bluuebunny/crossref_metadata_2025 huggingface dataset:
Immunohistochemical localization of choline acetyltransferase during development and in Chats mutants of Drosophila melanogaster
Authors: Gorczyca MG , Hall JC
Published: March 2018

The distribution of choline acetyltransferase (CAT) in the nervous system of Drosophila melanogaster was determined by indirect immunohistochemical procedures using a monoclonal antibody specific to the enzyme. Immunoreactivity was first detected in the nervous system of 16 hr embryos, and increased considerably by the end of embryogenesis. Neuropil was preferentially stained, though cell bodies could also be observed. Staining was prominent in the CNS of all 3 larval instars but decreased substantially during the mid-pupal stage. Prior to eclosion, the level of immunoreactivity increased and the adult staining pattern became discernible. In the adult brain, staining was extensive, with numerous structures, such as the optic lobes and mushroom bodies, staining strongly. The adult thoracic ganglia were also moderately immunoreactive. These results imply a wide distribution of cholinergic neurons in the CNS of Drosophila. Immunoreactivity was also determined for 2 temperature-sensitive CAT mutants, Chats1 and Chats2. These files exhibit reduced CAT activity at permissive temperature, 18 degrees C, which eventually falls to undetectable levels after incubation at nonpermissive temperature, 30 degrees C. Chats2 mutants, after incubation at either 18 or 30 degrees C displayed virtually no staining. This result indicated that the immunoreactivity observed in wild-type flies was specifically associated with the enzyme encoded by the Cha gene. The intensity of staining in Chats1 mutants incubated at 18 degrees C appeared greater than in control flies, even though CAT enzyme activity in Chats1 is lower. This suggests that the enzyme molecule itself is structurally altered in Chats1 mutants. After incubation at 30 degrees C, staining in Chats1 flies decreased but did not disappear.
Growth cone morphology varies with position in the developing mouse visual pathway from retina to first targets
Authors: Bovolenta P , Mason C
Published: March 2018

We have labeled the growth cones of retinal ganglion cell axons with HRP in intact mouse embryos. This has allowed us to visualize growth cone morphology during outgrowth along an entire CNS pathway from origin to target; to ask whether growth cone forms, and thus behaviors, differ at various points along the pathway; and to study the relationships of growth cones with the cellular environment. During the major period of axon outgrowth between embryonic day (E) 12 and 15, growth cones in the optic nerve are highly elongated (up to 40 microns) and have lamellopodial expansions, but the majority lack the microspikes or filopodia characteristic of many growth cones. Within the optic chiasm (E13–15), most growth cones shorten and spread, and project several short filopodia. In the optic tract, growth cones become more slender and again lack filopodia, resembling sleeker versions of optic nerve growth cones. Near the first target region (lateral geniculate nucleus), growth cones with filopodia arise from individual axon lengths and turn medially toward the target. Within target regions, the branches of immature axon arbors are tipped by minute swellings rather than by the enlarged growth cones prevalent during outgrowth toward targets. Electron-microscopic analysis of identified labeled growth cones in the optic nerve reveal intimate interactions between growth cones and glia or other growth cones in the form of invaginating contacts. In the optic nerve, growth cones contact immature glial (neuroepithelial) cells somewhere along their length, and also envelop bundles of neurites. In the chiasm, single growth cones simultaneously relate to many different profiles. These results demonstrate that in this single pathway from origin to targets, growth cone morphology varies systematically with position along the visual pathway. During outgrowth, simple growth cones are prominent when axons follow well-defined common pathways, and more elaborate filopodial forms appear when growth cones diverge, as they turn or come to decision regions. Together with observations in vitro and in nonmammalian nervous systems in situ, these data serve as reference points for testing to what extent growth cone form reflects intrinsic factors and interactions with the environment.
Numbers and proportions of GABA-immunoreactive neurons in different areas of monkey cerebral cortex
Authors: Hendry SH , Schwark HD , Jones EG , Yan J
Published: March 2018

The number and proportion of neurons displaying GABA immunoreactivity were determined for 50-micron-wide columns through the thickness of 10 areas of monkey cerebral cortex, including the precentral motor area (area 4), 3 cytoarchitectonic fields of the first somatic sensory area (areas 3b, 1, and 2), 2 areas of parietal association cortex (areas 5 and 7), the first and second visual areas (areas 17 and 18), area 21 of the temporal lobe, and areas of the orbital and lateral frontal cortex. Methods of fixation and immunocytochemical processing were designed to maximize the number of stained cells in 15-micron-thick frozen sections and 1-micron-thick plastic sections. In 8 of the 10 areas the number and proportion of GABA-immunoreactive neurons per 50-micron-wide column were found to be the same (34–43 cells/column; 25% of the total neuronal population). Areas 17 and 3b differed. Area 17 contained 50% more GABA-immunoreactive neurons (52–66 cells/column) but more than twice the total number of neurons, so that the GABA cells made up less than 20% of the total. In 3 monkeys, the number and proportion of GABA- positive neurons per 50-micron-wide column in area 3b were smaller than in adjacent areas of sensorimotor cortex (26–42 cells/column; 19–22%). In 2 other monkeys, the number and proportion (34–43 cells/column; 24– 26%) were the same as in adjacent areas. Despite the similarity among most areas of monkey cortex, within some areas, the number of GABA- positive neurons per 50-micron-wide column varied as much as 30%. These variations form a significant, repeating pattern only in area 18, where narrow bands (150–200 micron wide) of relatively few stained cells alternated with either narrow or wide bands (600–700 micron wide) in which columns contained more cells. The GABA-immunoreactive neurons were unevenly distributed across layers, with every area containing large numbers and proportions of stained cells in layer II, and every area but area 4 displaying a second concentration in the principal thalamocortical recipient layers. In area 4, the number of GABA- positive neurons declined sharply from layer II to layer III and remained low through layer VI. For areas displaying the greatest intra- areal variability, only 1 or 2 layers contributed significantly to that variability (layer IV in area 3b, layers III and V in area 18, and layers II and III in area 17).(ABSTRACT TRUNCATED AT 400 WORDS)
A rhodopsin gene expressed in photoreceptor cell R7 of the Drosophila eye: homologies with other signal-transducing molecules
Authors: Zuker CS , Montell C , Jones K , Laverty T , Rubin GM
Published: March 2018

We have isolated an opsin gene from D. melanogaster that is expressed in the ultraviolet-sensitive photoreceptor cell R7 of the Drosophila compound eye. This opsin gene contains no introns and encodes a 383 amino acid polypeptide that is approximately 35% homologous to the blue absorbing ninaE and Rh2 opsins, which are expressed in photoreceptor cells R1–6 and R8, respectively. Amino acid homologies between these different opsins and other signal-transducing molecules suggest an important role for the conserved domains of rhodopsin in the transduction of extracellular signals.
Interactions between growth cones and neurites growing from different neural tissues in culture
Authors: Kapfhammer JP , Raper JA
Published: March 2018

We have previously used retinal and sympathetic explants to show that growth cones recognize and retract from specific neurites in culture (Kapfhammer et al., 1986). In an effort to determine the generality of this phenomenon and to see how many different neurite labels can be detected by it, we have studied interactions between individual growth cones and neurites extending from a variety of neural sources in vitro. Using most of the possible pairings between sympathetic, ciliary, dorsal root ganglion (DRG), retinal, and diencephalic neurons, we have found that in most instances: (1) Growth cones do not retract from neurites originating from the same tissue; (2) retinal growth cones do not retract from diencephalic neurites; (3) sympathetic, ciliary, and DRG growth cones, with one possible exception, do not retract from sympathetic, ciliary, or DRG neurites; (4) retinal growth cones retract from sympathetic, ciliary, and DRG neurites; (5) sympathetic, ciliary, and DRG growth cones retract from retinal neurites; and (6) sympathetic growth cones retract from diencephalic neurites. A simple hypothesis consistent with these results is that 2 labels exist--one associated with central neurites and another associated with peripheral neurites-- and that peripheral growth cones are programmed to retract from the central label and central growth cones are programmed to retract from sympathetic, ciliary, and DRG neurites; (5) symevant to the separation of the CNS and PNS during development.
The major pelvic ganglion: androgen control of postnatal development
Authors: Melvin JE , Hamill RW
Published: March 2018

Previously we reported the effects of postnatal castration on the postorganizational development of the sympathetic hypogastric ganglion (Hamill and Guernsey, 1983; Melvin and Hamill, 1986). “Postorganization” implies an activational role for gonadal hormones, in contrast to the permanent organizing effects that occur perinatally. We now report results that suggest that the major pelvic ganglion (PG), a mixed parasympathetic and sympathetic ganglion, is similarly regulated by testosterone during development. Choline acetyltransferase (CAT) and tyrosine hydroxylase (T-OH) activities were used to examine normal PG ontogeny. The normal development of these biochemical indices occurs primarily after day 10. Postnatal castration at 10–11 d of age completely prevented the postorganizational developmental increase of T- OH activity. At 12 postoperative weeks T-OH activity in castrates was approximately 6% that of the control animals (control, 2880 +/- 127 pmol/ganglion X hr; castrated, 161 +/- 16 pmol/ganglion X hr; p less than 0.001). In fact, by only 1 postoperative week, T-OH activity was already significantly reduced in castrated animals (control, 480 +/- 69 pmol/ganglion X hr; castrated, 179 +/- 6 pmol/ganglion X hr; p less than 0.001). CAT activity and total ganglion protein were also significantly reduced by 1 postoperative week. In contrast to T-OH activity, however, these indices continued to develop at diminished rates. By 12 postoperative weeks CAT activity and total ganglion protein in castrates were 30 and 50% of control values, respectively, resulting in a significant developmental abnormality in CAT-specific activity. Testosterone replacement reversed the castration-induced developmental deficits of T-OH and CAT activities.(ABSTRACT TRUNCATED AT 250 WORDS)
Increased dopamine efflux from striatal slices during development and after nigrostriatal bundle damage
Authors: Stachowiak MK , Keller RW , Stricker EM , Zigmond MJ
Published: March 2018

Dopaminergic control over striatal targets appears to be retained in rats sustaining lesions of the nigrostriatal dopamine (DA) system as long as 5–10% of that projection remains. Similarly, during postnatal development, dopaminergic control over striatal neurons matures well before the innervation of striatum by the nigrostriatal bundle is attained. These observations suggest that enhanced efficacy of dopaminergic transmission may compensate for hypoinnervation of striatum after lesions or during development. To examine this hypothesis, striatal slices were superfused with Krebs bicarbonate buffer and effluent was collected and analyzed for endogenous DA. Electrical field stimulation (2 Hz) continuously delivered to slices prepared from intact adult rats increased DA efflux to 3–5 times the prestimulation rate within 10 min. Efflux then fell to approximately twice the basal rate over the next 20 min. DA efflux was also examined using slices prepared from adult animals given 6-hydroxydopamine 2–3 weeks earlier, and from 7–10-d-old rat pups. In each group, striatal DA levels were 10–40% of adult control values. Nevertheless, stimulated DA efflux from these slices attained the same rate as that observed with intact, adult slices. Thus, fractional DA efflux from these slices was several times higher than the control rate by the end of the stimulation period. This increased DA efflux appeared to be a consequence of both increased release and decreased reuptake of DA, as the fractional DA efflux from control striatal slices could not be increased to the rate seen in hypoinnervated slices using nomifensine (10 microM), an inhibitor of DA efflux.(ABSTRACT TRUNCATED AT 250 WORDS)
Sodium channels in normal and regenerated feline ventral spinal roots
Authors: Querfurth HW , Armstrong R , Herndon RM
Published: March 2018

Regenerated and remyelinated nerve fibers have shorter internodes and thus more nodes than normal mature fibers. This requires either a decrease in the number of sodium channels per node or an increase in the number of channels per fiber or both. The purpose of this investigation was to determine what happens to sodium channel number, as estimated by 3H-saxitonin (STX) binding, in regenerated fibers and to relate this to nodal number. Five adult cats underwent cryoaxotomy of ventral root levels L5, L6, L7, and S1 on the left side. After regeneration for 16–45 weeks, binding parameters were determined. On the right (control) side, binding was consistent with that in unoperated animals (b = 1.3, Bmax = 10.2 +/- 0.4 fmol/mg wet, Kd = 0.6 +/- 0.1 nM). However, the regenerated nerves showed a 3.5-fold increase in maximal binding (b = 1.3, Bmax = 36.1 +/- 0.5, Kd = 0.45 +/- 0.4). Computer-aided histologic analysis of the regenerated roots revealed a decrease in fiber size; a significant decrease in internodal length for fibers in a given size class; and a 1.35-fold increase in total fibers per root. These factors account for a 2.36-fold increase in nodes per milligram (wet). The number of STX binding sites per regenerated node was calculated to be 1.95 X 10(6) (1.31, 3.07, 95% confidence limits), whereas it was 1.26 X 10(6) (0.78, 2.02) for the control roots. The difference was not significant (p greater than 0.05). It is concluded that, in regeneration, the increase in nodal number is accompanied by an increase in sodium channels, so that the number of channels per node is normal or slightly increased. There is a marked increase in channels per fiber and an even greater increase in channels per anterior horn cell. The implications of these data for nodal reorganization in remyelination are discussed.
Physiological consequences of a peptide cotransmitter in a crayfish nerve-muscle preparation
Authors: Bishop CA , Wine JJ , Nagy F , O'Shea MR
Published: March 2018

The pentapeptide proctolin is colocalized with a conventional, conductance-increasing neurotransmitter in 3 of 5 excitatory motoneurons that innervate a posture-related tonic flexor muscle of the crayfish. It is released from these neurons in response to nerve impulses. Nanomolar concentrations of proctolin superfused on the tonic flexor muscle act postsynaptically to potentiate tension generated by a given level of depolarization. Proctolin alone has no detectable effect on muscle tension, nor does it alter the resting membrane potential of the muscle. Proctolin produces no detectable effect on the EPSPs of the 1 proctolinergic motoneuron that was examined. Neurally released proctolin can be selectively depleted from severed motor axons following prolonged, low-frequency stimulation; EPSPs reflecting conventional transmitter release are unaltered by this procedure. After proctolin depletion, tension generated by the motoneuron is greatly reduced. Taken together, these results indicate that the peptide secondary transmitter in this neuromuscular preparation is an important contributor to the magnitude of tension generated by the motoneuron, but since its effect is dependent on the depolarizing EPSPs of the conventional neurotransmitter, it does not contribute to the temporal aspects of tension generation. These aspects are controlled exclusively by the conventional neurotransmitter.
Origin of orientation-selective EPSPs in simple cells of cat visual cortex
Authors: Ferster D
Published: March 2018

According to current theory, orientation selectivity in cortical simple cells is critically dependent on intracortical synaptic inhibition. In particular, it is thought that IPSPs evoked by stimuli of the nonpreferred orientation are required to prevent a neuron from responding to a broadly tuned excitatory input at any but the preferred orientation. Yet EPSPs recorded in simple cells are in themselves highly orientation-selective. How is this possible, when excitation arises primarily from relay cells of the lateral geniculate nucleus (LGN), which are largely insensitive to orientation? In this paper, the properties of EPSPs are compared with the predictions of a model of geniculate excitation of simple cells. The model, which is derived from the suggestions of Hubel and Wiesel (1962), relies on the now familiar arrangement of the receptive fields of presynaptic geniculate cells into lines parallel to the axis of orientation of the postsynaptic cell. Several properties of the EPSPs observed in simple cells, and the orientation tuning of simple cells observed in extracellular experiments, can be accounted for without resorting to intracortical inhibition.
Immunological identification of a nicotinic acetylcholine receptor on bovine chromaffin cells
Authors: Higgins LS , Berg DK
Published: March 2018

Two probes previously shown to distinguish the nicotinic ACh receptor of chick ciliary ganglion neurons also recognize a component on the surface of bovine chromaffin cells in culture that displays the properties expected for the chromaffin nicotinic ACh receptor. The first probe is a monoclonal antibody, mAb 35, raised against ACh receptor from Electrophorus electric organ, and the second is an alpha- neurotoxin, Bgt 3.1, purified from B. multicinctus venom. mAb 35 binds specifically to a single class of high-affinity sites on the chromaffin cells in culture. Scatchard analysis indicates a KD of 2.1 +/- 0.2 nM for the binding and a Bmax of 1.6 +/- 0.1 X 10(4) mAb 35 sites per cell. The number of sites on the cells can be reduced through modulation by exposure of the cells to Bgt 3.1. The modulation can be blocked by the cholinergic ligands d-tubocurarine and carbamylcholine. Long-term exposure to the agonist carbamylcholine alone also reduces the number of mAb 35 binding sites. Bgt 3.1 inhibits nicotine-induced catecholamine release from the cells to the same extent and with the same concentration dependence that it modulates the number of mAb 35 sites on the cells. In addition, mAb 35 treatment of the cells causes a specific and almost complete blockade of nicotine-induced catecholamine release, apparently through a modulation of the receptor. These results indicate that the bovine chromaffin component recognized by mAb 35 and Bgt 3.1 is likely to be the nicotinic ACh receptor on the cells and that it has many similarities to ACh receptors on chick autonomic neurons.
Laminar and regional distributions of neurofibrillary tangles and neuritic plaques in Alzheimer's disease: a quantitative study of visual and auditory cortices
Authors: Lewis DA , Campbell MJ , Terry RD , Morrison JH
Published: March 2018

The number of Thioflavine S-positive neurofibrillary tangles (NFT) and neuritic plaques (NP) was determined in visual and auditory cortical regions of 8 patients with Alzheimer's disease. On both a regional and laminar basis, NFT exhibited very distinctive and consistent distribution patterns. The mean (+/- SEM) number of NFT in a 250-micron- wide cortical traverse was very low in area 17, primary visual cortex (0.9 +/- 1.0), increased 20-fold in the immediately adjacent visual association cortex of area 18 (19.7 +/- 3.6), and showed a further doubling in area 20, the higher-order visual association cortex of the inferior temporal gyrus (35.5 +/- 8.8). Similar differences in NFT number were present between primary auditory (1.6 +/- 0.5) and auditory association (18.9 +/- 5.4) regions. On a laminar basis, NFT were predominantly present in layers III and V, although there were striking regional differences in the proportion of NFT in these 2 layers. Layer III contained 79% of the NFT in layers III and V in area 18, 41% in area 20, and only 27% in area 22. In contrast, NP showed different, and less specific, regional and laminar distribution patterns. Total NP number was similar in the 3 visual areas, although there were marked regional differences in the type of NP present. Nearly 80% of the NP in area 17 was of the NPc type (i.e., contained a dense, brightly fluorescent core), whereas over 70% of the NP in both areas 18 and 21 was of the NPnc type (i.e., lacked a dense, brightly fluorescent core). NP were present in every cortical layer but were most numerous in layers III and IV. The distinctive distribution patterns of NFT are very similar to the regional and laminar locations of long corticocortical projection neurons in homologous regions of monkey neocortex. This association suggests that NFT reside in the cell bodies of a subpopulation of pyramidal neurons, namely, those that furnish long corticocortical projections. In contrast, the distribution patterns of NP suggest that multiple neuronal systems contribute to their formation.
Resolving GABAA/benzodiazepine receptors: cellular and subcellular localization in the CNS with monoclonal antibodies
Authors: Richards JG , Schoch P , Haring P , Takacs B , Mohler H
Published: March 2018

Monoclonal antibodies, raised against a purified GABAA/benzodiazepine receptor complex from bovine cerebral cortex, have been used to visualize the cellular and subcellular distribution of receptorlike immunoreactivity in the rat CNS, cat spinal cord, and bovine and postmortem human brain. Two different antibodies have been used for these studies; bd-17 recognizes the beta-subunit (Mr 55 kDa) in all the species tested, whereas bd-24 recognizes the alpha-subunit (Mr 50 kDa) of bovine and human but not rat and cat tissues. In bovine and human brain, both antibodies produced very similar staining patterns, indicating a homogeneous receptor composition, at least in the brain areas investigated. The general distribution and density of receptor antigenic sites in all tissues studied were very similar to that of benzodiazepine binding sites radiolabeled with 3H-Ro 15–1788 and of glutamate decarboxylase (GAD)-stained nerve terminals. The results demonstrate a very high receptor density (around neuronal cell bodies and processes or less discretely distributed) in the rat olfactory bulbs, cerebral cortex, ventral pallidum, islands of Calleja, globus pallidus, hippocampus, dentate gyrus, substantia nigra, geniculate nuclei, inferior colliculus, cerebellum, reticular formation, spinal cord, and retina. In contrast, no receptors could be detected in white matter, pineal, pituitary, adrenals, and superior cervical ganglia. Only among the cerebellar layers did we observe a conspicuous difference between the staining intensity and the radiolabeling. In bovine and postmortem human brain, e.g., hippocampus, dentate gyrus, cerebral cortex, and substantia nigra, the same close correlation between the immunohistochemical and radiohistochemical findings was observed. At the electron microscopic level, the immune reaction product in the rat substantia nigra and globus pallidus, for example, was localized to pre- and postsynaptic membranes of axodendritic and axosomatic synapses. Whether the presynaptic labeling represents GABA autoreceptors is discussed. In the near future, the monoclonal antibodies will be used in double-labeling experiments with GAD to identify those GABAergic projections that are modulated by benzodiazepine minor tranquillizers. Furthermore, they could also be used, in studies of postmortem human brain, to diagnose receptor dysfunction possibly associated with CNS disorders such as epilepsy.
Cell death during gangliogenesis in the leech: competition leading to the death of PMS neurons has both random and nonrandom components
Authors: Macagno ER , Stewart RR
Published: March 2018

The posteromedial, serotonin-containing (PMS) neurons are found in the ventral aspect of certain anterior segmental ganglia of adult leeches. With one exception, these cells are unpaired in all the ganglia where they are found. During early embryogenesis in Hirudo medicinalis, however, a bilateral pair of PMS neurons appears and differentiates in each of the 21 segmental ganglia (SG1-SG21). Over the next several days, one of the pair dies in SG3-SG21. Examination of the PMS neuron in any one of these segments reveals that either the right or the left cell remains, with equal probability, suggesting that the elimination of one of the pair is a random process. When unpaired PMS neurons are examined in pairs of adjacent ganglia, however, the cells are from opposite sides in the majority of cases (approximately 88%). This observation implies that the death of a PMS neuron in 1 ganglion strongly biases which member of the pair of PMS neurons degenerates in adjacent ganglia. Detailed examination of the sequence of degeneration shows that it begins at several separate loci in the nerve cord. We propose that the mechanism responsible for the death of one of the pair of PMS neurons in a segmental ganglion is competition between these 2 cells for some trophic factor, but that the outcome of this competition is predetermined if one of the PMS neurons in an adjacent ganglion has already begun to degenerate.
Glial-guided granule neuron migration in vitro: a high-resolution time- lapse video microscopic study
Authors: Edmondson JC , Hatten ME
Published: March 2018

To study neuronal migration, migrating granule neurons in microcultures prepared from early postnatal cerebellum have been analyzed with time- lapse, video-enhanced differential interference contrast microscopy. The morphology of migrating neurons resembles the elongated forms of migrating neurons described both in vivo and in vitro (Rakic, 1971; Hatten et al., 1984). The neuron closely apposes its soma along the glial fiber and extends a thickened leading process in the direction of migration. This leading tip is highly motile, with several filopodial extensions. Intracellular vesicular structures extend from the nucleus into the leading process of migrating neurons in vitro. Quantitation of the motions of migrating neurons revealed a saltatory pattern of advance along the glial fiber. Periods of cell soma movement at the rate of 56 +/- 26 micron/hr along the glial fiber are punctuated by periods during which the cell soma slows to a complete stop. The overall rate of migration is 33 +/- 20 micron/hr. The growing tip of the leading process rapidly extends and retracts, resulting in a net advance along the glial fiber. However, the periods of the extension and retraction of the leading process growing tip are not synchronized with the motions of the cell soma.
Local circuit interactions between oriens/alveus interneurons and CA1 pyramidal cells in hippocampal slices: electrophysiology and morphology
Authors: Lacaille JC , Mueller AL , Kunkel DD , Schwartzkroin PA
Published: March 2018

Electrophysiological and anatomical techniques were used to determine the role, in the hippocampal circuitry, of local circuit neurons located at the oriens/alveus border (O/A interneurons). Intracellular recording from these cells showed that their response characteristics were clearly nonpyramidal: high input resistance, short membrane time constant, short-duration action potential, pronounced, brief afterhyperpolarizations (AHP), and nondecremental firing during intrasomatic depolarizing current pulses. Intracellular Lucifer yellow (LY) injection and subsequent fluorescence microscopy confirmed their nonpyramidal nature. O/A interneuron somata were bipolar or multipolar; their dendrites projected mostly parallel to the alveus, except for 1 or 2 processes that turned perpendicularly, and ascended through stratum oriens and pyramidale and into radiatum. Their axons were seen to branch profusely in stratum oriens and pyramidale. Simultaneous intracellular recordings from O/A interneurons and CA 1 pyramidal cells showed that pyramidal cells directly excite these interneurons. Major hippocampal afferents also directly excited the O/A interneurons. In a small number of interneuron-pyramidal pairs, stimulation of the O/A interneuron directly inhibited pyramidal cells. In one case, reciprocal connections were observed: The pyramidal cell excited the interneuron, and the interneuron inhibited the pyramidal cell. In 1 interneuron-to- interneuron pair, an inhibitory connection from O/A interneuron to stratum pyramidale interneuron was also observed. With intracellular HRP injections into O/A interneurons and subsequent electron microscopy, we observed that O/A interneuron axons made contacts with pyramidal and nonpyramidal cells. HRP-filled symmetric synaptic contacts were found on pyramidal cell dendrites and somata. HRP-filled axons also made contacts with pyramidal cell initial segments. HRP- filled O/A interneuron axon contacts were also found on nonpyramidal cell dendrites in stratum oriens. These electrophysiological and anatomical results suggest that O/A interneurons make synaptic contact with pyramidal cells and may mediate feedforward and feedback inhibition onto CA 1 pyramidal cells.
Mechanisms of postsynaptic plasticity: remodeling of the junctional acetylcholine receptor cluster induced by motor nerve terminal outgrowth
Authors: Yee WC , Pestronk A
Published: March 2018

Motor nerve terminal outgrowth (NTO) at neuromuscular junctions (NMJs) occurs rapidly in response to denervation changes in muscle. We have previously found that NTO can produce an elongation of the synaptic area of the NMJ as defined by cholinesterase-silver staining. In the present study, we examined the effects of NTO on a postsynaptic muscle membrane component, the usually stable cluster of acetylcholine receptors (AChRs) at the NMJ. NTO was evoked in rat soleus muscles using botulinum toxin. AChRs were demonstrated using immunocytochemistry or autoradiography of alpha-bungarotoxin binding. Our results show that NTO induces rapid elongation of the cluster of AChRs at the NMJ within 7 d of treatment with botulinum toxin. The growth in the size of the AChR clusters was accompanied by an increase in the number of AChRs/NMJ. No elongation of AChR clusters was seen following surgical denervation, suggesting that cluster growth is related to NTO and not to denervation changes in muscle per se. Growth of NMJ-AChR clusters appeared to result primarily from 2 processes: insertion of new AChRs into the NMJ membrane and, surprisingly, redistribution of preexisting NMJ-AChRs. These results show that NTO can cause rapid changes in the normally stable cluster of AChRs at the NMJ. Motor nerve terminals provide a strong and anatomically precise control of AChRs at the NMJ.
Effect of neurotensin on neurons in the periaqueductal gray: an in vitro study
Authors: Behbehani MM , Shipley MT , McLean JH
Published: March 2018

The effect of neurotensin (NT) on periaqueductal gray (PAG) neurons was measured with extra- and intracellular recording methods in an in vitro preparation. Neurons excited by NT were heterogeneously distributed, being preferentially located in a region of PAG rich in NT-like fiber immunoreactivity. The majority of the responsive neurons were located in regions surrounding the aqueduct and the medial part of PAG. In 90% of the neurons, application of NT produced excitation that lasted for more than 2 min, while in the remaining cells, the excitatory effect lasted for less than 1 min. An inhibitory action of NT was rarely observed. Intracellular recordings showed that NT produced a depolarization leading to an increase in the spontaneous activity and multiple spiking with only a slight decrease in membrane resistance. The excitatory effect of NT was observed in neurons that were maintained in a solution containing cobalt. These results support the hypothesis that NT action on PAG neurons is due to the depolarization of the membrane and that this effect is mediated through a postsynaptic mechanism.
Characterization of big dynorphins from rat brain and spinal cord
Authors: Xie GX , Goldstein A
Published: March 2018

To examine the processing of products of the dynorphin gene in the central nervous system, immunoreactive (ir) dynorphin (Dyn) A, Dyn B, Dyn A-(1-8), alpha- and beta-neo-endorphin (alpha- and beta-Neo) in rat brain and spinal cord were measured, using specific antisera after gel filtration and high-performance liquid chromatography (HPLC). Three peaks of Mr about 8, 4, and 2 kDa for ir-Dyn A and ir-Dyn B, and one peak of Mr less than 2 kDa for ir-Dyn A-(1-8), ir-alpha-, and ir-beta- Neo were found both in the brain and in the spinal cord. The 8 kDa peak was recognized by Dyn A and Dyn B antisera and, after hydrolysis by proline-specific endopeptidase, by beta-Neo antiserum. The 8 kDa peak was recognized by a monoclonal antibody against the amino terminal sequence Tyr-Gly-Gly-Phe of all opioid peptides and by an antiserum directed toward the carboxyl terminus of Dyn B, indicating that it contains, from the amino terminal tyrosine of neo-endorphin to the carboxyl-terminal threonine of Dyn B, all 3 opioid peptide regions in the prodynorphin. By means of proline-specific endopeptidase hydrolysis, we also found a big dynorphin precursor (Mr approximately equal to 26 kDa) in both brain and spinal cord.
Modulation of the lobster pyloric rhythm by the peptide proctolin
Authors: Hooper SL , Marder E
Published: March 2018

The modulation of the pyloric network of the stomatogastric ganglion (STG) of the lobster Panulirus interruptus by the neuropeptide proctolin is described. First, the effects of proctolin on the pyloric motor patterns were characterized in terms of frequency and phase relations. Pyloric cycle frequency and lateral pyloric (LP) neuron activity increased and ventricular dilator (VD) neuron activity decreased with increasing concentrations (10(-9)-10(-6) M) of applied proctolin. Next, the effects of proctolin on the individual neurons that constitute the pyloric network were determined. Identified neurons were isolated from chemical and electrical presynaptic inputs by using pharmacological agents (Marder and Eisen, 1984a) and/or photoinactivation following Lucifer yellow injection (Miller and Selverston, 1979). Proctolin increased the amplitude and frequency of bursts produced by isolated pacemaker anterior burster (AB) neurons. Isolated LP and pyloric (PY) neurons responded to proctolin with increases in activity only when they were at or above threshold. All other pyloric neurons were unaffected. To determine how the direct effects of proctolin on isolated neurons resulted in the observed changes in frequency and phase relations in the motor pattern of the intact pyloric circuit seen in proctolin, individual neurons were deleted from the circuit. A comparison of proctolin's effects on isolated neurons with those on the intact network shows that the synaptic connectivity among neurons directly affected by proctolin and those unaffected by it shapes the network's response to proctolin.
Regional distribution of astrocytes with intense immunoreactivity for glutamate dehydrogenase in rat brain: implications for neuron-glia interactions in glutamate transmission
Authors: Aoki C , Milner TA , Sheu KF , Blass JP , Pickel VM
Published: March 2018

The principally mitochondrial enzyme glutamate dehydrogenase (GDH) exhibited low-intensity, uniform immunoreactivity in neurons and intense heterogeneous labeling of glial cells of rat brain. Simultaneous peroxidase labeling for GDH and immunoautoradiography for glial fibrillary acidic protein (GFAP) confirmed the astrocytic localization of the enzyme. Immunoreactivity in astrocytes, but not in neurons, required the presence of Triton X-100 as a solubilizing agent. Most of the intensely labeled glial processes were localized to regions previously reported as containing moderate to high densities of binding sites for the excitatory amino acids, L-glutamate or L-aspartate, and glutamatergic fibers. These included several forebrain regions, such as the superficial layers of the rostral neocortex, dorsal neostriatum, nucleus accumbens, septohippocampal nucleus, intralaminar thalamic nuclei, and external capsules. However, the central gray of the midbrain, the nuclei of the reticular formation, brain stem regions projecting to the cerebellum, and cranial nuclei of the trigeminal and vagal nerves also exhibited intense glial labeling for GDH, even though some of these regions are known to receive only weak glutamatergic projections. A second factor determining the distribution of GDH appeared to be neuronal activity, as assessed by correspondence with reported high densities of cytochrome oxidase. We conclude that GDH enriched in glial populations exists in a subcellular compartment distinct from that of neurons and may serve as one of the enzymes involved in glutamatergic transmission. Deficiencies of glial GDH and the consequent cytotoxic effects of high levels of excitatory amino acids may contribute to a number of neurodegenerative disorders.
On the role of cortical area V4 in the discrimination of hue and pattern in macaque monkeys
Authors: Heywood CA , Cowey A
Published: March 2018

Cortical visual area V4 in macaque monkeys has a large proportion of neurons that are sensitive to the wavelength or to the color of light. We tested its role in hue discrimination by removing it in macaque monkeys trained to discriminate small differences in hue. Hue discrimination thresholds were permanently elevated in 4 macaque monkeys in which V4 was removed bilaterally. In contrast, there was no impairment in achromatic intensity thresholds tested in an identical manner. However, the discrimination of pattern and orientation was also conspicuously impaired, indicating that area V4 is not concerned solely with processing information about wavelength. The multiple defect is consistent with evidence that V4 provides the major cortical visual input to the temporal lobe, where a large range of visual properties is registered. The performance of monkeys with V4 ablation was compared with that of unoperated control monkeys and monkeys with removal of cortex in the banks and floor of the rostral superior temporal sulcus (STS). Removal of STS had only slight effects on pattern discrimination and none of hue discrimination. To control for the possible effects of inadvertent damage to the visual radiations when removing V4, the lateral striate cortex was partially ablated bilaterally in a control monkey. This had no effect on any discrimination, despite producing more retrograde damage to the lateral geniculate nuclei than in any monkey with V4 ablation. The visual disorder following removal of visual area V4 strikingly resembles the clinical disorder of mild cerebral achromatopsia with associated apperceptive agnosia for objects and patterns.
Monoclonal antibodies specific for fixative-modified aspartate: immunocytochemical localization in the rat CNS
Authors: Madl JE , Beitz AJ , Johnson RL , Larson AA
Published: March 2018

Aspartate is a putative excitatory amino acid neurotransmitter that is widely distributed in the CNS. To study its distribution, monoclonal antibodies were produced against beta-L-aspartyl-L-aspartate (beta-Asp- Asp) conjugated to keyhole limpet hemocyanin (KLH) using glutaraldehyde- borohydride. Three monoclonal antibodies, Asp1-3, were obtained with high degrees of selectivity for aldehyde-fixed aspartate. The immunocytochemical staining pattern of rat CNS was found to be similar for all 3 antibodies but differed in some regions from staining patterns produced by Glu1, a monoclonal antibody with high selectivity for a form of amide-linked glutamate. Tissue staining produced by Asp1- 3 could be inhibited using aspartate conjugated to carrier proteins. Staining by Asp1 and Asp2 was also inhibited by free small molecules containing aspartate. Specificity of the 3 antibodies was evaluated by enzyme-linked immunoassay (ELISA) as follows: (1) reactivity of antibodies for conjugates of small molecules coated on ELISA plates; (2) ability of free small molecules to inhibit reactivity of antibodies for beta-Asp-Asp/KLH coated on ELISA plates; and (3) ability of conjugates to inhibit reactivity of antibodies for beta-Asp-Asp/KLH coated on ELISA plates. In all 3 types of assays, Asp1 and Asp2 displayed strong reactivity for small molecules and conjugates containing aspartate and little reactivity for small molecules and conjugates containing glutamate or GABA. Asp3 was highly reactive with conjugates containing aspartate using both directed and inhibition ELISA assays. For all 3 antibodies the precise staining pattern varied with the fixative used. Following glutaraldehyde fixation, dense immunocytochemical staining was observed in cerebral cortical neurons, some cerebellar granule cells, hippocampal pyramidal cells, and neurons of the inferior olivary nucleus. In addition, some putative GABAergic neurons, e.g., cerebellar basket and stellate cells, appeared to be stained. In general, acrolein fixation resulted in a more selective staining pattern in the CNS. For example, putative GABAergic neurons were no longer immunoreactive nor were hippocampal pyramidal cells.
Characterization of an inhibitor of neuronal plasminogen activator released by heart cells
Authors: Pittman RN , Patterson PH
Published: March 2018

A basic understanding of growth cone dynamics and developmental events involving growth cones requires an understanding of the function and regulation of molecules associated with and released by growth cones. Rat sympathetic neurons in culture release a urokinase-like plasminogen activator from their distal processes and/or growth cones (Pittman, 1985a). When sympathetic neurons are grown in cocultures with heart cells, however, plasminogen activator activity is not detected. The absence of plasminogen activator activity in cocultures of sympathetic neurons and heart cells appears to be due to the release of an inhibitor of plasminogen activator by heart cells. This inhibitor has a molecular weight of approximately 50 kDa in the presence of SDS and apparent molecular weights of approximately 50 and greater than 2000 kDa under native conditions. A significant fraction of the large- molecular-weight form of the inhibitor is converted to the smaller form following treatment with heparinase. Extremely stable complexes of 68 and 80 kDa are formed between the heart inhibitor and the plasminogen activator, urokinase, such that the complexes withstand boiling in SDS/mercaptoethanol. The data are consistent with the formation of an 80 kDa urokinase-inhibitor complex in the presence of heparan sulfate proteoglycan and a 68 kDa complex in the absence of heparan sulfate proteoglycan. A highly purified preparation of the heart inhibitor produces a 2- to 3-fold increase in neurite outgrowth from sympathetic neurons. These data indicate that the activity of the plasminogen activator released by sympathetic neurons can be regulated by a normal target tissue and that this regulation may result in increased neurite outgrowth from the neurons.
Light and electron microscopic localization of a cell surface antigen (NG2) in the rat cerebellum: association with smooth protoplasmic astrocytes
Authors: Levine JM , Card JP
Published: March 2018

Immunofluorescence and immunoperoxidase techniques were used to localize a cell surface chondroitin-sulfate proteoglycan antigen, termed NG2, in the developing and adult rat cerebellum. In the adult, both polyclonal and monoclonal anti-NG2 antibodies labeled cells throughout the cerebellar cortex, with the labeled cells being especially prominent in the molecular layer. The labeled cells had small, irregularly shaped cell bodies from which thin highly branched processes radiated in a stellate array. The NG2-labeled cells were not labeled with antibodies against glial fibrillary acidic protein (GFAP), vimentin, or S-100 protein, intracellular markers for astrocytes. However, electron microscopic immunocytochemical analysis of NG2 immunoreactive cells revealed a cell morphology consistent with that of protoplasmic astrocytes. Labeled cell bodies contained a thin rim of organelle-poor cytoplasm surrounding a euchromatic nucleus. Thick processes originating from the cell soma tapered to form thin branches with highly irregular surface contours that extended between adjacent neuronal elements. The labeled processes did not form synapses in the neuropil, and no synaptic profiles onto anti-NG2-labeled cell bodies or processes were observed. Thus, we conclude that the NG2 antigen is a cell surface marker for a class of smooth protoplasmic astrocytes. Immunoreactive cells were seen in the developing cerebellum beginning at embryonic day 16. The number of labeled cells increased during the early stages of cerebellar development, reaching a peak at about postnatal day (PND) 4 or 5 and declining thereafter. In the developing cerebellum, labeled cells lying within the forming molecular layer resembled the cells seen in the adult, whereas cells lying deeper within the folia had an immature appearance with fewer processes and less branching. This apparent gradient of morphological maturation suggests that an interaction with parallel fibers in the developing molecular layer may play a role in the terminal cytodifferentiation of the NG2-labeled smooth protoplasmic astrocytes.