bluuebunny/crossref_metadata_2025 huggingface dataset:
Incorporation of 3H-amino acids into proteins in a partially purified fraction of axoplasm: evidence for transfer RNA-mediated, post- translational protein modification in squid giant axons
Authors: Ingoglia NA , Giuditta A , Zanakis MF , Babigian A , Tasaki I , Chakraborty G , Sturman JA
Published: March 2018

Transfer RNA (tRNA) has been demonstrated to be present in axons of both invertebrates and the higher vertebrates, but nothing is known of its role in the metabolism of the axon. The present experiments were performed to determine whether tRNA functions in axons as a participant in post-translational protein modification of endogenous proteins. RNA was extracted from the axoplasm of squid giant axons and incubated with a variety of 3H-amino acids, aminoacyl-tRNA synthetases (obtained from squid optic lobe), and an appropriate reaction mixture. All of the amino acids tested were bound to an RNA fraction, but this reaction did not occur when samples were incubated in the presence of ribonuclease or in the absence of axoplasmic RNA. When radioactive RNA was chromatographed by polyacrylamide gel electrophoresis, the radioactivity comigrated with known tRNA markers, suggesting the presence of 3H-aminoacylated tRNA. Aminoacylation of RNA could also be demonstrated by incubating fresh axoplasm with labeled amino acids and a reaction mixture, minus exogenous aminoacyl-tRNA synthetases. These findings indicate the presence in axoplasm of a variety of species of aminoacyl-tRNAs as well as their corresponding synthetase enzymes. In the latter experiment no radioactivity was found associated with the protein fraction. This was also the finding when 3H-aminoacylated tRNA was either injected directly into the axon or incubated with extruded axoplasm. Thus, under the conditions described above, there is no evidence of transfer of amino acids from tRNA to proteins. In other experiments, axoplasm was pooled to a volume of 50 to 100 microliters, homogenized gently, and centrifuged at 150,000 X g for 1 hr. Some of the high speed supernatant was incubated with labeled amino acids and an appropriate reaction mixture, and the remainder was passed through an S-200 Sephacryl column before incubation with the same reaction mixture. There was no incorporation of amino acids into protein in the high speed supernatant fraction. However, in the S-200 purified fraction 3H-labeled Arg, Lys, Tyr, Leu, and Asp were all incorporated into proteins in amounts of 44, 30, 7, 5 and 3.5 times heat-inactivated controls. The reaction is not inhibited by Ca2+ or Ca2+-activated proteases, but appears to be dependent on the presence of tRNA. The addition of amino acids to protein is not protein synthesis since the reactions occurred in a partially purified fraction of the 150,000 X g supernatant, a fraction devoid of ribosomes and free amino acids.(ABSTRACT TRUNCATED AT 400 WORDS)
Synaptic control of excitability in isolated dendrites of hippocampal neurons
Authors: Masukawa LM , Prince DA
Published: March 2018

The apical dendrites of CA1 pyramidal cells were isolated from their cell bodies by making cuts through proximal stratum radiatum of transverse hippocampal slices from the guinea pig. This lesion separated the distal apical dendritic elements from the somata, basal dendrites, and 50 to 100 microns of the proximal apical dendritic tree. Orthodromic stimuli in stratum radiatum evoked excitatory synaptic responses in isolated dendrites, but no phasic inhibitory components could be detected. In spite of this surgically produced disinhibition, orthodromic stimuli did not elicit burst activity at the resting membrane potential. However, isolated dendrites and intact dendrites could generate multiple slow spike activity when directly stimulated with depolarizing current pulses. When isolated dendrites were depolarized by DC current, excitatory postsynaptic potentials could evoke subthreshold intrinsic slow depolarizations, or repetitive slow spikes, similar to responses elicited by depolarizing current pulses alone. After exposure to bicuculline (5 microns), both intact and isolated dendrites generated bursts of activity following synaptic activation. A possible mechanism for this action of bicuculline is blockade of a residual GABA-mediated inhibition which was not expressed as a postsynaptic hyperpolarization in isolated dendrites. This bicuculline-sensitive event was capable of depressing dendritic excitability in the absence of the recurrent inhibitory synaptic input and was very effective in controlling burst activity. Our results indicate that the dendritic electrical behavior is dependent on a complex interaction between synaptic and voltage-sensitive events.
Target selection by surgically misdirected optic fibers in the tectum of goldfish
Authors: Meyer RL
Published: March 2018

This study tested the capacity of regenerating optic fibers to read tectal markers and thereby grow to their appropriate tectal loci when initial position, optic pathway, and interfiber interactions are eliminated as useful cues. The stability of these markers with long- term optic denervation of the tectum was also examined. In adult goldfish optic fibers innervating lateroposterior optic tectum were dissected free of tectum and inserted into the medial anterior region of the opposite “host” tectum. Normally, fibers at this position either innervate medial anterior tectum or follow the medial division of the optic pathway into medioposterior tectum. Host tectum was denervated of all other optic fibers by enucleating its contralateral eye either at the time of the deflection or at various times up to 18 months prior to deflection. The regeneration of these deflected fibers into host tectum was examined by autoradiography and electrophysiology at 1 to 11 months later. At the insertion site deflected fibers split into two groups of roughly equal size. One group directly entered the optic layers of medial tectum and grew posterolaterally across the medial half of tectum into the lateral half. The second group followed an almost direct path to the lateral tectum, sometimes traversing through the deep cell layers of tectum in which optic fibers are not usually found. These fibers subsequently entered the optic layers at the lateral edge of tectum and grew posteriorly. This second path was not seen in controls in which optic fibers from medioposterior tectum were similarly deflected. Instead growth was almost entirely posteriorly directed. On the average by 1.5 months deflected lateroposterior fibers were preferentially distributed in the lateral half of the tectum. Densitometric measurements indicated nearly a 4-fold difference in lateroposterior compared with medial posterior labeling. By contrast, controls in which medial posterior fibers were deflected had 4 times more grains medially than laterally. There was also a posterior over anterior preference, but this was weak. There was no suggestion that long periods of optic denervation prior to deflection or long postoperative periods after deflection of lateroposterior fibers diminished the lateral over medial preference. These findings support the idea that stable tectal markers exist which are differentially read by medial and lateral optic fibers. However, in no case was the innervation by deflected fibers as selective as in the normal projection.(ABSTRACT TRUNCATED AT 400 WORDS)
Increased glucose metabolism during long-duration recurrent inhibition of hippocampal pyramidal cells
Authors: Ackermann RF , Finch DM , Babb TL , Engel J
Published: March 2018

The locally subnormal brain metabolism observed in some experiments utilizing the Sokoloff 2-deoxyglucose (2-DG) method has often been attributed to postsynaptic inhibition despite the fact that inhibitory postsynaptic potentials are themselves caused by energy-requiring mechanisms. To explore this issue, neurophysiologically confirmed long-duration recurrent inhibition of hippocampal pyramidal unit firing was induced by low frequency (2 to 4 Hz) stimulation of the fornix for 60 min following intravenous infusion of [14C]-2-DG. The resulting autoradiograms showed that long-duration suppression of pyramidal cell firing was accompanied by distinctly increased hippocampal 2-DG uptake, particularly in the stratum pyramidale, which contains a dense plexus of inhibitory interneuronal terminals upon pyramidal cells. Both the pyramidal inhibition and the increased 2-DG uptake were confined to the ipsilateral hippocampus in animals with previously severed fornices and hippocampal commissures. In a second series of rats, the excitatory entorhinohippocampal “perforant path” (PP) was stimulated at low frequency (2 to 9 Hz) following 2-DG administration. At 2 to 4 Hz, each PP stimulation resulted in a brief burst of pyramidal unit firing followed by short-duration firing suppression; this result was associated with paradoxically decreased 2-DG uptake in the ipsilateral stratum molecular. By contrast, 7 to 9 Hz entorhinal stimulation induced PP-mediated excitation immediately followed by powerful intrinsic hippocampal inhibition, evidenced by prolonged pyramidal unit suppression after each stimulation. This suppression was accompanied by increased 2-DG uptake in the dentate stratum molecular and hippocampal stratum pyramidale. Thus it appeared that even with entorhinal stimulation, hippocampal 2-DG uptake was more closely associated with long-duration recurrent inhibition than with transient pyramidal excitation. Therefore, although it still remains possible that regions of hypometabolism observed in some previous 2-DG studies may actually reflect mild inhibition, other mechanisms such as disfacilitation are more likely mechanisms for this metabolic pattern.
Regional variations in excitability of barnacle neurons
Authors: Krauthamer V , Ross WN
Published: March 2018

Optical recording techniques using voltage-sensitive dyes were used to examine the initiation and propagation of action potentials within neurons of the supraesophageal ganglion of the giant barnacle, Balanus nubilus. When a neuron was stimulated with current injection into the soma, the site of spike initiation, determined as the location with the earliest time-to-peak, was usually in the axon, 100 to 200 micron from the soma. The soma spike was broader and often later, suggesting that the cell body was less excitable than the axon. The action potential was narrowest at the site of initiation and then widened before propagating uniformly down the axon. In most cases, somatically stimulated action potentials and electrotonic pulses propagated into the dendrites with little delay or change of shape, indicating that the electrotonic length of these processes was great. Several different kinds of experiments indicated that some dendrites of these cells are excitable. These included the observations that (a) spikes could be made to initiate earlier in the dendrites than in the axonal region to which they were connected, and (b) action potentials sometimes decremented less than subthreshold pulses along dendritic processes. However, in other cases a decline in amplitude and a widening of the action potential demonstrated passive propagation into the dendrites, suggesting that not all dendrites are equally excitable.
Demonstration of retrogradely transported endogenous nerve growth factor in axons of sympathetic neurons
Authors: Palmatier MA , Hartman BK , Johnson EM
Published: March 2018

The presence of retrogradely transported endogenous nerve growth factor (NGF) in sympathetic nerves of the guinea pig was demonstrated directly by fluorescent and peroxidase immunohistochemistry in the ligated superior postganglionic nerve of the superior cervical ganglion. Fixed, frozen sections of previously ligated nerve were incubated with either rabbit antiserum against guinea pig NGF (gpNGF), rabbit antibodies against gpNGF purified on a mouse NGF (mNGF) affinity column, the portion of rabbit antiserum against gpNGF that did not bind to the mNGF affinity column, or nonimmune rabbit serum. Positive staining on the peripheral side of the ligation was obtained only with unfractionated antiserum against gpNGF and with purified antibodies against gpNGF. The staining properties of the various antiserum preparations correlated with their ability to block gpNGF- and mNGF-induced neurite outgrowth in the embryonic chick dorsal root ganglion bioassay and in the PC12 bioassay. Homogenates of superior postganglionic nerve supported growth of embryonic chick dorsal root ganglia and differentiation of PC12 cells. This support was blocked by the specific antisera against NGF used in the immunohistochemistry experiments. These experiments demonstrate that endogenous NGF, presumably released by peripheral target tissues, is retrogradely transported in vivo.
Reduction of experimentally induced neuronal death in spinal ganglia of the chick embryo by nerve growth factor
Authors: Hamburger V , Yip JW
Published: March 2018

Extirpation of the wing bud in 2-day chick embryos results in a conspicuous degeneration of neurons in both populations of brachial dorsal root ganglia (DRG). Daily injections of 1 to 6 micrograms of nerve growth factor (NGF), beginning at 4 1/2 days of incubation, rescued all small, late differentiating (DM) neurons and approximately 50% of large, early differentiating (VL) neurons, which would have died otherwise. The fact that NGF is an effective substitute for the hypothetical trophic maintenance factor for DRG which is normally produced by limb tissues strengthens our belief that NGF is identical with this factor. The control experiment, i.e., wing extirpation without NGF injections, revealed an inconsistency with previous data. Experiments on a number of different neuronal units had shown rather consistently that the period of experimentally induced neuron degeneration, caused by removal of the target, is synchronous with the period of normally occurring neuronal death in the same neuronal unit. This synchrony rule is violated by the VL population of brachial DRG. In this unit, the peak of degeneration resulting from wing bud extirpation occurs considerably earlier than the peak of normally occurring neuronal death. The competition hypothesis for the explanation of neuronal death had been based, in part, on the synchrony rule. We discuss the question of whether the deviation from the synchrony rule observed in our material represents a serious challenge to the competition hypothesis.
Dynamics of intracellular calcium and its possible relationship to phasic transmitter release and facilitation at the frog neuromuscular junction
Authors: Stockbridge N , Moore JW
Published: March 2018

We have developed a mechanistic model for intracellular influx, diffusion, and efflux of calcium, and we compare its predictions to the dynamics of transmitter release at the frog's motor nerve terminal. The model includes a square wave influx of calcium, a linear or saturable pump for the efflux of calcium, and slow diffusion of calcium within the terminal due to rapid equilibrium binding of the major portion of the influx to fixed, nonsaturable sites in the cytoplasm. Transmitter release is taken as proportional to the fourth power of the calcium concentration in a region within 100 A of the surface membrane. The model predicts phasic release of transmitter with a time course similar to that of the endplate current--it turns on with a lag, rises rapidly to a peak, and then declines more slowly. It also predicts facilitation, the increased transmitter release in response to a second stimulus for many milliseconds after the initial transmitter release has ceased. This facilitation is similar to that reported at the frog neuromuscular junction in its initial amplitude and time course.
Glial fibrillary acidic protein-like immunoreactivity in the iris: development, distribution, and reactive changes following transplantation
Authors: Bjorklund H , Dahl D , Olson L , Seiger A
Published: March 2018

Using immunohistochemistry with antisera raised against the glial fibrillary acidic protein (GFA), we have studied the appearance and distribution of GFA-like immunoreactivity in whole mounts of rodent iris and in sectioned cat and cow iris. In the adult rat iris, a dense plexus of GFA-positive fibers was seen in both the dilator plate and the sphincter. The fluorescent fibers formed large meandering bundles and a dense irregular network of thinner fibers. In the sphincter, mainly thinner fibers were seen. Thin fibers were also seen winding around blood vessels in the dilator plate. In adult mouse iris, the GFA- positive fibers had a quite different distribution with a few radially oriented fiber bundles superimposed on a more regular network of thinner fibers. Adult guinea pig irides showed still another pattern of GFA-positive fibers with a low number of bundles and thinner fibers forming a sparse irregular network. In thicker fiber bundles of all three rodent species, as well as at branching sites of the thinner fibers, negative or weakly fluorescent swellings surrounded by GFA-like immunoreactivity were present. These structures probably represent the cellular origin of the GFA-positive fibers. Thick, strongly fluorescent fiber bundles, as well as numerous thinner fibers, were seen in sections of cat and cow iris. Prenatally, fibers were visualized at embryonic day 18 in the rat. In these irides as well as in irides from 21-day-old embryos and 1-day-old pups, most fibers were organized in a gradually increasing system of thin meandering fiber bundles that showed limited branching. At postnatal day 6, a more mature network of thinner fibers had developed between the now more numerous fiber bundles. No obvious increase or decrease in the amount of GFA-positive fibers was seen in irides grafted to the anterior eye chamber of adult rat recipients examined 1 and 6 days after grafting. However, in these irides, as well as in the host irides, strongly fluorescent spider-like cells with short branching processes and a negative nucleus were seen. These cells were more numerous and more strongly fluorescent in grafted irides as compared to recipient irides and in the 6-day iris grafts as compared to the 2-day grafts. In all probability, the GFA-positive fibers and cells forming a network in adult irides from different species and in embryonic and grafted rat irides represent Schwann cells and their processes. The cellular origin of the spider-like cells in the iris grafts is less clear.
Rules of order in the retinotectal fascicles of goldfish
Authors: Stuermer CA , Easter SS
Published: March 2018

Individual fascicles of retinal axons were labeled in the goldfish tectum with horseradish peroxidase (HRP). The contralateral retina was later processed for HRP histochemistry to mark the cells that had axons in the fascicles. Labeled cells were found in a partial half anulus in ventral hemiretina, centered on the optic disk. The distance of the partial anulus from the disk depended on which tectal fascicle had been labeled; the more rostrocentral the fascicle, the smaller was the annular radius. The angular subtense of the partial anulus with respect to the disk depended on where (along its tectal course) the fascicle had been labeled; the more rostral the label site, the longer was the angular subtense. These results were interpreted in the context of retinotectal growth, and it was inferred that the axons followed two rules: (1) grow in along the edge of the tectum and (2) exit and terminate in order, axons from temporal retina first, nasal retina last. These rules would produce a retinotopic projection in peripheral tectum, but they require that some of the terminals already in place must shift as the tectum grows.
Does timing of axon outgrowth influence initial retinotectal topography in Xenopus?
Authors: Holt CE
Published: March 2018

The question of whether timing is involved in generating the topographic organization of the earliest embryonic projection from the retina to the tectum has been examined in Xenopus laevis. First, the normal schedule of axonal outgrowth from the retina to the tectum was characterized. Groups of axons originating from either dorsal or ventral extremes of the retina were labeled by in vitro incubation of sectors (one-quarter to one-third) of eye primordia in [3H]proline and their time courses of outgrowth were determined using light microscope autoradiography. Comparisons of the growth of dorsal and ventral nerve fiber populations showed that those from the dorsal retina leave the eye first, grow along the optic pathway, and reach the tectum roughly 6 hr ahead of those from the ventral retina. This stereotyped sequence of outgrowth schedules the development of the initial retinotectal map: first the ventrolateral tectum receives input from the dorsal retina (stage 37/38), and then the dorsomedial part receives input from ventral retina (stage 40). Second, to test whether the accurate timing of axon outgrowth and target invasion defines the spatial ordering of the earliest connections, the normal schedule of retinal fiber outgrowth was altered by substituting dorsal halves of young stage 21 eye primordia, labeled with [3H]proline, for those in older stage 27 embryos. These heterochronic transplants resulted in retarded outgrowth from the dorsal retina such that the original pioneer fibers reached the tectum at least 9 hr later than normal, arriving after ventral retinal fibers, thereby reversing the normal sequence of tectal invasion. Despite this, the initial pattern of tectal innervation remained spatially normal. It is concluded that the retinotectal map is generated not by the temporal sequence of retinal axon outgrowth but, rather, by selective means of neuronal recognition.
Axonal pathfinding in the absence of normal pathways and impulse activity
Authors: Harris WA
Published: March 2018

Retinal axons were challenged to grow to their targets both along abnormal pathways and in the absence of impulse activity. Eye primordia were first transplanted from normal to ectopic sites in axolotl embryos. Most of the hosts were genetically eyeless, others were enucleated normal embryos. These axolotl embryos were then parabiotically joined to California newt embryos. Both operations were completed by stage 28, which is before axons have left the eye. The result of the parabiosis was a paralysis of the “eyeless” axolotl twin due to the newt's tetrodotoxin (TTX), while the newt twin remained normally active. When the axolotl twin reached early larval stage, about 1 week later, the projection from the silent transplanted retina was assessed using horseradish peroxidase (HRP) injections into the retina, after which the animals were killed and prepared histologically to reveal the presence of HRP in neuronal processes. The results from 17 such cases show normal topographic retinotectal projections: the dorsal retina projecting to the ventrolateral tectum, and the ventral retina projecting to the dorsomedial tectum. Unusual pathways were often taken to achieve these destinations. Control animals, both normal axolotl larvae developing alone and normal axolotl larvae parabiosed to newts, also showed the normal retinotectal projection patterns. These results indicate that the retinal projections in the experimental group were basically normal. Thus, fibers need neither impulse activity nor a particular pathway to navigate to their correct targets during development. Both factors can be eliminated simultaneously, yet retinal axons still find their way to the tectum and make an ordered map. This indicates that other factors, such as the chemoaffinity mechanisms proposed by Sperry (Sperry, R.W. (1963) Proc. Natl. Acad. Sci. U.S.A. 50: 703–710), may play a more major role in axonal pathfinding in this system.
Regional differences in the neuronal cytoskeleton
Authors: Drake PF , Lasek RJ
Published: March 2018

Regional differences in the neuronal cytoskeleton were investigated in the giant neurons of Aplysia. Using SDS-PAGE, we have compared the proteins which comprise the cytoskeletons of cell bodies and axons. Separate populations of cell bodies and axons were collected and the proteins stained by the Coomassie brilliant blue method. Individual identified cell bodies, with long segments of their axons attached, were isolated, and the proteins were labeled with the [125I]Bolton- Hunter reagent. The proteins which are stably associated with the cytoskeleton were obtained by extracting the neuronal material in a physiological buffer containing Triton X-100. As a correlative measure to the biochemical analyses, electron microscopy was performed on the cell body and axonal fractions. Our results demonstrate that the composition and biochemical properties of the cytoskeletal proteins in the neuron cell bodies differ from those associated with axons. Specifically, the amount of neurofilament proteins, designated NF60 and NF65 , is 5 times more abundant in the axon than in the cell body. The relative amounts of actin and tubulin are comparable in these two regions of the neuron. In addition, the ratio of NF60 and NF65 is different in the cell body and axon. The cell bodies contain proportionally more NF60 than the axons. However, the physical properties of the tubulin in the cell body, as measured by relative solubility, differ from that of the axon. The substantial differences between the composition of the cytoskeleton of the cell bodies and axons of Aplysia suggests that at least two distinct cytoskeletal networks exist in these neurons, one specific for the cell body and the other specific for the axon.
Actions of procaine on specific nociceptive cells in leech central nervous system
Authors: Johansen J , Yang J , Kleinhaus AL
Published: March 2018

The effects of the local anesthetic, procaine, on the electrophysiological properties of the leech neurons responding to nociceptive stimuli (N cells) were examined in isolated segmental ganglia in Macrobdella . In the N cell situated laterally in ganglia 7 to 19, procaine produced a depolarization; whereas in the N cell situated medially, it caused a hyperpolarization. These changes in membrane potential were accompanied by a decrease in input resistance, persisted in solutions containing 20 mM MgCl2, and were reversible after drug washout. The depolarization induced by procaine in the lateral N cell was abolished in Na-free solutions. The hyperpolarization produced by procaine in the medial N cell persisted in low chloride solutions and was enhanced by about 30% when the K concentration was reduced to one-tenth of its control value. Therefore, it seems likely that an increase in the resting K conductance was a contributing factor to this hyperpolarization. Procaine greatly prolonged the action potential of the lateral but not the medial N cell. This effect was not solely a consequence of the membrane potential change produced by the drug. Ganglia 5, 6, 20, and 21 contained two rather than four cells with N-like properties. These neurons were N-like by virtue of the shape of their action potential and their morphological similarities to the N cells found in ganglia 7 to 19, as well as by their selective sensitivities to nociceptive mechanical stimulation of the skin. The cells in ganglia 5 and 6 responded to gut stimulation, as did the medial N cells in ganglia 7 to 19.(ABSTRACT TRUNCATED AT 250 WORDS)
Carbon dioxide sensitivity of dye coupling among glia and neurons of the neocortex
Authors: Connors BW , Benardo LS , Prince DA
Published: March 2018

Among neocortical astrocytes and neurons, intracellular injection of the fluorescent dye Lucifer Yellow CH into single cells will often label multiple adjacent cells. It is possible that this intercellular dye movement occurs through gap junctions, which in several systems are sensitive to cytoplasmic acidification. In the present study we tested the effect of increased CO2 levels, a treatment expected to decrease intracellular pH (pHi), on the prevalence of glial and neuronal dye coupling in neocortical slices. When CO2 levels were raised to 40% or 50%, dye coupling among astrocytes was completely abolished. Under the same conditions, the prevalence of dye coupling among neocortical neurons of adult guinea pigs was significantly reduced to 18.7%, compared to the control level of 32.3%. Dye coupling among immature rat neocortical neurons, which normally occurs at relatively high rates, was not measurably affected by CO2 levels up to 50%. The results suggest that coupling between cortical glia is very sensitive to cytoplasmic acidification. The relative insensitivity of neuronal coupling to CO2 may indicate either that the conductance of neuronal gap junctions is only weakly affected by pHi or that neuron-to-neuron dye passage occurs via other types of intercytoplasmic pathways.
Identification and characterization of voltage-sensitive calcium channels in neuronal clonal cell lines
Authors: Freedman SB , Dawson G , Villereal ML , Miller RJ
Published: March 2018

Voltage-sensitive calcium channels ( VSCCs ) have been identified in three clonal cells. These are the neuroblastoma X Chinese hamster brain hybrid ( NCB -20), the neuroblastoma X glioma hybrid (NG108–15), and the neuroblastoma ( N4TG1 ). Depolarization of NCB -20 cells with 50 mM KCl or 50 microM veratridine (VE) produced a 2- to 3-fold increase in net 45Ca2+ uptake. In NCB -20 cells, this voltage-sensitive 45Ca2+ uptake was inhibited selectively by organic calcium antagonists such as nitrendipine, cinnarizine, verapamil, and diltiazem (IC50 values = 6.4, 750, 1800, and 4500 nM, respectively). High K+-induced uptake was unaffected by 4-aminopyridine, tetraethylammonium, and tetrodotoxin (TTX), whereas VE-induced 45Ca2+ uptake was completely blocked by 3 microM TTX. In contrast to NCB -20 cells, NG108–15 cells showed a much smaller response to depolarizing stimuli. Following differentiation of NG108–15 cells by chronic treatment with 10 microM prostaglandin E1 and 50 microM 3-isobutyl-1-methylxanthine, depolarization induced a large increase in voltage-sensitive 45Ca2+ uptake. This induction was apparent after 24 hr and increased linearly for 96 hr. VSCC activity was also induced by 1.5% dimethyl sulfoxide and by other agents that increase intracellular cAMP, such as forskolin (1 microM) and cholera toxin (1 microgram/ml). Voltage-sensitive 45Ca2+ uptake in differentiated NG108–15 cells was inhibited by nitrendipine, D-600, and diltiazem (IC50 values = 7, 690, and 1600 nM). Our results suggest that VSCCs in neuronal clonal cell lines can be altered by cellular differentiation. In contrast to those VSCCs involved in neurotransmitter release, the VSCCs described here appear to be blocked by organic calcium channel antagonists at very low concentrations.
Calcium/calmodulin-dependent protein phosphorylation in the nervous system of Aplysia
Authors: DeRiemer SA , Kaczmarek LK , Lai Y , McGuinness TL , Greengard P
Published: March 2018

An afterdischarge in the bag cell neurons of Aplysia was previously shown to be associated with calcium entry into these cells and with changes in the phosphorylation state of at least two bag cell proteins (BC-I and BC-II). We have now investigated the role of calcium plus calmodulin (Ca/CaM) in the control of phosphorylation of Aplysia nervous system proteins, including those of the bag cell neurons. In cell-free preparations of Aplysia CNS, we demonstrated Ca/CaM- stimulated protein phosphorylation that could be inhibited by the calmodulin-blocking drugs R24571 , trifluoperazine, chlorpromazine, and W7 . A number of substrate proteins for Ca/CaM-dependent protein phosphorylation with Mr values from 17,000 to 310,000 were consistently observed in homogenates of the Aplysia CNS. In the bag cells, we found that a major substrate for Ca/CaM-dependent protein phosphorylation was the bag cell-specific, Mr = 21,000 protein (BC-II). BC-I (Mr = 33,000), on the other hand, appeared not to be a substrate for a Ca/CaM- dependent protein kinase. We found that there are a minimum of two Ca/CaM-dependent protein kinases in the Aplysia nervous system. These enzymes were distinguished on the basis of their subcellular distribution and their ability to phosphorylate distinct sites on synapsin I, an exogenous neuronal protein from vertebrates. Phosphorylation by one of these kinases (calmodulin kinase I) was on a site recovered in an Mr = 10,000 proteolytic fragment of synapsin I, and phosphorylation by the other (calmodulin kinase II) was on a site recovered in an Mr = 30,000 fragment. The predominant enzyme in the Aplysia CNS, as in the mammalian nervous system, was calmodulin kinase II.(ABSTRACT TRUNCATED AT 250 WORDS)
Analysis of the apparent biphasic axonal transport kinetics of fucosylated glycoproteins
Authors: Goodrum JF , Morell P
Published: March 2018

Following intraocular injection of [3H]fucose, which labels many glycoproteins of retinal ganglion cells, the accumulation of transported radioactivity arriving at the superior colliculus (nerve terminals) peaks within a few hours and decays with a time course of hours. Then, over a period of several days, radioactivity again accumulates at the superior colliculus and then decays with a half-life of days. The second peak also represents fast transported material since it occurs almost simultaneously along the optic nerve and tract as well as at the nerve endings. Such data have been interpreted as evidence for both a group of rapidly released, rapidly transported glycoproteins (first peak) and a group of slowly released but rapidly transported glycoproteins (second peak). We investigated this supposition by studying in more detail the metabolism of some individual fucosylated proteins in both the retina and superior colliculus. We noted that much of the radioactivity incorporated in fucosylated glycoproteins at the retina was rapidly metabolized (with a turnover on the order of hours), while the remainder of the fucosylated moieties had a metabolic half-life on the order of days. This was also true of the metabolic behavior of several individual glycoproteins, selected for study because they are major components of the group committed to transport and accumulating in two waves at the superior colliculus. In other experiments we injected [35S]methionine intraocularly and examined the metabolism in the retina and the kinetics of transport to the superior colliculus of the peptide backbone of these same individual proteins. In contrast to the two waves of accumulation of radioactivity from [3H]fucose, accumulation of radioactivity of the peptide backbone of the same glycoproteins was monophasic. Our explanation of these data involves the presence of two types of fucose moieties on the peptides. One group of fucose moieties is labile and is lost from the peptide backbone over a period of hours. Other fucose moieties are approximately as metabolically stable as the peptide backbones to which they are attached. The actual peptide backbones of the glycoproteins are committed to rapid transport over a period of several days. Thus, the first (and most prominent) peak of transported radioactivity in [3H]fucosylated glycoproteins does not represent a discrete phase of transport but, rather, is the summation of kinetics of gradual arrival of proteins and the rapid drop in their specific radioactivity as the more labile moieties of [3H]fucose are lost.(ABSTRACT TRUNCATED AT 400 WORDS)
Substance P-containing pathways to avian sympathetic preganglionic neurons: evidence for major spinal-spinal circuitry
Authors: Davis BM , Cabot JB
Published: March 2018

The origin of substance P-like immunoreactivity (SPLI) in the neuropil of sympathetic preganglionic neurons (SPNs) was investigated in pigeon (Columba livia). Previous investigators had suggested that a major SPLI- containing projection to SPNs arose from caudal brainstem (Johansson, O., T. Hokfelt, B. Pernow, S. L. Jeffcoate, N. White, H. W. M. Steinbusch, A. A. J. Verhofstad, P. C. Emson, and E. Spindel (1981) Neuroscience 6: 1857–1881; Gilbert, R. F. T., P.C. Emson, S. P. Hunt, G. W. Bennett, C. A. Marsden, B. E. B. Sandberg, H. W. M. Steinbusch, and A. A. J. Verhofstad (1982) Neuroscience 7: 69–87; Helke, C. J. J. J. Neil, V. J. Massari, and A. D. Loewy (1982) Brain Res. 243: 147–152) and, furthermore, that the bulbospinal fibers and terminals in the SPN neuropil which contained SPLI also contained serotonin-like immunoreactivity (5-HTLI) (Johansson, O., T. Hokfelt, B. Pernow, S. L. Jeffcoate, N. White, H. W. M. Steinbusch, A. A. J. Verhofstad, P. C. Emson, and E. Spindel (1981) Neuroscience 6: 1857–1881; Gilbert, R. F. T., P.C. Emson, S. P. Hunt, G. W. Bennett, C.A. Marsden, B. E. B. Sandberg, H. W. M. Steinbusch, and A. A. J. Verhofstad (1982) Neuroscience 7: 69–87). In the present study, various spinal lesions were made and the SPLI and 5-HTLI content of thoracic spinal cord was examined using immunohistochemistry. Interruption of descending bulbospinal fibers by cervical hemisection had no demonstrable effect on SPLI in the SPN neuropil, while 5-HTLI was almost totally depleted in the half of the spinal gray (including the SPN cell column) ipsilateral to the hemisection. Following a complete thoracic transection, SPLI was only depleted 2 to 3 mm rostral and caudal to the lesion, while normal SPLI staining was present in the remainder of the spinal cord. 5-HTLI was totally depleted caudal to a thoracic transection. Dorsal rhizotomy of three to six sequential spinal segments significantly depleted SPLI in the dorsal horn but had no effect on SPLI in the region of SPNs. Major depletion of SPLI within the SPN cell column was only seen when portions of thoracic spinal cord were isolated by complete transections or unilateral hemisections. Finally, evidence was found for intraspinal SPLI-containing fiber systems. These results demonstrate that the majority of SPLI in the SPN cell column in the pigeon is probably of intraspinal origin. The data also confirm that 5-HTLI is contained in axons and terminals arising from cell bodies of supraspinal origin.
Afferent specific localization of muscarinic acetylcholine receptors in cingulate cortex
Authors: Vogt BA
Published: March 2018

The laminar distribution of acetylcholine receptors in rat cingulate cortex and their localization to axons of neurons in the anterior thalamic nuclei (ATN) were evaluated with the muscarinic antagonist [3H]propylbenzilylcholine mustard (PrBCM) in vitro. Specific binding of PrBCM in granular area 29 was heterogeneous, with a 57% variation from the highest binding in layer Ia to the lowest in layer II-III. In contrast, binding in area 24 was homogeneous, with only a 14% variation. The heterogeneity of PrBCM binding almost exactly duplicated the distribution of termination of ATN afferents in layers I to IV of area 29c. Four experiments indicated that 50% of the excess binding in layers Ia and IV was due to axonal receptor sites. First, ATN lesions abolished 41% and 27% of total specific binding in layers Ia and IV, respectively. Second, an undercut procedure that totally deafferented layers I to Va showed changes similar to those following ATN lesions, suggesting that other afferents to these layers may not have muscarinic receptors associated with them. Third, the sequence of losses in receptor binding and acetylcholinesterase (AChE) activity was evaluated 2, 3, 5, 9, and 14 days following ATN lesions. Since AChE was present in ATN axons, as evidenced by early postlesion losses, the correlation of both losses as well as previous analyses of axon degeneration in this cortex confirmed that these receptors were in axons. Fourth, binding peaks in layers Ia and IV remained in area 29c following destruction of virtually all neurons with the neurotoxin ibotenic acid. This is the first evidence that the activity of a major neocortical thalamic afferent may be regulated by axonal acetylcholine receptors.
Temperature effects on spontaneous and evoked quantal size at the frog neuromuscular junction
Authors: Van der Kloot W , Cohen IS
Published: March 2018

The amplitudes, time integrals, and half-decay times of miniature endplate currents (MEPCs) and of endplate currents (EPCs) were measured in preparations in low Ca2+-high Mg2+ Ringer, in which the probability of the evoked release of more than one quantum was low. The measurements were made at 5 to 7, 11, and 15 degrees C. There is no consistent difference in these properties of evoked and spontaneously released quanta at any of the temperatures. I. S. Cohen and W. Van der Kloot ((1983) J. Physiol. (Lond.) 336: 335–344) compared the effects of temperature changes on MEPCs and on the endplate currents evoked by set amounts of iontophoretically applied ACh. The results suggested that the quantity of ACh/quantum was relatively unaffected by changes in temperature. This conclusion can now be extended to quanta released by nerve stimulation.
Localization of active sites along the myelinated goldfish Mauthner axon: morphological and pharmacological evidence for saltatory conduction
Authors: Funch PG , Wood MR , Faber DS
Published: March 2018

njections of Lucifer Yellow (LY) and horseradish peroxidase (HRP) were made within the myelin sheath of the goldfish Mauthner axon to determine the domains of individual oligodendrocytes. Long segments of the myelin sheath were stained with both markers. The lengths and locations of these sheath segments were analyzed in whole mount preparations (LY) or in reconstructions of serial vibratome sections (HRP). The termination sites of individual myelin sheaths, relative to gross anatomical landmarks of the brain, were consistent within and between all fish studied. In particular, the average locations of the termination sites were separated by 2.2 to 2.6 mm and corresponded to the brain regions where active site foci have been previously localized electrophysiologically. Individual sheath segments generally spanned the entire distance between adjacent active sites. The node-internode- node structure of the Mauthner axon that is suggested by these findings was further tested by ejecting tetrodotoxin (TTX) at various discrete rostral-caudal locations just outside the fiber. Large all-or-nothing components of the antidromic action potential were rapidly blocked (within seconds) only when the TTX ejections were made within a few hundred micrometers of the active site foci. The amplitudes of these blocked components are also consistent with predictions based upon previous electrophysiological analyses which demonstrated an active site spacing of 2.2 to 2.6 mm, a space constant of 5.0 mm, and a safety factor of 6 for impulse propagation. It is concluded from these morphological, pharmacological, and electrophysiological observations that the Mauthner axon possesses nodes separated by 2.2 to 2.6 mm and that a single oligodendrocyte spans an internodal region. Although nodal ultrastructure remains to be described, these results rule out the possibility that each of the short (approximately 50 micron), closely spaced (average separation = 155 micron) axon collaterals is a site of action current production.
The histochemical localization of cytochrome oxidase in the retina and lateral geniculate nucleus of the ferret, cat, and monkey, with particular reference to retinal mosaics and ON/OFF-center visual channels
Authors: Kageyama GH , Wong-Riley MT
Published: March 2018

The histochemical localization of cytochrome oxidase within the normal retina and lateral geniculate nucleus (LGN) of cats, ferrets, and monkeys revealed that distinct layers, types of cells, and portions of neurons are more intensely stained than others. The dark staining of photoreceptor inner segments and cone pedicles and the light staining of photoreceptor outer segments, somata, and rod spherules demonstrates that different segments of the same cell may have disparate but distinct levels of oxidative enzyme activity. In tangential sections of retina, regular mosaic arrays were evident for each of several darkly reactive retinal components, such as cone inner segments, cone pedicles, and horizontal cells. In the cat and ferret, regular mosaic arrays were also formed by metabolically distinguishable populations of ganglion cells. Ia and IIa ganglion cells (OFF-; Nelson, R., E. V. Famiglietti, Jr., and H. Kolb (1978) J. Neurophysiol. 41: 472–483) were more darkly reactive than the other classes. The darker staining of sublamina a of the inner plexiform layer (OFF-; Famiglietti, E. V., Jr., and H. Kolb (1976) Science 194: 193–195) in the cat and ferret retina, as well as sublamina A′ and A1′ of the ferret LGN (OFF-; Stryker, M.P., and K.R. Zahs (1983a) J. Neurosci. 3: 1943–1951) suggest that, under typical rearing conditions, the OFF-channels may be metabolically more active than the ON-channels in these species. In Macaca and Saimiri, darker staining was observed in sublamina b of the inner plexiform layer (ON-; Famiglietti, E.V., Jr., and H. Kolb (1976) Science 194: 193–195) and laminae 1, 2, and 6 of the LGN, implying that, under similar rearing conditions, a different pattern is observed. The dark staining of many large retinal ganglion cells, as well as most of the larger LGN neurons (presumed Y/Y-like), in all species studied is evidence that the Y/Y-like pathway is also highly active.
Critical periods for effects of monocular deprivation: differences between striate and extrastriate cortex
Authors: Jones KR , Spear PD , Tong L
Published: March 2018

The critical period of susceptibility to effects of monocular deprivation was compared in striate cortex and the lateral suprasylvian (LS) visual area of cortex. Twenty-three cats received monocular lid suture for a period of 4 weeks beginning at 4, 12, 18, 26, or 35 weeks of age or as adults. Immediately following the deprivation, single cell recordings were carried out in both cortical areas of each cat. Recordings also were made from five normally reared control cats. For both striate and LS cortex, early monocular deprivation had marked effects on neuronal ocular dominance, including an increased percentage of cells dominated by the nondeprived eye, a decreased percentage of cells dominated by the deprived eye, and a decreased percentage of binocularly driven cells. In both cortical areas, these effects were maximal in animals deprived at 4 weeks of age. Both areas then showed similar monotonic declines in effects of the deprivation following onsets from 4 to 18 weeks of age. However, in older animals there were clear differences in the effects of monocular deprivation on LS and striate cortex. In LS cortex, the monotonic decline in effects continued until 26 weeks of age, and no significant abnormalities were present in animals deprived at 26 weeks of age or older. In striate cortex, however, the effects of monocular deprivation remained relatively constant following onsets from 18 to 35 weeks of age, and significant abnormalities in all measures of ocular dominance were present when deprivation was begun as late as 35 weeks of age. Within- animal comparisons indicated that the greater effects of monocular deprivation on striate cortex than on LS cortex were present in every cat deprived at 26 or 35 weeks of age. Neither cortical area showed significant abnormalities following monocular deprivation in adult cats. These results indicate that the critical period for effects of the same regime of monocular deprivation is over sooner in LS cortex (between 18 and 26 weeks of age) than in striate cortex (after 35 weeks of age). This observation has important implications for an understanding of the sites and mechanisms of effects of visual deprivation and the mechanisms that control critical periods of development.
Neural control of hatching: fate of the pattern generator for the leg movements of hatching in post-hatching chicks
Authors: Bekoff A , Kauer JA
Published: March 2018

Chicks from 0 to 61 days post-hatching were gently folded into the hatching position and placed in artificial glass eggs. Within 0 to 2 min they began to produce a behavior that qualitatively resembled normal hatching. Furthermore, quantitative analysis of electromyographic records showed that under these conditions the intralimb and interlimb leg motor output patterns produced during each hatching episode (the episode motor program) were those typical of normal hatching. The only major change associated with increasing post- hatching age was a gradual increase in inter-episode interval. Therefore, we conclude that the neural pattern-generating circuitry which produces the motor program for the leg movements of hatching remains functional in post-hatching chicks despite the fact that, under normal conditions, hatching behavior is never used again.