bluuebunny/crossref_metadata_2025 huggingface dataset:
Orientation of neurite growth by extracellular electric fields
Authors: Patel N , Poo MM
Published: March 2018

Extracellularly applied steady electric fields of 0.1 to 10 V/cm were found to have marked effects on the neurite growth of single dissociated Xenopus neurons in culture: (1) neurites facing the cathode showed accelerated growth, while the growth of those facing the anode was reduced. Neurites growing relatively perpendicular to the field axis were prompted to curve toward the cathode. (2) More neurites appeared to be initiate from the cathodal side of the cell. (3) The number of neurite-bearing neurons per culture and the average neurite length were increased. These effects are absent in cultures treated with electric fields of similar strength but alternating polarity and cannot be attributed either to a gradient of extracellular diffusible substances or to the flow of culture medium produced by the field. The field effects are reversible: (1) removal of the electric field resulted in the loss of neurite orientation in a few hours and (2) reversal of the polarity of the electric field led to a rapid reversal in the neurite orientation. To determine the cellular loci of these field effects, we treated the neurons with a number of pharmacological agents or altered their ionic environments. Incubation with concanavalin A (Con A) was found to abolish these filed effects completely. Since the binding of Con A to the neuronal surface was shown to prevent field-induced accumulation of the Con A receptors toward the cathodal side of these neurons, our finding is accumulation of the Con A receptors toward the cathodal side of these neurons, our finding is consistent with the notion that cathodal accumulation of growth-controlling surface glycoproteins by the field is the underlying mechanism of the field-induced orientation of neurite growth toward the cathode.
Patterns and kinetics of neurite extension from sympathetic neurons in culture are age dependent
Authors: Argiro V , Johnson MI
Published: March 2018

Long term (2- to 3-week) cultures of superior cervical ganglia (SCG) were established from rats and rat embryos ranging in age from 15 days of gestation (E15) to 279 days postnatal (P279). Cultures were grown on a collagen substratum and fed a serum-containing medium with added nerve growth factor. Radial outgrowth of neurites was measured as a function of time for up to 2 to 3 weeks. Computer-aided analysis generated estimates of onset time, initial rate, and subsequent changes in the rate of growth of these neurites. The explants from perinatal rats showed the fastest growth onset time (5 to 13 hr), fastest initial rate of growth (370 to 660 microns/d), and a decline in growth rate during the first 2 weeks in culture. The outgrowth from these perinatal explants was composed of many small fascicles. Neurites from the prenatal explants (E15 to E20) began to grow within 22 hr in vitro. Their rate of growth was lower initially (150 to 300 microns/d) but increased to equal the perinatal explant initial rate before again falling to an intermediate level (200 to 300 microns/d). The outgrowth from prenatal explants contained fewer larger fascicles. Postnatal explants had low initial rates of growth (70 to 176 microns/d) but exhibited an increasing growth rate in vitro, again approaching an intermediate rate of 200 to 250 microns/d after 2 to 3 weeks. Neurite outgrowth from the postnatal explants was delayed by an amount roughly correlated with the age of the animal advancing postnatal age but reached an asymptote of about 50 to 150 microns/d at about P30. The outgrowth was initially sparse but became denser with time in culture. Thus, in a culture system in which medium composition and growth substratum are held constant, marked differences can be observed in pattern, latency, initial rate, and subsequent changes in rate of neurite extension among SCG explants from different ages of rats and rat embryos.
The role of the tectal column in facilitation of amphibian prey- catching behavior: a neural model
Authors: Lara R , Arbib MA , Cromarty AS
Published: March 2018

We used computer analysis of differential equations to study the properties of a family of models of a unit of neural circuitry in the amphibian tectum, the tectal column. Computer experiments were used to discriminate among various hypotheses and to suggest new experiments. Particular attention was paid to physiological data on facilitation of amphibian prey-catching behavior which led us to model the facilitation in terms of dynamic activity in the tectal column rather than in terms of synaptic modification.
The blocking action of baclofen on excitatory transmission in the rat hippocampal slice
Authors: Olpe HR , Baudry M , Fagni L , Lynch G
Published: March 2018

The mode of action of baclofen on the physiology of the rat hippocampus was investigated by studying its effect on electrophysiological responses in the hippocampal slice preparation and by measuring biochemical parameters related to glutamate uptake, binding, and release. Baclofen inhibited, in a dose-dependent fashion, the dendritic field potentials in field CA1 produced by stimulation of the Schaffer commissural fiber system. The drug was inactive in this respect at concentrations of 10 and 100 nM but consistently reduced the amplitude of both the dendritic field potential and the population spike at a concentration of 1 microM. At a concentration of 25 microM, baclofen virtually abolished the dendritic and cell body responses to afferent stimulation. Recovery of field potentials required between 7 and 10 min following the addition of 1 microM baclofen. The levorotatory form of baclofen was much more potent in suppressing synaptic responses than was the dextrorotatory enantiomer. Baclofen, at a concentration of 5 microM, strongly antagonized the excitation of pyramidal neurons evoked by iontophoretically applied glutamate. The antagonism of the glutamate effect was much reduced when the slices were maintained in low calcium, high magnesium perfusion medium. Moreover, under low calcium conditions, baclofen did not interfere with the effects of bath-applied glutamate on antidromically elicited responses. Baclofen did not affect the Na+-dependent or Na+-independent binding of [3H]glutamate to crude synaptic membrane fractions from the hippocampus. However, at a concentration of 1 microM, it markedly inhibited potassium-induced release of [3H]glutamate from hippocampal synaptosomes. Taken together, the present results strongly suggest that baclofen suppresses synaptic responses in the Schaffer commissural fiber system of the hippocampus by blocking the release of an excitatory amino acid transmitter.
The arrangement of axons according to fiber diameter in the optic tract of the cat
Authors: Guillery RW , Polley EH , Torrealba F
Published: March 2018

The topographic distribution of axons according to diameter classes in the optic tract of the cat has been studied by light and electron microscopic methods. The subdivision of the tract into a ventral and a dorsal sector, with the former containing the thickest fibers of the tract, has been confirmed. In addition, there is a ventrolateral marginal zone of the tract which contains many of the finest axons and there are gradients of fiber diameters definable for thick and intermediate axons. The gradients show the thinner fibers lying rostral to the thicker fibers. The major points demonstrable by fiber diameter measurements have been confirmed by counts of myelin lamellae. Comparison of fiber distributions in the optic tract with the distributions in the optic nerve on the one hand and with the geniculate terminations on the other shows that, in the region of the optic chiasm, retinofugal axons are sorted not only according to their crossed or uncrossed pathway but also according to their fiber diameter class and thus are presorted in terms of their final destinations.
The effects of microstimulation and microlesions in the ventral and dorsal respiratory groups in medulla of cat
Authors: Speck DF , Feldman JL
Published: March 2018

The responses in respiratory outflow resulting from microstimulation and successive microlesions of the dorsal (DRG) and ventral (VRG) respiratory groups of neurons in the brainstem were studied in anesthetized, paralyzed, artificially ventilated cats. Microstimulation (2 to 120 Hz; 5 to 50 microA; 100 musec pulse duration) at almost every point within the DRG or VRG produced a bilateral short latency inhibition of phrenic nerve activity which had an onset latency of 4 to 9 msec and a duration of 4 to 24 msec. This global stereotyped phrenic inhibition was elicited by single pulses and often was accompanied by a postinhibitory excitation. In 48% (92/193) of the stimulation trials, trains of stimulus pulses during inspiration decreased the duration of inspiration. In 25% of the expiratory microstimulation trials, expiratory duration was increased and in 11%, expiration was shortened markedly by trains of pulses. Single shocks delivered to the right VRG or DRG produced a short latency excitation in the ipsilateral recurrent laryngeal nerve (RRL). This RRL excitation had an onset latency of 2 to 5 msec and a duration of 3 to 15 msec. Evidence suggests that the RRL excitation is due to a paucisynaptic activation of expiratory motoneurons in the caudal VRG. This activation is synchronous with the inhibition of inspiratory neurons in DRG and VRG. Despite the powerful short latency effects of microstimulation in VRG and DRG, extensive bilateral destruction of these neuronal populations had only modest effects on respiratory rhythm, while it decreased (or abolished) respiratory outflow in phrenic and recurrent laryngeal nerves. The combined results of the microstimulation and microlesion portions of this study suggest that a region (or regions) outside of the DRG and VRG might be important in the control of the respiratory pattern and that the DRG and VRG are important in determining the depth of inspiration; their role in generating respiratory rhythm needs to be critically re-examined.
The sites of origin brain stem neurotensin and serotonin projections to the rodent nucleus raphe magnus
Authors: Beitz AJ
Published: March 2018

The combined horseradish peroxidase retrograde transport-peroxidase- antiperoxidase immunohistochemical procedure was utilized in the present study to ascertain the sites of origin of serotonin and neurotensin projections to the rodent nucleus raphe magnus. The major serotonin inputs to the raphe magnus arise from the B-8 and B-9 groups of Dahlstrom and Fuxe (Dahlstrom, A., and K. Fuxe (1964) Acta Physiol. Scand. Suppl. 232 62: 1–55), the nucleus reticularis paragigantocellularis, and the nucleus reticularis gigantocellularis pars alpha. Neurotensinergic projections to the raphe magnus originate predominantly from the periaqueductal gray, the nucleus solitarius, the dorsal and ventral parabrachial nuclei, and the nucleus cuneiformis. The periaqueductal gray and the nucleus paragigantocellularis were found to provide both a neurotensin and a serotonin projection to this raphe nucleus. The present results indicate that several brain stem nuclei, which have been implicated previously in endogenous analgesia mechanisms, provide serotonergic and neurotensinergic input to the nucleus raphe magnus.
Effects of gamma-aminobutyric acid agonist and antagonist drugs on local cerebral glucose utilization
Authors: Palacios JM , Kuhar MJ , Rapoport SI , London ED
Published: March 2018

The [14C]2-deoxy-D-glucose method of Sokoloff et al. (Sokoloff, L., M. Reivich, C. Kennedy, M.H. Des Rosiers, C. S. Patlak, K. D. Pettigrew, O. Sakurada, and M. Shinohara (1977) J. Neurochem. 28:897–916) weas used to study local cerebral glucose utilization (LCGU) in rats treated with gamma-aminobutyric acid (GABA) agonist (muscimol and 4,5,6,7- tetrahydroisoxazolo[5,4-C]pyridin-3-ol, THIP) and antagonist (bicuculline) drugs. It was of interest to determine if the pattern of LCGU responses to GABA agonists and antagonists administered systemically in vivo would reflect the known distributions of markers for central GABAergic synapses. The patterns of LCGU responses to muscimol and THIP generally were similar. Most brain regions showed dose-dependent decreases in LCGU; others showed no effects; but the red nucleus showed an increase. The GABA antagonist bicuculline produced convulsions and variable LCGU responses, depending on the time of administration. Bicuculline also partially antagonized the depressant effects of muscimol of LCGU. The magnitudes and distribution of in vivo cerebral metabolic responses to specific GABA agonists were not correlated simply with markers for GABAergic synapses. This lack of correlation indicates that additional factors, such as neural circuitry, regulate the LCGU responses to GABAergic drugs.
Immunocytochemical and ultrastructural differentiation between Met- enkephalin-, Leu-enkephalin-, and Met/Leu-enkephalin-immunoreactive neurons of feline gut
Authors: Larsson LI , Stengaard-Pendersen K
Published: March 2018

Selective immunocytochemical procedures, applied at the light and electron microscopical level, have resulted in the identification of three types of enkephalin-immunoreactive nerves in the feline gastrointestinal tract, including neurons containing either Met- or Leu- enkephalin immunoreactivity or those simultaneously storing both Met- and Leu-enkephalin immunoreactants. The three different types of enkephalin nerves show a similar distribution in the gut with the exception of the small arteries which frequently appear to be innervated only by Met-enkephalin-immunoreactive terminals. Electron microscopical identification using optimally fixed, osmicated, and contrasted tissue revealed ultrastructural differences between the three types of nerve terminals. Thus, the terminals contained variable proportions of large granular and large opaque p-type vesicles. Large granular vesicles were associated predominantly with Met-enkephalin immunoreactions. It is presently impossible to decide whether the different p-type vesicles reflect a continuous processing of a precursor contained within them or whether they are truly different in peptide content. Thus, granular vesicles had cores of either high or medium electron density. Medium electron-dense granular vesicles were sometimes difficult to distinguish from large opaque vesicles suggesting the existence of possible transitional forms. All types of enkephalin terminals also contained small clear (“cholinergic-like”) vesicles which were not immunoreactive to enkephalins. These data indicate that multiple enkephalin precursors/enkephalin-containing peptides occur in the gut or, alternatively or additionally, that a common Met/Leu-enkephalin precursor is processed differently in different neurons.
Synthesis an accumulation of putative neurotransmitters by cultured neural crest cells
Authors: Maxwell GD , Sietz PD , Rafford CE
Published: March 2018

The events mediating the differentiation of embryonic neural crest cells into several types of neurons are incompletely understood. In order to probe one aspect of this differentiation, we have examined the capacity of cultured quail trunk neural crest cells to synthesize, from radioactive precursors, and store several putative neurotransmitter compounds. These neural crest cultures develop the capacity to synthesize and accumulate acetylcholine and the catecholamines norepinephrine and dopamine. In contrast, detectable but relatively little synthesis and accumulation of 5-hydroxytryptamine gamma- aminobutyric acid, or octopamine from the appropriate radiolabeled precursors were observed. The capacity for synthesis and accumulation of radiolabeled acetylcholine and catecholamines is very low or absent at 2 days in vitro. Between 3 and 7 days in vitro, there is a marked rise in both catecholamine and acetylcholine accumulation in the cultures. These findings suggest that, under the particular conditions used in these experiments, the development of neurotransmitter biosynthesis in trunk neural crest cells ijs restricted and resembles, at least partially, the pattern observed in vivo. The development of this capacity to synthesize and store radiolabeled acetylcholine and catecholamines from the appropriate radioactive precursors coincides closely with the development of the activities of the synthetic enzymes choline acetyltransferase and dopamine beta-hydroxylase reported by others.
Single channel currents activated by gamma-aminobutyric acid, muscimol, and (-)-pentobarbital in cultured mouse spinal neurons
Authors: Jackson MB , Lecar H , Mathers DA , Barker JL
Published: March 2018

The patch electrode technique was used to record single channel current pulses in tissue-cultured mouse spinal cord neurons. In agreement with earlier noise studies, channels activated by gamma-aminobutyric acid (GABA), muscimol, and (-)-pentobarbital were found to have equal unit conductances. The kinetics of channel closing were studied by analyzing the distributions of open state lifetimes. Channels activated by (-)- pentobarbital and muscimol had longer mean open times than channels activated by GABA. As a result, the kinetics of (-)-pentobarbital- and muscimol-activated channels could be studied in greater detail. Most observed open state lifetime distributions were not exponential but contained an excess of short duration events. A sum of two exponential functions gave a much better fit than a single exponential function to most observed open state lifetime distributions. A critical comparison of noise analysis with single channel recording shows that the fast process responsible for the rapid closures would be very difficult to detect in a noise experiment. The channel noise is dominated by the slower process, and as a result, the relaxation time of the slower kinetic component derived from single channel studies is close to the mean open state lifetime derived from noise measurements. The observation of a faster process points toward either an additional population of channels or a scheme for the channel closing transition which is not a simple first order process.
Retinal projections in tyrosinase-negative albino cats
Authors: Creel D , Hendrickson AE , Leventhal AG
Published: March 2018

Retinal projections were examined in two tyrosinase-negative albino cats using autoradiographic techniques. Cats from this colony have pink eyes; their retinal pigment epithelium, ciliary body, and iris epithelium are completely devoid of melanin pigment. Test breeding for five generations indicates that these cats are true albinos (cc). The most striking feature of the albino cats' visual pathways was a reduction in ipsilateral input which was more severe than that reported for Siamese cats. The only evidence of ipsilateral input to the laminated dorsal lateral geniculate nucleus of the albino was a small lateral normal segment and a small projection to the lateral portion of lamina C1. Ipsilateral projections to the medial interlaminar nucleus, retinal recipient zone of the pulvinar complex, ventral lateral geniculate nucleus, superior colliculus, and pretectum also were reduced. Ipsilateral projections to the suprachiasmatic nucleus appeared to be normal in the albino cats studied. Our findings indicate that, compared to the normal cat as well as to the tyrosine-positive Siamese cat, the tyrosinase-negative albino has reduced ipsilateral retinal projections. The albino cat is a model system analogous to tyrosinase-negative albinism found in mammals.
Visualization at the mouse neuromuscular junction of a submembrane structure in common with Torpedo postsynaptic membranes
Authors: Sealock R
Published: March 2018

To test for submembrane structures common to postsynaptic membranes of muscle and electric tissue, mouse sternomastoid muscles were fixed in the presence of tannic acid under conditions such that a heavy layer of densely staining material was built up on plasma membranes. In this section electron micrographs of such muscles, the hydrophobic portion of the bilayer and some membrane-associated structures appear in strong negative contrast. At the endplate, acetylcholine receptor-rich membrane regions appear as a broad (14–15-nm) image approximately bisected by the thin (approximately 2-nm) image of the bilayer. The extracellular half of the image arises from the receptor protein. The cytoplasmic half contains a distinct bar of contrasted material which is precisely co-extensive with the receptor image Quantitative and direct visual comparisons show that the image is almost indistinguishable from that obtained by a similar method applied to isolated postsynaptic membranes from electric tissue of Torpedo californica (Sealock, R. (1982) J. Cell Biol. 92:514–522). In the latter case, the bar probably arises from the 43,000-dalton protein, a major peripheral membrane protein component of the isolated membranes. The results thus suggest that the 43,000-dalton protein in muscle and electric tissue forms similar structures and, conversely, that the structure in the isolated membranes retains the essential features of its in situ state.
Distribution of protein I and regulation of its state of phosphorylation in the rabbit superior cervical ganglion
Authors: Nestler EJ , Greengard P
Published: March 2018

The distribution and regulation of the state of phosphorylation of Protein I have been studied in the rabbit superior cervical sympathetic ganglion. The data indicate that the ganglion contains two pools of Protein I: a presynaptic pool that represents 60% of the total ganglion Protein I and a postsynaptic pool that represents 40% of the total ganglion Protein I. The state of phosphorylation of presynaptic Protein I, but not that of postsynaptic Protein I, is regulated by nerve impulse conduction, by dopamine, and by a high K+ concentration. Studies of the extracellular calcium requirements for Protein I phosphorylation, as well as peptide-mapping analyses of Protein I, suggest that the effects of nerve impulse conduction and of a high K+ concentration are mediated through the activation of calcium-dependent protein kinases and that the effect of dopamine is mediated through the activation of cyclic AMP-dependent protein kinase. The total amount of postsynaptic Protein I, but not that of presynaptic Protein I, is decreased by short periods of exposure to cycloheximide, a protein synthesis inhibitor. It is proposed that Protein I located in presynaptic nerve terminals plays a functional role in those terminals and that the Protein I located in cell bodies is newly synthesized and en route to nerve terminals.
Substance P: ionic basis for depolarizing responses of mouse spinal cord neurons in cell culture
Authors: Nowak LM , Macdonald RL
Published: March 2018

Intracellular recording methods were used to investigate the ionic basis for the postsynaptic actions of substance P (SP) on mouse spinal cord neurons grown in primary dissociated cell culture. SP and an analog, eledoisin-related peptide (ERP), were applied to single neurons by pressure ejection of peptide-containing solutions from blunt (2- to 10-micrometers) glass micropipettes. SP and ERP had similar excitatory actions, increasing spontaneous activity and depolarizing neurons by decreasing membrane conductance. Depolarizing responses were not inverted by intracellular injection of chloride ions, suggesting that SP responses did not result from decreased chloride conductance. SP and ERP responses were not abolished by extracellular tetraethylammonium ions (TEA+) but were reduced or eliminated by intracellular TEA+, suggesting that SP reduced a potassium conductance (gK). Finally, SP and ERP responses were larger when neurons were depolarized and smaller when the cells were hyperpolarized, and extrapolated reversal potentials for the peptide responses were 10 to 30 mV more negative than resting membrane potential. Thus, it was concluded that SP depolarized spinal cord neurons by decreasing a membrane potassium conductance. However, SP and ERP response polarity was not clearly reversed even at membrane potentials more negative than the expected potassium equilibrium potential. Moreover, extrapolated reversal potentials (RPeS) of SP responses varied linearly with the logarithm of extracellular potassium concentration ([K+]o) as predicted by the Nernst equation for potassium in [K+]o concentrations of 10, 15, 20, and 40 mM but were more depolarized than predicted for [K+]o concentrations of 1 and 5 mM. Since reduction of extracellular sodium by choline substitution did not alter the deviation from the Nernst equation for potassium, it was concluded that SP decreased a voltage- dependent potassium conductance which was absent at very negative potentials, present at resting membrane potential, and activated by membrane depolarization. Thus, SP decreases a conductance which appears similar to the muscarine-sensitive potassium conductance in sympathetic ganglion neurons (Brown, D. A., and P. R. Adams (1980) Nature 283: 673–676).
Neurite outgrowth and protein synthesis by PC12 cells as a function of substratum and nerve growth factor
Authors: Fujii DK , Massoglia SL , Savion N , Gospodarowicz D
Published: March 2018

Numerous studies have implied that enhanced cell-substratum adhesion plays a role in neurite outgrowth by neuronal cells. Using an extracellular matrix (ECM) produced by cultured corneal endothelial cells, we have investigated attachment and de novo neurite outgrowth by the pheochromocytoma cell line, PC12. PC12 cells were found to attach more rapidly and efficiently to the ECM than to plastic or collagen- coated surfaces. An extensive but temporary (5- to 10-day) neurite outgrowth occurred in the absence of nerve growth factor (NGF) when cells were on the ECM. However, long term neurite survival and further elongation required NGF. Our findings are consistent with the hypothesis that protein(s) in the ECM has an important role in neurite outgrowth. Thus, NGF may not so much initiate neurite outgrowth as it stabilizes neurites. ECM and NGF also were found to modulate cellular protein synthesis by PC12 cells. On ECM, a decreased synthesis of many high molecular weight proteins (Mr greater than 85,000) was observed in comparison to that of cells on collagen-coated dishes. The presence of neurites (in the presence of absence of NGF) as associated with the induction of the synthesis of a cellular protein (Mr = 55,000 to 56,000 and pI of 5.6). NGF was found to increase markedly the synthesis of two secreted proteins, while it drastically reduced the synthesis of all others regardless of the substratum upon which the cells were maintained.
Spatial summation and conduction latency classification of cells of the lateral geniculate nucleus of macaques
Authors: Marrocco RT , McClurkin JW , Young RA
Published: March 2018

Cells in the lateral geniculate nucleus (LGN) of the macaque monkey were investigated with microelectrodes in an attempt to develop an overall classification scheme. We classified cells in the parvocellular (P) and magnocellular (M) layers according to (non)linearity of spatial summation, shock latency, and chromatic organization of center and surround. We also measured the spatial and temporal tuning to counterphasing and drifting sine wave gratings and tested for periphery effects. Our results showed that no strict laminar segregation existed for any cell property studied. Our results can be summarized as follows: 1. Most P layer cells showed a linear summation (98%) and color-opponent responses (80%), while other cells showed a nonlinear summation (Y-cells, 2%) and broad band responses (28%). In contrast, 37% of the M layer cells were linear summators and the remainder were nonlinear. Therefore, there are overlapping distributions of X- and Y- cells in P and M layers but not a strict segregation. 2. P layer cells had longer shock latencies than M layer cells. X-cells conducted more slowly (2.4 +/- 0.7 msec) than Y-cells (1.6 +/- 0.8 msec), but there were overlapping distributions. Latency shortened gradually, rather than abruptly, with increasing depth. 3. The first harmonic of X- and Y- cell responses was maximally sensitive to spatial frequencies of about 2 cycles/deg. Each type of cell modulated about a mean rate to a drifting grating, although Y-cells had higher distortion than X-cells. Response amplitudes to drifting gratings were higher for MX- and MY- than for PX-cells. No DC elevation to high spatial frequencies was seen. Spatial bandwidths averaged 2 to 5 octaves. X-cells were maximally tuned to temporal frequencies around 11 Hz, and Y-cells were tuned to about 19 Hz;. temporal bandwidths for both averaged 2.8 octaves. 4. Periphery effects were detected in 4% of the X-cells and 25% of the Y-cells. 5. These data indicate that gradual changes occur between dorsal and ventral layers: summation changes from linear to nonlinear; conduction latencies shorten; peak temporal tuning increases; response amplitudes increase; the periphery effect becomes more prevalent. Spatial tuning does not change. No strict laminar segregation or specificity exists for any of the properties that we studied.
Progestin receptor levels in rat hypothalamic and limbic nuclei
Authors: Parsons B , Rainbow TC , MacLusky NJ , McEwen BS
Published: March 2018

We have utilized a method to minimize cytosol progestin receptor loss during freezing in order to localize the quantify estrogen-inducible progestin receptors in individual nuclei of the female rat brain. Ovariectomized females received estradiol benzoate (20 micrograms for 3 days) or vehicle prior to sacrifice. All animals were perfused with cold distilled H2O containing the cryoprotective compound, dimethyl sulfoxide (DMSO; 10% (v/v)). Thirty-one nuclei or brain regions were removed from frozen sections (300 micrometers) according to the method of Palkovits (Palkovits, M. (1973) Brain Res. 59: 449–450) and were assayed in vitro using a synthetic radioligand, [3H]R5020. In ovariectomized animals perfused with DMSO, a basal level (1 to 8 fmol/mg of protein) of progestin receptors was observed in a variety of preoptic, hypothalamic, and limbic structures. Moreover, estrogen treatment induced high levels (24 to 49 fmol/mg of protein) of progestin receptors in regions of the preoptic area of hypothalamus which contain high levels of estrogen receptors. These regions included the medial, periventricular, and superchiasmatic nuclei of the preoptic area, the periventricular anterior hypothalamus, the ventromedial nucleus, and the arcuate-median eminence. Moderate levels (2 to 8 fmol/mg of protein) of progestin receptors were induced by estrogen in other hypothalamic and limbic structures, including the anterior and lateral hypothalamus, the bed nucleus of the stria terminalis, the cingulate cortex, the medial amygdaloid nucleus, and the CA subfield of the hippocampus. By contrast, some areas, such as the caudate-putamen and the supraoptic nucleus, were devoid of both estrogen-inducible and uninduced progestin receptors. These results support the hypothesis that progesterone action in the central nervous system is mediated by cytosol receptors in discrete brain regions and provide the first quantitative map of progestin binding in a vertebrate brain.
Electrophysiology of isolated hippocampal pyramidal dendrites
Authors: Benardo LS , Masukawa LM , Prince DA
Published: March 2018

In order to study the electrical properties of dendritic membranes independent of the effects of somatic potentials, intracellular recordings in guinea pig hippocampal slices were obtained from the dendrites of CA1 pyramidal neurons (HPCs) which had been isolated from their somata by cuts made through the proximal stratum radiatum. Spikes and subthreshold membrane responses to intracellular current pulses were compared in intact and isolated dendrites and in the residual portions of neurons whose apical dendrites had been severed (“isolated somata”). Isolated dendrites generated both fast, QX-314-sensitive, sodium-mediated spikes, and slow higher threshold spikes which were QX- 314-resistant and presumably mediated by Ca2+. Depolarization of “isolated somata” ordinarily evoked only fast (Na+) spikes, but presumed Ca2+ spikes could be elicited after exposure to QX-314 (a local anesthetic). Anomalous inward rectification was depressed by QX- 314 in somata but not in dendrites, suggesting that the ionic basis for subthreshold as well as regenerative conductances was different at different sites on the neuron. The dendritic membrane in CA2 HPCs thus generates both Na+- and CA2+-mediated spike potentials and a subthreshold response which probably is mediated primarily by CA2+. Attempts to describe the integrative functions of these neurons must take into account the variety of conductances which are activated nonuniformly in somata and dendrites by changes in membrane potential.
Formation of postsynaptic specializations induced by latex beads in cultured muscle cells
Authors: Peng HB , Cheng PC
Published: March 2018

An early event in the formation of neuromuscular junctions is an accumulation of acetylcholine receptors (AChRs) in the postsynaptic membrane. This concentration of receptors is essential for the establishment of the synaptic transmission. Previously, we showed that latex beads coated with positively charged polypeptide molecules can induce the formation of AChR clusters at the bead-muscle contacts (Peng, H. B., P.-C. Cheng, and P. W. Luther (1981) Nature 292: 831–834). In this study, we demonstrate that, in addition to AChR clusters, a set of postsynaptic specializations normally present at the adult neuromuscular junction also develops at the bead-muscle contacts in the absence of nerve. These specializations include the membrane-associated cytoplasmic density and its associated thin (6- to 8-nm) filaments, clusters of intramembranous particles, membrane invaginations, the basal lamina, and 60- to 100-nm coated vesicles. Our results indicate that the mechanism for the formation of the postsynaptic specializations is preprogramed in the muscle and can be triggered by an interaction with polypeptide-coated latex beads. Thus, this bead- muscle system can be used as a simple model for understanding the postsynaptic development during synaptogenesis.
Intracellular identification of central noradrenergic and serotonergic neurons by a new double labeling procedure
Authors: Aghajanian GK , Vandermaelen CP
Published: March 2018

The purpose of this study was to ascertain the identity of presumed noradrenergic or serotonergic neurons recorded by single cell techniques in the mammalian brain. A double labeling method was developed in which intracellular injections of a red fluorescing dye (ethidium bromide) could be co-localized with the formaldehyde-induced green fluorescence of norepinephrine or yellow fluorescence of serotonin. By this method, neurons of the rat locus coeruleus that display a characteristic activation-inhibition response to noxious stimuli were confirmed to be noradrenergic; the slow, rhythmically firing neurons of the dorsal raphe nucleus were confirmed to be serotonergic.
Trans-synaptic modulation via muscarinic receptors of serotonin- containing small intensely fluorescent cells of superior cervical ganglion
Authors: Hadjiconstantinou M , Potter PE , Neff NH
Published: March 2018

Verhofstad et al. (Verhofstad, A. A. J., H. W. M. Steinbusch, B. Penke J. Varga, and H. W. J. Joosten (1981) Brain Res. 212: 39–49) have reported that serotonin of the rat superior cervical ganglion is contained in a distinct and separate population of small intensely fluorescent (SIF) cells. We provide evidence that the serotonin- containing SIF cells are modulated, in part, by preganglionic cholinergic neurons. For example, administration of the muscarinic agonists carbachol or oxotremorine increases the content of serotonin, and the increase induced by oxotremorine is blocked by atropine. Treatment with atropine alone or decentralization of the ganglion lowers the content of serotonin. From the rate of accumulation of serotonin in the ganglion after the administration of the monoamine oxidase inhibitor pargyline, it appears that the rate of formation of serotonin is increased after oxotremorine treatment. Reserpine, p- chlorophenylalanine, or fluoxetine treatment reduces the content of serotonin in the ganglion, suggesting that the SIF cell system has properties similar to those of serotonergic neurons of the brain. We postulate that the serotonin-containing SIF cells of the rat superior cervical ganglion participate in local circuit modulation of ganglionic transmission by receiving preganglionic information via muscarinic receptors.
Monoclonal antibodies to choline acetyltransferase: production, specificity, and immunohistochemistry
Authors: Levey AI , Armstrong DM , Atweh SF , Terry RD , Wainer BH
Published: March 2018

Immunohistochemical localization of choline acetyltransferase (ChAT) in cholinergic neurons has been difficult to achieve because of problems encountered in producing specific antisera. Here we describe the production and characterization of several distinct monoclonal antibodies to ChAT. Each of the monoclonal antibodies exhibits one of three general patterns of cross-species reactions; one pattern shows reactivity limited mainly to bovine ChAT, a second pattern shows reactivity only to ChAT from higher mammals including humans, and the third pattern shows reactivity to ChAT from all mammals tested. The antibodies bound specifically to two closely related bovine proteins of 68,000 and 70,000 daltons using the Western blotting technique. One of the antibodies was used to localize immunohistochemically known cholinergic structures in the rat brain, including motor neurons, basal forebrain neurons, and neostriatal neurons.
Pathfinding by neuronal growth cones in grasshopper embryos. I. Divergent choices made by the growth cones of sibling neurons
Authors: Raper JA , Bastiani M , Goodman CS
Published: March 2018

We are interested in how the growth cones of identified neurons navigate in the central nervous system of the grasshopper embryo. The behavior of identified growth cones was observed as a function of developmental time by (i) periodically removing embryos from synchronized clutches of eggs and (ii) filling identified neurons in the central nervous system with the fluorescent dye Lucifer Yellow. We examined the first six progeny of the identified neuronal precursor cell, neuroblast 7–4: the Q1, Q2, G, C, Q5, and Q6 neurons. Their growth cones extend sequentially upon the same route across the midline of the posterior commissure of the developing ganglionic neuropil. However, after reaching the contralateral side of the neuropil, their growth cones diverge from each other at reproducible, cell-specific choice points. By focusing our attention on one such choice point, at which the growth cone of the G neuron turns anteriorly while the growth cone of its sibling C neuron turns posteriorly, we hope to elucidate the environmental cues which guide individual growth cones.
Transneuronal regulation of tyrosine hydroxylase expression in olfactory bulb of mouse and rat
Authors: Baker H , Kawano T , Margolis FL , Joh TH
Published: March 2018

Peripheral afferent input regulates the expression of dopaminergic properties in a population of local circuit intrinsic neurons of the rodent olfactory bulb. Lesions of the olfactory receptor neurons produced in the mouse by intranasal irrigation with either ZnSO4 or Triton X-100 and in the rat by surgical deafferentation or axotomy are associated with a decrease in the levels of dopamine (DA), the DA metabolite 3,4-dihydroxyphenylacetic acid (DOPAC), the activity of the enzyme tyrosine hydroxylase (TH), bulb weight and an increase in norepinephrine (NE) levels in the olfactory bulb. The anatomical correlates of the biochemical sequelae of deafferentation of olfactory bulb DA neurons were studied using immunohistochemical techniques to localize TH. Within 3 to 4 weeks all lesions produced a dramatic and uniform reduction in TH staining of the juxtaglomerular DA neurons and their processes which was paralleled by a reduction in DA and DOPAC levels and bulb weight. Seven weeks following reversible chemical lesions produced by Triton X-100, DA and DOPAC levels and tissue weight as well as TH staining in the juxtaglomerular neurons returned to control levels. These observations suggested that DA neurons remained present even when not demonstrable with TH antibodies. Additional evidence for the continued presence of the DA neurons was the ability of the olfactory bulbs from both lesioned mouse and rat to synthesize DA from exogenously administered L-3,4-dihydroxyphenylalanine (l-DOPA). These data suggested that the decrease in DA levels and TH staining in the olfactory bulb following lesions of the olfactory receptor neurons were produced by transneuronal mechanisms since there was no direct injury of the bulb. Furthermore, the demonstration that following reinnervation, catecholamine synthetic capacity is restored suggests that the juxtaglomerular dopamine neurons remain in the bulb and that afferent receptor input is required for expression of TH enzyme.
Control of intercellular communication by voltage dependence of gap junctional conductance
Authors: Harris AL , Spray DC , Bennett MV
Published: March 2018

The junctional conductance between coupled amphibian blastomeres exhibits a high degree of voltage dependence, as previously described in voltage clamp studies (Spray, D.C., A.L. Harris, and M.V.L. Bennett (1981) J. Gen. Physiol. 77: 77–95; Harris, A.L., D.C. Spray, and M.V.L. Bennett (1981) J. Gen. Physiol. 77: 95–117). The present study examines the properties which this voltage dependence confers on electrotonic coupling between cells. The effects of applied pulses and ramps of current are studied experimentally and are modeled by calculation. During sufficiently large current pulses applied to one cell of a pair, the cells uncouple and then recouple after termination of the pulses. Ramps of current applied to one of the cells can give voltage-current (V-I) relations with a region of hysteresis within which the cells are stably coupled or stably uncoupled depending on previous history. Intrinsically generated currents are able to cause bistability of coupling in the absence of externally applied current. Calculations from the parameters of junctional conductance defined under voltage clamp fully account for these findings and illustrate how junctional and nonjunctional conductances affect the V-I relations in the region of bistability. Recordings from several cells within a small group show that boundaries of intercellular communication can be altered by applied current, a finding that also can be accounted for by voltage dependence of junctional conductance. The “Appendix” examines quantitatively the criteria required for bistability of coupling and the relevance of bistability for intercellular signaling. The plasticity of coupling which the voltage dependence of junctional conductance confers on cells offers an intriguing mechanism by which patterns of intercellular communication could be determined and changed in developing tissues.
Metabolism of acetylcholine receptors on embryonic amphibian muscle
Authors: Brehm P , Yeh E , Patrick J , Kidokoro Y
Published: March 2018

The functional properties of acetylcholine receptors are altered during the development of Xenopus myotomal muscle. A similar change also occurs in dissociated cell cultures of embryonic myotomal muscle. Metabolism of acetylcholine receptors was examined in order to ascertain whether the turnover rate changed at a time when the functional changes were detected. Incorporation of newly synthesized receptors occurred at a constant rate over a 24-hr period. The degradation of receptors followed first order kinetics with a half-life of 52 hr. Incorporation and degradation occurred at approximately equal rates such that 1 to 1.5% of the total receptor number was replaced every hour at equilibrium. Addition of neural tube cells to muscle cell culture, which altered the functional properties of channels, had no effect on either the incorporation or the degradation rate. We conclude that the changes in the functional properties of receptors occur independent of changes in the turnover rate of receptors.
alpha-Bungarotoxin labeling and acetylcholinesterase localization at the Mauthner fiber giant synapse in the hatchetfish
Authors: Day JW , Hall DH , Hall LM , Bennett MV
Published: March 2018

Autoradiographic and histochemical techniques have been used to characterize further the pharmacology of transmission at the Mauthner fiber giant synapse of the South American hatchetfish. [125I]alpha- Bungarotoxin was applied to hatchetfish medullae and a standard autoradiographic procedure was carried out on 3- to 4-microns sections of glutaraldehyde-fixed tissue. All Mauthner fiber giant synapses, as identified by light microscopic criteria, had closely associated silver grains. Labeling was blocked by d-tubocurarine. Glutaraldehyde-fixed slices of hatchetfish medulla were stained histochemically for acetylcholinesterase; all giant synapses that could be identified in the light microscope showed heavy deposits of reaction product. Staining was blocked by diisopropyl-fluorophosphate, which inhibits both pseudocholinesterase and acetylcholinesterase, but was not blocked by tetraisopropylpyrophosphoramide, a specific pseudocholinesterase inhibitor. This evidence strongly supports the suggestion that the Mauthner fiber giant synapse is nicotinic cholinergic.
Effects of spinal tractotomy on spatial sequence recognition in macaques
Authors: Vierck CJ , Cohen RH , Cooper BY
Published: March 2018

Five Macaca speciosa monkeys were trained to discriminate between the order of two tactile stimulus presentations (proximal-distal vs. distal- proximal) to the lateral calf. Psychophysical thresholds for tactile sequence recognition were obtained by varying the distance between the spots stimulated. Stabilized preoperative thresholds for stimulation on the lateral calf averaged 10.4 mm, which is comparable to point localization thresholds for sequential contact of the proximal limbs of humans. Complete interruption of the ipsilateral fasciculus gracilis produced elevations of thresholds within the first 3 postoperative weeks to an average of 3.6 times the control values. With training over 6 postoperative months, normal discrimination recovered. In contrast to the effects of dorsal column (DC) lesions, interruption of the lateral and ventral columns ipsilaterally or contralaterally produced no deficit or a mild and transient deficit. Monkeys with isolation of either the ipsilateral dorsolateral column (ILC) or the contralateral ventral quadrant (CVQ) exhibited substantial and enduring deficits. Thresholds were elevated an average of 5 times the control values, and recovery was not observed or was incomplete after months of training. In contrast, isolation of the ipsilateral dorsal column produced little or no threshold elevation. The results of isolating lesions complement the series of primary lesions to each cord sector, showing that spatiotactile sequence recognition for the hindlimb is served more critically by the dorsal columns than by the lateral columns.