bluuebunny/crossref_metadata_2025 huggingface dataset:
Transfected rat high-molecular-weight neurofilament (NF-H) coassembles with vimentin in a predominantly nonphosphorylated form
Authors: Chin SS , Liem RK
Published: March 2018

A fully encoding cDNA for the high-molecular-weight rat neurofilament protein (NF-H) has been isolated from a lambda gt11 library, sequenced and subcloned into eukaryotic expression vectors. Sequence analysis shows that rat NF-H has an overall homology of 72 and 88% with human and mouse NF-H, respectively. The head and rod domains are almost entirely identical, and the divergences are due to differences in the long C-terminal extensions of the molecule. The consensus phosphorylation sequence for neurofilaments Lys-Ser-Pro (KSP) is present 52 times. The predicted molecular mass of the protein is 115 kDa, 42% lower than that observed by SDS-PAGE. Upon transfection into vimentin-containing fibroblasts, such as L tk-, L929, and 3T6 cells, NF- H is seen distributed with vimentin by light and electron microscopic examinations indicating that copolymers of NF-H and vimentin are formed in these cells. Only a negligible proportion of the cells is positive when stained with a number of antibodies directed against phosphorylated NF-H epitopes. This is in contrast with the middle molecular weight NF protein (NF-M) transfected into L tk- and L929 cells, which can readily be detected by antibodies against phosphorylated neurofilament epitopes. The mobilities of the transfected protein on 1- and 2-dimensional gels confirm that NF-H is predominantly in a nonphosphorylated form. These results indicate that phosphorylation of NF-H, but not NF-M, on the KSP sequence is due to protein kinases, which are not present in fibroblasts and are presumably NF-H specific. The stable NF-H-expressing cell lines can therefore be used to study these putative neurofilament kinases in vitro and in vivo.
Oligodendrocytes arrest neurite growth by contact inhibition
Authors: Bandtlow C , Zachleder T , Schwab ME
Published: March 2018

We have used video time-lapse microscopy to analyze in vitro the interactions of growth cones of newborn rat dorsal root ganglion cells with dissociated young rat CNS glial cells present in the cultures at low density. To provide optimal conditions for neurite extension, cells were grown on laminin and in NGF-supplemented medium. Our initial observation showed that there are 2 subpopulations of growth cones differing in their growth rate on laminin (averages of 12 and 45 microns/hr). When these growth cones encountered astrocytes, they maintained their normal configuration and growth velocity. They subsequently grew along or on top of astrocytes. In some cases, however, fast-moving growth cones showed a slight reduction in their growth rate. When growth cones countered oligodendrocytes, however, firm filopodial contact was sufficient to induce a rapid and long- lasting arrest of the growth cone motility, often followed by a collapse of the growth cone structure. One third of the paralyzed growth cones were observed to retract. Growth arrest and growth cone collapse were strictly dependent on membrane contact between neurons and oligodendrocytes. This contact inhibition phenomenon was exclusively found with differentiated oligodendrocytes and could be prevented by the monoclonal antibody IN-1 directed against neurite growth inhibitors NI-35 and NI-250 (Caroni and Schwab, 1988b). These results confirm previous findings that the neurite growth inhibitor proteins are important in axon outgrowth. Further, the inhibition of neurite growth exerted by oligodendrocytes is a contact-mediated phenomenon that can be triggered by the tip of growth cone filopodia.
The neural site of binocular rivalry relative to the analysis of motion in the human visual system
Authors: Wiesenfelder H , Blake R
Published: March 2018

Neural processing is disrupted during suppression phases of binocular rivalry, as evidenced by the temporary invisibility of an otherwise complex, high-contrast visual stimulus. This paper investigates the locus of this disruption relative to the processing of information about image motion. In one experiment, observers tracked binocular rivalry between a stationary textured field and a plaid composed of 2 drifting cosine gratings, with the angle between components varied to produce different pattern speeds. (Plaid speed is given by the ratio of the component speed to the cosine of the angle between the 2 directions of motion.) Predominance of the moving plaid increased with pattern speed, even though the speed of the individual components remained constant. Control measures verified that this influence of plaid speed was not attributable to specific component orientations. Information about coherent motion influences the rivalry process, implying that the site of coherent motion analysis, presumably the middle temporal area (MT), received input during dominance phases of rivalry. A second experiment investigated the effect of suppression on the processing of complex, nonlinear motion. Observers tracked rivalry phases for a rotating spiral, then indicated the duration of the subsequently perceived spiral aftereffect (SAE) for both rivalry and nonrivalry conditions. The SAE was reduced when adaptation occurred under the rivalry condition, with aftereffect duration proportional to the total duration of spiral visibility during adaptation. Earlier work places rivalry after the site of the linear motion aftereffect, and the present results show that rivalry suppression occurs prior to the site of spiral motion processing.(ABSTRACT TRUNCATED AT 250 WORDS)
Different classes of glutamate receptors and GABA mediate distinct modulations of a neuronal oscillator, the medullary pacemaker of a gymnotiform electric fish
Authors: Kawasaki M , Heiligenberg W
Published: March 2018

Gymnotiform electric fish generate distinct communicatory signals by modulating the rate of their electric organ discharges (EODs). Each EOD is triggered by a command pulse from the medullary pacemaker nucleus (PN), which contains pacemaker cells and relay cells. The firing rate of this nucleus is modulated by inputs from the diencephalic prepacemaker nucleus (PPN). The NMDA receptor blocker APV and the kainate/quisqualate receptor blocker CNQX, administered to the PN, suppress different types of modulations, indicating that different classes of glutamate receptors mediate the generation of different modulations. A comparison of the 2 genera, Hypopomus and Eigenmannia, reveals that sustained modulations, such as smooth rises in the rate of pacemaker cell firing and the selective silencing of the relay cells (only observed in Hypopomus), are mediated by NMDA receptors, whereas the brief and rapid acceleration, called “chirp” or “decrement burst,” is mediated by kainate/quisqualate receptors. Application of the GABA blocker bicuculline reveals that the 2 genera differ in the mechanism by which they slow the firing rate of their pacemaker. Whereas Hypopomus uses GABAergic inhibition to slow down and ultimately silence its pacemaker cells, Eigenmannia reduces tonic, APV-sensitive excitation originating from its PPN and lacks GABAergic inhibition in the PN.
Colocalization of neurotensin receptors and of the neurotensin- degrading enzyme endopeptidase 24-16 in primary cultures of neurons
Authors: Chabry J , Checler F , Vincent JP , Mazella J
Published: March 2018

This paper compares the localization of neurotensin receptors and of endopeptidase 24–16, a peptidase likely involved in the inactivation of neurotensin in primary cultures of neurons. Neurotensin binding sites were radiolabeled with 125I-Tyr3-neurotensin, whereas endopeptidase 24– 16 was stained by immunohistochemical techniques using a monospecific polyclonal antibody. Endopeptidase 24–16 is present in 80–85% of the nondifferentiated neurons. The proportion of immunoreactive neurons decreased during maturation to reach 35–40% after 4–8 d of culture. By contrast, neurotensin receptors were not detectable in nondifferentiated cells and appear during maturation. Specific 125I- Tyr3-neurotensin labeling is maximal after 4 d of culture and is located on about 10% of differentiated neurons. Double-labeling experiments show that about 90% of cortical, hypothalamic, and mesencephalic neurons bearing the neurotensin receptor also contained endopeptidase 24–16, supporting the hypothesis that one of the functions of endopeptidase 24–16 is the physiological inactivation of neurotensin. However, the presence of endopeptidase 24–16 on numerous neurons that do not contain neurotensin receptors also suggests that the enzyme could be involved in the degradation and/or maturation of other neuropeptides.
Patterns of glutamate immunoreactivity in the goldfish retina
Authors: Marc RE , Liu WL , Kalloniatis M , Raiguel SF , van Haesendonck E
Published: March 2018

Postembedding silver-intensified immunogold procedures reveal high levels of glutamate immunoreactivity in “vertical” elements of the goldfish retina: (1) Red-sensitive and green-sensitive cones display strong glutamate immunoreactivity, especially in their synaptic terminals, but blue-sensitive cones are poorly immunoreactive. (2) All type Mb (on-center) and Ma (off-center) mixed rod-cone bipolar cells and all identifiable cone bipolar cells are highly glutamate immunoreactive. We find no evidence for bipolar cells that lack glutamate immunoreactivity. (3) The majority of the somas in the ganglion cell layer and certain large cells of the amacrine cell layer resembling displaced ganglion cells are strongly glutamate immunoreactive. (4) Despite their high affinity symport of acidic amino acids, the endogenous levels of glutamate in Muller's cells are among the lowest in the retina. (5) GABAergic neurons possess intermediate levels of glutamate immunoreactivity. Quantitative immunocytochemistry coupled with digital image analysis allows estimates of intracellular glutamate levels. Photoreceptors and bipolar and ganglion cells contain from 1 to 10 mM glutamate. The bipolar and ganglion cell populations maintain high intracellular glutamate concentrations, averaging about 5 mM, whereas red-sensitive and green-sensitive cones apparently maintain lower levels. Importantly, photoreceptor glutamate levels are extremely volatile, and in vitro maintenance is required to preserve cone glutamate immunoreactivity in the goldfish. GABAergic horizontal and amacrine cells contain about 0.3–0.7 mM glutamate, which matches the values predicted from the Km of glutamic acid decarboxylase. Muller's cells and non-GABAergic amacrine cells contain less than 0.1 mM glutamate. Though Muller's cells are known to possess potent glutamate symport, they clearly possess equally potent mechanisms for maintaining low intracellular glutamate concentrations.
The posterior nervous system of the nematode Caenorhabditis elegans: serial reconstruction of identified neurons and complete pattern of synaptic interactions
Authors: Hall DH , Russell RL
Published: March 2018

Serial-section electron microscopy has been used to reconstruct the cellular architecture of the posterior nervous system of the nematode Caenorhabditis elegans. Each of 40 neurons in the tail of the adult hermaphrodite can be reproducibly and unambiguously identified by a set of morphological features, including cell body position, fiber geometry and size, and staining properties. A complete list of synapses has been assembled for 2 isogenic animals, and these lists are compared with a third isogenic animal reconstructed by White et al. (1986). The set of neurons and their pattern of synaptic interactions is simple and reproducible. Most of the cells are involved in sensory transduction or in local signal processing to relay signals via a few interneurons to motoneurons and thence to body muscles. Because the tail neurons are well separated and fairly reproducible in position, the hermaphrodite tail lends itself to laser-ablation studies of sensory processing (cf. Chalfie et al., 1985). Most of the synapses in the tail are concentrated in the preanal ganglion. Among the approximately 150 synapses there, about 85% are dyadic chemical synapses. The dyadic synapses are involved in reproducible patterns that have several interesting features. Most neurons synapse onto a few preferred pairs of target cells, in patterns that suggest a combinatorial model of synapse specification that may be open to genetic analysis. Furthermore, most dyadic contacts A----B,C fit a pattern in which the 2 postsynaptic partners are involved elsewhere in unidirectional synapses B----C. Thus, the dyadic synapse may serve to diverge sensory signals into parallel pathways, which then reconverge. This divergence/reconvergence pattern eventually directs processed sensory signals to the ventral cord interneurons PVCL and PVCR. About 80–90% of the synapses fall into repeated classes of synapses. Many of the remaining synapses are widely scattered and irreproducible from one animal to the next. Some of these contacts may be developmental mistakes reflecting a degree of “noise” in synapse specification (Waddington, 1957).
Photoinactivation of the crayfish segmental giant neuron reveals a direct giant-fiber to fast-flexor connection with a chemical component
Authors: Fraser K , Heitler WJ
Published: March 2018

The escape tail flip of the crayfish is “commanded” by 2 sets of giant- fiber (GF) interneurons. In each hemisegment, these drive the motor giant (MoG) abdominal flexor motor-neuron through a monosynaptic electrical connection, but the remaining 8 or 9 fast-flexor (FF) motorneurons receive most of their input via a disynaptic electrical pathway through the segmental giant (SG) neuron. We have investigated a monosynaptic GF-FF pathway, which operates in parallel to the disynaptic GF-SG-FF pathway, by using dye-mediated photoinactivation to remove the SGs from the tail-flip circuit. SG photoinactivation involves an initial broadening of the spike, leading to a long- duration, massively depolarized plateau. This is followed by loss of spike capability, a gradual reduction in the resting potential, and eventual total loss of electrical responsiveness. After bilateral photoinactivation of the SGs, a spike in one set of GFs, the medial giants (MGs), produces little if any effect in FFs in any ganglion. A spike in the other set, the lateral giants (LGs), produces an EPSP in FFs with a declining anterior-to-posterior segmental gradient in amplitude. These differences in LG and MG outputs, which are obscured in the intact circuit by the common MG/LG-SG-FF pathway, give clues to a probable early evolutionary form of the circuit. The LG-FF connection in anterior ganglia has a significant electrical component. However, it also has an apparent monosynaptic chemical component, as revealed by the response to saline containing cadmium ions, and to cooling the preparation. This is the first physiological evidence for chemical output from a crayfish GF.
Synaptic transmission between individual pyramidal neurons of the rat visual cortex in vitro
Authors: Mason A , Nicoll A , Stratford K
Published: March 2018

Synaptic transmission between pairs of neurons in layer 2/3 of in vitro slices from the rat visual cortex was studied by dual intracellular recording. The intrinsic electrophysiological properties of these neurons suggested that they were pyramidal cells. More than 1/3 of the total number of synaptically connected neurons were stained by intracellular iontophoresis of biocytin, and all had pyramidal morphology. Postsynaptic potentials (PSPs) were evoked by single action potentials (elicited by current injection) in presynaptic cells. Measurements of PSP latency, amplitude, and shape were made on spike- triggered averages. Forty-eight synaptic connections were found out of a possible total of 549, equivalent to a probability of about 0.09. For these 48 connections, the distance between the 2 impalement sites ranged from 50 to 340 microns. All PSPs were depolarizing at rest (-74 +/- 5 mV, mean +/- SD), and all 14 of the PSPs that were also recorded with the postsynaptic cell depolarized to around action potential threshold (about -55 mV) remained depolarizing. PSPs had short latencies (1.2 +/- 0.6 msec, mean +/- SD), suggesting that they were mediated by monosynaptic pathways. Peak amplitudes of the averaged PSPs varied widely (range, 0.05–2.08 mV), but the majority were less than 0.5 mV. PSPs decayed exponentially with time constants that were correlated with, but slightly longer than, the membrane time constants of the postsynaptic cells measured using injected current pulses. Four cell pairs were connected reciprocally, and 6 examples of convergent input were found in which a single cell was postsynaptic to more than 1 presynaptic cell. Trial-to-trial fluctuations in PSP amplitude were analyzed for 16 synaptic connections. The amplitude of the PSP evoked by a presynaptic action potential fluctuated more than could be accounted for by the background noise.
Acetylcholine release from the rabbit retina mediated by NMDA receptors
Authors: Linn DM , Massey SC
Published: March 2018

The cholinergic amacrine cells of the rabbit retina may be labeled with 3H-choline, and the activity of the cholinergic population may be monitored by following the release of 3H-ACh. In magnesium-free medium, the glutamate analog NMDA caused massive ACh release, up to 50x the basal efflux. Magnesium blocked the NMDA-evoked release of ACh with an IC50 of 151 microM. The NMDA-evoked release of ACh was unchanged in glycine-free medium or in the presence of 500 microM glycine. However, the block of NMDA-evoked release by 7-chlorokynurenic acid (7-Cl-Kyn) was reversed by exogenous glycine. This suggests that the NMDA receptors mediating ACh release possess an allosteric glycine binding site, but under normal conditions, it is saturated by endogenous glycine. Submaximal doses of NMDA were used to determine the potency of NMDA antagonists and their specificity was established with submaximal doses of other glutamate agonists. DL-2-amino-7-phosphonoheptanoate (DL- AP-7) was a competitive NMDA antagonist, with an IC50 of 33 microM and (+)5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801) was a noncompetitive NMDA antagonist, with an IC50 of 10.6 nM. Neither antagonist blocked the light-evoked release of ACh from the retina. Furthermore, light stimulation did not activate the use-dependent block characteristic of MK-801, indicating that the endogenous transmitter did not open the NMDA channel. These results suggest that NMDA receptors do not mediate the physiological input to cholinergic amacrine cells in the rabbit retina.
Intracellular Ca2+ suppressed a transient potassium current in hippocampal neurons
Authors: Chen QX , Wong RK
Published: March 2018

The effects of intracellular Ca2+ (Ca2+i) on K+ currents in hippocampal cells were examined using acutely isolated cells obtained from adult guinea pigs. Whole-cell voltage-clamp recordings were carried out in a configuration that allowed a continuous perfusion of the intracellular medium. Recording media were made to block inward currents and allowed selective activation of K(+)-dependent outward currents. Voltage- dependent outward currents consisted of an initial rapidly decaying component followed by a sustained component. The time constant of decay of the transient current was about 25 msec, and previous studies (Numann et al., 1987) showed that the kinetic and pharmacological properties of this current closely resembled the A current recorded in invertebrate neurons (Connor and Stevens, 1971; Thompson, 1982). Intracellular perfusion of hippocampal cells with a solution containing elevated Ca2+ (about 4.5 x 10(-4) M) elicited outward currents at the holding potential (-45 to -55 mV) and produced changes in voltage- dependent K+ currents. The transient outward current (IA) activated by depolarization was suppressed with increases in Ca2+i. Delayed, sustained K+ currents were greatly potentiated. Data also showed that, among the 3 effects elicited by Ca2+i, suppression of IA was most sensitive to Ca2+i elevation. Previous results (Numann et al., 1987) showed that IA had a lower threshold (about -45 mV) than sustained currents (about -40 mV). By using low levels of depolarization (-40 mV), IA can be selectively activated, and the suppressive effect of Ca2+i on IA was confirmed on the kinetically isolated IA.(ABSTRACT TRUNCATED AT 250 WORDS)
NGF rescues substance P expression but not neurofilament or tubulin gene expression in axotomized sensory neurons
Authors: Wong J , Oblinger MM
Published: March 2018

A reduction in the supply of retrogradely transported NGF has been proposed as a possible signal for the axotomy response in dorsal root ganglion (DRG) neurons. Components of the axotomy response that have previously been well characterized in axotomized DRG cells include changes in cytoskeletal gene expression and changes in the expression of neurotransmitters/neuromodulators such as substance P. In this study, we examined the role of NGF in the axotomy response by examining protein synthesis and mRNA levels of the low-MW neurofilament protein (NF-L) and beta-tubulin in DRG cells at 1, 7, and 12 d after axotomy with and without continuous administration of exogenous NGF. We also examined substance P levels in the DRG by immunocytochemistry under the same experimental conditions. Sciatic nerves of adult male rats were unilaterally transected at the midthigh level, and the proximal nerve stumps were placed into Silastic tubes connected to osmotic minipumps that were filled with biologically active NGF. NGF (0.5 mg/ml in saline) was continuously infused (0.5 microliter/hr) onto the proximal stumps of transected sciatic nerves for 1–12 d. Control animals were prepared in an identical fashion except that the nerves were treated with saline alone. At death, DRGs were removed from the animals; the L4 experimental DRGs (axotomized) and contralateral L4 DRGs (uninjured) were used immediately for protein synthesis experiments, while the experimental and contralateral L5 DRGs were fixed in 4% paraformaldehyde and subsequently used for in situ hybridization and immunocytochemistry. From another set of experimental animals, the L4 and L5 DRGs were harvested and used for total RNA isolation and RNA blotting experiments. Immunocytochemical studies using a polyclonal antibody to substance P showed that the immunodetectable levels of this peptide decreased to undetectable levels in DRG neurons after axotomy and saline administration. However, in axotomized neurons treated with NGF, the level of immunodetectable substance P did not decrease, but instead, increased over even that present in normal DRG neurons. Pulse labeling of DRGs with 35S-methionine:cysteine followed by 2-dimensional (2D) gel electrophoresis and fluorography revealed that the synthesis of neurofilament (NF) proteins was decreased, while that of tubulin was increased, 12 d after sciatic nerve transection. NGF administration to axotomized neurons did not alter this pattern. Quantitative analysis of in situ hybridizations of DRG neurons and RNA blot analysis with cDNA probes specific for NF-L and beta-tubulin mRNAs showed that NGF treatment of axotomized DRGs did not significantly affect cytoskeletal gene expression at the mRNA level.(ABSTRACT TRUNCATED AT 400 WORDS)
Stable clathrin: uncoating protein (hsc70) complexes in intact neurons and their axonal transport
Authors: Black MM , Chestnut MH , Pleasure IT , Keen JH
Published: March 2018

We have studied the organization of clathrin during its transport in axons. Using immunoprecipitation techniques we have confirmed earlier findings that clathrin is transported as part of slow component b, but we also detect small amounts of clathrin in fast component. As fast component is known to correspond to the transport of membraneous material, including coated vesicle membrane components, our findings suggest that some clathrin in axons undergoes transport in the form of coated membranes and that a portion of the clathrin delivered to axons and axon terminals arrives by way of fast component. The organizational form of clathrin in slow component b (SCb) was examined in more detail, as it is thought to represent a non-membrane-associated species, is relatively long-lived, and at any instant represents the major transport species in axons. We used nondenaturing immunoprecipitation methods with stringent wash procedures to identify other SCb proteins that interact with clathrin. The immunoprecipitates contained major labeled bands that corresponded to clathrin heavy and light chains, along with a prominent 70-kDa band and several minor bands that ranged in apparent Mr from 70,000 to 150,000; the 70-kDa band was shown to be the ATP-dependent uncoating protein by two-dimensional gel electrophoresis. A very similar profile of polypeptides was also immunoprecipitated from extracts of cultured neurons. The results from a variety of control immunoprecipitations, including the use of antisera preadsorbed with purified clathrin trimers or clathrin light chains, indicate that coprecipitation of clathrin and uncoating protein with the other 70,000–150,000-Da polypeptides from SCb reflects specific interactions. Including exogenous uncoating protein in the lysis buffer had no detectable effect on the levels of endogenous uncoating protein recovered in the immunoprecipitates, indicating that complexes of clathrin, uncoating protein, and the other coimmunoprecipitating SCb protein existed in the intact neurons prior to lysis. Finally, a specific and functional association is further supported by the release of uncoating protein, but not the other 70,000– 150,000-Da polypeptides, from the immunoprecipitated complexes on the addition of ATP. Collectively, these observations provide the first direct evidence of interaction between clathrin and uncoating protein in intact cells, lend strong support to the concept that uncoating protein plays an intimate role in clathrin dynamics within cells, and reveal a family of 70,000–150,000-Da polypeptides that form a stable nonmembranous association with clathrin in intact cells.
Mutual inhibition among neural command systems as a possible mechanism for behavioral choice in crayfish
Authors: Edwards DH
Published: March 2018

Mutual inhibition among behavioral command systems frequently has been suggested as a possible mechanism for switching between incompatible behaviors. Several neural circuits in crayfish that mediate incompatible behaviors have been found to interact through inhibition; this accounts for increased stimulus threshold of one behavior (e.g., escape tailflip) during performance of others (eating, walking, defense). To determine whether mutual inhibition between command systems can provide a mechanism that produces adaptive behavior, I developed a model crayfish that uses this mechanism to govern its behavioral choices in a simulated world that contains a predator, a shelter, and a food source. The crayfish uses energy that must be replaced by eating while it avoids capture by the predator. The crayfish has seven command systems (FORAGE, EAT, DEFENSE, RETREAT, ESCAPE, SWIM, HIDE) that compete through mutual inhibition for control of its behavior. The model crayfish was found to respond to changing situations by making adaptive behavioral choices at appropriate times. Choice depends on internal and external stimuli, and on recent history, which determines the pattern of those stimuli. The model's responses are unpredictable: small changes in the initial conditions can produce unexpected patterns of behavior that are appropriate alternate responses to the stimulus conditions. Despite this sensitivity, the model is robust; it functions adaptively over a large range of internal and external parameter values.
Metabolic and contractile protein expression in developing rat diaphragm muscle
Authors: Kelly AM , Rosser BW , Hoffman R , Panettieri RA , Schiaffino S , Rubinstein NA , Nemeth PM
Published: March 2018

Progressive changes in myosin isozyme expression and in energy- generating enzyme activities were followed in the diaphragm and, for comparison, in axial and appendicular muscles of rats from 18 d gestation to maturity. Native myosins were characterized by pyrophosphate gel electrophoresis. Myosin heavy-chain (MHC) isozymes were measured with ELISA using monoclonal antibodies and were localized by immunocytochemistry. RNA transcripts for the MHCs were demonstrated on Northern blots and by RNase protection assays. Quantitative activities of malate dehydrogenase (MDH), beta-hydroxyacyl CoA dehydrogenase (beta OAC), 1-phosphofructokinase (PFK), lactate dehydrogenase (LDH), creatine kinase (CK), and adenylokinase (AK) were measured in muscle homogenates and in individual fibers by fluorometric pyridine nucleotide-dependent assays. Compared to limb muscles, expression of neonatal myosin in the diaphragm is precocious. Neonatal MHC mRNA is prominent in the diaphragm at 19 d gestation, and neonatal myosin is the major MHC isoform present at birth. Slow and fast IIa MHCs are also present at birth. Transcripts for IIa MHC are detectable in the diaphragm at 21 d gestation and are upregulated at birth. Comparable signal for IIa MHC mRNA is not found in the gastrocnemius until 10 d postpartum. Adult fast IIb MHC mRNA was detected only as a faint signal at 30–40 d in the diaphragm and then disappeared. Results indicate that a separate phenotype, the IIx type, matures late in diaphragmatic development. The activities of enzymes representing all of the major energy pathways are higher in the fetal diaphragm than in the fetal hindlimb muscles. For example, beta OAC had sixfold higher activity in the diaphragm than in the extensor digitorum longus (EDL) muscle at birth, activity in the diaphragm than in the extensor digitorum longus (EDL) muscle at birth.
Synaptic transmission in decentralized axons of rock lobster
Authors: Parnas I , Dudel J , Atwood HL
Published: March 2018

Axons of the lobster deep abdominal extensor muscles were cut, and the resulting effects on their synaptic properties were observed. Decentralized axons continued to conduct action potentials and to release transmitter for at least a year after the cut. In controls, the single quanta were brief, and their decay phase could be fit by a single exponent, with a time constant of about 2 msec. Quanta of “cut axons” were slower, and their decay phase could not be fit by a single exponent. At midamplitude, the duration of the cut-axon quanta varied between 1.6 and 5.8 msec, as opposed to 0.6–2.8 msec in controls. Synaptic delay histograms were taken as a measure of time course of evoked release. In controls, evoked release lasted less than 10 msec at 14 degrees C. In cut axons, release lasted up to 10 times longer. The duration of release was not affected by tetrodotoxin, membrane depolarization, or hyperpolarization. It appears that the basic mechanism that controls the time course of evoked release is altered in degenerating terminals.
Relation of cortical cell orientation selectivity to alignment of receptive fields of the geniculocortical afferents that arborize within a single orientation column in ferret visual cortex
Authors: Chapman B , Zahs KR , Stryker MP
Published: March 2018

Neurons in the primary visual cortex of higher mammals are arranged in columns, and the neurons in each column respond best to light-dark borders of particular orientations. The basis of cortical cell orientation selectivity is not known. One possible mechanism would be for cortical cells to receive input from several lateral geniculate nucleus (LGN) neurons with receptive fields that are aligned in the visual field (Hubel and Wiesel, 1962). We have investigated the relationship between the arrangement of the receptive fields of geniculocortical afferents and the orientation preferences of cortical cells in the orientation columns to which the afferents provide visual input. Radial microelectrode penetrations were made into primary visual cortex of anesthetized adult sable ferrets. Cortical cells were recorded throughout the depth of the cortex, and their orientation preferences were determined. Cortical cell responses were then eliminated by superfusion of the cortex with either kainic acid (Zahs and Stryker, 1988) or muscimol. After the drug treatment, responses from many single units with distinct receptive fields were recorded. These responses were presumed to be those of geniculocortical afferents, because they had the response properties characteristic of LGN neurons, and because they could be recorded only in cortical layers that receive geniculate input. In 16 of 18 cases, the afferent receptive fields recorded in a single penetration covered an elongated region of visual space. In these penetrations, the best-fit line through the centers of the afferent receptive fields generally paralleled the preferred orientation of cortical cells recorded at the same site in cortex. These results are consistent with the Hubel and Wiesel (1962) model for the construction of oriented visual cortical receptive fields from geniculate inputs with aligned receptive fields.
Gap junctions between cultured astrocytes: immunocytochemical, molecular, and electrophysiological analysis
Authors: Dermietzel R , Hertberg EL , Kessler JA , Spray DC
Published: March 2018

The properties of astroglial gap junction channels and the protein that constitutes the channels were characterized by immunocytochemical, molecular biological, and physiological techniques. Comparative immunocytochemical labeling utilizing different antibodies specific for liver connexin 32 and connexin 26 and antibodies to peptides corresponding to carboxy-terminal sequences of the heart gap junction protein (connexin 43) indicates that the predominant gap junction protein in astrocytes is connexin 43. The expression of this connexin in cultured astrocytes was also established by Western and Northern blot analyses. Cultured astrocytes expressed connexin 43 mRNA and did not contain detectable levels of the mRNAs encoding connexin 32 or connexin 26. Further, the cells contained the same primary connexin 43 translation product and the same phosphorylated forms as heart. Finally, electrophysiological recordings under voltage-clamp conditions revealed that astrocyte cell pairs were moderately well coupled, with an average junctional conductance of about 13 nS. Single-channel recordings indicated a unitary junctional conductance of about 50–60 pS, which is of the same order as that found in cultured rat cardiac myocytes, where the channel properties of connexin 43 were first described. Thus, physiological properties of gap junction channels appear to be determined by the connexin expressed, independent of the tissue type.
Hyperalgesia during acute opioid abstinence: evidence for a nociceptive facilitating function of the rostral ventromedial medulla
Authors: Kaplan H , Fields HL
Published: March 2018

Naloxone-precipitated opioid abstinence is associated with enhancement of reflex responses to noxious stimulation (hyperalgesia). The present experiments in lightly anesthetized rats were designed to determine (1) whether neurons in the rostral ventromedial medulla (RVM) contribute to this enhancement, and (2) whether this enhancement is due to removal of an inhibitory modulatory influence or to activation of a facilitatory influence. In the first experiment, 10 micrograms of morphine was microinjected into the RVM; subsequent administration of naloxone (1 mg/kg, i.v.) shortened tail-flick latency. This is evidence that a synaptic action of opioids within the RVM can contribute to hyperalgesia. In the second experiment, systemic administration of morphine (2 mg/kg, i.v.) was followed by systemic administration of naloxone (1 mg/kg, i.v.), which produced a significant hyperalgesia that could be markedly attenuated by microinjection of 10 micrograms lidocaine into the RVM. That inactivation of RVM reduces the hyperalgesia indicates that the CNS is capable of generating a facilitating action on nociceptive transmission. Previous studies from this laboratory have indicated that a population of RVM neurons, on- cells, shows increased activity during opioid abstinence. The present experiments support the hypothesis that RVM on-cells exert a facilitating influence on nociceptive transmission.
Olfactory bulb granule cell aggregates: morphological evidence for interperikaryal electrotonic coupling via gap junctions
Authors: Reyher CK , Lubke J , Larsen WJ , Hendrix GM , Shipley MT , Baumgarten HG
Published: March 2018

Anaxonic interneurons of the granule cell type in the mammalian main olfactory bulb (MOB) are characterized by prominent membrane specializations, which include reciprocal, interdendritic chemical and electrical synapses; however, the latter are thought to be restricted to the external plexiform layer (EPL) and connect granule-mitral- and granule-tufted-cell dendrites (Landis et al., 1974). The present study focuses on interperikaryal membrane specializations between tangentially oriented aggregates of granule microneurons in the lamina granularis interna (IGL). Both infraprimate (Rattus norvegicus, Gerbillus perpallidus) and primate species (Callithrix jacchus) were studied using the following methods: (1) transmission electron microscopy (TEM), (2) freeze-fracture analysis, (3) light and TEM immunohistochemistry using affinity-purified antibodies directed against the connexin-32 or connexin-43 carboxy tail fragment of the gap- junction protein (GJP), and (4) intracellular Lucifer yellow injections in fixed tissue (LYF technique). Freeze-fracture replicas of the MOB- IGL showed that adjacent granule cell perikarya have numerous particle aggregates on the cytoplasmic membrane; in terms of their structure and arrangement, such particles are characteristic of gap junctions. The existence of junctional membranes was substantiated by application of antibodies against GJP demonstrating punctate immunoreactivity, frequently confined to the interperikaryal plasmalemmae of granule cells in the IGL and their dendritic processes in the EPL. Upon TEM analysis, GJP-like immunoreactivity was additionally found in membranous organelles, including Golgi apparati and associated vesicular components. In order to test the permeability of identified membrane specializations, the LYF technique was used, which resulted in bright fluorescence of the perikaryal and dendritic components of the transsomatically injected neuron and staining of neighboring neurons with similar morphology. These findings imply that small molecules can diffuse across the interperikaryal membrane specializations. The existence of gap junctions between granule cell perikarya suggests that there is a significant, low-resistance electrical transmission between aggregated granule cells. This coupling might permit synchronization of neural discharge among small aggregates of these neurons. Gap junctions between granule cells may also serve signaling functions associated with the protracted period of granule cell development.
The effect of tau antisense oligonucleotides on neurite formation of cultured cerebellar macroneurons
Authors: Caceres A , Potrebic S , Kosik KS
Published: March 2018

Tau, a microtubule-associated protein (MAP) enriched in axons, may have a role in the generation and maintenance of an axonal morphology. Neurons from embryonic day 15 rat cerebellum in culture elaborate two morphologically distinct neurite populations--one with nontapering, elongated axonlike neurites and the other with tapered dendritelike neurites that branch frequently and are selectively stained with antibodies to MAP2. Tau antisense oligonucleotides were utilized in two ways: (1) continuous application of antisense every 24 hr for variable periods of time or (2) application of antisense that was delayed until neurite differentiation was underway. In both cases, 24 hr after the administration of the antisense, tau protein was not detected immunocytochemically. When the antisense was given continuously directly after plating, the neurites persisted as simple minor outgrowths. When antisense was added 72 hr after plating, axonlike neurites were lost, while the remaining neurites continued to grow and increase in complexity. We concluded that the initial establishment of an elongated axonlike neurite is a prerequisite for further neurite maturation; however, once the axon is established, the remaining neurites are able to grow independently and assume a tapered dendritelike appearance.
(D-Ala2)deltorphin II: D1-dependent stereotypies and stimulation of dopamine release in the nucleus accumbens
Authors: Longoni R , Spina L , Mulas A , Carboni E , Garau L , Melchiorri P , Di Chiara G
Published: March 2018

In order to investigate the relative role of central delta- and mu- opioid receptors in behavior, the effects of (D-Ala2)deltorphin II, a natural delta-opioid peptide, and PL017, a beta-casomorphin derivative specific for mu receptors, were compared after local intracerebral and intraventricular administration. Intracerebral infusion of the two peptides was done bilaterally in the limbic nucleus accumbens and in the ventral and dorsal caudate putamen of freely moving rats through chronic intracerebral cannulas. After intra-accumbens infusion, the two peptides elicited marked but opposite behavioral effects: while (D- Ala2)deltorphin II evoked dose-dependent motor stimulation characterized by locomotion, sniffing, and oral stereotypies, PL017 elicited motor inhibition with rigidity and catalepsy. These effects were site specific because they could not be evoked from the ventral or from the dorsal caudate. Low doses of naloxone (0.1 mg/kg, s.c.) blocked the effects of PL017 but not those of (D-Ala2)deltorphin II, which instead were reduced by high doses of naloxone (1.0 mg/kg) and by the putative delta-antagonist naltrindole; this drug failed to affect the catalepsy induced by PL017. Therefore, while (D-Ala2)deltorphin II effects were delta-mediated, PL017 effects were mu-mediated. Blockade of dopamine D1 receptors by SCH 23390 abolished (D-Ala2)deltorphin II effects, while blockade of dopamine D2 receptors by raclopride or by haloperidol was without effect. Local application by reverse dialysis of (D-Ala2)deltorphin II (5 microM) to the accumbens resulted in a naloxone-sensitive increase of extracellular dopamine concentrations; these effects could not be evoked from the caudate, nor by PL017 in the accumbens. Intracerebroventricular administration of (D-Ala2)deltorphin II or of PL017 elicited behavioral effects qualitatively similar to those obtained from the accumbens.
Vestibular-nerve inputs to the vestibulo-ocular reflex: a functional- ablation study in the squirrel monkey
Authors: Minor LB , Goldberg JM
Published: March 2018

Advantage was taken of differences in the electrical excitability of vestibular-nerve fibers to characterize the afferent input to the canal- related vestibulo-ocular reflex. Large anodal (inhibitory) currents, when delivered to both ears, result in a selective, reversible ablation of irregular afferents. Their background discharge and responses to head rotations are temporarily abolished. The same currents have less effect on the background discharge and no effect on the rotational sensitivity of regular afferents. Eye movements were evoked by head rotations in alert monkeys. The ablating currents did not alter the ocular responses to sinusoidal head rotations in yaw or pitch planes. Responses to rapid changes in head velocity were similarly unaffected. It is concluded that irregular afferents do not make a net contribution to the reflex. Slow-phase eye movements evoked by unilateral galvanic currents are consistent with this conclusion. The results are incorporated into a systems model of the reflex. There are three conclusions from the model: (1) the signal to motoneurons consists of the sum of three components related, respectively, to head velocity, eye position, and filtered eye velocity; (2) regular afferents provide the best match to the dynamic requirements of the reflex; and (3) the central pathways responsible for all three signal components receive regular inputs.
Stretched and upside-down maps of auditory space in the optic tectum of blind-reared owls; acoustic basis and behavioral correlates
Authors: Knudsen EI , Esterly SD , du Lac S
Published: March 2018

Vision during early life plays an important role in calibrating sound localization behavior. This study investigates the effects of visual deprivation on sound localization and on the neural representation of auditory space. Nine barn owls were raised with eyelids sutured closed; one owl was congenitally anophthalmic. Data from these birds were compared with data from owls raised with normal visual experience. Sound localization behavior was significantly less precise in blind- reared owls than in normal owls. The scatter of localization errors was particularly large in elevation, though it was abnormally large in both dimensions. However, there was no systematic bias to the localization errors measured over a range of source locations. This indicates that the representation of auditory space is degraded in some way for blind- reared owls, but on average is properly calibrated. The spatial tuning of auditory neurons in the optic tectum was studied in seven of the blind-reared owls to assess the effects of early visual deprivation on the neural representation of auditory space. In normal owls, units in the optic tectum are sharply tuned for sound source location and are organized systematically according to the locations of their receptive fields to form a map of auditory space. In blind-reared owls, the following auditory properties were abnormal: (1) auditory tuning for source elevation was abnormally broad, (2) the progression of the azimuths and elevations of auditory receptive fields across the tectum was erratic, and (3) in five of the seven owls, the auditory representation of elevation was systematically stretched, and in the two others large portions of the representation of elevation were flipped upside down. The following unit properties were apparently unaffected by blind rearing: (1) the sharpness of tuning for sound source azimuth, (2) the orientation of the auditory representation of azimuth, and (3) the mutual alignment of the auditory and visual receptive fields in the region of the tectum representing the area of space directly in front of the animal. The data demonstrate that the brain is capable of generating an auditory map of space without vision, but that the normal precision and topography of the map depend on visual experience. The space map results from the tuning of tectal units for interaural intensity differences (IIDs) and interaural time differences (ITDs; Olsen et al., 1989).
Neuropil threads of Alzheimer's disease show a marked alteration of the normal cytoskeleton
Authors: Perry G , Kawai M , Tabaton M , Onorato M , Mulvihill P , Richey P , Morandi A , Connolly JA , Gambetti P
Published: March 2018

Abnormal neurites, neuropil threads, are a widespread and characteristic lesion of Alzheimer's disease likely to play a major role in the cognitive impairment of this disease. Contrary to normal neurites, neuropil threads contain straight and paired helical filaments that contain the microtubule-associated protein tau and ubiquitin. It is not known whether these abnormal filaments are added to or replace the normal cytoskeleton. In this study, we examined the fine structure of neuropil threads and carried out a morphometric analysis of the neurofilaments and abnormal filaments contained in the neuropil threads by using an antiserum to tau and colloidal gold immuno- electron microscopy. Almost 70% of the neuropil threads contained straight or paired helical filaments with no neurofilaments. The total number of filaments in each neuropil thread remained essentially unchanged either when straight or paired helical filaments were present alone or when they coexisted either together or with neurofilaments. When the three types of filaments were expressed as a proportion of the total, a linear inverse correlation was found between neurofilaments and straight filaments as well as between straight and paired helical filaments. Approximately 10% of the neuropil threads were found to be myelinated axons. It is concluded that straight filaments are likely to replace neurofilaments, that they in turn might be replaced by paired helical filaments, and that this process occurs in axons as well as dendrites.