations in somatic [Ca2+]i, suggesting the expression of some Ca2+ channels in the somatic membrane. Caffeine (10 mM) released Ca2+ from intracellular stores in about one-half of the cultured neurons. This effect could be repeated if the stores were first reloaded by a depolarization-induced elevation in [Ca2+]i. The effects of caffeine were reduced by prolonged application of ryanodine (10 microM). We were also able to demonstrate that the caffeine-sensitive Ca2+ stores could regulate electrophysiological events in some cells, altering patterns of spontaneous activity. Furthermore, in the presence of caffeine, [Ca2+]i signals induced by an evoked spike train were larger and accompanied by long-lasting after hyperpolarizations. We conclude that in addition to providing a releasable pool of Ca2+, the caffeine-sensitive stores also influence cellular events by their contribution to Ca2+ buffering.
A neural-specific GAP-43 core promoter located between unusual DNA elements that interact to regulate its activity
Authors: Nedivi E , Basi GS , Akey IV , Skene JH
Published: April 2018

In an effort to identify cis-acting elements that respond to signals controlling different stages of neural differentiation, we have analyzed the promoter and surrounding regulatory sequences of the rat GAP-43 gene. Expression of this gene is both neural specific and, within neurons, strongly modulated by signals related to axon integrity. Expression analysis in cell lines and primary rat cortical cultures demonstrates that neural-selective gene expression can be directed by a 386 base pair GAP-43 promoter fragment that contains canonical TATA and CCAAT box consensus sequences. A short region of homology with other neural-specific genes, identified upstream of the core promoter, is not essential for selective expression in neuronal cells. Within cortical cell cultures, expression is strongly modulated by two interacting elements on either side of the promoter, each of which contains a sequence with the potential to adopt an unusual DNA conformation. While each of these flanking elements reduces expression when added alone to the core promoter, each counteracts the negative influence of the other when both elements are present.
Acetylcholine synthesis and release is enhanced by dibutyryl cyclic AMP in a neuronal cell line derived from mouse septum
Authors: Blusztajn JK , Venturini A , Jackson DA , Lee HJ , Wainer BH
Published: April 2018

Cholinergic properties of the SN56.B5.G4 cell line derived from the fusion of neurons of the mouse postnatal day 21 septum and the murine neuroblastoma cell line N18TG2 were investigated and correlated with morphological differentiation. In basal serum-containing growth medium, few cells developed neurites. Neurite extension occurred in cells grown for 2 d with forskolin or dibutyryl cAMP (dbcAMP) but not with butyrate. In cells treated with these compounds, the activity of ChAT and ACh content were two- to threefold higher relative to controls. The cells synthesized ACh from choline taken up by the sodium-dependent high-affinity transport. Forskolin-, dbcAMP-, and butyrate-treated cells (but not the controls) were capable of spontaneous and depolarization-evoked ACh release. The results indicate that the morphological and the neurochemical aspects of SN56.B5.G4 cell differentiation are independently regulated.
Proctolin activates an inward current whose voltage dependence is modified by extracellular Ca2+
Authors: Golowasch J , Marder E
Published: April 2018

The pentapeptide proctolin modulates the activity of the rhythmic pattern generators in the crustacean stomatogastric nervous system. Proctolin strongly excites the lateral pyloric and the inferior cardiac neurons of the stomatogastric ganglion (STG), causing them to fire extended high-frequency bursts of action potentials (Hooper and Marder, 1987; Nusbaum and Marder, 1989a,b). We now report that proctolin depolarizes these cells maximally at membrane potentials close to the threshold for action potential generation. In voltage clamp, proctolin evokes an inward current, carried at least partially by Na+, that shows strong outward rectification. Removal of extracellular Ca2+ markedly increases the amplitude of the proctolin-evoked current and linearizes its current-voltage curve. The properties of the proctolin current make it ideally suited to contribute to the activity-dependent modulation of the pyloric network of the STG.
Neurite growth on different substrates: permissive versus instructive influences and the role of adhesive strength
Authors: Lemmon V , Burden SM , Payne HR , Elmslie GJ , Hlavin ML
Published: April 2018

Growing axons use environmental cues to guide them to their targets. One class of cues is thought to be adhesion molecules on cells and in the extracellular matrix that axons interact with as they grow to their targets. In choosing between two possible pathways, the relative adhesiveness of the two substrates could be an important factor in controlling neurite growth. We conducted experiments in vitro to study how naturally occurring adhesion molecules influence neurite growth. Neurite growth rates, the degree of neurite fasciculation, the choices neurites make between two substrates, and the relative adhesiveness of different substrates were examined. We found that the relative adhesiveness of a substrate was a poor predictor of either axon growth rate or the degree of fasciculation. Furthermore, neurites showed little selectivity between three different naturally occurring substrates, L1, N-cadherin, and laminin. These results suggest that some adhesion molecules may serve as permissive substrates in that they can define axonal pathways but they do not provide information about which path to take at a choice point or about which direction to go along the path. Finally, these results suggest that substrates in vivo may not exert their effects on axon guidance principally via relative adhesiveness.
Extinction of fear-potentiated startle: blockade by infusion of an NMDA antagonist into the amygdala
Authors: Falls WA , Miserendino MJ , Davis M
Published: April 2018

Data derived from in vitro preparations indicate that NMDA receptors play a critical role in synaptic plasticity in the CNS. More recently, in vivo pharmacological manipulations have suggested that an NMDA- dependent process may be involved in specific forms of behavioral plasticity. All of the work thus far has focused on the possible role of NMDA receptors in the acquisition of responses. However, there are many examples in the behavioral literature of learning-induced changes that involve the reduction or elimination of a previously acquired response. Experimental extinction is a primary example of the elimination of a learned response. Experimental extinction is well described in the behavioral literature, but has not received the same attention in the neurobiological literature. As a result, the neural mechanisms that underlie this important form of learning are not at all understood. In the present experiments, the fear-potentiated startle paradigm was employed to begin to investigate neural mechanisms of extinction. The results show that infusion of the NMDA antagonist D,L-2- amino-5-phosphonovaleric acid (AP5) into the amygdala, a limbic structure known to be important for fear conditioning, dose-dependently blocked extinction of conditioned fear. Control experiments showed that the blockade of extinction was neither the result of the permanent disruption of amygdaloid function nor the result of decreased sensitivity of the animals to the conditioned stimulus. Infusion of AP5 into the interpositus nucleus of the cerebellum, a control site, did not block extinction. Finally, intra-amygdala infusion of a selected dose of the non-NMDA antagonist 6-cyano-7-nitroquinoxaline-2,3-dione did not block extinction of conditioned fear. These results, together with a previous report from our laboratory (Miserendino et al., 1990), demonstrate the importance of the amygdala in the elaboration of conditioned fear and suggest that an NMDA-dependent process might underlie the extinction of conditioned fear.
The embryonic form of neural cell surface molecule (E-NCAM) in the rat hippocampus and its reexpression on glial cells following kainic acid- induced status epilepticus
Authors: Le Gal La Salle G , Rougon G , Valin A
Published: April 2018

The neural cell adhesion molecule (NCAM) changes at the cell surface during development, from highly sialylated forms (embryonic or E-NCAM) to three size classes of less sialylated proteins with apparent molecular mass of 180, 140, and 120 kDa (adult NCAM). In the nervous system, E-NCAM has been localized in developing tissues, where it is thought to play a role in the structuring of neuronal groups and tissue pattern formation. In the present study a monoclonal antibody that specifically detects E-NCAM was used in immunoblot and immunohistochemical procedures. In developing rat hippocampus, E-NCAM cell expression was found to change according to a precise pattern and persisted until 1 month after birth. It was closely associated with the mossy fiber system, an area known for its sprouting propensity. In adult rats, although immunoreactivity considerably decreases and becomes undetectable by immunoblot analysis, E-NCAM was still found to be associated with a few pyramidal-shaped cells in the innermost part of the dentate gyrus. In order to acquire some insight into potential histogenetically plastic functions of E-NCAM, in another series of experiments adult rats were treated with kainic acid, a powerful excitotoxic and convulsant glutamate analog eliciting status epilepticus. When these animals were examined for E-NCAM expression, an intense labeling was found associated with glial-like cells, particularly in the hippocampal formation, and corresponding approximately to the reactive gliosis, as confirmed by staining with anti-glial fibrillary acidic protein antibodies. This expression was detectable from about 3 d following kainic acid administration and persisted for at least 12 weeks; it developed according to an observable spatiotemporal distribution pattern. In animals submitted to amygdala kindling, a nonlesional model of secondarily generalized epilepsy, no such reexpression of E-NCAM was observed. Our observations imply that polysialylation may be a means of identifying neuronal structures capable of plasticity in the CNS. Moreover, intense reexpression of E-NCAM could be a marker of reactive gliosis following brain damage.
Calcium/calmodulin-activated phosphodiesterase expressed in olfactory receptor neurons
Authors: Borisy FF , Ronnett GV , Cunningham AM , Juilfs D , Beavo J , Snyder SH
Published: April 2018

We show that calmodulin-dependent phosphodiesterase (CAM-PDE) is selectively expressed in mature olfactory receptor neurons within the olfactory mucosa. Immunocytochemical staining reveals neuronal immunoreactivity that is most pronounced within cilia, dendritic knobs, and axon bundles. Neither sustentacular cells nor basal cells display immunoreactivity. The extent of loss of neuronal immunoreactivity following bulbectomy resembles loss of the neuronal population. High- affinity CAM-PDE activity in olfactory cilia is fivefold greater than in brain, when assayed at low micromolar cAMP. This activity is depleted in turbinates following bulbectomy. Olfactory mucosal PDE activity is composed of a minimum of two major forms. In the absence of Ca(2+), rolipram-sensitive PDE comprises 65% of total activity. Following stimulation by Ca2+, CAM-PDE activity is elevated sixfold to become the predominant form, thereby increasing total activity 300%, with half-maximal effect at 1 microM Ca2+. We propose that Ca2+ stimulation of CAM-PDE may be necessary for termination of olfactory signals.
Increased GABAA receptor binding in superficial layers of cingulate cortex in schizophrenics
Authors: Benes FM , Vincent SL , Alsterberg G , Bird ED , SanGiovanni JP
Published: April 2018

Recent investigations of postmortem brain from schizophrenic patients have revealed reduced numbers of neurons in several different corticolimbic brain regions. In the prefrontal and anterior cingulate cortices, more specific decreases in the numbers of interneurons, but not pyramidal cells, have been reported to occur preferentially in layer II. Based on this latter finding, a loss of inhibitory basket cells leading to a compensatory upregulation of the GABAA receptor has been hypothesized to occur in schizophrenic patients and to be a contributory factor in the pathophysiology of this disorder. We now report the results of a high-resolution quantitation of GABAA receptor binding in anterior cingulate cortex of postmortem specimens from normal and schizophrenic cases. The results indicate a preferential increase in bicuculline-sensitive 3H-muscimol binding on neuronal cell bodies of layers II and III, but not layers V and VI, of the schizophrenic cases. There was no difference in the size of neurons in any of the layers examined when the control and schizophrenic groups were compared. The neuropil of layer I also showed significantly greater GABAA binding in schizophrenics. The differences seen in the schizophrenic group did not appear to be the result of exposure to antipsychotic medication because one patient who was medication naive and a second who had received minimal exposure to antipsychotic drugs also showed elevated GABAA receptor binding. Since information processing depends on corticocortical integration in outer layers I- III, a disturbance of inhibitory activity in these superficial layers of limbic cortex may contribute to the defective associative function seen in schizophrenia.
Spatiotemporal patterns of expression of NGF and the low-affinity NGF receptor in rat embryos suggest functional roles in tissue morphogenesis and myogenesis
Authors: Wheeler EF , Bothwell M
Published: April 2018

We show here that NGF and its low-affinity receptor (p75NGFR) are expressed during rat embryogenesis at sites that are known to have important roles in tissue morphogenesis and myogenesis. The developing skin of the maxilla, the mandible, and the limb showed very similar patterns of NGF and p75NGFR expression. However, NGF and p75NGFR expression in the developing limb initiated at the limb bud stage and was concentrated at proximal and distal developmental sites that have been reported to be involved in limb morphogenesis. Expression at the proximal/distal ends of the limb persisted throughout limb development, with some of the highest levels of expression occurring at the limb axillary sites, which were not highly innervated. We have also found p75NGFR expression at sites of mesenchymal/epithelial interactions in several developing organs that do not appear to have an adjacent source of NGF and may therefore be sites that bind and respond to the other members of the NGF family (brain-derived neurotrophic factor and neurotrophin-3). These organs include the lung, testes, and kidney, where expression of p75NGFR occurred during the morphogenesis of specific epithelial structures and was coexpressed with the cell adhesion molecule NCAM. In addition, we found that NGF and p75NGFR were expressed during myogenesis. p75NGFR was observed in myoblast cells expressing MyoD1, a myoblast differentiation marker, and NGF transcripts in cells just adjacent to the developing myoblasts. When the myoblasts differentiate into myotubes, p75NGFR and MyoD1 cease to be expressed and the adjacent cells concomitantly cease to be make NGF. However, NGF and p75NGFR were not present in the early muscle precursor cells of the myotome of the somites but were observed in the dermatome and sclerotome, respectively. These results suggest that NGF and p75NGFR have functional roles in developmental processes that affect morphogenesis and cell differentiation.
Vital imaging of glomeruli in the mouse olfactory bulb
Authors: LaMantia AS , Pomeroy SL , Purves D
Published: April 2018

We have monitored the pattern of identified glomeruli in the olfactory bulbs of newborn, juvenile, and adult mice over intervals of several hours to several weeks. Our purpose was to assess the development and stability of these complex units in the mammalian brain. Glomeruli can be observed by vital fluorescent staining and laser-scanning confocal microscopy without causing acute or long-term damage to brain tissue. Repeated observation of bulbs in the same animals between birth and 3 weeks of age showed that this region of the brain develops by progressive addition of these units to the original population. This increment occurs by the genesis of smaller new glomeruli between larger existing ones; no elimination of glomeruli was observed during this process. Finally, no addition (or loss) of glomeruli occurred in adult animals over a 2 week interval; once established, the number, size, and pattern of glomeruli are evidently stable.