bluuebunny/crossref_metadata_2025 huggingface dataset:
Structure and physiology of developing neuromuscular synapses in culture
Authors: Takahashi T , Nakajima Y , Hirosawa K , Nakajima S , Onodera K
Published: March 2018

The structure and function of developing neuromuscular synapses in culture have been investigated. We used neuromuscular junctions formed by coculturing dissociated muscle cells and dissociated neurons obtained from Xenopus embryos. After recording nerve-evoked endplate potentials (e.p.p.s) and spontaneously occurring miniature endplate potentials (m.e.p.p.s) from a given junction, the same specimen was investigated for electron-microscopic histology. We surveyed almost the total area of the junctional region by making serial sections. Even in preparations cocultured for only a short time (4–11 hr), both e.p.p.s and m.e.p.p.s could be obtained. The junctional region of these early synapses revealed a simple structure. The presynaptic terminals contained smooth-surfaced clear vesicles, but there were no presynaptic specializations such as active zones. The width of the synaptic cleft was variable, with predominance of narrow regions (10–30 nm), and there was no basal lamina inside the cleft. When the coculture time was 1 d or longer, the junctional area started to show structural features resembling a mature neuromuscular synapse. In the presynaptic terminal there were active zones, consisting of the presynaptic density and an accumulation of vesicles near the density. In many junctions, the postsynaptic membrane showed densities and thickenings, with a widened synaptic cleft, that contained basal lamina. It is known that growth cones, prior to making neuromuscular junctions, can release the transmitter substance with a very long latency if stimulated repetitively. In contrast, e.p.p.s with short latencies can be evoked by single stimuli soon after the growth cones attach to muscle cells. However, our data did not reveal any structural changes to account for such functional changes.(ABSTRACT TRUNCATED AT 250 WORDS)
Spatial and temporal selectivity in the suprasylvian visual cortex of the cat
Authors: Zumbroich TJ , Blakemore C
Published: March 2018

We recorded from single units in the medial and lateral banks of the posterolateral suprasylvian visual cortex (PMLS/PLLS) of the cat. The responses to drifting high-contrast gratings of optimum orientation and direction of motion, but varying in spatial and temporal frequency, were examined quantitatively for a sample of cells, whose receptive fields covered a wide range of eccentricities. The optimum spatial frequencies (average about 0.2 cycles/deg) were low compared to the values reported for striate cortex but similar to those for area 18. The mean spatial bandwidth (about 2 octaves) was slightly broader than that of cells in other cortical visual areas. The cut-off spatial frequencies (“acuities”) covered a wide range, from 0.05 to 2.1 cycles/deg, similar to those of cells in area 18. Responses to drifting sinusoidal gratings were usually dominated by an unmodulated elevation of discharge, although some modulation occurred at the temporal frequency of drift, especially at low spatial frequencies. Modulated responses were relatively stronger in PMLS than in PLLS. For those cells that responded to flashed stimuli, stationary, contrast-modulated gratings presented at different spatial positions typically evoked small responses at the fundamental frequency (dependent on spatial phase) and a larger component at the second harmonic of temporal frequency, with no overall “null-position.” The optimum spatial frequency was usually higher than would be predicted by simple summation within the dimensions of the receptive field. Thus, neurons in PMLS and PLLS, like complex cells in areas 17 and 18, behave nonlinearly and their spatial selectivity is determined by “subunits” smaller than their receptive fields. The range of preferred temporal frequencies ranged from less than 2.5 Hz to more than 10 Hz. In their temporal selectivity neurons in PMLS resembled cells in area 17, with little attenuation at low temporal frequencies, whereas there was a tendency for cells in PLLS to prefer higher temporal frequencies, as is common in area 18.
Characterization of somatomedin/insulin-like growth factor receptors and correlation with biologic action in cultured neonatal rat astroglial cells
Authors: Han VK , Lauder JM , D'Ercole AJ
Published: March 2018

The role of somatomedin/insulin-like growth factors (Sm/IGFs) in neural growth and development is not clearly defined. To characterize Sm/IGF receptors and to correlate binding with the biologic actions of Sm/IGFs in a homogeneous population of neural cells, we isolated and studied a nearly pure population of cultured astroglial monolayers derived from cerebral cortices of neonatal rats. Binding of radiolabeled Sm/IGFs was specific, saturable, and reversible, with 90% of the binding occurring within 6 hr of incubation at 4 degrees C. Competitive binding studies with Sm-C/IGF I yielded curvilinear Scatchard plots, while studies with IGF II and multiplication stimulating activity (MSA) yielded linear plots, suggesting that 125I-Sm-C/IGF I binds to more than 1 receptor species, and 125I-IGF II and 125I-MSA bind to one only. These findings were supported by affinity-labeling studies with radiolabeled Sm/IGFs using disuccinimidyl suberate as a cross-linking agent. Sm-C/IGF I appeared to bind to both type I and II Sm/IGF receptors, because cross- linked 125I-Sm-C/IGF I-receptor complexes with molecular weight (Mr) of greater than 300,000 (300K) and 130K (type I receptor) were observed under nonreducing and reducing conditions, respectively, as were 220 and 260K complexes (type II receptor) under the same respective conditions. 125I-IGF II and 125I-MSA, however, bound only to the Mr 220 and 260K moieties under nonreducing and reducing conditions, respectively, suggesting that these peptides bind only to the type II receptor. Competitive binding studies of the cross-linked moieties were consistent with this interpretation. In contrast, 125I-insulin bound poorly to astroglia (less than 0.5% specific binding), and cross- linking studies could not definitely distinguish among low-affinity binding to the type I Sm/IGF receptor, binding to a paucity of insulin receptors, or both. In addition, by combining autoradiography to localize 125I-Sm/IGFs binding on astroglial cells and immunocytochemistry with anti-glial fibrillary acidic protein to identify the cell type, we have demonstrated cell-surface binding and apparent internalization of radiolabeled Sm/IGFs. Concurrent studies of Sm/IGF stimulation of 3H-thymidine incorporation revealed that these cells were most sensitive to Sm-C/IGF I, followed by IGF II and MSA, and insulin. MSA and IGF II, however, were the most potent followed by Sm-C/IGF I and then insulin. Half-maximal stimulations of 3H-thymidine incorporation corresponded closely with half-maximal binding displacement for Sm-C/IGF I and less so for IGF II and MSA.(ABSTRACT TRUNCATED AT 400 WORDS)
Response properties of nociceptive and low-threshold mechanoreceptive neurons in the hamster superior colliculus
Authors: Larson MA , McHaffie JG , Stein BE
Published: March 2018

There are many somatosensory neurons in the hamster superior colliculus (SC); some respond to innocuous tactile stimuli, while others respond either preferentially, or solely, to noxious stimuli. Yet, there are little quantitative data describing the responses of these neurons. We sought to provide such information by relating stimulus intensity to the magnitude of the neural response using controlled innocuous and noxious mechanical and thermal stimuli. Of 122 somatosensory SC neurons studied in urethane-anesthetized hamsters, the majority (52%) had low- threshold mechanoreceptive properties (LT). LT neurons had force thresholds less than 1 gm, adapted rapidly to maintained stimuli, and did not respond with higher numbers of impulses to noxious mechanical or thermal stimuli. A smaller, though substantial, proportion of neurons (45%) responded either preferentially, or solely, to noxious stimuli. A few neurons (3%) were inhibited by either light tactile or noxious mechanical stimuli. Two populations of nociceptive neurons were found and classified either as wide dynamic range (WDR) neurons (n = 25), those that responded to gentle mechanical, noxious mechanical, and/or thermal stimuli; or nociceptive-specific (NS) neurons (n = 30), those that responded solely to high-intensity mechanical or noxious thermal stimuli. WDR neurons responded monotonically to increases in the intensity of innocuous mechanical stimuli, and displacement- response relationship for this population was a slightly negatively accelerating power function with an exponent of 0.785. However, the thermal stimulus-response relationships (to graded skin temperatures) of both WDR and NS neurons were positively accelerating power functions with exponents of 2.3 and 2.5 (r2 = 0.988), respectively. These values are consistent with both electrophysiological data from dorsal horn nociceptive neurons and from human psychophysical results using the same range of thermal stimuli. These experiments demonstrate that SC neurons are capable of signaling not only the presence and location of a noxious stimulus but its intensity as well. Presumably, these neurons play a significant role in the animal's reactions to potentially harmful stimuli. The partial laminar segregation of WDR and NS neurons may reflect different involvements of particular nociceptive subtypes in the various overt responses mediated by the SC.
Large depolarization induces long openings of voltage-dependent calcium channels in adrenal chromaffin cells
Authors: Hoshi T , Smith SJ
Published: March 2018

Single Ca2+-channel currents in bovine adrenal chromaffin cells were studied with the patch-clamp technique using Ba2+ as the charge carrier. Depolarizing pulses to voltages less than +10 mV from holding voltage of -60 mV elicited short openings with a mean life time of less than 1 msec. Depolarization to more positive voltages elicited longer openings with a mean life time of about 3 msec in addition to the short openings similar to those observed at less positive voltages. Following large depolarizing prepulses, 2 types of “tail” openings, one with a mean duration of less than 1 msec and the other with a mean duration of 4 msec, were observed. In the presence of a dihydropyridine BAY K 8644, openings with a mean duration of more than 12 msec were present. Depolarization-induced long openings and BAY K 8644-produced long openings differed in the first latency and open-time properties. The results could be explained in terms of multiple open states of one type of Ca2+ channel. A kinetic model with at least 2 open states is required to explain activation of Ca2+ channels in chromaffin cells.
Development of neurotransmitter metabolism in embryos of the leech Haementeria ghilianii
Authors: Glover JC , Stuart DK , Cline HT , McCaman RE , Magill C , Stent GS
Published: March 2018

We have investigated the development of neurotransmitter metabolism in embryos of the glossiphoniid leech Haementeria ghilianii. The neurotransmitter content of dissected embryonic tissues was measured by means of radioenzymatic assays, while the presence of neurotransmitters in individual identified neurons was detected by means of immunocytochemical and monoamine histofluorescence techniques. The capacity for synthesis of neurotransmitters was measured by incubating dissected embryonic tissues in radiolabeled neurotransmitter precursors. A specific neurotransmitter uptake system present in some neurons was detected by means of an autoradiographic technique. At an early stage of development of the nervous system, when most neurons are just beginning process outgrowth, the nerve cord acquires the capacity to synthesize ACh, 5-HT, and GABA from their immediate precursors, and contains ACh. Moreover, 5-HT-immunoreactive neurons and neurons that are capable of GABA uptake can be identified. Dopamine-containing neurons are first detected by their histofluorescence at a slightly later stage, after process outgrowth is under way. As development continues, the content of and capacity for synthesis of these neurotransmitters increase, as does the number of neurons capable of GABA uptake. During the earlier stages of development, ACh content exceeds 5-HT content, which in turn exceeds dopamine content. By the end of embryogenesis, however, 5-HT and dopamine contents have greatly increased relative to ACh content, with 5-HT content exceeding ACh content by a factor of 2. Of the neurotransmitters thus far studied, 5- HT is present in the highest amount in the juvenile and adult nerve cord. Our results indicate that in the development of the leech nervous system neurotransmitter metabolism is one of the first neuronal characters to differentiate and that the subsequent levels of the different neurotransmitters are differentially regulated.
Simultaneous optical recording of activity from many neurons during feeding in Navanax
Authors: London JA , Zecevic D , Cohen LB
Published: March 2018

Optical methods for monitoring changes in membrane potential have been used to measure action potential activity in the buccal ganglion of an opisthobranch mollusc, Navanax inermis, while the animal was feeding. During feeding activity was detected in 10–15% of the approximately 200 neurons present in the ganglion. Control experiments carried out to determine the completeness of the optical recording showed that activity in at least 70% of the neurons could be detected. Thus, in certain invertebrate ganglia, it is possible to make a reasonably complete recording of the neuron activity responsible for generating relatively complex behaviors.
Activators of adenylate cyclase and cyclic AMP prolong calcium- dependent action potentials of mouse sensory neurons in culture by reducing a voltage-dependent potassium conductance
Authors: Grega DS , Macdonald RL
Published: March 2018

The effects of compounds that activate adenylate cyclase and of cAMP on calcium-dependent action potentials recorded from mouse dorsal root ganglion neurons were assessed. Application of compounds that stimulate the adenylate cyclase system (forskolin, cholera toxin, and prostaglandin E1) increased action potential duration with an associated decrease in afterhyperpolarization. An adenylate cyclase inhibitor, 2′,5′-dideoxyadenosine, partially inhibited the responses to forskolin and cholera toxin. cAMP analogs mimicked the effect of forskolin, and the phosphodiesterase inhibitor theophylline enhanced the response to forskolin. Following intracellular injection of the potassium channel blocker cesium, the forskolin response was reduced. Forskolin did not significantly alter resting membrane potential or conductance. The action potential responses to forskolin were voltage dependent, being reduced when the membrane was held at less negative (less than -50 mV) potentials. The data suggest that activators of adenylate cyclase and cAMP prolong calcium-dependent action potentials by blocking a voltage-dependent potassium conductance that is responsible, in part, for action potential repolarization and that inactivates at membrane potentials less negative than -50 mV.
Pharmacology of retinotectal transmission in the goldfish: effects of nicotinic ligands, strychnine, and kynurenic acid
Authors: Langdon RB , Freeman JA
Published: March 2018

The goal of this study was to evaluate different neurotransmitters and their receptors that might be involved in retinotectal transmission in the goldfish. Sections of tectum were isolated and maintained in vitro while pharmacological agents were administered via the tissue bath. Field potentials were elicited by electrical stimulation of the optic nerve and recorded at 50 micron depth intervals, and profiles of current source densities (CSDs) were computed from the second spatial derivatives of these potentials. The preparations were treated with low [Ca2+]/high [Mg2+] media, various cholinergic agonists and antagonists, eserine, strychnine, or kynurenic acid, via the tissue bath. Prior to treatment, depth profiles of these in vitro field potentials and CSDs closely resembled those previously reported in vivo, including 2 prominent sink-source pairs with their sinks in the superficial optic neuropil, followed by a smaller and more prolonged sink-source pair of opposite polarity. These were rapidly and reversibly eliminated by low [Ca2+]/high [Mg2+] bathing media, and substantially reduced by 0.5 or 1.0 mM kynurenic acid. By contrast, d-tubocurarine (d-TC; up to 0.16 mM) reduced peak response amplitudes by less than 40%, eliminated the third sink-source pair, and more than doubled the duration of decay of sink-source pairs 1 and 2 in a concentration-dependent manner. Strychnine had a similar action to d-TC but was slightly more potent. The time course and amplitudes of responses were not much affected by the following nicotinic agonists or antagonists (concentrations in microM): mecamylamine, 50; dihydro-beta-erythroidine, 50; nicotine, 200; tetramethylammonium, 500; ACh (protected by eserine, 20), 200; alpha-bungarotoxin, 2 microM for 2.5 hr, and 0.4 microM for up to 10.5 hr; and lophotoxin, 32 microM for up to 94 min. Eserine (20 microM) and carbachol (200 microM) increased peak response amplitudes by up to 80% within 5–10 min, and amplitudes remained elevated during 20–33 min of continued treatment. The onset of the effects of d-TC, strychnine, and kynurenic acid began in 5–10 min and was completed in 30 min or less, indicating that test substances could adequately penetrate into the interior of the isolated sections of tectum. The failure of these cholinergic ligands to prevent postsynaptic responses indicates that excitatory retinotectal transmission does not depend on an intact nicotinic (or other cholinergic) system, as previously proposed. The action by kynurenic acid suggests the involvement of an excitatory amino acid neurotransmitter in retinotectal transmission.(ABSTRACT TRUNCATED AT 400 WORDS)
Voltage-dependence of glycine-activated Cl- channels: a potentiometer for inhibition?
Authors: Faber DS , Korn H
Published: March 2018

Single- and double-electrode voltage-clamp techniques have been employed in situ to analyze Mauthner cell inhibitory synaptic responses produced both by activation of the recurrent collateral network and by direct intracellular stimulation of single presynaptic interneurons. The results demonstrate that the synaptically evoked glycinergic postsynaptic currents exhibit a striking voltage sensitivity. Specifically, the time constant of the decay of the synaptic conductance is increased by depolarization and decreased by hyperpolarization. This parameter is exponentially related to membrane potential, changing e-fold for a 45 mV potential shift, regardless of the degree of intracellular chloride loading or the magnitude of the underlying synaptic conductance. In addition, the amplitude of this inhibitory conductance change is decreased by membrane hyperpolarizations of 15 mV or more. Computer modeling demonstrates that the voltage dependence of the kinetics of the synaptic response may serve to enhance the magnitude and duration of inhibitory responses appreciably in the face of increased excitation.
Circling behavior following unilateral microinjections of cocaine into the medial prefrontal cortex: dopaminergic or local anesthetic effect?
Authors: Morency MA , Stewart RJ , Beninger RJ
Published: March 2018

Dopaminergic projections to the medial prefrontal cortex have been implicated in cocaine reinforcement; therefore, it was of interest to examine the locomotor effects of acute administration of cocaine to this area. Circling behavior was assessed following injections of 1.0 microliter of cocaine in doses of 0 (0.9% saline), 25, 50, and 100 micrograms/microliters into the medial prefrontal cortex of rats prepared with chronic unilateral guide cannulae. Animals were scored during four 5 min intervals of a 60 min test session that began with the central injection and placement in a flat circular arena. Cocaine was found to produce dose-dependent contraversive circling, an effect previously seen with the dopamine (DA) agonists LY 14 1865 and (+)- amphetamine, suggesting a unilateral stimulant effect. However, since cocaine has potent local anesthetic properties that have been reported to produce behavioral effects and also to inhibit the reuptake of norepinephrine and 5-HT, it was important to demonstrate that the directional bias was a dopaminergic effect. Intra-frontocortical microinjections of the local anesthetic procaine (10, 100, and 1000 micrograms in 0.5 microliter) did not induce circling. Sulpiride (0.001–10.0 micrograms in 0.5 microliter), a DA antagonist specific for the D- 2 receptors, produced ipsiversive circling in a dose-dependent manner in rats treated with (+)-amphetamine (1.5 mg/kg, i.p.). In addition, sulpiride (1.0 micrograms in 0.5 microliter) blocked the circling behavior induced by cocaine (50 micrograms in 0.5 microliter) when administered into the medial prefrontal cortex 15 min prior to the cocaine injection. These results provide further evidence for an excitatory influence of mesocortical DA on motor control.
Blockade of Ca2+ and K+ currents in bag cell neurons of Aplysia californica by dihydropyridine Ca2+ antagonists
Authors: Nerbonne JM , Gurney AM
Published: March 2018

The effects of dihydropyridine calcium antagonists on whole-cell Ca2+ and K+ currents in the neurosecretory bag cells of the marine mollusc Aplysia californica have been investigated. Nifedipine and nisoldipine blocked bag cell Ca2+ currents with effects similar to those seen previously on Ca2+ currents in cardiac muscle: both compounds appeared to interact with Ca2+ channels when they were closed, open, and inactivated. Also, as seen in cardiac cells, nifedipine apparently binds with higher affinity to Ca2+ channels when they are inactivated than when they are either closed or open. Nifedipine and nisoldipine also inhibited 2 outward K+ currents in bag cells: the “delayed rectifier” (IK) and the “A” (IA) currents. Half-maximal blockade of Ca2+ currents occurred at approximately 1.4 microM nifedipine, compared to approximately 3–5 microM for half-maximal blockade of IK and IA. The effects of these compounds on bag cell Ca2+ and K+ currents are interpreted and discussed here in terms of the “modulated receptor” model of drug action. In contrast, however, no measurable effects of nifedipine or nisoldipine were seen on Ca2+ (and/or K+) currents in several vertebrate neuronal cell types. Our results suggest that there are likely to be structural and/or conformational variations in Ca2+ channels in different cells, tissues, and/or species and also that, in some cells, Ca2+ and K+ channels might be structurally similar. These findings also suggest, therefore, that if dihydropyridine binding is used to identify Ca2+ channels, care should be taken to ensure that binding correlates closely with the Ca2+ channels of interest.
Cell lineage, cell death, and the developmental origin of identified serotonin- and dopamine-containing neurons in the leech
Authors: Stuart DK , Blair SS , Weisblat DA
Published: March 2018

The nervous system of the glossiphoniid leech includes segmentally iterated neurons that contain serotonin (5-HT) and dopamine. These have been investigated in Helobdella triserialis, Theromyzon rude, and Haementeria ghilianii. Five types of 5-HT neurons are identified by immunocytochemistry in the abdominal ganglia of the ventral nerve cord: the bilaterally paired Retzius, anteromedial, ventrolateral and dorsolateral neurons, and the unpaired posteromedial (pm) neuron. Three types of bilaterally paired dopamine neurons are identified by glyoxylic acid-induced fluorescence in the segmental body wall: MD, LD1, and LD2. Each left or right half of the segmental complement of the leech nervous system is known to develop from 6 distinct ectodermal primary blast cells (ns, nf, o, p, qs, and qf). To identify the blast cells of origin of the 5-HT and dopamine neurons, fluorescent cell lineage tracers were injected into the various precursors of the blast cells in early (stage 6) embryos. The embryos were then raised until their 5-HT and dopamine neurons could be scored (stage 11) for the presence or absence of lineage tracer. We find that the Retzius, anteromedial, and posteromedial 5-HT neurons are derived from the ns blast cell, while the ventrolateral and dorsolateral 5-HT neurons are derived from the nf blast cell. The unpaired pm 5-HT neuron arises as one of a bilateral pair of neurons, of which one later dies. Whether the left or right pm neuron survives in any given ganglion is the consequence of some form of competitive interaction between cells derived from the left and right n primary blast cells, possibly between the left and right pm neurons themselves. We find that, of the dopamine neurons, the LD1 neuron is derived from the o blast cell, the LD2 neurons from the p blast cell, and the MD neuron from one of the 2 kinds of q blast cells. These results show that the 5-HT and dopamine neurons arise from 5 different primary blast cells in a highly determinate manner, and they support the view that cells of a similar phenotype need not be closely related in the developmental cell lineage tree.
Regional heterogeneity in the distribution of somatostatin-28- and somatostatin-28(1-12)-immunoreactive profiles in monkey neocortex
Authors: Campbell MJ , Lewis DA , Benoit R , Morrison JH
Published: March 2018

The distribution of the prosomatostatin-derived peptides (PSDP), somatostatin-28 and somatostatin-28(1–12), in the cynomolgus monkey (Macaca fascicularis) neocortex was characterized in quantitative immunohistochemical studies of 3 visual areas (V1, primary visual cortex; V2, the adjacent visual association area; and AIT, a visual association area in anterior inferior temporal cortex), 2 auditory areas (AI, primary auditory cortex; and T1, an adjacent auditory association area) and anterior cingulate cortex (Area 24). The results of similar quantitative analyses in 3 homologous areas in rat neocortex (primary visual, primary auditory, and anterior cingulate) are also presented. Primate cortical areas differed significantly in both density and laminar distribution of PSDP-immunoreactive profiles. Area 24, the most densely labeled area, had nearly 6 times as many PSDP- immunoreactive neurons as V1. Both auditory areas contained approximately two-thirds the number of PSDP-immunoreactive neurons found in Area 24; however, both had nearly 4 times as many immunoreactive neurons as V1. The 3 visual areas showed incremental increases in the number of PSDP-immunoreactive neurons; V2 contained nearly twice and AIT nearly 3 times the number of immunoreactive neurons present in V1. Both the supra- and infragranular layers were densely labeled in Area 24 and Area T1, however, in AI, V1, V2, and AIT the infragranular layers were relatively sparsely labeled. In contrast to the regional heterogeneity found in the primate neocortex, the distribution of immunoreactive neurons was quite uniform across the 3 rat cortical areas. The rat cortical areas contained substantially fewer immunoreactive neurons than most of the monkey cortical areas, and a majority of these immunoreactive neurons were located in the infragranular layers. These findings suggest that the regional specialization of primate neocortex involves the selective distribution of PSDP-immunoreactive neurons. They also suggest that chemically specified intrinsic organization of neocortex is not likely to be uniform across species or across cortical areas in the primate. The distinctive regional distribution patterns of PSDP-immunoreactive profiles appear to parallel that of the long corticocortical projections (contralateral and distant ipsilateral projections), suggesting an association between these presumed inhibitory interneurons and this important extrinsic system.
Long-term modifiability of anomalous and delayed rectification in guinea pig inferior olivary neurons
Authors: Yarom Y , Llinas R
Published: March 2018

Delayed and anomalous rectification was studied in inferior olivary (I.O.) neurons in guinea pig brain stem slices maintained in vitro. Hyperpolarization of the I.O. cell beyond rest membrane potential was accompanied by anomalous rectification (AR). This consisted of 2 parts: an instantaneous and a time-dependent component. The “instantaneous” component was blocked by bath addition of Ba2+ or Cs+ and demonstrated inactivation following prolonged hyperpolarization. The time-dependent component, referred to as the gK(ol), was blocked by harmaline in concentrations of 0.1 mg/ml or by substitution of Co2+, Cd2+, or Mn2+ for Ca2+ in the bath. The gK(ol) was blocked by extracellular Cs+ but not by Ba2+. Delayed rectification (DR), consisting of 2 distinct components, was observed after membrane depolarization by more than 10 mV with respect to rest (usually at -65 mV). One of the components of the DR was found to be quite similar to the classical gK. It did not demonstrate significant inactivation with membrane potential change and was reduced by Ba2+ or tetraethylammonium (TEA). A second component of the DR demonstrated voltage-dependent inactivation and was thus referred to as gK(inact). This inactivation determined by current-clamp measurements had a sigmoidal time course, with approximately a 1 sec onset latency and a half-time to peak of 7 sec. The inactivation of gK(inact) outlasted current injection for tens of seconds to several minutes, depending on the duration and amplitude of the preceding depolarization. During this period, I.O. neurons could be easily activated and demonstrated full dendritic spikes following current injection or excitatory synaptic input that had previously been subthreshold for spike initiation. The inactivation component of the DR was removed by prolonged membrane hyperpolarization beyond rest. gK(inact) was blocked by 4-aminopyridine (4-AP; 100 microM) but not by Ba2+. This inactivation was dependent on the presence of extracellular Ca2+ or Ba2+. Addition of Co2+ or Cd2+ to the bath did not block gK(inact) but did prevent its inactivation. The modulatory effects of these different membrane conductances on the integrative properties of I.O. neurons are described. The long duration of the inactivation of DR and AR is considered as the basis for a dynamic long-term modulation of the electroresponsive and integrated properties of I.O. neurons.
Synapsin I in PC12 cells. I. Characterization of the phosphoprotein and effect of chronic NGF treatment
Authors: Romano C , Nichols RA , Greengard P , Greene LA
Published: March 2018

PC12 cells contain a synapsin I-like molecule. Several serum and monoclonal antibodies raised against bovine brain synapsin I bind to and precipitate this molecule, demonstrating immunochemical similarity between the brain and PC12 species. PC12 synapsin I, like brain synapsin I, is a phosphoprotein: It is phosphorylated in intact cells and, when partially purified, serves as a substrate for several synapsin I kinases. PC12 cell synapsin I is structurally similar to brain synapsin I as shown by peptide mapping of 35S-methionine-and 32P- phosphate-labeled molecules from the 2 sources. Chronic NGF treatment of the cells induces a significant increase in the amount of synapsin I relative to total cell protein, measured either by immunolabeling or incorporation of 35S-methionine. The synapsin I present in untreated PC12 cells migrates predominantly as a singlet and that present in cells treated chronically with NGF as a doublet in SDS-PAGE.
Synapsin I in PC12 cells. II. Evidence for regulation by NGF of phosphorylation at a novel site
Authors: Romano C , Nichols RA , Greengard P
Published: March 2018

NGF treatment of PC12 cells caused a rapid increase in the state of phosphorylation of synapsin I. This phosphorylation of synapsin I is accompanied by a decrease in its electrophoretic mobility on SDS-PAGE. Phosphopeptide fingerprint analysis of the synapsin I revealed that this phosphorylation occurred on a particular phosphopeptide, designated peptide N. Phosphoserine was the only phosphoamino acid detected in peptide N. Partially purified PC12 synapsin I was a substrate for several protein kinases known to be capable of phosphorylating brain synapsin I, but none of these kinases phosphorylated synapsin I on peptide N. The results suggest that the NGF-stimulated phosphorylation of synapsin I may be mediated by a novel protein kinase.
Immunocytochemical and electrophysiological differentiation of rat cerebellar granule cells in explant cultures
Authors: Hockberger PE , Tseng HY , Connor JA
Published: March 2018

We have used a combination of immunocytochemical and electrophysiological measurements to monitor the differentiation of cerebellar granule cells in vitro. We present immunocytochemical evidence showing that several characteristic features of developing rat cerebellar tissue were retained in postnatal explant cultures. Most notably the cultures expressed radiating GFAP-positive (Bergmann) glia processes, proliferating NSE-negative neuroblasts, and migrating NSE- positive granule cells. The latter were subdivided into 3 developmental stages--i.e., immature, intermediate, and mature granule cells, based upon cell differences in location from the explant, intensity of NSE staining, excitability, and the amplitude of voltage-dependent conductances. Immature cells were identifiable during the first week in culture and were located up to 140 micron from the explant. These cells stained lightly for NSE and displayed conductances of insufficient magnitude to generate action potentials. Intermediate cells were present after 1–2 weeks in culture and were located up to 500 micron from the explant. These cells were also NSE positive and were characterized by the presence of soma action potentials. Intermediate cells displayed 3 large voltage-dependent conductances: a transient, TTX-sensitive inward current; a delayed, TEA-sensitive outward current; and a transient, 4AP-sensitive outward current. Mature cells were present after 1 month in culture and, like intermediate cells, were no more than 500 micron from the explant. However, mature cells stained more intensely for NSE, and the somata of these cells were devoid of voltage-dependent conductances (although axonal currents were usually present). These results indicate that granule cells undergo a stereotypic sequence of differentiation in postnatal explant cultures. These stages may correspond to developmental changes in granule cells during migration into the (internal) granular cell layer in vivo.
Synaptic sites on reinnervated nerve cells visualized at two different times in living mice
Authors: Purves D , Lichtman JW
Published: March 2018

Synaptic boutons on the surface of identified autonomic ganglion cells were visualized by methylene blue staining at intervals of 1–2 months following denervation to assess whether regenerating axon terminals reoccupy original synaptic sites. The distribution of synapses observed on the same neuronal cell bodies was almost always different in appearance after reinnervation. These results are at odds with the conclusions of earlier workers, who have argued that mammalian neurons bear a fixed number of synaptic sites, which are reoccupied during reinnervation.
Mu-conotoxins share a common binding site with tetrodotoxin/saxitoxin on eel electroplax Na channels
Authors: Yanagawa Y , Abe T , Satake M
Published: March 2018

The binding characteristics of conotoxin GIIIA purified from the venom of a marine snail, Conus geographus, with regard to electroplax membranes from Electrophorus electricus were studied using a radiolabeled monopropionyl derivative of the toxin (3H-Pr-CGIIIA). 3H- Pr-CGIIIA bound specifically to a single class of saturable binding sites in electroplax membranes with a dissociation constant of 1.1 +/- 0.2 nM and a maximal binding capacity of 11 +/- 2 pmol/mg of protein. The latter value was similar to the number of specific binding sites (10 +/- 2 pmol/mg of protein) for 3H-lysine-tetrodotoxin (3H-Lys-TTX). Monopropionyl CGIIIA and CGIIIA had similar inhibitory effects on the binding of 3H-Lys-TTX (1 nM) to electroplax membranes with IC50 values of 3.5 and 0.9 nM, respectively. The association and dissociation of 3H- Pr-CGIIIA and electroplax membranes were much slower than those of 3H- Lys-TTX and the membranes. Mu-Conotoxins (CGIIIA and CGIIIB) and guanidinium toxins (TTX and saxitoxin) inhibited 3H-Pr-CGIIIA (1 nM) binding to electroplax membranes with IC50 values of 0.6, 1.1, 7.1, and 2.2 nM, respectively. However, several other kinds of neurotoxins and local anesthetics known to interact with Na channels did not affect 3H- Pr-CGIIIA binding. These findings indicate that mu-conotoxins must be classified in the same group of Na channel inhibitors as guanidinium toxins, since they competed with guanidinium toxins for binding sites on the Na channel. The peptide mu-conotoxins should be useful in studies on the functional and structural domains of Na channel proteins.
Reduced visual acuity in both eyes of monocularly deprived kittens following a short or long period of reverse occlusion
Authors: Murphy KM , Mitchell DE
Published: March 2018

While there can be substantial recovery of vision during reverse occlusion in the deprived eye of kittens that have been monocularly deprived from birth, in many situations this visual improvement does not appear to be retained following the introduction of binocular visual input. This study examines the consequences of periods of reverse occlusion whose onset and duration would be thought to maximize the opportunity for good and permanent recovery of vision in the initially deprived eye. Twenty kittens were monocularly deprived from near birth to either 4, 5, or 6 weeks of age and then reverse occluded for periods that were either very brief (9–18 d) or very long (9–12 weeks). In the former situation, binocular visual experience was introduced when the visual cortex was most susceptible to environmental modification, which would be thought to maximize the opportunity for the eventual development of good vision in both eyes. On the other hand, the long periods of reverse occlusion might be expected to lead to a permanent recovery of vision in only the initially deprived eye, since binocular visual input would have been introduced at a time when the cortex was no longer very plastic. Surprisingly, the end result of both these extreme regimens of reverse occlusion was the same, namely, a severe bilateral amblyopia in which the acuity of both eyes was permanently reduced to levels that were as low as 1/10, but more typically between 1/3 and 1/4 of normal values. Contrast sensitivity functions measured on 2 of the cats that received a brief period of reverse occlusion revealed deficits in contrast sensitivity of about an order of magnitude at all spatial frequencies. These observations have important implications for the nature of the physiological and anatomical changes that occur in the visual cortex during reverse occlusion.
Peripheral specification of sensory neurons transplanted to novel locations along the neuraxis
Authors: Smith CL , Frank E
Published: March 2018

Thoracic dorsal root ganglia in bullfrogs contain sensory neurons that innervate the skin of the trunk and have synaptic connections in the dorsal horn of the spinal cord. The ganglion that innervates the forelimb contains, in addition to cutaneous afferents, many muscle afferents that project more ventrally in the spinal cord and make monosynaptic connections with motoneurons. In the present study, we have transplanted thoracic sensory neurons to the brachial level in tadpoles to discover whether they can innervate forelimb muscles and, if so, whether they form central connections characteristic of forelimb muscle afferents. The ganglion that normally supplies the forelimb was removed from tadpoles and replaced with 2 thoracic ganglia. After the tadpoles completed metamorphosis, the peripheral and central connections of the transplanted thoracic sensory neurons were examined with anatomical and electrophysiological techniques. When the ganglia were transplanted at stage XIV or earlier, transplanted sensory neurons innervated the forelimb and projected into the brachial spinal cord. Electrical stimulation of forelimb muscle nerves evoked impulses in the dorsal root, indicating that some centrally projecting sensory neurons were muscle afferents. Furthermore, muscle afferents were also activated by stretching muscles which suggest that they terminated on spindles. HRP labeling of the central projections revealed that transplanted sensory neurons terminated at sites characteristic of both cutaneous and muscle afferents. The pattern of synaptic connections was assessed by recording intracellularly from motoneurons. Stimulation of muscle afferents produced monosynaptic EPSPs in motoneurons. As in normal frogs, triceps muscle afferents projected more strongly to triceps motoneurons than to subscapularis and pectoralis motoneurons, while subscapularis afferents projected to all 3 types of motoneurons. Thus, the transplanted sensory neurons formed central connections appropriate to their novel peripheral targets. These observations suggest that interactions between sensory neurons and their targets may be important in determining their central connections.
The impact of the NINCDS on the neurosciences: an essay written for the centennial of the NIH
Authors: Tower DB
Published: March 2018

This essay, written as a contribution to the June 1987 participation by the NINCDS in the celebration of the centennial of the NIH (dated from the founding of the Hygienic Laboratory at Staten Island in 1887), discusses the impact of the NINDB/NINCDS on the neurosciences. Contrasts are drawn with the status of basic and clinical neurosciences in the late 19th century and prior to 1950, when the NINDB was established. This impact has involved its role as advocate and focal point for the support of research and research training in the neurosciences; the training of research and academic personnel and the funding of research projects, both significant factors in the postwar proliferation of methods and technologies; and the catalyzing of the steady progress in scientific knowledge, as reflected in examples from various facets of the programs of the neurological institute, in a continuum from the very basic and fundamental to the very clinical and pathological neurosciences.
Circadian rhythmicity restored by neural transplant. Immunocytochemical characterization of the graft and its integration with the host brain
Authors: Lehman MN , Silver R , Gladstone WR , Kahn RM , Gibson M , Bittman EL
Published: March 2018

It is well established that overt circadian rhythms are permanently disrupted following lesions of the hamster hypothalamic suprachiasmatic nucleus (SCN). In the present study, we show that implantations of brain grafts containing the fetal SCN reestablish circadian rhythms of locomotor activity in adult hamsters previously made arrhythmic by SCN lesions. The restoration of free-running rhythms in conditions of constant darkness is correlated with the presence in the graft of neuropeptides normally present in the SCN of unlesioned hamsters, including vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), and vasopressin (VP). In several recipients, grafts were found to receive retinal input, and appeared to send efferents into the host brain. Not all functions of the SCN were reinstated by the graft: animals with restored locomotor rhythms did not show gonadal regression in the absence of light, and failed to synchronize (entrain) to light intensities to which SCN-intact animals responded.
Phorbol esters: voltage-dependent effects on calcium-dependent action potentials of mouse central and peripheral neurons in cell culture
Authors: Werz MA , Macdonald RL
Published: March 2018

The beta-phorbol esters 12–O-tetradecanoylphorbol-13-acetate (TPA) and phorbol 12,13-dibutyrate (PDBu), which activate protein kinase C, were applied to mouse dorsal root ganglion (DRG) and cerebral hemisphere neurons grown in primary dissociated cell culture. Phorbol esters did not modify the membrane potential or input resistance of either type of neuron. To assess the effects of beta-phorbol esters on voltage- dependent conductances, the effects of PDBu and TPA on action potentials evoked from these neurons were determined. The neurons were bathed in a solution containing 5 mM tetraethylammonium and action potentials that contained sodium and calcium components were evoked. When applied at resting membrane potential and at more negative potentials, PDBu and TPA reversibly increased action potential duration. The alpha-phorbol ester 4-alpha-phorbol, which does not activate protein kinase C, did not modify action potential duration. The effects of the beta-phorbol esters, however, were voltage- dependent. When the neurons were depolarized to membrane potentials less negative than -50 mV, PDBu and TPA reduced action potential duration. The effects of both PDBu (10 nM-1 microM) and TPA (100 pM-100 nM) on action potential duration were dose-dependent. The prolongation of action potentials produced at large negative potentials may be due to a reduction in voltage-and/or calcium-dependent potassium conductance, since the prolongation was associated with a reduction in the potassium-dependent afterhyperpolarization; following membrane depolarization in control solution, action potential duration was increased for several minutes, while the afterhyperpolarization was reduced and, following this prolongation, phorbol esters no longer prolonged the action potentials.(ABSTRACT TRUNCATED AT 250 WORDS)
Tactile discrimination of shape: responses of rapidly adapting mechanoreceptive afferents to a step stroked across the monkey fingerpad
Authors: LaMotte RH , Srinivasan MA
Published: March 2018

Responses of rapidly adapting Meissner corpuscle mechanoreceptive afferent fibers (RAs) to steps of varying shape stroked across the distal fingerpad were recorded from anesthetized monkeys. A series of flat plates were used, each having an increase in thickness (a step) in the middle so that one-half of the plate was thicker than the other. The cross-sectional shape of the step approximated that of a half-cycle sinusoid, 0.5 mm high. The width (half-cycle wavelength) of the sinusoidal step was varied from 0 to 3.13 mm, producing a series of step shapes that differed in steepness and curvature. These steps could be broadly categorized into 2 groups, “steep” and “gradual.” Each step was stroked back and forth under constant compressional force, using a servocontrolled mechanical stimulator. The RA's response to a step provided a spatial pattern of action potentials in which the occurrence of each impulse corresponded to a position of the step on the skin. This response consisted of a single “burst” of impulses to the sinusoidal portion of the step. Changes in stroke direction, step shape, or velocity of stroking primarily affected the RA discharge rate during the burst, and, less consistently, the spatial width of the burst. For a given step shape and stroke velocity, the discharge rate was greater for strokes from the low to the high side of the step than for strokes from the high to the low side. Discharge rate was greater for steep than for gradual steps and, for a given step, it increased with stroke velocity. All the major features of the responses were interpreted as being due predominantly to the sensitivity of the RA to vertical velocity at the most sensitive spot on its receptive field, together with a sensitivity to the rate of change in skin curvature at that spot. RA discharge rate distinguished not only the gross differences between steep and gradual steps, but also some of the finer differences in sharpness among steep steps. From a comparison with the human capacity for tactile discrimination of the steps, it was concluded that RAs, through their discharge rates, provide primarily “intensive” information about the sharpness of shapes.