bluuebunny/crossref_metadata_2025 huggingface dataset:
Non-neuronal cell-conditioned medium regulates muscarinic receptor expression in cultured sympathetic neurons
Authors: Smith KE , Kessler JA
Published: March 2018

Regulation of muscarinic receptor expression was examined in cultured sympathetic neurons of the neonatal rat superior cervical ganglion. Receptor concentration was determined by measuring binding of the muscarinic antagonist 1-quinuclidinyl[phenyl-4-3H] benzilate (3H-QNB). 3H-QNB bound to one apparent class of noninteracting sites on sympathetic neuron membranes with a Kd of 28.9 pM and a Bmax of 2.91 pmol/mg protein. Binding increased as a linear function of tissue protein and was saturable. The number of receptors per milligram protein increased approximately 6-fold during 16 d of culture, and receptor numbers were down-regulated by treatment with the agonist carbachol. These observations suggested that measurement of 3H-QNB binding would provide a reliable estimate of muscarinic receptor number on cultured sympathetic neurons. To determine whether nonneuronal cells produce soluble factors that influence muscarinic receptor expression, the effects of treatment with rat fibroblast-conditioned medium (RFCM) were examined. Exposure of sympathetic neurons to 50% RFCM resulted in a 57% decrease in muscarinic receptor numbers without a change in the apparent Kd. The decrease in 3H-QNB binding in response to RFCM was dose-related, with a minimum dose of 15% RFCM required to observe a significant effect. In contrast to the carbachol-induced down- regulation, the reduction in binding after RFCM treatment was not prevented by atropine, indicating that the effect was not mediated by acetylcholine stimulation of muscarinic receptors. Binding of 125I- alpha-bungarotoxin, which labels a nonfunctional membrane site distinct from nicotinic receptors, was not altered by treatment with RFCM, indicating the selectivity of the change in membrane muscarinic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
Guidance of regenerating motor axons in larval and juvenile bullfrogs
Authors: Lee MT , Farel PB
Published: March 2018

The segmental distribution of regenerating bullfrog motor axons was mapped in advanced tadpoles and juvenile frogs by stimulating selected muscle nerves and recording from the distal ends of the 3 lumbar ventral roots (VRs) that innervate the hindlimb. When motoneurons were axotomized by VR transection, they reestablished their original innervation fields, rarely, if ever, growing beyond the territory normally supplied by their spinal segment. However, when motoneurons were axotomized in the spinal nerves at the level of the hindlimb plexus, some of them regenerated into limb nerves that lay outside the axons' normal segmental boundaries, and many regenerated into the medial femoral cutaneous nerve, a pathway normally limited to sensory axons. These observations suggest that the ultimate destinations of regenerating axons are largely determined by structures the axons encounter as they penetrate the distal nerve stumps. Thus, axons regenerating from a severed VR grow into that root's own distal stump and reinnervate the hindlimb in a manner that is segmentally appropriate; axons transected near the plexus have access to the pathways of sensory, as well as motor, axons in all 3 lumbar segments, and establish innervation fields that are inappropriate for their segment of origin and their motor function.
Binocularly deprived cats: binocular tests of cortical cells show regular patterns of interaction
Authors: Ohzawa I , Freeman RD
Published: March 2018

If an animal is prevented from receiving visual experience during an early developmental phase, pronounced dysfunctions are observed. Physiological tests reveal gross abnormalities in the striate cortex. Cells in visual cortex are either unresponsive of their response characteristics are erratic. Although fewer than normal numbers of binocular cells are found in cats reared with binocular lid suture, a population remains that can be activated by stimulation through either eye. We have studied cortical cells in binocularly deprived cats in order to specify monocular and binocular response characteristics. The primary hypothesis we have examined is that the abnormal response properties of these cells are a result of an irregular structure or substructure of the receptive fields. Kittens were binocularly lid- sutured soon after birth, and were studied physiologically at ages between 7 and 11 months. Standard techniques were used to record from single cells in striate cortex. Drifting gratings were presented to either eye or to both eyes together. In the latter case, the relative interocular phase was varied between the gratings so that the retinal disparity of the stimuli was changed. We confirmed the expected finding that most cells were either unresponsive or erratic in their response. Of the cells that responded, monocular tuning functions for orientation and spatial frequency of the stimulus were often irregular. However, even in these cases, binocular interaction patterns of cortical responses were nearly always highly regular and displayed phase- specific profiles. A model is presented that explains this finding and suggests how binocular deprivation may result in disorganized receptive- field structure.
Different projections of the central amygdaloid nucleus mediate autonomic and behavioral correlates of conditioned fear
Authors: LeDoux JE , Iwata J , Cicchetti P , Reis DJ
Published: March 2018

The purpose of the present study was to determine whether lesions of areas projected to by the central amygdaloid nucleus (ACE) would disrupt the classical conditioning of autonomic and/or behavioral emotional responses. The areas studied included 3 projection targets of the ACE: the lateral hypothalamic area (LH), midbrain central gray (CG) region, and bed nucleus of the stria terminalis (BNST). Lesions were made either electrolytically or by microinjection of ibotenic acid, which destroys local neurons without interrupting fibers of passage. Two weeks later, the animals were classically conditioned by pairing an acoustic stimulus with footshock. The next day, conditioned changes in autonomic activity (increases in arterial pressure) and emotional behavior (“freezing,” or the arrest of somatomotor activity) evoked by the acoustic conditioned stimulus (CS) were measured during extinction trials. Electrolytic and ibotenic acid lesions of the LH interfered with the conditioned arterial pressure response, but did not affect conditioned freezing. Electrolytic lesions of the rostral CG disrupted conditioned freezing but not conditioned changes in arterial pressure. Ibotenic acid injected into the rostral CG reduced neither the arterial pressure nor the freezing response. Injection of ibotenic acid in the caudal CG, like electrolytic lesions of the rostral CG, disrupted the freezing, but not the arterial pressure response. Injection of ibotenic acid into the BNST had no effect on either response. These data demonstrate that neurons in the LH are involved in the autonomic, but not the behavioral, conditioned response pathway, whereas neurons in the caudal CG are involved in the behavioral, but not the autonomic, pathway. Different efferent projections of the central amygdala thus appear to mediate the behavioral and autonomic concomitants of conditioned fear.
Inositol trisphosphate releases intracellularly stored calcium and modulates ion channels in molluscan neurons
Authors: Fink LA , Connor JA , Kaczmarek LK
Published: March 2018

Stimulation of the bag cell neurons of Aplysia triggers a long-lasting afterdischarge in these cells. In vivo, such a discharge causes the onset of a sequence of reproductive behaviors. We have found that treatments that trigger discharges in vitro stimulate the hydrolysis of phosphoinositides in the bag cell neurons, as measured by increased incorporation of 3H-inositol into fractions containing membrane lipids and water-soluble inositol phosphates. The electrophysiological effects of inositol trisphosphate, one of the products of phosphoinositide turnover that has been shown to mobilize intracellular calcium in non- neuronal cells, were investigated using isolated bag cell neurons in cell culture. Microinjection of inositol trisphosphate into cultured bag cell neurons caused a transient hyperpolarization of the membrane (approximately 35 sec), together with an increase in conductance. This effect of inositol trisphosphate was abolished by 50 mM tetraethylammonium ions. Inositol trisphosphate also reduced the amplitude of action potentials. Injection of calcium ions directly into bag cell neurons mimicked these responses seen after inositol trisphosphate injection. Using the cell-attached patch-clamp technique in conjunction with inositol trisphosphate microinjection, we observed that inositol trisphosphate evoked increases in the activity of a channel carrying outward current at the resting potential and more positive potentials. The estimated slope conductance of the channel modulated by inositol trisphosphate was approximately 40 pS, and its reversal potential was close to that predicted for potassium ions. The increased opening of this channel in response to inositol trisphosphate injection appeared to result from a transient shift of its voltage- dependence to more negative potentials. In a few cases, inositol trisphosphate injection also elicited an increase in the activity of a channel passing inward current at rest. Direct measurements of changes in intracellular calcium in response to inositol trisphosphate were made using digital imaging of isolated neurons loaded with the fluorescent calcium indicator fura-2. These revealed that injection of inositol trisphosphate significantly elevated intracellular calcium levels, and that this inositol trisphosphate-induced rise in cytosolic calcium was not affected by removal of extracellular calcium. In contrast to the effects of trains of action potentials in calcium- containing media, which produced increases in calcium primarily in neurites, the inositol trisphosphate-induced elevation of calcium appeared more localized to the somata of these neurons.(ABSTRACT TRUNCATED AT 400 WORDS)
Distribution and phosphorylation of the growth-associated protein GAP- 43 in regenerating sympathetic neurons in culture
Authors: Meiri KF , Willard M , Johnson MI
Published: March 2018

Sympathetic neurons regenerating in culture were studied in order to gain further insight into the intracellular distribution and phosphorylation of GAP-43, a protein that has been suggested to have a role in axonal outgrowth and neuronal plasticity (Willard et al., 1987). Superior cervical ganglion neurons from embryonic rats were highly reactive with a polyclonal antibody against the growth- associated protein GAP-43 soon after they were placed in culture on a laminin substrate. As these neurons extended neurites, the distribution of GAP-43 reactivity changed. The cell body became progressively less reactive, whereas the growth cone at the tip of the growing neurite reacted strongly. The pattern of immunofluorescence was punctate both in the growth cone and the adjacent neurite, but appeared more diffusely distributed in the cell body. The antibody reacted only with cells that had been subjected to treatment that permeabilized the plasma membrane. When antibody was supplied in the medium of growing neurons, it neither bound to the cells nor altered normal neurite initiation or elongation. Of the different types of cells in these cultures, the antibody reacted only with neurons; it did not react with Schwann cells or fibroblasts. The stimulation of protein kinase C in these cultures resulted in a 7-fold stimulation of the phosphorylation of a protein of similar electrophoretic mobility to GAP-43. These observations demonstrate that GAP-43 is neuron-specific, is present throughout the neuron but at higher levels in the growth cone, and is a major substrate of protein kinase C. The high concentration of GAP-43 in the growth cones may necessitate its increased synthesis in neurons with elongating axons. Its location and phosphorylation by kinase C suggest that it could perform a function in the growth cone that is modulated by extracellular signals, such as those used in pathfinding or in the control of axonal elongation.
Local role of Ca2+ in formation of veils in growth cones
Authors: Goldberg DJ
Published: March 2018

A previous study that used high-resolution video (VEC-DIC) microscopy to examine axonal growth cones of Aplysia giant neurons growing in culture had demonstrated that growth occurs by the extension of veils of membrane between filopodia and the subsequent morphological transformation of these veils, in place, into the swollen, organelle- filled central region of the growth cone and then into the cylindrical axon. The possible involvement of Ca2+ in this sequence of events was now examined using VEC-DIC microscopy. Reduction of [Ca2+]o from the normal level of 11 to 1.3 mM or below or the addition of 20 mM Co2+, which blocks Ca2+ channels, caused a large decrease in the area of immature veil (flat and with few organelles) in the growth cone within minutes. Ba2+, 20 mM, which flows well through Ca2+ channels, and 5 microM A23187, a Ca2+ ionophore, caused new immature veil to form in the presence of reduced [Ca2+]o. Maturation of veil into central region was not inhibited by reduced [Ca2+]o. In fact, the disappearance of immature veil was often the result partly, or entirely, of continued veil maturation in the absence of formation of new veil. The next step in maturation, conversion of the central region to cylindrical axon, was also probably not inhibited by reduced [Ca2+]o. Ca2+ was microapplied to large growth cones that had lost their veils by exposure to reduced [Ca2+]o. There was a strong tendency for the first, or only, incidence of veil formation to occur near the micropipette, the rest of the perimeter of the growth cone remaining quiescent. It is concluded that intracellular Ca2+ plays a role in veil formation and that the site of the Ca2+-dependent step is close to the site of veil formation. If this step is exocytosis, veil forms where there is net addition of membrane. Whether a change in [Ca2+]i, rather than some other factor, normally directly triggers veil formation remains uncertain, but, if it does, then the site of formation, which will strongly influence the direction of axon growth, is probably determined by focal changes in [Ca2+]i within the growth cone.
Depolarization of nonmyelinated fibers of the rat vagus nerve produced by activation of protein kinase C
Authors: Rang HP , Ritchie JM
Published: March 2018

The effect of activation of protein kinase C by phorbol esters has been studied on the nonmyelinated (C) fibers of the rat vagus nerve. Grease- gap recording at room temperature was used to monitor changes in the resting and action potentials. Effects of phorbol esters on the rate of efflux of 86Rb and 14C-guanidinium were also measured. The active isomer beta-phorbol 12,13-dibutyrate (PDBu), applied for 10 min at concentrations of 10 nM to 3 microM, caused a slowly developing depolarization, which persisted after the drug was washed out. The action potential was concomitantly reduced. These effects did not occur with the inactive isomer alpha-phorbol 12,13-didecanoate. The PDBu- induced depolarization was reduced by about 75% if Na+ was replaced by the impermeant cation N-methyl-(+)-glucamine (NMG); the residual effect was almost abolished if the nerves were presoaked in a solution containing gluconate in place of Cl-. It was concluded that increases in conductance mainly to Na+ and Cl- were responsible for the depolarization. The response was unaffected by tetrodotoxin or calcium- channel blockers. Omission of Ca2+, surprisingly, enhanced the PDBu- induced depolarization 3–5-fold; furthermore, addition of 2 mM Ca2+ following a PDBu-induced depolarization recorded in Ca2+-free solution caused a pronounced repolarization. This effect of Ca2+ occurred also with Sr2+ and Ba2+, but not with other divalent cations or with La3+. Divalent cations known to block Ca channels inhibited the repolarizing action of Ca2+. These results suggested that Ca2+ acts intracellularly, either to block Na channels opened by PDBu or to activate protein phosphatases. The PDBu-induced response in Ca2+-free solution was increased 2-fold by a reduction in pH from 7.4 to 6.5. Under normal conditions the nerve was reversibly depolarized by this pH change; after PDBu this pH sensitivity was enhanced, and depolarization occurred at a less acidic pH. PDBu caused a 3–4-fold increase in the rate of efflux of 86Rb (a marker for K+ ions) and of 14C-guanidinium (a marker for Na+ ions) from preloaded nerves. These effects, in contrast to the depolarization, were transient.(ABSTRACT TRUNCATED AT 400 WORDS)
Nerve growth factor enhances dendritic arborization of sympathetic ganglion cells in developing mammals
Authors: Snider WD
Published: March 2018

Recent observations have suggested that the dendritic arbors of sympathetic ganglion cells may be regulated by interactions with their peripheral targets (Voyvodic, 1987a; Yawo, 1987). In order to assess a potential mechanism for such interactions, I have investigated the effects of the target-derived trophic molecule for sympathetic ganglion cells on the development of dendrites in the rat superior cervical ganglion. Systemic treatment of neonatal animals with NGF for 1 or 2 weeks results in a striking expansion of ganglion cell dendritic arbors, as revealed by intracellular staining with HRP. During this period, neurons in treated animals extend new primary dendrites, and the length and branching of existing dendrites are increased compared to age-matched controls. These results support the idea that targets may regulate ganglion cell arbors via elaboration of NGF, and suggest an explanation for the correlation between animal size and dendritic complexity noted in several recent studies (Purves and Lichtman, 1985a; Snider, 1987; Voyvodic, 1987a).
Selective attention in an insect auditory neuron
Authors: Pollack GS
Published: March 2018

Previous work (Pollack, 1986) showed that an identified auditory neuron of crickets, the omega neuron, selectively encodes the temporal structure of an ipsilateral sound stimulus when a contralateral stimulus is presented simultaneously, even though the contralateral stimulus is clearly encoded when it is presented alone. The present paper investigates the physiological basis for this selective response. The selectivity for the ipsilateral stimulus is a result of the apparent intensity difference of ipsi- and contralateral stimuli, which is imposed by auditory directionality; when simultaneous presentation of stimuli from the 2 sides is mimicked by presenting low- and high- intensity stimuli simultaneously from the ipsilateral side, the neuron responds selectively to the high-intensity stimulus, even though the low-intensity stimulus is effective when it is presented alone. The selective encoding of the more intense (= ipsilateral) stimulus is due to intensity-dependent inhibition, which is superimposed on the cell's excitatory response to sound. Because of the inhibition, the stimulus with lower intensity (i.e., the contralateral stimulus) is rendered subthreshold, while the stimulus with higher intensity (the ipsilateral stimulus) remains above threshold. Consequently, the temporal structure of the low-intensity stimulus is filtered out of the neuron's spike train. The source of the inhibition is not known. It is not a consequence of activation of the omega neuron. Its characteristics are not consistent with those of known inhibitory inputs to the omega neuron.
Proteolytic processing of human brain alpha spectrin (fodrin): identification of a hypersensitive site
Authors: Harris AS , Morrow JS
Published: March 2018

The processing of brain spectrin (fodrin) by calcium-dependent proteases at the postsynaptic membrane has been postulated to be one of the central molecular mechanisms underlying long-term potentiation (LTP). The effect of such processing on the structure and function of brain spectrin, and on spectrin's ability to organize or otherwise regulate receptor function remains unclear. To address these issues, human and bovine brain spectrin were digested under mild conditions with several proteases, and the resulting cleavage fragments analyzed by 2-dimensional chymotryptic 125I peptide mapping. These studies identify an underlying protease-resistant domain structure reminiscent of, yet distinctly different from, human erythroid spectrin. More importantly, fodrin is unusual for the presence of a single, proteolytically hypersensitive site in the center of the alpha subunit, which is the favored site of action by many proteases, including the calcium-dependent neutral proteases. This proteolytically hypersensitive site is a unique feature of alpha nonerythroid spectrin since it is absent from human erythrocyte spectrin and appears to be the site at which the molecule is processed in vivo. In addition, on the basis of gel overlay techniques, it appears that the hypersensitive site is also the site at which calmodulin binds to the alpha-subunit in a calcium-dependent manner. These studies thus establish at the molecular level 2 calcium-dependent mechanisms by which brain spectrin function might be regulated and provide a conceptual and methodological framework for further investigation into the function of this important molecule.
Nature and fate of proliferative cells in the hippocampal dentate gyrus during the life span of the rhesus monkey
Authors: Eckenhoff MF , Rakic P
Published: March 2018

The nature of proliferative cells in the subgranular zone (SGZ) of the hippocampal region and the fate of their progeny was analyzed by 3H- thymidine (3H-TdR) autoradiography combined with immunocytochemistry at the light and electron microscopic levels in 18 rhesus monkeys ranging in age from late gestation to 17 years. Our analysis indicates that, during the last quarter of gestation and the first 3 postnatal months, the SGZ produces both glial and neuronal cells. These 2 major classes of cells originate from the 2 precursor lines and, following their mitotic division, migrate to the granular layer. During the juvenile period (4–6 months of age), neuronal production tapers off and most postmitotic cells remaining within the SGZ differentiate into glial elements. In postpubertal animals (3 years and older), the 3H-TdR- labeled cells in the dentate gyrus belong to several non-neuronal classes. The largest group was immunoreactive to the glial fibrillary acidic protein (GFAP) at both the light and electron microscopic levels, indicating their astrocytic nature. The remaining 3H-TdR- labeled, GFAP-negative cells had ultra-structural characteristics of either microglia, oligodendroglia, or their progenitory stem cells. Therefore, there is a continuing addition and/or turnover of the glial cells in the dentate gyrus of sexually mature monkeys, but, in contrast to the massive neurogenesis reported in adult rodents, the production of new neurons could not be detected after puberty. The significance of a stable population of neurons in the hippocampal formation of mature primates is discussed in relation to its possible function in memory.
Presynaptic inhibition of identified wind-sensitive afferents in the cercal system of the locust
Authors: Boyan GS
Published: March 2018

The paired cerci located at the tip of the locust abdomen bear a large number of wind-sensitive filiform hairs, each of which sends an axon via the cercal nerve into the terminal ganglion of the CNS. The filiform afferents fire bursts of action potentials when their hairs are displaced by wind or mechanical stimuli. Filiform axon terminals in the CNS are depolarized concomitantly with the discharge of another type of unit (a primary afferent-depolarizing, or PAD, unit) recorded in the cercal nerve. The instantaneous spike frequency of PAD unit discharges matches the evoked depolarization very closely, and during such depolarizations spike amplitudes in the filiform afferent terminals are reduced by up to 55%. Depolarizing current pulses injected into the axonal terminals of an identified filiform afferent evoke spikes that are blocked by the PAD unit, probably via an intercalated interneuron. The PAD unit makes a monosynaptic connection with only one of the 4 giant interneurons (GIN 2) on each side of the terminal ganglion, and indirect connections with 2 others. Depolarizing current pulses injected into the neuropilar segments of GINs evoke fewer spikes when the PAD unit is active, consistent with the PAD unit's mediation of conductance changes in postsynaptic cells. Iontophoretic injection of Lucifer yellow shows the PAD unit to be an afferent with axon terminals overlapping those of filiform afferents and posteriorly directed branches of interneurons such as GIN 2 in the CNS. Passive movements of a cercus, monitored with a position transducer, show that the PAD unit fires discrete bursts during cercal displacement. The PAD unit most probably has its soma and dendrites in tissue spanning the cercal base. By responding to cercal movements sufficient to also activate filiform hairs, and by mediating conductance changes in both the presynaptic terminals of filiform afferents and the postsynaptic membranes of interneurons, the PAD unit desensitizes a pathway to movement-generated afferent input, and ensures that the locust remains sensitive to external wind stimuli.
Flexible body dynamics of the goldfish C-start: implications for reticulospinal command mechanisms
Authors: Eaton RC , DiDomenico R , Nissanov J
Published: March 2018

As a model for learning how reticulospinal networks coordinate movement, we have analyzed the function of the Mauthner (M-) neurons in the escape response of the goldfish. We used water displacements of 3–6 micron to elicit C-start escape responses. These responses consist of 2 fundamental movements that grade into each other: Stage 1 lasts 15–40 msec and rotates the body 30 degrees-100 degrees about the center of mass; stage 2 is an axial acceleration that moves the center of mass 2– 6 cm. Combined, the 2 stages result in trajectory turns ranging from 15 degrees to 135 degrees. Thus, these data show that M-initiated C-starts are not fixed movement patterns. The durations of stage 1 body muscle EMGs were correlated with turn angles achieved during stage 1. Since variable stage 1 EMGs are not seen when the M-cell is triggered by itself, other circuits, independent of the M-cell, must control the extent of the initial turn, and consequently escape trajectory. Furthermore, turning angles of stages 1 and 2 were correlated, allowing escape trajectory to be predicted, on average, 26 msec after movement started. This suggests that the commands for escape trajectory should be organized by the end of stage 1. In concert with this, the time of onset of the stage 2 EMG preceded the stage 2 onset by a range with a mean of 28.4 msec, typically putting the stage 2 command at the beginning of stage 1 movement. Thus, stage 2 initiation does not require motion-dependent feedback. Our findings indicate that the Mauthner cell initiates the first of a series of motor commands that establish the initial left-right decision of the escape sequence from the side of the stimulus, whereas parallel circuits simultaneously organize the command controlling the escape angle.
MAP2 is a component of crossbridges between microtubules and neurofilaments in the neuronal cytoskeleton: quick-freeze, deep-etch immunoelectron microscopy and reconstitution studies
Authors: Hirokawa N , Hisanaga S , Shiomura Y
Published: March 2018

Microtubules (MT) and neurofilaments (NF) are linked by frequent crossbridges in situ. In order to answer the question of what makes these crossbridges, we performed the immunogold procedure on rat spinal cord motor neurons using an affinity-purified polyclonal antibody against rat brain MAP2 and gold-labeled anti-rabbit IgG goat IgG. A quick-freeze, deep-etch technique (QF-DE) in conjunction with decoration with anti-MAP2 antibody and ferritin-labeled second antibody was also used. In motor neuron dendrites crossbridges were clearly displayed between MTs and NFs by QF-DE. These crossbridges were revealed in thin sections as fuzzy filamentous structures between MT and NF. Gold particles studded the fuzzy structures associated with MT. Many such structures connected MTs to NFs. Furthermore, antibody complexes containing ferritin were localized on the crossbridges between MTs and NFs by the QF-DE study. In addition, we performed reconstitution experiments. We isolated 70 kDa (L) protein of neurofilaments from calf spinal cords and assembled L to form neurofilaments in vitro. MAP2 bound these neurofilaments according to both SDS-PAGE and QF-DE electron microscopy of the pellets of suspensions containing L proteins and MAP2. When we added tubulin to this suspension and polymerized it in the presence of taxol, neurofilaments were crosslinked with microtubules by MAP2 crossbridges. Hence, from these 2 approaches we concluded that MAP2 is a component of crossbridges between MTs and NFs in the neuronal cytoskeleton in vivo and in vitro.
Methylenedioxyamphetamine (MDA) and methylenedioxymethamphetamine (MDMA) cause selective ablation of serotonergic axon terminals in forebrain: immunocytochemical evidence for neurotoxicity
Authors: O'Hearn E , Battaglia G , De Souza EB , Kuhar MJ , Molliver ME
Published: March 2018

The psychotropic amphetamine derivatives 3,4-methylenedioxyamphetamine (MDA) and 3,4-methylenedioxymethamphetamine (MDMA) have been used for recreational and therapeutic purposes in man. In rats, these drugs cause large reductions in brain levels of serotonin (5-HT). This study employs immunocytochemistry to characterize the neurotoxic effects of these compounds upon monoaminergic neurons in the rat brain. Two weeks after systemic administration of MDA or MDMA (20 mg/kg, s.c., twice daily for 4 d), there is profound loss of serotonergic (5-HT) axons throughout the forebrain; catecholamine axons are completely spared. Regional differences in drug toxicity are exemplified by partial sparing of 5-HT axons in hippocampus, lateral hypothalamus, basal forebrain, and in some areas of neocortex. The terminals of 5-HT axons are selectively ablated, while axons of passage and raphe cell bodies are spared. Thickened preterminal fibers exhibit increased staining due to damming-up of neurotransmitter and other axonal constituents. Fine 5- HT axon terminals are extremely vulnerable to these drugs, whereas terminal-like axons with large varicosities survive, raising the possibility that some 5-HT axons may be resistant to the neurotoxic effects. At short survivals, visualization of greatly swollen, fragmented 5-HT axons provides anatomic evidence for degeneration of 5- HT projections. The results establish that MDA and MDMA produce structural damage to 5-HT axon terminals followed by lasting denervation of the forebrain. Both drugs have similar effects, but MDA produces a greater reduction of 5-HT axons than does MDMA at the same dosage. The selective degeneration of 5-HT axons indicates that these drugs may serve as experimental tools to analyze the organization and function of 5-HT projections. Caution should be exercised until further studies determine whether these compounds may be hazardous in man.
Synuclein: a neuron-specific protein localized to the nucleus and presynaptic nerve terminal
Authors: Maroteaux L , Campanelli JT , Scheller RH
Published: March 2018

We used an antiserum against purified cholinergic synaptic vesicles from Torpedo and expression screening to isolate a cDNA clone encoding synuclein, a 143 amino acid neuron-specific protein. A cDNA clone was also isolated from a rat brain cDNA library that encodes a highly homologous 140 amino acid protein. The amino terminal 100 amino acids of both proteins are comprised of an 11 amino acid repeating unit that contains a conserved core of 6 residues. The synuclein gene is expressed only in nervous system tissue, not in electric organ, muscle, liver, spleen, heart, or kidney. In the electric organ synapse Torpedo synuclein-immunoreactive proteins are found in 3 major molecular-weight classes of 17.5, 18.5, and 20.0 kDa. In the neuronal cell soma the 17.5 kDa species is predominant and immunoreactivity is localized to a portion of the nuclear envelope.
The otolithic origin of the vertical vestibuloocular reflex following bilateral blockage of the vertical semicircular canals in the rabbit
Authors: Barmack NH , Pettorossi VE
Published: March 2018

The influence of bilateral plugs of the anterior and posterior semicircular canals (ASCs, PSCs) on the vertical vestibuloocular reflex (VVOR-0) of the rabbit, oscillated about the longitudinal axis maintained in a horizontal orientation, was studied. Bilateral plugs of either the ASCs or PSCs reduced the gain of the VVOR-0 evoked by stimulus frequencies above 0.005 Hz, but left a residual gain of the VVOR at higher stimulus frequencies that could be attributed to the remaining vertical canals and utricular otoliths. Bilateral plugs of both the ASCs and PSCs totally eliminated the canal component of the VVOR-0, as determined from measurements made when the animal was oriented at 90 degrees (“nose up”) with respect to the earth horizontal axis, eliminating changes in the gravitational vector acting on the utricular maculae (VVOR-90). Bilateral plugs of both the ASCs and PSCs also eliminated the nonlinear increase in gain observed at intermediate frequencies in intact rabbits when oscillated about a supine orientation (VVOR-180). The contribution of the utricular otolith to the VVOR-0 in rabbits with both the ASCs and PSCs plugged was compared with the predicted otolith contribution based on the assumption of linear summation of semicircular canal and utricular signals. The assumption of linear summation appears to be incorrect. Bilateral plugs of the ASCs and PSCs increased the latency and the time constant of vertical eye movements evoked by step-roll stimulation about the longitudinal axis maintained in a horizontal orientation.
Sex and regional differences in the incorporation of neurons born during song learning in zebra finches
Authors: Nordeen EJ , Nordeen KW
Published: March 2018

In zebra finches only males sing, and several nuclei controlling song contain more neurons in adult males than in females. The ontogeny of sex differences in neuron number differs across song regions and overlaps with song learning in males. We examined the development of neuron number in several song regions in both sexes. We then determined whether neurons are born and incorporated into song nuclei as sex differences in neuron number emerge, and whether sex or regional differences in the insertion of such neurons may account for differences in the development of these areas. Males add neurons to hyperstriatum ventralis pars caudalis (HVc) and Area X between 20 and 55 d of age. In females there is no change in HVc neuron number during this time, and Area X never appears as a distinct nucleus. In both sexes, 3H-thymidine administration between 20 and 30 d results in neuronal labeling at 55 d in HVc and the region of Area X. However, in these areas the incidence of labeled neurons is higher in males than in females. In contrast to HVc and Area X, sex differences in neuron number in the robustus nucleus of the archistriatum (RA) and the magnocellular nucleus of the neostriatum (MAN) emerge because males retain neurons that are lost in females between 20 and 55 d of age. Accordingly, RA and MAN neurons are not labeled following 3H-thymidine administration between 20 and 30 d of age. These data indicate that sex and regional differences in the ontogeny of song nuclei are related to differences in the incorporation of neurons born during song learning.
Classical conditioning of the eye withdrawal reflex in the green crab
Authors: Abramson CI , Feinman RD
Published: March 2018

Eye withdrawal in the green crab, Carcinus maenas was conditioned by pairing a mild vibration to the carapace as a conditioned stimulus (CS) with a puff of air to one of the eyes as an unconditioned stimulus (US). Animals subjected to repeated pairings showed an increased probability of eye retraction during CS presentation. Significantly less responding was found in several control groups subjected to backward conditioning, unpaired stimuli, stimuli alone, or simply time in the apparatus. Although conditioned animals showed few responses to CS alone after 24 hr, retention could be demonstrated by acquisition that was much more rapid on day 2 than on day 1. Conditioning could also be effected in the eye when it was restrained, a result consistent with reports in the literature that this reflex does not require proprioceptive feedback. Because the neuromuscular circuitry of eye withdrawal is already well defined in Carcinus, this is a promising candidate for studying the neuronal basis of classical conditioning.
Glutamate-positive corticocortical neurons in the somatic sensory areas I and II of cats
Authors: Conti F , Fabri M , Manzoni T
Published: March 2018

Combined retrograde transport-immunocytochemical experiments were carried out on cats to study the morphology, laminar distribution, and percentages of corticocortical projecting neurons of somatosensory area I (SI) and II (SII) showing immunoreactivity to an antiserum raised against the amino acid glutamate (Glu). A previously characterized anti- Glu serum (Conti et al., 1987a, b; Hepler et al., 1987) was used in conjunction with HRP. This tracer was injected either in SI to label retrogradely neurons in ipsilateral SII (SII-SI association neurons) and contralateral SI (SI-SI callosal neurons) or in SII to label retrogradely neurons in ipsilateral SI (SI-SII association neurons) and contralateral SII (SII-SII callosal neurons). In sections from SI and SII processed for simultaneous visualization of Glu and HRP (Bowker et al., 1982), and containing the cells from which every one of the 4 corticocortical projections arise, 3 types of labeled neurons were observed: (1) single-labeled neurons showing the homogeneous brown immunoreaction product of Glu (Glu-positive neurons); (2) single- labeled neurons containing the granular black reaction product of retrogradely transported HRP (Glu-negative, association or callosal neurons); and (3) double-labeled neurons in which both the black HRP granules and the brown immunostaining were present (Glu-positive, association or callosal neurons). Double-labeled neurons were all pyramidal in shape and were distributed intermingled with Glu-negative corticocortical neurons in all layers of SI and SII known to give rise to association and callosal projections. Counts from 25-micron-thick sections showed that of 432 association and callosal neurons sampled from SI and SII, 214 (49.5%) were Glu-negative and 218 (50.5%) Glu- positive. In counts carried out on 5-micron-thick sections, the percentage of Glu-positive corticocortical neurons raised to about 70%. The 2 populations of single- and double-labeled corticocortical neurons showed no difference in their perikaryal cross-sectional areas. The present results show that a large fraction of association and callosal neurons of SI and SII are immunoreactive for Glu, and, therefore, these neurons probably use this excitatory amino acid, or a closely related compound, as neurotransmitter.
Disconnection of the amygdala from visual association cortex impairs visual reward-association learning in monkeys
Authors: Gaffan EA , Gaffan D , Harrison S
Published: March 2018

Cynomolgus monkeys (Macaca fascicularis) were trained in a task that assessed their ability to associate visual stimuli with food reward. Acquisition of stimulus-reward associations was measured under 2 conditions, a 2-stimuli acquisition condition and a 1-stimulus acquisition condition. On each trial in the 2-stimuli condition, the positive (correct) and negative (incorrect) stimuli were presented side by side and the animal chose one by touching it; if the choice was correct, a food reward was dispensed. On each trial in the 1-stimulus condition, either the positive or the negative stimulus was presented alone; if the stimulus was the positive, it was followed by reward delivery, regardless of the animal's response to it, and if it was the negative, it was not followed by reward delivery. Thus, reward delivery was contingent upon the animal's response to the stimuli in the 2- stimuli condition but not in the 1-stimulus condition. The effect of acquisition trials under these 2 conditions was measured, in both conditions, by the animal's subsequent choice when presented with the 2 stimuli side by side. Following preoperative training in this task, the animals were first subjected to unilateral ablation of the inferotemporal cortex. This operation had little effect on the animals' learning ability. Then, the amygdala was ablated in the hemisphere contralateral to that in which the unilateral inferotemporal ablation had been carried out. This combination of crossed unilateral lesions of the amygdala and of the inferotemporal cortex, which disconnects the amygdala from the output of visual association cortex, produced a profound impairment in stimulus-reward-associative learning.(ABSTRACT TRUNCATED AT 250 WORDS)
Micropruning: the mechanism of turning of Aplysia growth cones at substrate borders in vitro
Authors: Burmeister DW , Goldberg DJ
Published: March 2018

Growth cones of Aplysia californica neurons were observed with video- enhanced contrast-differential interference contrast (VEC-DIC) microscopy as they turned at a border between poly-L-lysine-treated and untreated glass. Growth cones that turned generally developed 2 distinct active areas of filopodial and veil formation, much in the way of growth cones undergoing branching. Both active areas advanced, but turning of the neurite occurred through the selective resorption of the incipient branches developing on the untreated substrate. Thus, micropruning of developing regions of the growth cone, rather than the asymmetric extension of filopodia or veils, was primarily responsible for directing neurite growth. We present the hypothesis that abrupt turns by growing neurites are mediated by 2 sets of signals, one causing growth cone splitting, and a second set regulating the survival of the separate branches.
Capsaicin-induced ion fluxes in dorsal root ganglion cells in culture
Authors: Wood JN , Winter J , James IF , Rang HP , Yeats J , Bevan S
Published: March 2018

Capsaicin is a pungent pain-producing compound found in plants of the capsicum family; it exerts excitatory, desensitizing, and toxic effects on a subset of sensory neurons, including the polymodal nociceptor population. We have carried out a quantitative study of capsaicin- induced fluxes of sodium, guanidine, calcium, rubidium, and chloride ions in cultures of neonatal and adult rat DRG neurons, in conjunction with the use of a histochemical stain that identifies capsaicin- sensitive neurons by means of cobalt uptake. Those cells that take up cobalt in a capsaicin-dependent manner (EC50 = 0.2 microM) represent about 50% of the total neuronal population derived from neonatal DRGs on short-term culture. Overnight treatment of cultures with 2 microM capsaicin leads to the loss of the cobalt-staining subpopulation. The capsaicin-insensitive neurons contain immunoreactive neurofilament epitopes that are present in fewer than 10% of capsaicin-sensitive neurons. This observation provides indirect evidence that the sensitive cells correspond to the small, dark B-type neurons, which are negative for neurofilament immunoreactivity in vivo. A capsaicin-dependent calcium uptake (EC50 = 0.2 microM), as measured by 45Ca incorporation, is shown by a DRG neuronal subpopulation that, like the cobalt-staining population of DRG neurons, is lost after overnight capsaicin treatment (2 microM). Capsaicin application leads to the accumulation of millimolar levels of calcium within a few minutes. Cadmium and other divalent cations block capsaicin-induced calcium uptake, but little or no inhibition is seen with organic calcium channel antagonists. Mitochondria, rather than the endoplasmic reticulum, are the probable destination of the internalized calcium, because ruthenium red inhibits calcium uptake (IC50 = 0.05 microM), whereas methylxanthines are inactive. The subset of sensory neurons that takes up calcium also releases 86Rb when exposed to capsaicin (EC50 = 0.06 microM). No efflux of 36Cl ions could be induced by capsaicin. These cells also show a capsaicin-induced uptake of 22Na or 14C guanidine (EC50 = 0.06 microM). In contrast, chick DRG cells in culture showed no capsaicin-induced calcium or cobalt uptake. Primary cultures of rat superior cervical ganglion neurons and Schwann cells, and a number of neuronal cell lines, also failed to respond to capsaicin, as judged by the calcium, cobalt, or guanidine uptake assays.
The effect of unilateral naris occlusion on cell dynamics in the developing rat olfactory epithelium
Authors: Farbman AI , Brunjes PC , Rentfro L , Michas J , Ritz S
Published: March 2018

The effect of unilateral naris occlusion on the cellular dynamics of developing olfactory epithelium was investigated in postnatal rats. Nares of rat pups, at 1, 5, and 10 d postnatally, were cauterized; after a 30 d survival period, the olfactory mucosa was examined histologically, and quantitative estimates were made of total number of receptor neurons (together with basal cells), supporting cells, and epithelial thickness. In each group, there were significant differences between occluded and patent sides in total numbers of neurons and in epithelial thickness but no difference in number of supporting cells. The differences were greater in the animals that had been occluded for 1–30 d than in the 5–35 or 10–40 d groups, suggesting that the earlier postnatal days are more sensitive to the effects of occlusion. We evaluated the number of mature olfactory neurons by staining immunohistochemically with an antibody against olfactory marker protein. There were no differences in the number of mature receptor neurons between the occluded and non-occluded sides, indicating the effect of naris occlusion was on the neurons in the immature and/or the “almost mature” population. Using 3H-thymidine autoradiography, we determined that there was a 40% reduction in the rate of neurogenesis in the animals occluded 1–30 d after birth. Further, the rate of cell proliferation in nasal respiratory epithelium declined by approximately the same amount.(ABSTRACT TRUNCATED AT 250 WORDS)
Muscarine and t-LHRH suppress M-current by activating an IAP- insensitive G-protein
Authors: Pfaffinger P
Published: March 2018

The control of M-current by muscarinic ACh receptors and luteinizing hormone releasing hormone (LHRH) receptors was studied in dialyzed frog sympathetic ganglion neurons. M-current was recorded in dialyzed cells without run-down or changes in its biophysical properties and could be reversibly suppressed by muscarine and teleost LHRH (t-LHRH). However, dialysis with internal solutions lacking ATP or substituting with APP(NH)P caused the loss of M-current, suggesting that dephosphorylation suppresses the activity of M-channels. M-current over- recovers after agonist addition and removal to a size 30% larger than control, as if latent channels are activated during the recovery. Dialysis of cells with the G-protein activators GTP gamma S, fluoride, and aluminum fluoride causes loss of M-current. G-protein activation by receptors was confirmed by dialysis with low concentrations of GTP gamma S in competition with GTP. This prevents the rapid loss of M- current, but addition of muscarine or t-LHRH caused irreversible loss of M-current, suggesting that both transmitter receptors do suppress M- current by activating a G-protein. Suppression of M-current was not affected by treatment with 0.1 microgram/ml pertussis toxin (IAP) for 24–48 hr. In addition, based on the lack of IAP-specific labeling of frog sympathetic neuron membrane proteins, no IAP-sensitive G-proteins are present in these cells. These results indicate that an IAP- insensitive G-protein couples muscarinic and LHRH receptors to the suppression of M-current.