bluuebunny/crossref_metadata_2025 huggingface dataset:
The effects of monensin and of puromycin on transport of membrane components in the frog retinal photoreceptor. II. Electron microscopic autoradiography of proteins and glycerolipids
Authors: Matheke ML , Holtzman E
Published: March 2018

Monensin converts the Golgi apparatus of rod photoreceptors into distended vacuoles, similar to those seen in other monensin-treated cell types, and leads to the accumulation of [3H]leucine in the distended vacuoles. As evaluated by quantitative, electron microscopic autoradiography, transport of newly made proteins--both to the outer segments and to the presynaptic terminals--is inhibited. These effects suggest that the Golgi apparatus is involved in transport in both principal directions within the highly polarized photoreceptors, a matter of interest since there seems to be only a single, extensive, Golgi apparatus in the cell body. Seemingly there are two distinguishable “sorting” routes, for proteins, out of the Golgi apparatus and, for the terminals, an additional non-Golgi route. Accumulation of newly made glycerolipids in the outer segments and terminals is less affected by monensin than is accumulation of new proteins, and glycerolipid accumulation is little affected by puromycin, an inhibitor of protein synthesis. These latter findings suggest that the routes or mechanisms of assembly of newly made lipids into membranes in the photoreceptors are at least partially dissociable from those for newly made proteins.
Defects in specific associations between astroglia and neurons occur in microcultures of weaver mouse cerebellar cells
Authors: Hatten ME , Liem RK , Mason CA
Published: March 2018

The neurological mutation weaver is characterized by defects in granule cell migration along Bergmann glial processes and by subsequent death and disposition of granule cells. Immunocytochemical localization of antisera raised against purified glial filament protein (AbGF) and transmission electron microscopy were used to visualize specific associations between granule neurons and astroglia in microcultures of cerebellar cells dissociated from normal (+/+), heterozygous (+/wv), and homozygous (wv/wv) B6CBA-w mouse cerebella. In microcultures of cells dissociated from normal B6CBA-Aw-J-wv (+/+) cerebella, staining with AbGF closely resembled results previously reported for cells taken from C57BL/6J (+/+) tissue. Two forms of stained astroglia were seen, one with a larger perikaryon and shorter processes, among which 12 to 20 unstained cells nestled; and another with a smaller cell soma and longer processes, along which a few unstained cells were seen. The first resembled astrocytes of the internal granular layer and white matter, the second Bergmann astroglia. In microcultures of heterozygous animals (+/wv), the number of granule cells was reduced slightly. Many stained astroglia resembled those from +/+ cerebella, but others had thickened processes and enlarged terminal “endfeet.” Granule cells associated with both forms of stained astroglia. Cultures from homozygous weaver (wv/wv) cerebella contained very few, if any, granule cells and did not exhibit specific neuronal/glial interactions characteristic of +/+ cells. Both forms of stained wv/wv glial cells had enlarged cell somata giving rise to stunted processes, suggesting that both Bergmann glia and astrocytes are affected by the weaver gene. By both immunocytochemical staining and electron microscopy, accumulation and tangling of glial filaments were seen. Immunocytochemical staining of weaver cerebellar tissue with AbGF also revealed abnormalities in astrocytes of the internal granular layer and white matter in addition to previously reported defects in Bergmann glia. Agglutination with five plant lectins, each with a different carbohydrate-binding specificity, revealed that postnatal weaver cerebellar cells agglutinate with the lectins concanavalin A and the wheat germ agglutinin, suggesting the persistence postnatally of embryonic cell surface elements on postnatal weaver cells.
Lateral hypothalamic modulation of oral sensory afferent activity in nucleus tractus solitarius neurons of rats
Authors: Matsuo R , Shimizu N , Kusano K
Published: March 2018

This study was designed to examine whether the sensory afferents from the anterior part of the tongue are modulated by activity of the lateral hypothalamic area (LHA) at the level of the nucleus tractus solitarius (NTS) in rats. The electrical activity of the NTS neurons was recorded extracellularly, and they were classified as gustatory, thermal, or mechanical neurons in accordance with their responsiveness to tongue stimulation by taste solutions, by warm and cool water, and by stroking the tongue surface. Sixty-two percent of the neurons were polysynaptically activated by electrical stimulation of the LHA. When a single conditioning stimulus of the LHA was applied prior to the test electrical stimulation of the tongue at various conditioning-test intervals, the activity of the gustatory neurons was facilitated by 30 to 80% of their control level for a period of approximately 20 to 150 msec. The activity of the mechanical and thermal neurons was suppressed for a 50- to 300-msec period. These results demonstrate the existence of modulatory effects from the LHA on the NTS neurons, which receive sensory information from the tongue.
In vivo biosynthesis and transport of oxytocin, vasopressin, and neurophysins to posterior pituitary and nucleus of the solitary tract
Authors: White JD , Krause JE , McKelvy JF
Published: March 2018

A growing body of literature suggests that oxytocin (OXY) and arginine- vasopressin (AVP), in addition to their neuroendocrine roles, may serve as neuromodulators within the central nervous system of mammals. The present study investigated the biosynthesis of OXY, AVP, and their associated neurophysins in the paraventricular nucleus of the hypothalamus (PVN) and the transport of these peptides to the neural lobe and the nucleus of the solitary tract (NTS) in the brainstem of the rat. Rats were cannulated bilaterally in the PVN, and 24 hr later a 2-hr pulse of [35S]cysteine was administered using an Alzet minipump delivery system. After a 10-hr chase period, the neural lobe was removed and the PVN and NTS were punched. Tissue homogenates were adsorbed to and eluted from octadecyl-silica cartridges and analyzed by linear high performance liquid chromatography (HPLC) gradient elution, chemical and enzymatic modification, and exponential gradient HPLC elution followed by linear HPLC gradient analysis using an ion-pairing buffer system. This rigorous approach has allowed us to identify 35S- labeled material which co-purifies with OXY and AVP from the PVN, neural lobe, and NTS samples. Specific transport of the nonapeptides to the NTS was demonstrated when a unilateral transection of the hypothalamic fibers resulted in a unilateral depletion of the radiolabeled peptides from the NTS samples. Additionally, each of the neurophysins was purified from the neural lobe and NTS samples after linear HPLC gradient analysis, ion-pairing buffer linear gradient analysis, then tryptic digestion followed by exponential gradient HPLC analysis.(ABSTRACT TRUNCATED AT 250 WORDS)
Electrophysiological mechanisms of kainic acid-induced epileptiform activity in the rat hippocampal slice
Authors: Fisher RS , Alger BE
Published: March 2018

Depression of GABA-mediated IPSPs has been proposed to be a crucial factor in the onset of epileptiform activity in most models of epilepsy. To test this idea, we studied epileptiform activity induced by bath application of the excitatory neurotoxin kainic acid (KA) in the rat hippocampal slice. Repetitive field potential firing, spontaneous or evoked, occurred during exposure to KA. Intracellular records from 52 CA1 pyramidal cells during changes from control saline to saline containing 1 microM KA indicated that KA depolarized cells an average of about 5 mV and caused a 15% decrease in input resistance. Action potentials and current-induced burst afterhyperpolarizations did not change significantly. In several cells the tonic effects of KA were preceded by a transient phase of sporadic, spontaneous depolarizations of 2 to 10 mV and 50 to 200 msec duration. These phasic depolarizations were blocked by hyperpolarization. The major effect of 1 microM KA was a depression of synaptic potentials. Initially, KA depressed fast GABA- mediated IPSPs and slow, non-GABA-mediated late hyperpolarizing potentials to 23% and 40% of control values, respectively. IPSP depression correlated closely with onset of burst potential firing in response to synaptic stimulation. Similar observations were made on six cells from the CA2/3 region, although these cells were affected by lower doses of KA. The mechanism of IPSP depression was studied by using KCl-filled electrodes to invert spontaneous IPSPs and make them readily visible. In nine CA1 cells the rate and amplitude of spontaneous IPSPs transiently increased but then decreased in conjunction with evoked IPSP depression. Possible KA effects on postsynaptic GABA responses were investigated by applying GABA locally to cells. KA did not significantly affect GABA responses. Prolonged exposure of CA1 cells to KA in doses of 1 microM or higher depressed intracellularly and extracellularly recorded EPSPs and all field potential activity. This depression was not apparently due to depolarization block in CA1, however. We conclude that KA induces epileptiform activity in hippocampus principally by a presynaptic block of IPSP pathways.
Substantial reduction of cortical noradrenaline by lesions of adrenergic pathway does not prevent effects of monocular deprivation
Authors: Daw NW , Robertson TW , Rader RK , Videen TO , Coscia CJ
Published: March 2018

We tested the theory that depletion of noradrenaline reduces the plasticity of the visual cortex in kittens by using another method of depletion. Lesions were made in the lateral hypothalamus to interrupt fibers in the dorsal noradrenergic bundle going from the locus ceruleus to the telencephalon. The lesions were induced at approximately 3 1/2 weeks of age in kittens; approximately 2 weeks later one eye was sutured shut, and about 10 days after that cells were recorded in the visual cortex. The location of the lesions was verified histologically, and the effect of the lesions was verified by noradrenaline analyses (high pressure liquid chromatography-electrochemistry) of samples from the visual cortex. The noradrenaline content of the visual cortex was reduced by 70 to 90%. However, the majority of cells recorded in the visual cortex could not be driven through the eye that had been sutured closed. The ocular dominance histograms for cells in the visual cortex were indistinguishable from those of kittens that were monocularly deprived for a similar period in the “critical period” and that had normal amounts of noradrenaline in their visual cortex. Therefore, we conclude that reduction of the noradrenaline content of the visual cortex by 70 to 90% is insufficient, by itself, to prevent the physiological changes that occur in the visual cortex after monocular deprivation.
Quantitative lineage analysis of the frog's nervous system. I. Lineages of Rohon-Beard neurons and primary motoneurons
Authors: Jacobson M , Moody SA
Published: March 2018

After injection of horseradish peroxidase into single blastomeres in Xenopus embryos at 2- to 512-cell stages, all of the descendants could be traced and counted at tailbud stages. Progenitors of Rohon-Beard neurons and of primary spinal motoneurons were identified, and all neurons of each type that originated from individual progenitors were counted. From these data were derived the geometric mean of the number (N) of Rohon-Beard neurons (or primary motoneurons) that descended from a single progenitor at each generation from the first to ninth, the mean number of progenitors of each generation, and the probability that, following mitosis, daughter cells continue in the Rohon-Beard (or primary motoneuron) lineages (the continuation probability). On a log- log plot a straight line fitted to the N values intercepts the abscissa at the 13th generation for Rohon-Beard progenitors and the 16th generation for primary motoneuron progenitors. This indicates the number of generations before the entire set of each type of neuron is finally produced. The Rohon-Beard neurons and primary motoneurons each originated from a separate group of progenitors at the 9th generation (512-cell stage), but those progenitors gave rise to mesodermal, endodermal, and ectodermal cells in addition to neurons. Nevertheless, the continuation probability significantly greater than 0.5 shows that there is a bias toward recruitment of progenitors into lineages leading to production of those two types of neurons.(ABSTRACT TRUNCATED AT 250 WORDS)
Prenatal development of functional connections in the cat's retinogeniculate pathway
Authors: Shatz CJ , Kirkwood PA
Published: March 2018

The development of functional connections between the axons of retinal ganglion cells and the neurons of the dorsal lateral geniculate nucleus (LGNd) of fetal and neonatal cats was studied using an in vitro assay. Extracellular microelectrode recordings of single units were made from histologically identified sites in the LGNd of isolated diencephalon preparations between embryonic day 39 ( E39 ) and postnatal day 2 (P2). (Gestation is 65 days in the cat.) Postsynaptic units activated by electrical stimulation of one or both optic nerves were found at all ages tested from E39 onwards. Over 90% of the units studied in the fetal preparations received convergent excitation from both optic nerves, compared with roughly half of the units studied in the neonatal optic nerves, compared with roughly half of the units studied in the neonatal preparations. Inhibition was detected in the LGNd of the neonatal preparations, but in only the oldest of the fetal preparations ( E59 ). This physiological change from predominantly convergent excitation to an adult-like mixture of excitation and inhibition seen at birth coincides with the change from mixed to segregated afferent input from the two eyes seen anatomically ( Shatz , C. J. (1983) J. Neurosci . 3: 482–499). These results indicate that attainment of the adult pattern of retinogeniculate connectivity involves the elimination of already functional synapses.
Acetylcholine metabolism in rat spinal cord cultures: regulation by a factor involved in the determination of the neurotransmitter phenotype of sympathetic neurons
Authors: Giess MC , Weber MJ
Published: March 2018

Acetylcholine metabolism has been studied in sister cultures of E13 rat spinal cord cells cultured for 1 to 3 weeks with or without conditioned medium (CM) from rat skeletal muscle cells. Spinal cord cells grown with CM synthesized and accumulated 3 to 4 times more [3H]ACh from [3H]choline than cultures grown without CM. This effect of CM was accompanied by a comparable increase in CAT activity and could not be mimicked by increasing the density of the spinal cord cultures. A 2- to 3-fold increase in AChE activity was also observed in 2- to 3-week-old CM cultures, whereas the activity of lactate dehydrogenase was identical in cultures grown with and without CM. We have compared the effects of CMs from various non-neuronal cell cultures on [3H]ACh synthesis and storage by spinal cord cultures and by sympathetic neuron cultures. CM by skeletal muscle greater than skin fibroblasts greater than rat heart muscle greater than C6 glioma cells were the most active on both types of neuron cultures, whereas CM from rat brain, L6 myoblasts, mouse 3T3, and PYT21 fibroblasts was inactive on spinal cord cultures and only weakly active on sympathetic neurons. Serum-free CM from skeletal muscle was inactive on both types of neuron cultures. The CM factor active on spinal cord cultures has been purified several thousand-fold by using a four-step fractionation scheme which has previously led to a partial purification of the CM factor involved in the regulation of CAT, AChE, and catecholamine-synthesizing enzymes in sympathetic neuron cultures ( Swerts , J. P., A. Le Van Thai, A. Vigny , and M. J. Weber (1983) Dev. Biol. 100: 1–11). Moreover, a comparison of dose-response curves established with this purified material showed that it exerted its effects on spinal cord and on sympathetic neuron cultures in the same range of concentration. Thus, these results suggest that the same macromolecule is involved in the regulation of neurotransmitter phenotype in both types of cultures despite their different embryological origins.
Astrocytes in primary culture have chemically activated sodium channels
Authors: Bowman CL , Kimelberg HK , Frangakis MV , Berwald-Netter Y , Edwards C
Published: March 2018

The effects of two neurotoxins, veratridine and alpha-scorpion toxin II, on the resting potential (RMP) of identified astrocytes in primary cell culture were studied using standard electophysiological techniques. Veratridine caused either a series of transient depolarizations or a single sustained depolarization. alpha-Scorpion toxin (II), which alone had no effect on the RMP, increased the duration of the veratridine-induced transient depolarizations. The depolarization caused by veratridine, or veratridine plus alpha- scorpion toxin II, was reversibly inhibited by tetrodotoxin and was reversibly reduced in magnitude by reductions of the external sodium concentration. Cells that were depolarized by veratridine were shown to contain glial fibrillary acidic protein, a marker considered specific for astrocytes. The effect of long-term (10 sec) hyperpolarizing and depolarizing current pulses on the RMP were also studied using two microelectrodes, i.e., dual impalement. In the absence of veratridine, the RMP returned to its original value after termination of the hyperpolarizing current, and the current-voltage relationship was found to be linear over a wide range of membrane potentials. In the presence of veratridine, termination of the hyperpolarizing current produced a transient depolarization that was sensitive to tetrodotoxin. Depolarizing current pulses were without effect. These results show that astrocytes in primary culture have a chemically inducible Na+ channel that appears to be voltage-dependent.
Intracellular injection with horseradish peroxidase of physiologically characterized stellate and bushy cells in slices of mouse anteroventral cochlear nucleus
Authors: Wu SH , Oertel D
Published: March 2018

Nissl-stained tissue from brain slice preparations of the anteroventral cochlear nucleus of the mouse resembles tissue fixed in situ. Multipolar, spherical, globular, and granule cells can be distinguished after intracellular injection with horseradish peroxidase (HRP). Stellate cells have relatively large dendritic fields; their axons have collaterals which terminate within the cochlear nuclear complex. Bushy cells have smaller dendritic fields; where they can be seen, axons have no collaterals. Granule cells have few short dendrites; their very fine axons branch close to the cell body and could be followed only for short distances. Intracellular recordings from six stellate cells labeled by intracellular injection of HRP revealed that they have linear current-voltage relationships around the resting potential and that they respond to suprathreshold depolarization with large, regularly firing action potentials. Intracellular recordings from four bushy cells, also labeled by injection of HRP, showed that these cells have nonlinear current-voltage relationships around the resting potential and that they respond to suprathreshold depolarizations with only one or two small action potentials. The anatomical and physiological features of bushy cells reduce summing in time and space and make bushy cells well suited to preserve the firing patterns of auditory nerve inputs. The anatomical and physiological features of stellate cells, in contrast, allow summing in time and space.
The dynamics of nerve growth factor-induced neurofilament and vimentin filament expression and organization in PC12 cells
Authors: Lee VM , Page C
Published: March 2018

Monoclonal antibodies specific for neurofilament (NF) subunits or for vimentin filament (VF) protein were used to study nerve growth factor (NGF)-induced intermediate filament expression and organization in a rat pheochromocytoma cell line (PC12 cells). NGF induced increased amounts of NF subunits and VF protein, and they were, in part, differentially localized within PC12 cells. The relative expression of each of the NF subunits and of the VF protein was measured by an enzyme- linked immunosorbent assay which revealed that PC12 cells grown in the absence of NGF (PC12- cells) contained 3 times more VF than 68,000- dalton NF subunits and only barely detectable amounts of 150,000- and 200,000-dalton NF subunits. Exposure of these cells to NGF (PC12+ cells) increased the amount of 68,000-dalton NF subunits 8-fold, VF protein 3-fold, and 150,000- and 200,000-dalton NF subunits 2-fold. The ratio of VF to 68,000-dalton NF proteins in PC12- versus PC12+ cells decreased from 3.0 to 1.5. Both VF and 68,000-dalton NF subunits were arranged in juxtanuclear “ball-like” configurations and both were present in neurites of PC12+ cells. The distribution of 150,000- and 200,000-dalton NF subunits was diffuse with perinuclear stippling and only occasional, weakly fluorescent “balls.” After perturbation of the PC12 cell cytoskeleton, 68,000-dalton NF subunits and VF redistributed together and, thus, may exist as heteropolymers. Finally, the increased expression of NF and VF proteins was specific for NGF since they could not be induced by other hormones or “growth” factors. We conclude that PC12 cells constitute a model system for studies of NF and VF expression, assembly, and interactions.
Anatomical basis of specific connections between sensory axons and motor neurons in the brachial spinal cord of the bullfrog
Authors: Lichtman JW , Jhaveri S , Frank E
Published: March 2018

The anatomical basis for the specificity of the monosynaptic stretch reflex has been studied in the brachial spinal cord of bullfrogs. Sensory axons from the triceps brachii muscle innervate the corresponding triceps motoneurons but do not innervate two types of unrelated motoneurons (subscapularis and pectoralis) (Lichtman, J.W., and E. Frank (1984) J. Neurosci. 4: 1745–1753). Retrograde labeling of these three types of motoneurons with horseradish peroxidase (HRP) demonstrated that their cell bodies had overlapping distributions in the lateral motor column, and their dendrites all occupied the same region of the dorsal horn. In addition, triceps sensory axons aborized extensively in the dorsal horn throughout the brachial spinal cord, with no obvious predilection for the region of the triceps motoneurons. Thus, the physiological specificity of these sensory-motor connections was not apparent from the anatomical location of the sensory or motor neurons. However, by injecting single pairs of related or unrelated sensory and motor cells with HRP, we found that related pairs formed anatomical contacts with each other more frequently than unrelated sensory-motor pairs did. These observations suggest that the specificity of these connections is most likely the result of local interactions between sensory and motor processes.
Biosynthesis and axoplasmic transport of neurophysins in the hypothalamo-neurohypophysial system of Rana pipiens
Authors: Chang AC , Dellmann HD
Published: March 2018

Biosynthesis, axoplasmic transport, and storage of neurophysin in the amphibian (Rana pipiens) magnocellular peptidergic neurosecretory system were studied, and the results were compared with those reported in mammals. After injection of [35S]cysteine into the preoptic recess, light microscopic autoradiography provides evidence that neurons of the preoptic nucleus (PON) synthesize cysteine-rich proteins. The time course of appearance of these [35S]cysteine-labeled proteins in different regions of the hypothalamo-neurohypophysial system was studied by slab gel autoradiography. [35S]Cysteine-labeled proteins were found in the PON less than 1 hr postinjection, whereas a major labeled protein, tentatively identified as the neurophysin, first appeared in the infundibulum and neural lobe 4 hr after the injection. In addition, the labeled neurophysin persisted in the neural lobe throughout the entire observation period of 5 days. The minimum transport rate for neurophysin was calculated as 0.9 mm/hr (22 mm/day) at 25 degrees C. Two different neurophysins (with isoelectric points (pI) 4.9 +/- 0.1, 4.6 +/- 0.1, and Mr = 23,000, 20,100) may be resolved from the neural lobe extracts by isoelectric focusing and SDS- polyacrylamide gel electrophoresis, respectively. In addition to the neurophysin peaks, two radioactive peaks with pI 5.2 and 5.8 may be detected in the preoptic nucleus and the infundibulum as early as 30 min after [35S]cysteine injection. Preliminary conversion studies suggest a putative precursor role for the pI 5.2 protein. The results indicate that in the amphibian peptidergic neurosecretory system, the synthesis of cysteine-rich neurophysin by the preoptic neurons, the transport through the infundibulum, and the storage in the neural lobe proceed similarily to their mammalian counterparts.
Characterization of glycolipids synthesized in an identified neuron of Aplysia californica
Authors: Sherbany AA , Ambron RT , Schwartz JH
Published: March 2018

Because radioactive precursors can be injected directly into the cell body or axon of R2, a giant, identified neuron of the Aplysia abdominal ganglion, it was possible to show that glycolipid is synthesized in the cell body, inserted into membranes along with glycoprotein, and then exported into the axon within organelles that are moved by fast axonal transport. After intrasomatic injection of N-[3H]-acetyl-D- galactosamine, five major 3H-glycolipids were identified using thin layer polysilicic acid glass fiber chromatography. At least two of the lipids are negatively charged. Analysis of 32P-labeled lipid from the abdominal ganglion revealed the presence of 2-aminoethylphosphonate, indicating that these polar substances are sphingophosphonoglycolipids. The major 3H-glycolipids synthesized in R2 are similar to a family of phospholipids isolated from the skin of A. kurodai, previously characterized by Araki et al. (Araki, S., Y. Komai, and M. Satake (1980) Biochem J. 87: 503–510). Since sialic acid is absent in Aplysia as in other invertebrates, these polar glycolipids may function like gangliosides in vertebrates. The polar 3H-glycolipids are synthesized and incorporated into intracytoplasmic membranes solely in the cell body. Direct injection of the labeled sugar into the axon revealed no local synthesis or exchange of glycolipid. Moreover, there was no indication for transfer from glial cells into axoplasm. Although the incorporation of N-[3H]-acetyl-D-galactosamine into glycolipid is not affected by anisomycin, an effective inhibitor of protein synthesis, the export into the axon of membranes containing the newly synthesized lipid is completely blocked by the drug.(ABSTRACT TRUNCATED AT 250 WORDS)
Pharmacological properties of isolated horizontal and bipolar cells from the skate retina
Authors: Lasater EM , Dowling JE , Ripps H
Published: March 2018

Retinal neurons were enzymatically and mechanically dissociated from adult skate retinas and maintained in cell culture for up to 14 days. Intracellular recordings were made from isolated horizontal and bipolar cells while neurotransmitters were applied via pressure ejection. L- Glutamate, quisqualate, kainate, and gamma-amino-butyric acid (GABA), when applied to horizontal cells, produced large (60 to 70 mV), long- lasting depolarizations. These responses appear to consist of at least two components: a graded depolarization and a Ca++-dependent regenerative component. As regards bipolar cells, L-glutamate and its analogues depolarized about 30% of the cells tested, while GABA hyperpolarized most of these neurons. Both agents acted on bipolar cells by increasing conductance. Repeated applications of L-glutamate, quisqualate, kainate, and GABA to horizontal cells produced no desensitization, but in these circumstances the glutamate analogues, kainate and quisqualate, induced certain morphological changes, most notably a retraction of cell processes and the appearance of blebs on the cell surface.
Antibodies against filamentous components in discrete cell types of the mouse retina
Authors: Drager UC , Edwards DL , Barnstable CJ
Published: March 2018

Three monoclonal antibodies have been raised against the ganglion cell layer of the adult mouse retina. The first antibody, R3, labeled optic axons in the inner retina, and with colchicine pretreatment somata and dendrites of large ganglion cells could be seen. A small number of other processes, including fibers projecting to the retina from elsewhere (efferent fibers), were also labeled in the inner retina. In the outer plexiform layer R3 stained the axonless class of horizontal cells. R3 recognized a 185,000- to 200,000-dalton polypeptide which is most probably the heaviest of the neurofilament subunits. Antibodies R4 and R5 labeled filamentous components mainly in glia and cells of mesenchymal origin. The antigens appeared in most but not quite all locations morphologically closely related to the intermediate filament protein vimentin. In the retina both antibodies labeled strongly the regularly spaced Muller glia. The astroglia of the optic fiber layer was stained with R5 but not R4. Although the two antigens were in general not expressed in neurons, they were both present in axonless horizontal cells in the outer plexiform layer, coexisting with neurofilaments in this neuron.
Topographical distribution of decrements and recovery in muscarinic receptors from rat brains repeatedly exposed to sublethal doses of soman
Authors: Churchill L , Pazdernik TL , Jackson JL , Nelson SR , Samson FE , McDonough JH
Published: March 2018

[3H]Quinuclidinyl benzilate binding to rat brain muscarinic receptors decreased after repeated exposure to soman, a potent organophosphorus cholinesterase inhibitor. The topographical distribution of this decrement was analyzed by quantitative receptor autoradiography. After 4 weeks of soman, three times a week, quinuclidinyl benzilate binding decreased to 67 to 80% of control in frontal and parietal cortex, caudate-putamen, lateral septum, hippocampal body, dentate gyrus, superior colliculus, nucleus of the fifth nerve, and central grey. Minor or no decreases were observed in thalamic or hypothalamic nuclei, reticular formation, pontine nuclei, inferior colliculus, nucleus of the seventh nerve, and cerebellum. Scatchard analyses of saturation curves using frontal cortex sections from soman-treated rats revealed a decrease in maximal quinuclidinyl benzilate binding from that in control rats and a return toward control levels by 24 days without any significant change in affinity. These brain areas showing significant decrements in muscarinic receptors recovered with a similar time course. An estimate of the time for 50% recovery for some of the brain areas was 14 days for superior colliculus, 16 days for cortex, and 19 days for hippocampal body. The application of quantitative receptor autoradiography to analyze receptor alterations has been valuable in localizing the telencephalon as a region more susceptible to change in receptor concentration.
Initial expression of neurofilaments and vimentin in the central and peripheral nervous system of the mouse embryo in vivo
Authors: Cochard P , Paulin D
Published: March 2018

The appearance of neurofilaments (NFs) and vimentin (Vim) in the nervous system of the mouse embryo was documented using immunohistochemical techniques. The three NF protein subunits appear early and simultaneously in central and peripheral neurons at 9 to 10 days of gestation. The onset of NF expression is concomitant with axon elongation and correlates extremely well with neurofibrillar differentiation and, in the case of autonomic ganglia, with the expression of adrenergic neurotransmitter properties. In the central and peripheral nervous system, NF expression is preceded by that of Vim, and both types of intermediate filaments coexist within the same cell for a short period of time.
Monoclonal antibodies against the alpha-bungarotoxin-binding protein of chick optic lobe
Authors: Betz H , Pfeiffer F
Published: March 2018

The nicotinic acetylcholine (ACh) receptor probe alpha-bungarotoxin (alpha-Butx) binds with high affinity to a membrane protein of the vertebrate central nervous system. To characterize further this putative neuronal ACh receptor, we have prepared monoclonal antibodies (mAbs) against the alpha-Butx-binding protein of chick optic lobe. Mice were immunized with affinity-purified protein preparations which were estimated to be 10 to 20% pure. Spleen cells from an immunized mouse were fused with the mouse myeloma cell line X63-Ag 8.653. From this fusion, 14 stable hybridoma lines were isolated which produce mAbs against the chick neuronal alpha-Butx-binding protein. Most of the antibodies inhibited alpha-Butx-binding to membrane fractions and/or detergent extracts of chick optic lobe. Some of the mAbs cross-reacted with the alpha-Butx-binding protein of the rat pheochromocytoma cell line PC12. However, none of the mAbs bound to a significant extent to the nicotinic ACh receptor of chick skeletal muscle or of Torpedo californica electric organ. All antibodies specifically isolated a polypeptide of Mr = 57,000 (+/- 2,000) from radiolabeled neuronal protein preparations. The present data show that these mAbs constitute useful tools for the further molecular and functional characterization of the neuronal alpha-Butx-binding protein.
Demonstration of postsynaptic opioid modulation of thalamic projection neurons by the combined techniques of retrograde horseradish peroxidase and enkephalin immunocytochemistry
Authors: Ruda MA , Coffield J , Dubner R
Published: March 2018

Thalamic projection neurons represent a major source of nociceptive information from the dorsal horn to higher centers of the neuraxis. The synaptic relationship between thalamic projection neurons and the opioid peptide enkephalin (ENK) was examined at the light (LM) and ultrastructural (EM) level using the combined techniques of retrograde transport of horseradish peroxidase and ENK immunocytochemistry. Utilizing two different chromogens to develop the peroxidase reaction product, the two labeled neural elements could be readily distinguished at the LM level in the same tissue section. In the medullary, cervical, and lumbar levels of the dorsal horn of both the cat and monkey, at least 30% of the thalamic projection neurons in lamina I were observed at the LM level to be contacted by ENK-immunoreactive varicosities. In lamina V, approximately 50% of the thalamic projection neurons received ENK contacts. Since some neurons were not observed to receive a dense ENK innervation on their somata and proximal dendrites, these data suggest that there may be different functional types of thalamic projection neurons. At the EM level, the ENK-immunoreactive varicosities were observed to form asymmetrical synaptic contacts on the labeled somata and proximal dendrites of the projection neurons. In all cases, the ENK varicosities were morphologically similar and contained round or oval agranular vesicles and a few dense-core vesicles. These observations suggest that ENK acts to a substantial degree on postsynaptic opiate receptors located on thalamic projection neurons.
An electron microscopic analysis of the trans-synaptic effects of peripheral nerve injury subsequent to tooth pulp extirpations on neurons in laminae I and II of the medullary dorsal horn
Authors: Gobel S
Published: March 2018

To examine the effects of peripheral nerve injury on second-order neurons in laminae I and II of the medullary dorsal horn, tooth pulps of all mandibular teeth in adult cats on one side were extirpated. This procedure severed and removed the receptors and terminal branches of the primary trigeminal neurons which innervate the tooth pulps of these teeth. The empty pulp chambers were then filled with dental cement to prevent regeneration. At 30 and 60 days postoperatively, membrane-lined cavities had formed inside many of the small-caliber dendrites of second-order neurons in laminae I and II. Cavity formation occurred mainly in dendritic shafts less than 2 micron in diameter and involved dendrites with synaptic vesicles as well as those without synaptic vesicles. The cavities extensively hollowed out these dendrites, often occupying more than half the cross-sectional diameter of the shafts and extending for appreciable distances in the long axis of the shaft. The process of cavitation ultimately resulted in the destruction of the affected dendrites. Many cavities became patent to the intercellular space with the cavity membrane establishing continuity with the dendritic membrane. Many cavities often formed in a single dendrite, and such severely cavitated dendrites became reduced to a trabeculated shell which ultimately fragmented into several small pieces. The presence of synaptic connections from a number of different kinds of axonal endings, including scalloped and dome-shaped endings, was not sufficient to prevent cavitation. The actual severing of synaptic connections on the cavitated dendrite appeared to be a relatively late event in the process since small pieces of dendritic debris could still be found clinging to their axodendritic synapses. Evidence that dendrites were being lost from the neuropil was most readily apparent in many of the disrupted glomeruli in lamina II in which many of the scalloped depressions in the central axonal endings that normally contained small dendrites were empty. Many central axonal endings remained in synaptic contact with only a single dendrite which often showed signs of cavitation. Such central endings showed only subtle remaining traces of their normal scalloped contours. This study demonstrates that injury to the distal branches of primary trigeminal neurons which innervate tooth pulps resulted in trans-synaptic degenerative changes in the dendritic arbors of second-order neurons which destroyed fine-caliber higher order dendrites.
Comparative immunohistochemical and biochemical analysis of pancreatic polypeptide-like peptides with special reference to presence of neuropeptide Y in central and peripheral neurons
Authors: Lundberg JM , Terenius L , Hokfelt T , Tatemoto K
Published: March 2018

Antisera raised against porcine neuropeptide Y (NPY) and peptide YY (PYY) were characterized with regard to immunohistochemical staining, cross-reactivity to several pancreatic polypeptide (PP)-related peptides, and radioimmunoassayable tissue levels in the rat and pig. The NPY antiserum (102B) reacted with nerves in many areas of both the central and peripheral nervous systems, but it did not stain endocrine cells of the pancreas or intestine. No evidence for any cross- reactivity of the NPY antiserum with related peptides of the PP family, such as avian PP, bovine PP, PYY, gamma-MSH, FMRF-amide, or avian PP (31–36), was obtained. The NPY antiserum was N-terminally directed, and regional levels of NPY as seen by radioimmunoassay paralleled well the occurrence of NPY-immunoreactive structures seen in the immunohistochemical study. High pressure liquid chromatography analysis revealed that the NPY-immunoreactive material from cerebral cortex and vas deferens had elution profiles similar to those of standard porcine NPY. The PYY antiserum mainly stained endocrine cells in the pancreas and intestine as well as a small neuron system in the brainstem of the rat. Although this antiserum had a slight cross-reactivity to NPY in radioimmunoassay, the neuronal PYY staining was separate from that of NPY. High levels of PYY were found in the intestine, and levels above the threshold were also seen in the dorsal vagal complex of the rat. The other antisera investigated (raised against avian PP, bovine PP, gamma-MSH, and FMRF-amide) caused neuronal staining that was abolished by preabsorption with NPY. This was also seen even if no detectable cross-reactivity with NPY was found in radioimmunoassay. These latter antisera also stained endocrine cells in the pancreas and intestine with complex cross-reactivity relationships, suggesting the presence of intestinal PP-like peptides in addition to PYY and NPY.
Evidence implicating substantia nigra in regulation of kindled seizure threshold
Authors: McNamara JO , Galloway MT , Rigsbee LC , Shin C
Published: March 2018

We studied the effects of microinjected drugs and brainstem lesions on motor and limbic seizures in the kindling model of epilepsy. The duration of motor seizures was determined by timing the colonic and tonic movements of the extremities. The duration of limbic seizures was determined by measuring afterdischarge recorded on the electroencephalogram. Bilateral microinjection of a gamma-aminobutyric acid (GABA) agonist, muscimol, into the area of the substantia nigra (SN) markedly suppressed both motor and limbic seizures induced by stimulation of amygdala, olfactory structures, or lateral entorhinal cortex. Microinjection of saline did not suppress seizures. The suppressive effect of muscimol: (i) dissipated after several hours and was dependent on dose; (ii) was due to an elevation of the seizure threshold, since typical seizures could be elicited with electrical current far exceeding the threshold; and (iii) exhibited spatial specificity since muscimol injections 1 to 2 mm dorsal to the SN or into neocortex did not suppress the seizures. The actions of muscimol were probably mediated by its GABA agonist properties, since microinjection of an irreversible inhibitor of GABA transaminase (gamma- vinyl GABA) into the area of the SN also suppressed kindled seizures. Destruction of brainstem structures was produced by microinjection of the neurotoxin, N-methyl-D,L-aspartate. Seizures were markedly suppressed in animals with bilateral destruction of the SN but not in animals in which the SN was spared bilaterally. We interpret the data to indicate that the SN is the site at which the GABA agonists and lesions act to raise the threshold for kindled seizures. The suppression of limbic seizures indicates that this brainstem nucleus can regulate the intrinsic neuronal excitability of hemispheric sites.
[3H]-beta-endorphin binding in rat brain
Authors: Houghten RA , Johnson N , Pasternak GW
Published: March 2018

The binding of [3H]-beta-endorphin to rat brain homogenates is complex. Although Scatchard analysis of saturation studies yields a straight line, detailed competition studies are multiphasic, suggesting that even at low concentrations of the compound, the 3H-ligand is binding to more than one class of site. A portion of [3H]-beta-endorphin binding is sensitive to low concentrations of morphine or D-Ala2-Leu5- enkephalin (less than 5 nM). The inhibition observed with each compound alone (5 nM) is the same as that seen with both together (each at 5 nM). Thus, the binding remaining in the presence of both morphine and the enkephalin does not correspond to either mu or delta sites. The portion of [3H]-beta-endorphin binding that is inhibited under these conditions appears to be equally sensitive to both morphine and the enkephalin and may correspond to mu1 sites. Treating membrane homogenates with naloxonazine, a mu1 selective antagonist, lowers [3H]- beta-endorphin binding to the same degree as morphine and D-Ala2-Leu5- enkephalin alone or together. This possible binding of [3H]-beta- endorphin to mu1 sites is consistent with the role of mu1 sites in beta- endorphin analgesia and catalepsy in vivo.
Cyclic guanosine 3':5'-monophosphate mimics the effects of light on a circadian pacemaker in the eye of aplysia
Authors: Eskin A , Takahashi JS , Zatz M , Block GD
Published: March 2018

Environmental light regulates the phase of a circadian oscillator in the eye of Aplysia. We are attempting to define the events involved in transmitting light information from the environment to the circadian pacemaking mechanism in the eye. In this paper, we present several lines of evidence that cyclic guanosine 3′:5′-monophosphate (cGMP) is involved in the photic entrainment pathway. Light increases the level of cGMP in eyes without having detectable effects on cyclic adenosine 3′:5′-monophosphate (cAMP). An analogue of cGMP, 8-bromoguanosine 3′:5′- cyclic monophosphate (cGMP), can shift the phase of the circadian rhythm from the eye; the phase response curves for light and for 8- bromo cGMP are indistinguishable. Neither 8-bromo cAMP nor 8-bromo 5′- GMP mimics the effect of light or of 8-bromo cGMP on the rhythm. Light and 8-bromo cGMP appear to use convergent mechanisms for entrainment since the effects of these two treatments are nonadditive. Also, low Na+ solutions antagonize the effects of both treatments. Finally, the kinetics of phase shifting by 8-bromo cGMP are similar to the kinetics of phase shifting by light. In addition to perturbing the circadian rhythm, 8-bromo cGMP increases the frequency of spontaneous optic nerve impulses. The pattern of nerve impulses during 8-bromo cGMP treatment is the same as the pattern of impulses produced by light. The excitatory effect of 8-bromo cGMP, the low Na+ blockade of the effects of 8-bromo cGMP, and the involvement of membrane depolarization in phase shifting by light suggest that depolarization mediates the effect of 8-bromo cGMP on the rhythm.(ABSTRACT TRUNCATED AT 250 WORDS)
Morphology, distribution, and synaptic relations of somatostatin- and neuropeptide Y-immunoreactive neurons in rat and monkey neocortex
Authors: Hendry SH , Jones EG , Emson PC
Published: March 2018

Neurons in the monkey and rat cerebral cortex immunoreactive for somatostatin tetradecapeptide (SRIF) and for neuropeptide Y (NPY) were examined in the light and electron microscope. Neurons immunoreactive for either peptide are found in all areas of monkey cortex examined as well as throughout the rat cerebral cortex and in the subcortical white matter of both species. In monkey and rat cortex, SRIF-positive neurons are morphologically very similar to NPY-positive neurons. Of the total population of SRIF-positive and NPY-positive neurons in sensory-motor and parietal cortex of monkeys, a minimum of 24% was immunoreactive for both peptides. Most cell bodies are small (8 to 10 micron in diameter) and are present through the depth of the cortex but are densest in layers II-III, in layer VI, and in the subjacent white matter. From the cell bodies several processes commonly emerge, branch two or three times, become beaded, and extend for long distances through the cortex. The fields formed by these processes vary from cell to cell; therefore, the usual morphological terms bipolar, multipolar, and so on do not adequately characterize the full population of neurons. Virtually every cell, however, has at least one long vertically oriented process, and most processes of white matter cells ascent into the cortex. No processes could be positively identified with the light microscope as axons. The processes of the peptide-positive neurons form dense plexuses in the cortex. In each area of monkey cortex, SRIF-positive and NPY-positive processes form a superficial plexus in layers I and II and a deep plexus in layer VI. These plexuses vary in density from area to area. All appear to arise from cortical or white matter cells rather than from extrinsic afferents. In some areas such as SI and areas 5 and 7, the superficial plexus extends deeply into layers III and IV; and in area 17, two very prominent middle plexuses occur in layers IIIB through IVB and in the upper one-third of layer V; these are separated by layer IVC, a major zone of thalamic terminations, which contains very few SRIF- or NPY-positive processes. The density of the plexuses is greater for NPY-positive processes than for SRIF-positive processes in all areas. In the rat, the plexuses do not display a strict laminar organization but generally are densest in the supragranular layers (I to III) and decline steadily in the deeper layers.(ABSTRACT TRUNCATED AT 400 WORDS)