bluuebunny/crossref_metadata_2025 huggingface dataset:
Medial preoptic sexual dimorphisms in the guinea pig. I. An investigation of their hormonal dependence
Authors: Byne W , Bleier R
Published: March 2018

The guinea pig exhibits sexually dimorphic patterns of cell density and distribution throughout the medial preoptic area, a region that has been shown to be involved in the regulation of sexually differentiated behavioral and endocrinological reproductive functions (Bleier et al., 1982). The most prominent sex differences involve 2 components of the medial preoptic nucleus (MPN), an anteriorly placed compact subnucleus (MPNa) that is twice as large in females as in males, and a centrally placed compact subnucleus that occupies an approximately 10-fold greater volume in males than in females and corresponds to the sexually dimorphic nucleus described in the rat by Gorski et al. (1978). In the present study the sex differences in both of these cell groups were shown to be unaffected by neonatal gonadectomy and postnatal hormonal manipulations. In contrast, MPNa volume was significantly decreased and MPNc volume significantly increased in genotypic females exposed to testosterone propionate (TP) on gestational days 28-37 or 28-65 but not 38-65. All 3 prenatal TP treatments administered to females significantly increased mounting frequencies and suppressed lordosis, ovulation, and the positive-feedback effects of estrogen and progesterone on luteinizing hormone release. Thus, the volumetric sex differences in MPNa and MPNc alone do not seem to be sufficient to account for the sex differences in the functions of the medial preoptic region. It is, therefore, suggested that androgens continue to exert organizational influences upon the developing brain after cytoarchitectonic patterns have been determined.
Molecular cloning of a 2',3'-cyclic nucleotide 3'-phosphodiesterase: mRNAs with different 5' ends encode the same set of proteins in nervous and lymphoid tissues
Authors: Bernier L , Alvarez F , Norgard EM , Raible DW , Mentaberry A , Schembri JG , Sabatini DD , Colman DR
Published: March 2018

Antibodies raised to a mixture of the 46 and 48 kDa rat CNS 2′,3′- cyclic nucleotide 3-phosphodiesterases (CNPs) recognized apparently identical proteins in peripheral nervous system (PNS), thymus, and circulating blood lymphocytes. These antibodies were used to identify, in a rat brain phage lambda gt11 expression library, cDNA clones encoding beta-galactosidase-CNP fusion proteins, some of which showed CNP activity. In RNA blots, the subcloned CNP cDNA inserts hybridized to mRNAs of approximately 2400 and approximately 2800 nucleotides (nts), and to a approximately 2500 nt mRNA from thymus. Several nonexpressing CNP cDNAs were identified by plaque hybridization, and the mRNA transcribed in vitro from one of these cDNAs (pCNP7) encoded a complete 46 kDa CNP polypeptide. Examination of the deduced amino acid sequence revealed an apparent homology to cAMP binding sites in several other proteins. A 373 bp segment from the 5′ end of this pCNP7 hybridized only to the 2800 nt nervous system mRNAs, thus revealing that not all CNP mRNAs share the same 5′-ends. Genomic DNA blots probed with CNP cDNAs suggest that there is a single gene which can be alternatively spliced to produce the various mRNA transcripts in the nervous and lymphoid tissues.
Modulation of activity of the striatal dopaminergic system during the hibernation cycle
Authors: Kilduff TS , Bowersox SS , Faull KF , Zeller-DeAmicis L , Radeke CM , Ciaranello RD , Heller HC , Barchas JD , Dement WC
Published: March 2018

To evaluate how the activity of a well-established neurotransmitter pathway is modulated by a behavioral state, 3H-spiperone binding sites and dopamine (DA) and DA metabolite concentrations were measured in the striata of ground squirrels in 5 phases of the hibernation cycle. Whereas levels of striatal DA and its deaminated metabolite DOPAC did not change significantly, the concentrations of the O-methylated- deaminated metabolite, homovanillic acid (HVA), decreased in all phases of hibernation relative to euthermia. Striatal 3H-spiperone binding sites declined across the hibernation cycle in parallel with the reduction of HVA concentration; receptor binding affinity was unchanged by arousal state. In conjunction with previously reported findings, these results are consistent with the hypothesis that hibernation is associated with a down-regulation of the postsynaptic D2 receptors secondary to increased extracellular DA concentration and reduced DA degradation.
Release of cortical catecholamines by visual stimulation requires activity in thalamocortical afferents of monkey and cat
Authors: Marrocco RT , Lane RF , McClurkin JW , Blaha CD , Alkire MF
Published: March 2018

Catecholamine (CA) release was measured in vivo in the monkey and cat visual cortices electrochemically. Stereate-modified, graphite-paste electrodes were used to monitor changes in norepinephrine and dopamine release. Micromolar changes in CA concentration were obtained by stimulation of the eye with nonspecific (strobe) or specific (oriented bars, radial gratings) stimuli. CA release depended on which eye was illuminated. Electrodes passed tangentially through the striate area recorded release following visual stimulation of one eye or the other in succession, and the shift in eye dominance occurred at about 500 microns intervals. The magnitude of CA release was highly correlated with the ocular dominance of neuronal activity measured with tungsten microelectrodes. Light-stimulated release was not recorded in monkey area V2, V4, or somatosensory area 1, but was recorded in cat V2, suggesting that the presence of LGN afferents is associated with CA release. Results are discussed in terms of the role of geniculate activity and the specific role of CAs in cortical information processing.
Glutamic acid decarboxylase cDNA: nucleotide sequence encoding an enzymatically active fusion protein
Authors: Kobayashi Y , Kaufman DL , Tobin AJ
Published: March 2018

Glutamic acid decarboxylase (GAD;E.C. 4.1.1.15) catalyzes the production of GABA, the major inhibitory neurotransmitter in the mammalian brain. We recently isolated a lambda gt-11 recombinant, lambda-GAD, that contains the cDNA for GAD from feline brain (Kaufman et al., 1986). Interestingly, the beta-galactosidase-GAD fusion protein encoded by lambda GAD is enzymatically active, catalyzing the conversion of glutamate to CO2 and GABA. Here we report the nucleotide sequence of feline GAD cDNA. It consists of 2265 bases, with a continuous open reading frame of 625 codons. The derived sequence contains the sequence Asn-Pro-His-Lys, which is identical to sequence at the pyridoxal phosphate-binding site of porcine DOPA decarboxylase (Bossa et al., 1977). The first ATG sequence in the open reading frame begins at nucleotide residue 118. The 585 codons 3′ to this putative initiation site predict an amino acid composition, N-terminal residue, and molecular size consistent with published characterizations of GAD.
Identification and purification of two precursors of the insect neuropeptide adipokinetic hormone
Authors: Hekimi S , O'Shea M
Published: March 2018

Our objective was to establish a system for the investigation of neuropeptide biosynthesis in an insect. To achieve this we developed an in vitro organ culture system for the corpora cardiaca (CC) of the locust (Schistocerca gregaria). The CC are the neurosecretory structures containing the adipokinetic hormones AKH I and AKH II. Tritiated amino acids were added to the glands, and we studied the development of label in newly made proteins using size-exclusion and reverse-phase liquid chromatography. We performed pulse-chase and pulse- translation block experiments using 3H-tryptophan as tracer. We also raised an antiserum to a synthetic AKH analog and used this in combination with liquid chromatography to identify 2 precursor polypeptides, P1 and P2. Size-exclusion chromatography indicated the presence of a major component of the CC of about 8 kDa that incorporates 3H-tryptophan before AKH I and II. Moreover, in both pulse- chase and pulse-translation block experiments we showed that label is transferred from this 8 kDa component into AKH I. We call this component proAKH; it is recognized by anti-AKH serum in a radioimmunoassay (RIA). Further fractionation of tritium-labeled proAKH by reverse-phase chromatography yielded 2 polypeptides, P1 and P2. Both are AKH-immunoreactive and contain 3H-tryptophan after in vitro pulse labeling, and both are proposed precursors of AKH I. The in vitro system we have developed may be a model system for the study of the processes of neuropeptide biosynthesis and its regulation in an intact neurosecretory tissue of an insect.
Octopamine- and cyclic AMP-stimulated phosphorylation of a protein in Limulus ventral and lateral eyes
Authors: Edwards SC , Battelle BA
Published: March 2018

The biogenic amine octopamine (OCT) fulfills most of the criteria as a neurotransmitter of efferent fibers that project to lateral and ventral eyes of the horseshoe crab, Limulus polyphemus. OCT is synthesized by and released from the efferent fibers, and OCT mimics many of the effects of endogenous efferent activity. OCT stimulates an increase in intracellular adenosine 3′,5′-monophosphate (cAMP) in both ventral and lateral eyes, and many of the physiological effects of OCT in these eyes appear to be mediated via cAMP-dependent mechanisms. Here we show that OCT, acting apparently through an OCT-specific receptor, stimulates the increased phosphorylation of a protein with an apparent molecular weight of 122 kDa in both ventral and lateral eyes. This protein is also phosphorylated in response to 8-bromo cAMP and forskolin, suggesting that its phosphorylation involves activation of a cAMP-dependent protein kinase. We present evidence that the 122 kDa protein may be widely distributed in the Limulus visual system but that its phosphorylation in intact tissue in response to OCT, or agents acting through cAMP, may be restricted to portions containing photoreceptor cell bodies. The 122 kDa protein is quantitatively a major cellular protein in the photoreceptor cell body enriched portions of the ventral eye, its isoelectric point is between pH 6.2 and 6.4, and it is associated with both cell membranes and the cytoplasm. The function of this protein is not yet known. It may be important in mediating one or more of the effects of octopamine on Limulus vision.
Induction of a neural phenotype in a serotonergic endocrine cell derived from the neural crest
Authors: Barasch JM , Mackey H , Tamir H , Nunez EA , Gershon MD
Published: March 2018

The thyroid parafollicular cell is an endocrine cell derived from the neural crest that stores 5-hydroxytryptamine (5-HT). In common with serotonergic neurons, but in contrast to 5-HT-storing cells that are not neurectodermal derivatives, parafollicular cells also contain a specific 5-HT binding protein. Despite this similarity to serotonergic neurons, parafollicular cells in situ were found to express an endocrine phenotype with few neural characteristics. Thus, the cells costore 5-HT with calcitonin, not calcitonin gene-related peptide (CGRP), which is the product of the calcitonin gene expressed in neurons, and they do not contain neurofilaments. The ability of adult parafollicular cells to respond to microenvironmental perturbations by expressing neuronal characteristics was examined. Sheep thyroid glands were dissociated, and parafollicular cells were purified by affinity chromatography. The purified parafollicular cells were grown in culture on a variety of substrates in the presence or absence of the beta subunit of nerve growth factor (beta-NGF). Parafollicular cells survived in culture for at least a week but retained a roughly spherical shape. Nevertheless, a subset of the cultured parafollicular cells began to display CGRP immunoreactivity. The addition of beta-NGF to the cultured parafollicular cells induced a number of them to extend neurites and increased the proportion of cells in which CGRP immunoreactivity could be found. Neurite-bearing parafollicular cells appeared not to survive for more than 2 d. While their survival was not enhanced when they were grown on collagen, polylysine, laminin, or reconstituted basal lamina, parafollicular cells that had extended neurites in response to beta-NGF survived for at least a week when cocultured with an explant of aneuronal chick hindgut. The effect of the gut was local and only those neurite-bearing parafollicular cells that were growing in direct contact with the explant survived. The thyroid parafollicular cell therefore resembles another crest-derived endocrine cell, the adrenal chromaffin cell, in being able to manifest neural properties in culture. For the parafollicular cell these neural properties include the processing of RNA encoded by the calcitonin gene to express CGRP and neurite outgrowth in response to beta-NGF.
Suppression of oscillatory activity in crustacean pyloric neurons: implication of GABAergic inputs
Authors: Cazalets JR , Cournil I , Geffard M , Moulins M
Published: March 2018

Generation of rhythmic pyloric motor output in the crustacean stomatogastric ganglion results from synaptic connections and cellular properties of a 14-cell network of pyloric neurons. These cellular properties are under the influences of modulatory inputs, which act, for the most part, in an activating mode, i.e., they enhance the bursting properties of the pyloric neurons and/or their ability to express their regenerative properties. Here we attempt to demonstrate that the pyloric motor output is also under the control of suppressive afferent inputs that are able to stop the pyloric rhythm in a long- lasting manner. Immunohistochemistry, using GABA antibodies, indicates that GABAergic-like fibers are present in both the stomatogastric ganglion and its afferent nerve. Bath-applied GABA suppresses spontaneous pyloric rhythmic activity. This is due to an inability of the pyloric pacemakers to express their bursting properties. The suppressive effect of GABA is blocked by picrotoxin and mimicked by muscimol. Isolating the pyloric neurons from all descending spiking influences with tetrodotoxin demonstrates that exogenously applied GABA acts directly on the pyloric neurons. To confirm the existence of a physiological suppressive system for the pyloric motor pattern, we show that the stimulation of an afferent nerve, known to contain GABA-like fibers, also causes the cessation of rhythmic activity and the inability of the pyloric neurons to express their bursting properties.
Excitatory amino acid-induced release of 3H-GABA from cultured mouse cerebral cortex interneurons
Authors: Drejer J , Honore T , Schousboe A
Published: March 2018

A newly developed continuous superfusion model was used for studies of 3H-GABA release from cultured mouse cerebral cortex neurons. It was found that a series of excitatory amino acids (EAAs) representing all receptor subtypes evoked Ca2+- dependent release of 3H-GABA from the neurons. Quisqualate was the most potent agonist tested, with an EC50 value of 75 nM. L-Glutamate, N-methyl-D-aspartate (NMDA), and kainate showed EC50 values of 12, 16 and 29 microM, respectively. The EAA- evoked 3H-GABA release could be blocked by a series of EAA antagonists. The highly selective NMDA antagonist D-2-amino-5-phosphonovaleric acid (D-APV) was found to block NMDA responses, whereas the nonselective antagonists cis-2,3-piperidine dicarboxylic acid (PDA) and gamma-D- glutamyl-aminomethyl sulphonic acid (GAMS) blocked responses to all agonists. NMDA responses were found to be sensitive to Mg+ blockade. EAA- as well as potassium-induced 3H-GABA release from the neurons could be detected as early as day 5 in culture. However, during the culture period up to 12 d, the responses to K+, quisqualate, and NMDA were increased. The ontogenetic development of binding sites for quisqualate, kainate, and NMDA in mouse cortex was studied using the radioligands 3H-alpha-amino-3-hydroxy-5-methyl-4-isoxasole propionate (3H-AMPA), 3H-kainate, and 3H-L-glutamate, respectively. The development of binding sites for the different EAA-receptor subtypes showed a good correlation with the development of neuronal 3H-GABA release evoked by the excitatory amino acids.(ABSTRACT TRUNCATED AT 250 WORDS)
Effects of an anxiogenic benzodiazepine receptor ligand on motor activity and dopamine release in nucleus accumbens and striatum in the rat
Authors: Brose N , O'Neill RD , Boutelle MG , Anderson SM , Fillenz M
Published: March 2018

The effects of the anxiogenic beta-carboline FG 7142 (N-methyl-beta- carboline-3-carboxylate) on motor activity and dopamine release in nucleus accumbens and striatum were measured in the rat. Changes in extracellular homovanillic acid (HVA) concentration, monitored by computer-controlled linear sweep voltammetry with carbon-paste electrodes, were used as an index of changes in dopamine release. An intraperitoneal injection of FG 7142 was followed by an inhibition of the nocturnal rise in motor activity and in dopamine release in nucleus accumbens, but not striatum. Two days after the drug injection, dopamine release in nucleus accumbens returned to control level and then increased on days 3-6 after the injection; there was no delayed change in motor activity or in striatal dopamine release. In parallel experiments using ex vivo changes in the ratio of 3,4- dihydroxyphenylacetic acid (DOPAC) to dopamine as an index of changes in dopamine turnover, a similar early depression and delayed increase of dopamine turnover in nucleus accumbens, with no change in striatum, was found after an intraperitoneal injection of FG 7142. Regression analysis of motor activity versus dopamine release showed a decrease in correlation between these 2 parameters for nucleus accumbens but not striatum after FG 7142 injection. These results suggest that the inverse benzodiazepine receptor agonist FG 7142 has a biphasic effect on dopamine release from mesolimbic neurons and support the hypothesis that dopamine release in nucleus accumbens and striatum has a modulatory effect on the control of motor activity but does not play a determining role in the regulation of movement.
Neuronal colocalization of peptides, catecholamines, and catecholamine- synthesizing enzymes in guinea pig paracervical ganglia
Authors: Morris JL , Gibbins IL
Published: March 2018

The patterns of colocalization of neuropeptides, catecholamines, and catecholamine-synthesizing enzymes were examined in principal neurons and nerve terminals in guinea pig paracervical ganglia using a double- labeling immunohistochemical procedure. A small proportion of nerve cell bodies (less than 10%) had the characteristics of catecholamine- synthesizing neurons and presumably were noradrenergic. Another 50% of the nerve cell bodies contained immunoreactivity (IR) to dopamine-beta- hydroxylase (DBH), but did not have any other characteristics of noradrenergic neurons; they did not contain detectable catecholamines, or IR to dopa decarboxylase (DDC) or tyrosine (TH) hydroxylase, nor did they take up exogenous catecholamines. Half of the catecholamine neurons had neuropeptide Y (NPY)-IR, and a small number (0.5% total neurons) had somatostatin (Som)-IR. Most of the non-noradrenergic neurons with DBH-IR (40–50% total neurons) contained IR for dynorphin (Dyn), NPY, and vasoactive intestinal peptide (VIP), and about half of them (20–25% total) also contained Som-IR. Ten to twenty percent of neurons contained IR to Som, but not to any other antigen examined here. Nerve terminals with substance P (SP)-IR or enkephalin (Enk)-IR were prominent in all ganglia. SP-IR fibers formed dense baskets only around those neurons with DBH/Dyn/NPY/VIP (+/- Som)-IR, while fibers with very bright Enk-IR were associated selectively with those neurons with Som-IR alone. In addition, most TH-IR nerve cell bodies were surrounded by NPY-IR varicose nerve fibers. In conclusion, this analysis of combinations of peptides and enzymes contained in principal neurons of the paracervical ganglia allows us to identify as many as 11 different neuron populations. The functional significance of the presence of the same neuropeptide (e.g., NPY) in different neuron populations is as yet unknown. Some of these classes of neurons are associated specifically with immunohistochemically distinct types of presynaptic nerve fibers, which suggests that different immunohistochemically defined classes of neurons represent different functional pathways.
Expression of two different tachykinin receptors in Xenopus oocytes by exogenous mRNAs
Authors: Harada Y , Takahashi T , Kuno M , Nakayama K , Masu Y , Nakanishi S
Published: March 2018

Three tachykinins, substance P (SP), neurokinin A (NKA), and neurokinin B (NKB), have been isolated from the mammalian nervous system. In accordance with the presence of multiple tachykinins, the existence of multiple tachykinin receptors has been suggested. These receptors differ in binding specificities for different tachykinins. However, it is not known whether these receptors are indeed made of different molecules or whether the same receptor molecule undergoes posttranslational modification at different destination tissues, thereby altering its binding specificity. We examined whether mRNAs isolated from different tissues may synthesize different types of tachykinin receptors in the same expression system. For this purpose, Xenopus oocytes were injected with poly (A)+ RNAs extracted from rat brain or bovine stomach, and their responses to different tachykinins were examined under voltage-clamp. On the basis of potency ranking of 6 tachykinin agonists, the receptor induced by rat brain mRNA was found to correspond to a tachykinin receptor subtype currently classified as the NK-1 (SP-P) receptor, whereas that synthesized by bovine stomach mRNA corresponded to the NK-2 (NK-A) receptor. Thus, each of the 2 receptors can be induced in the same expression system, depending upon the source of exogenous mRNA injected. Therefore, the difference in the nature of the 2 receptors does not seem to be due to the possible posttranslational modification alone. However, the ionic mechanisms underlying activation of the 2 receptors translated in oocytes were similar. It is likely that activation of the 2 receptors uses the same internal mediator in oocytes.
Ultrastructural analysis of the distribution of synaptic boutons from labeled preganglionic axons on rabbit ciliary neurons
Authors: Forehand CJ
Published: March 2018

The number of preganglionic inputs that innervate rabbit ciliary ganglion cells is directly correlated with the number of dendrites arising from each ganglion cell (Purves and Hume, 1981). In general, the innervation of multiply innervated ciliary neurons by individual preganglionic axons is regionally restricted to a portion of the postsynaptic surface that usually includes the cell body and some, but not all, of the dendrites (Forehand and Purves, 1984). These observations suggest that dendrites modulate convergence to each cell by providing relatively separate postsynaptic domains for individual inputs. To examine this possibility further, I have assessed the distribution of synaptic boutons from individually labeled preganglionic axons on ciliary ganglion cells at the ultrastructural level. The results show that at least a third of the dendrites of these neurons are contacted exclusively by synaptic boutons from a single preganglionic axon. However, at least half of the dendrites (and nearly all of the cell bodies) of multiply innervated ganglion cells are innervated by at least 2 different preganglionic axons. Moreover, synapses from 2 different inputs often coexist in close proximity on the postsynaptic surface. Thus, individual preganglionic axons do not require exclusive dominion over a particular part of a postsynaptic cell in order to maintain their connection with the cell. These results suggest that competitive interactions between the inputs to these cells occur between the sets of boutons arising from different inputs, rather than at the level of individual boutons.(ABSTRACT TRUNCATED AT 250 WORDS)
Inhibitory interactions between spiking and nonspiking local interneurons in the locust
Authors: Burrows M
Published: March 2018

Simultaneous intracellular recordings were made from pairs of spiking and nonspiking local interneurons in the metathoracic ganglion of the locust to search for interactions that might underlie tactile and proprioceptive reflexes of a leg. A spike in a spiking local interneuron is followed after a consistent latency (0.6 +/- 0.12 msec, mean +/- SD) by an IPSP in a particular nonspiking interneuron. The connection appears to be direct and chemically mediated. By contrast, manipulating the membrane potential of a nonspiking interneuron by injecting current through the recording electrode has no direct effect on a spiking local interneuron. The direct interactions between pairs of these local interneurons are thus one-way. If, however, the current injected into a nonspiking interneuron is sufficient to evoke a movement by exciting motor neurons, then the spiking interneuron can be excited or inhibited by the resulting reafference. The spiking local interneurons have excitatory regions in their receptive fields formed by arrays of exteroreceptors or by proprioceptors at specific joints. The inhibitory connections mean that the postsynaptic nonspiking interneurons have corresponding inhibitory regions to their receptive fields. Several spiking local interneurons with similar receptive fields may converge onto one nonspiking interneuron. Some nonspiking interneurons, however, have larger receptive fields than an individual spiking interneuron, again indicating convergence of inputs. The specificity of the inhibitory connections preserves the spatial representation of sensory information for use in particular reflexes. For example, touching hairs on the ventral femur evokes a reflex extension of the tibia. Spiking interneurons excited by these receptors inhibit a nonspiking interneuron that would cause the opposing and therefore unwanted flexion movement. Viewed in this behavioral context, the pattern of connections between the local interneurons forms the basis of the circuitry for the local reflex adjustments of posture and locomotion.
Dependence of an adenosine-activated potassium current on a GTP-binding protein in mammalian central neurons
Authors: Trussell LO , Jackson MB
Published: March 2018

Neurons in hippocampal and striatal cell cultures respond to adenosine with an inhibitory potassium current. This response disappears during whole-cell patch-clamp recording in which the cell is filled with minimal saline. We have found that this loss of sensitivity to adenosine can be prevented by including 100 microM GTP in the patch electrode filling solution. GDP is less effective than GTP in supporting the adenosine response, while GMP has little, if any, effect. Treatments known to inhibit GTP-binding proteins (G-proteins) block the adenosine-activated potassium current: The adenosine response is inhibited by including poorly metabolized analogs of guanine nucleotides along with GTP in the recording electrode. Diphosphate and triphosphate analogs appear to achieve this effect through different mechanisms. The adenosine response is also blocked by incubating cultures in islet-activating protein (pertussis toxin), an inhibitor of a class of G-protein. Thus, our data implicate a G-protein in the activation of a potassium current by adenosine. Intracellular ATP can increase the effectiveness of GMP, GDP, or low concentrations of GTP, suggesting that even during internal dialysis, neurons can maintain GTP levels through phosphotransferase reactions. Intracellular ATP also appears to suppress an outward current that is different from the adenosine-activated current. Raising intracellular cAMP levels either with bath-applied forskolin or by including a cAMP analog in the recording electrode did not alter the adenosine response. These results indicate that a G-protein is involved in the coupling between the adenosine receptor and a potassium channel, and that this coupling is not mediated by cAMP.
A transient postmamillary component of the rat fornix during development: implications for interspecific differences in mature axonal projections
Authors: Stanfield BB , Nahin BR , O'Leary DD
Published: March 2018

We have used anterograde and retrograde tracing techniques to study the development of the postcommissural fornix in albino rats. The fornix reaches the caudal hypothalamus a day or two before birth. Before fornix axons can be seen entering the mamillary nuclei, a prominent contingent of the fornix can be followed beyond the mamillary bodies. This postmamillary component continues to grow into the midbrain and pontine tegmentum during the first postnatal week as the projection to the mamillary nuclei is being elaborated. During the second and third postnatal weeks, the postmamillary component of the fornix becomes progressively smaller until it is completely eliminated. The cells of origin of this transient postmamillary component of the fornix are found within the subicular complex of the hippocampal region. Most, if not all, of the cells of origin of the postmamillary component of the fornix survive the period during which this projection is eliminated. Consistent with these observations is the interpretation that the axons of the fornix that eventually enter and arborize within the mamillary nuclei and are maintained in the adult, arise during development as interstitial collaterals from parent fibers, the distal portions of which are subsequently eliminated. While not present in adult rats, a postmamillary component of the fornix has been described in other species, such as the cat (Nauta, 1958). It would seem, therefore, that in this case the interspecific variations in projection patterns result from the differential elaboration and elimination of an initially quite similar pattern of connections.
Axonal signals regulate the differentiation of non-myelin-forming Schwann cells: an immunohistochemical study of galactocerebroside in transected and regenerating nerves
Authors: Jessen KR , Mirsky R , Morgan L
Published: March 2018

Little is known about the factors involved in directing and maintaining the divergent differentiation of the 2 major Schwann cell variants, myelin and non-myelin-forming cells, in peripheral nerves. There is strong evidence that the differentiation of myelin-forming cells depends critically on cell-cell signaling through contact with appropriate axons. In this paper we ask whether this remarkable dependence of the Schwann cell on axonal contact for full differentiation is unique to those cells that form myelin or whether axonal signaling is also an important factor in the differentiation of non-myelin-forming Schwann cells. Sciatic nerves or cervical sympathetic trunks of adult rats were either transected or crushed and the axons allowed to degenerate and, in the case of crushed nerves, to regenerate into the distal stump for periods of time varying from 2 d to 9 weeks. The distal stump of the nerve was excised at specific times, the Schwann cells dissociated and immunolabeled with antibodies to galactocerebroside. In the sciatic nerve, which contains a mixture of non-myelin-forming and myelin-forming Schwann cells, transection resulted in a loss of galactocerebroside expression from the surface of all the Schwann cells in the distal stump over a 9 week period, irrespective of their original phenotype. In crushed sciatic nerves, where axons were allowed to regrow into the distal stumps, the number of Schwann cells expressing immunohistochemically detectable quantities of galactocerebroside in the stump declined over the first 3 weeks, but by 9 weeks after crush the total percentage of galactocerebroside- positive cells in the nerve had risen to control levels.(ABSTRACT TRUNCATED AT 250 WORDS)
Two-stage expression of neurofilament polypeptides during rat neurogenesis with early establishment of adult phosphorylation patterns
Authors: Carden MJ , Trojanowski JQ , Schlaepfer WW , Lee VM
Published: March 2018

Monoclonal antibodies (mAbs) to rat neurofilament (NF) proteins NF-L, NF-M, and NF-H were used to examine the developmental programs of NF expression in rat embryos. The ability of these mAbs to recognize differentially phosphorylated states of NF-M and NF-H (Lee et al., 1987, the preceding paper) was exploited in order to examine the temporal and spatial patterns of NF phosphorylation during early neuronal development in vivo. NF proteins were first detected on the twelfth day postfertilization (E12) using NF-L- or NF-M-specific mAbs. By E13, the coexpression of NF-L and NF-M was widespread, reflecting dramatic increases of immunoreactivity to both subunits. Partial phosphorylation, denoted P[+], of NF-M was already present in perikarya and neurites of E12 neurons. Extensively phosphorylated, or P[+++], isoforms of NF-M appeared in E13 axons, thereby establishing a proximodistal gradient of NF phosphorylation during the earliest phase of NF expression. Immunoblots of tissue homogenates revealed that most NF-M of E13 embryos exists in a partially phosphorylated, or P[+], isoform. Unequivocal staining for NF-H first appeared at E15, a time at which NF-L and NF-M had already attained their adult patterns of immunocytochemical staining. Levels of NF-H were extremely low at E15 but could be detected in all of its differentially phosphorylated states, i.e., nonphosphorylated P[-], partly P[+], and highly P[+++] phosphorylated isoforms. P[+++] isoforms of NF-H were restricted to the distal portions of E15 axons, although staining of more proximal axons, like those in adult, was noted by E17. Immunoblots of E17 embryos revealed most NF-H as P[-] and P[+] isoforms. Quantities of immunoreactive NF-H increased very slowly and remained well below those of NF-M and NF-L for several weeks beyond birth. These results show that sequential forms of NFs are expressed by developing and maturing neurons throughout the nervous system. An “immature” form of NFs, composed of NF-M and NF-L, appears to function in establishing the neuronal phenotype and in initiating and maintaining neurite outgrowth. Addition of NF-H confers a “mature” state to the NF. This delayed expression of NF-H is a slow and graduated process that coincides in time with the stabilization of neuronal circuitries and may be important in modulating axonal events, such as the slowing of cytoskeletal transport and the growth of axonal caliber.
Catecholaminergic properties of cholinergic neurons and synapses in adult rat ciliary ganglion
Authors: Landis SC , Jackson PC , Fredieu JR , Thibault J
Published: March 2018

Parasympathetic neurons of the ciliary ganglion are innervated by preganglionic cholinergic neurons whose cell bodies lie in the brain stem; the ganglion cells in turn provide cholinergic innervation to the intrinsic muscles of the eye. Noradrenergic innervation of the iris is supplied by sympathetic neurons of the superior cervical ganglion. Using immunocytochemical and histochemical techniques, we have examined the ciliary ganglion of adult rats for the expression of cholinergic and noradrenergic properties. As expected, the postganglionic ciliary neurons possessed detectable levels of choline acetyltransferase immunoreactivity (ChAT-IR). Unexpectedly, many ciliary neurons also exhibited immunoreactivity for tyrosine hydroxylase (TH-IR). Some had dopamine beta-hydroxylase-like (DBH-IR) immunoreactivity, but none contained detectable catecholamines, even after treatment with nialamide and L-DOPA. A sparse plexus of fibers exhibiting faint TH-IR was present in the irises of acutely sympathectomized rats. The terminals of preganglionic axons in the ciliary ganglion exhibited not only immunoreactivity for ChAT, but also for TH and contained stores of endogenous catecholamine. Neither ciliary neurons nor their preganglionic innervation accumulated detectable stores of exogenous catecholamines. Rats sympathectomized as neonates by treatment with 6- hydroxydopamine subsequently had a greater proportion of neurons possessing detectable TH-IR in the ciliary ganglion; both the TH-IR perikarya and their axons in the iris were more intensely immunofluorescent. TH-IR was present in the ciliary neuron cell bodies of mouse, guinea pig, and ferret. These species, however, lacked detectable TH-IR or catecholamine stores in preganglionic terminals. These observations indicate that mature, functionally cholinergic neurons from 2 different embryonic origins, postganglionic ciliary neurons derived from the neural crest and preganglionic neurons derived from the neural tube, display several catecholaminergic properties.
Healing and growth of half-eye "compound eyes" in Xenopus: application of an interspecific cell marker
Authors: O'Gorman S , Kilty J , Hunt RK
Published: March 2018

Surgically constructed compound eyes have been widely used to examine the development of retinotectal projections in amphibians. Such studies have been limited, however, by the lack of an adequate cellular marker with which to assess the contributions of grafted and host tissues to the later larval and adult retina. We have followed the growth of graft- and host-derived ocular tissues in interspecific compound eyes prepared by orthotopic and heterotopic exchanges of half-eye fragments between pigmented Xenopus borealis and albino Xenopus laevis embryos. This genotypic combination allowed the growth of graft-derived choroid and pigment epithelium to be studied in the living animal, and permitted cell-by-cell resolution of graft- and host-derived neurons in quinacrine-stained paraffin sections. At mid- and late-larval stages graft-derived neurons occupied large and usually coherent territories of retina in all classes of orthotopic and heterotopic compound eyes. In successfully healed cases, sample means of the percentage of the total retina occupied by graft-derived neurons ranged from 43 to 51%. Graft-derived territories originated near the optic nerve head and extended into the germinal neuroepithelium at the retinal periphery. As compared to orthotopic compound eyes, graft-derived territories in all classes of heterotopic compound eyes were slightly smaller and significantly more variable. Despite this variability, the correlation between graft-derived pigment epithelial and retinal territories was quite good in heterotopic compound eyes. While graft-host borders were generally sharp, there was also evidence of local cell mixing over distances of several cell diameters in the inner nuclear and ganglion cell layers. Single displaced cells, isolated from other members of their genetic cohort by 5 or more cell diameters, were also present in the inner nuclear and ganglion cell layers.
Differential neuronal survival in the avian ciliary ganglion after chronic acetylcholine receptor blockade
Authors: Meriney SD , Pilar G , Ogawa M , Nunez R
Published: March 2018

We have described in the preceding 2 papers the development of the pharmacological and contractile properties of all targets of the ciliary ganglion: the iris and ciliary body (Pilar et al., 1987), and the choroidal coat (Meriney and Pilar, 1987). In this paper, we examine the chronic effects of ACh receptor (AChR) blockade on ciliary ganglion neuron survival. Nicotinic or muscarinic AChR blockers were administered daily to developing chicken embryos during the normal neuronal death period in the ciliary ganglion. The effects of the blockers on ganglionic and neuromuscular transmission were assessed, and neuronal survival was assayed by counting both the total number of ganglion neurons and the selectively HRP-labeled ciliary neurons after the normal neuronal death period. Blockade of ganglionic transmission decreases survival in both populations of neurons. Blockade of neuromuscular muscular transmission increases survival in the ciliary population, which innervates the striated iris and ciliary body muscle. In contrast, blockade of synaptic activity has various influences on the survival of the choroid population, which innervates the smooth muscle of the choroid coat. Smooth muscle muscarinic receptor blockade with atropine does not influence survival. At higher doses (which block ganglionic transmission), atropine decreases choroid survival. Survival of the choroid population is increased by nicotinic blockade with 75 micrograms alpha bungarotoxin (alpha BTX), but decreased by 12.5 micrograms alpha BTX. Two main conclusions arise from these studies. Activation of postsynaptic AChRs in both the ganglion and the periphery are important in the regulation of neuronal survival. These effects usually occur in opposite directions: Blockade of ganglionic transmission decreases neuronal survival, while paralysis of neuromuscular transmission increases neuronal survival. This embodies the “balance” hypothesis (Cunningham, 1982) for neuronal survival, which states that motoneurons must balance afferent and target interactions during a critical period after synapses are formed in both regions. The present observations support this hypothesis. However, although both ciliary and choroid neurons have been shown to depend on the presence of the periphery for survival, target muscle paralysis via AChR blockade rescues the ciliary neurons but does not influence survival in the choroid population. Target-dependent regulation of choroid neuron survival during the normal neuronal death period is clearly different from the regulation of ciliary neuron survival.
Optical recording of calcium and voltage transients following impulses in cell bodies and processes of identified leech neurons in culture
Authors: Ross WN , Arechiga H , Nicholls JG
Published: March 2018

Optical methods were used to examine the spread of electrical potentials and the distribution and time course of calcium transients in individual identified nerve cells isolated from the leech. A photodiode array detected voltage transients by measuring absorbance changes of cells stained with the voltage-sensitive dye RH155 added to the bath. Calcium transients were recorded by measuring absorbance changes of the dye arsenazo III, which had been injected into the cells. In addition, Lucifer yellow was injected to outline the some and processes. Calcium changes resulting from individual action potentials were recorded from N, P, and Retzius cells without averaging. Signals from T cells and anterior pagoda (AP) cells were weaker but could be detected with averaging. These results are in accord with previous studies on calcium contributions to action potentials in these cells. For all cells, larger or wider action potentials gave bigger signals. Calcium changes from each of a train of action potentials were of equal amplitude, showing no sign of facilitation. Calcium transients from Retzius cells that had formed chemical synapses with P cells had properties similar to those of isolated cells. We were also able to detect responses from prolonged subthreshold depolarizations to -40 mV from a hyperpolarized membrane potential (-65 mV). These signals rose throughout the duration of the pulse (1–2 sec). With the photodiode array we mapped the distribution of the calcium signals. The amplitudes from each pixel are proportional to the amount of calcium entering that element in response to the stimulating pulse, if the simplifying assumption is made that the calcium buffering of the cytoplasm is uniform throughout the cell. The largest signals were detected over the axon stump left from the cell isolation procedure. Large signals were also detected from the soma. Weak signals were detected from the processes of some cells. From many Retzius cells, no signals at all were detected from the newly formed processes. Using the photodiode array, we also recorded voltage transients from the cells. Signals were recorded from all over the arborization of the neuron, with no obvious variation in time course, showing that the entire cell, including fine slender processes and broad growth cones, was essentially isopotential. Combining these observations with the measured distribution of calcium transients in the same cell suggests that the density of calcium channels in most cells is less in the outgrowing processes than in the soma or axon stump.
The role of dopamine in intracranial self-stimulation of the ventral tegmental area
Authors: Fibiger HC , LePiane FG , Jakubovic A , Phillips AG
Published: March 2018

The role of dopaminergic (DA) neurons in brain stimulation reward produced by electrical stimulation of the ventral tegmental area (VTA) was investigated in the rat. In the first experiment, extensive 6- hydroxydopamine lesions of the ascending fibers of the mesotelencephalic DA projections resulted in significant changes in intracranial self-stimulation (ICS) rate-current intensity functions when the lesion was ipsilateral to the stimulating electrode. Similar contralateral lesions had no effect on these functions, thus ruling out lesion-induced performance deficits as being responsible for the decreases in ICS rates across the wide range of current intensities that occurred after the ipsilateral lesions. In the second experiment, ICS obtained from electrodes in the VTA resulted in significant increases in the DA metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the striatum, nucleus accumbens, and olfactory tubercle ipsilateral to the stimulating electrode. The ratios of DOPAC and HVA to DA, considered to be indices of DA utilization, were also increased in these brain regions ipsilateral to the electrode. No changes were observed in the contralateral striatum, nucleus accumbens, and olfactory tubercle. Similar increases were observed in stimulated “yoked” animals that received brain stimulation at identical rates and currents but did not lever-press for this stimulation. The third experiment examined the effects of lever- pressing for food on an FR8 schedule of reinforcement on DA utilization in the striatum, nucleus accumbens, and olfactory tubercle. Despite high rates of responding, no effects were observed on DOPAC:DA or HVA:DA ratios in these brain regions.
Serotonin-storing secretory granules from thyroid parafollicular cells
Authors: Barasch JM , Tamir H , Nunez EA , Gershon MD
Published: March 2018

The subcellular storage of 5-HT was studied in sheep thyroid parafollicular cells. These cells are neural crest derivatives and were investigated as a serotonergic model system. Light and electron microscopic immunocytochemistry was used to examine the distributions of 5-HT, 45 and 56 kDa forms of 5-HT binding protein (SBP), and calcitonin. A single type of parafollicular cell was found that contained calcitonin, 5-HT, and 45 kDa SBP but not 56 kDa SBP. The secretory granules of parafollicular cells all displayed calcitonin immunoreactivity, and many were also immunoreactive for 5-HT and 45 kDa SBP. Granules were isolated, first by size and then by density, on successive metrizamide gradients. These provided a granular fraction that was enriched in calcitonin, endogenous 5-HT, and 45 kDa SBP. Immunoblots confirmed the presence of 45 kDa SBP in the isolated granules and in suspensions of parafollicular cells that were purified by an affinity chromatographic technique. Parafollicular cell granules did not appear to contain substantial stores of ATP. Granules isolated on Percoll gradients were morphologically homogeneous and took up 3H-5- HT. The specificity of this uptake was confirmed by quantitative electron microscopic radioautography. The granular uptake of 3H-5-HT was inhibited by reserpine (10 microM). It is concluded that parafollicular cell granules are different from other amine-storing vesicles that do contain ATP; nevertheless, since parafollicular cell granules store 5-HT and have the same 45 kDa SBP as is found in serotonergic axon terminals, parafollicular cell granules may be analogous to the synaptic vesicles of serotonergic neurons.