bluuebunny/crossref_metadata_2025 huggingface dataset:
Insect olfactory neurons in vitro: morphological and physiological characterization of cells from the developing antennal lobes of Manduca sexta
Authors: Hayashi JH , Hildebrand JG
Published: March 2018

To facilitate studies of the development and membrane biophysics of cells in the central olfactory pathway of the moth Manduca sexta, we have dissociated neurons and glial cells from the antennal lobes (ALs) and cultivated them in the controlled environment of tissue culture. Cultures produced from cells of the lateral group of AL neurons alone are enriched in local interneurons (LNs), while cultures made from cells of the median and anterior groups of AL neurons contain projection neurons (PNs) but lack LNs. Cultures containing only PNs, but not cultures with both PNs and LNs, require a conditioning factor derived from a conspecific cell line in order to ensure survival. Under these conditions, we identify 5 types of cells in the “PN-only” cultures that are consistently observed and distinguishable on the basis of their morphology and characteristic whole-cell current profiles. In cultures that contain both PNs and LNs, we find 3 additional types of cells with neuronlike appearance. Whereas the PNs have whole-cell currents that suggest a neuronal phenotype, we have not yet observed similar currents in the 3 other types of cells. We tentatively identify the latter 3 cell types as LNs whose development has been arrested. The conditioning factor also has a trophic effect on phase-dark, multipolar cells that are observed in freshly dispersed cultures. Their morphology and whole-cell currents lead us to suggest that they are glial cells.
Na current in membrane blebs: implications for channel mobility and patch clamp recording
Authors: Milton RL , Caldwell JH
Published: March 2018

When suction was applied to loose patch clamp pipettes while recording from enzymatically dissociated muscle fibers, large membrane blebs formed within the pipettes. We initiated a study of these suction- induced blebs because ion channels in the blebs would complicate or possibly invalidate loose patch voltage clamp measurements of membrane current density. The low lateral mobility (Stuhmer and Almers, 1982) and steep gradients of Na channels at the end-plate and tendon (Caldwell et al., 1986) imply tight binding of Na channels to cytoskeletal elements and led us to expect few, if any, Na channels in the blebs. Bleb formation produced an increase in membrane capacitance, as expected from the increase in membrane area. Bleb formation also increased the Na current, indicating that the blebs contained Na channels. Assuming that the increased capacitance and Na current were due to lipid and Na channels moving from membrane outside the pipette, ejection of the bleb from the pipette was expected to bring the capacitance and Na current back to their original values. Capacitance did return to its original value, but Na current was lower than expected. The decrease in Na current is explained by Na channels moving from the patch membrane into the bleb. Normalization of bleb and patch Na current to their respective capacitances revealed that bleb membrane had a Na channel density almost 50% that of normal surface membrane. Thus, bleb membrane is neither devoid of proteins nor truly representative of the normal surface membrane from which it arose. It is enriched in membrane lipids and is relatively protein poor. Two conclusions can be drawn.(ABSTRACT TRUNCATED AT 250 WORDS)
Adaptations of synaptic form in an aberrant projection to the avian cochlear nucleus
Authors: Parks TN , Taylor DA , Jackson H
Published: March 2018

Surgical removal of the otocyst in chick embryos induces axons from the contralateral cochlear nucleus (nucleus magnocellularis, NM) to form, in addition to their normal endings in nucleus laminaris (NL), anomalous and persistent functional contacts in the ipsilateral NM (Jackson and Parks, 1988). We have examined how interaction between the abnormal synaptic partners during development influences the form of the axon terminal and its relation to the target neuron. In the light microscope, aberrant axon terminals labeled in vitro with HRP appear to form boutons quite unlike the large calycine endbulbs made by the normal cochlear nerve (CN) endings in NM. In the electron microscope, however, the anomalous endings appear embedded in the NM cells, something never seen normally in NM or NL. Morphometric analyses were performed on electron micrographs from NM and NL in animals aged embryonic day (E) 19 to posthatching day (P) 2 from which the right otocyst had been removed on E3 and in normal control animals. Aberrant endings appose 18% of the circumference of operated NM cells, versus 45% for CN axons in the normal NM at this age. The mean length of membrane apposition for the anomalous NM-to-NM endings was 215% greater than for normal NM-to-NL endings but 54% smaller than that in normal CN endings. These results support the idea that developmental interactions between synaptic partners can influence the form of the contact between the 2 neurons. The results also demonstrate, however, that formation of persistent and functional synapses with NM neurons throughout development is not sufficient to induce any axon to assume the calycine form of a cochlear nerve endbulb.(ABSTRACT TRUNCATED AT 250 WORDS)
Distribution and lateral mobility of glycine receptors on cultured spinal cord neurons
Authors: Srinivasan Y , Guzikowski AP , Haugland RP , Angelides KJ
Published: March 2018

Strychnine is one of the most potent antagonists of glycine-mediated inhibitory conductances in the mammalian spinal cord. In order to examine the distribution of glycine receptors (GlyRs) on neuronal cells, 2 novel fluorescent strychnine derivatives were synthesized and characterized chemically, spectroscopically, and biologically. Both compounds retain their biological activity after derivatization and are potent inhibitors of 3H-strychnine binding to bovine spinal cord membranes and membranes from rat spinal cord cultures. Using these fluorescent strychnine analogs, the cellular distribution and lateral mobility of GlyRs on cultured rat spinal cord neurons were studied by digital fluorescence imaging and photobleach recovery microscopy. On these neurons, even in the absence of observable synaptic contact and early in development GlyRs are predominantly localized to cell bodies with sparse labeling of neuritic processes. Although GlyRs are confined to the neuronal cell body, approximately 50% of the receptors are very mobile, with lateral diffusion coefficients of 1.15 +/- 0.05 x 10(-9) cm2/sec, a value which is characteristic of unrestricted protein lateral diffusion. However, the remaining fraction of these receptors are immobile on the neuronal cell body. More than 70% of the GlyRs distributed on neuronal processes are immobile, while 30% are laterally mobile, with diffusion rates of 5.50 +/- 0.1 x 10(-10) cm2/sec. The results indicate that even early in development GlyRs are expressed and segregated to the cell body, where they are confined within a domain that restricts their redistribution.
The lateral amygdaloid nucleus: sensory interface of the amygdala in fear conditioning
Authors: LeDoux JE , Cicchetti P , Xagoraris A , Romanski LM
Published: March 2018

Previous work has implicated projections from the acoustic thalamus to the amygdala in the classical conditioning of emotional responses to auditory stimuli. The purpose of the present studies was to determine whether the lateral amygdaloid nucleus (AL), which is a major subcortical target of projections from the acoustic thalamus, might be the sensory interface of the amygdala in emotional conditioning. Lesions were placed in AL of rats and the effects on emotional conditioning were examined. Lesions of AL, but not lesions of the striatum above or the cortex adjacent to the AL, interfered with emotional conditioning. Lesions that only partially destroyed AL or lesions placed too ventrally that completely missed AL had no effect. AL lesions did not affect the responses elicited following nonassociative (random) training. AL is thus an essential link in the circuitry through which auditory stimuli are endowed with affective properties and may function as the sensory interface of the amygdala during emotional learning.
Expression of long-term adaptation of synaptic transmission requires a critical period of protein synthesis
Authors: Nguyen PV , Atwood HL
Published: March 2018

The crayfish claw closer muscle is innervated by 2 distinct excitatory motoneurons, one tonic and the other phasic. The phasic motoneuron is relatively inactive and generates large EPSPs that normally depress rapidly with repetitive stimulation at moderate frequencies. Stimulation of the phasic motoneuron in vivo for 3 d at 5 Hz (2 hr/d) produced a marked adaptive shift in the neuromuscular synaptic response properties of the motoneuron: average initial EPSPs and depression of EPSPs were significantly reduced. We tested the hypothesis that neuronal protein synthesis is required for full expression of long-term adaptation (LTA). A reversible inhibitor of neuronal protein synthesis, cycloheximide (CHX), was injected into intact crayfish at various times prior to, during, or after each stimulation period. At a dosage of 5 micrograms/gm body weight, CHX inhibited the incorporation of [35S]- methionine into abdominal nerve cord protein for approximately 2 hr after administration (greater than 80% inhibition). Full expression of LTA was selectively blocked when CHX was administered 6 hr or 2 hr prior to each stimulation period. Both the reduction in initial EPSP amplitude and the resistance to synaptic depression were significantly attenuated. CHX administered at the onset of or at the end of each stimulation period did not affect the expression of LTA. Control experiments using unstimulated animals showed that neither chronic nor acute administration of CHX adversely affected the phasic axon's synaptic response properties. Our results suggest that full expression of neuronal LTA requires the presence of a pool of preexisting, short- lived (or rapidly utilized) protein(s). Depletion of such a pool prior to each stimulation period appears to interfere with subsequent induction of LTA.
Differential expression of ARPP-16 and ARPP-19, two highly related cAMP- regulated phosphoproteins, one of which is specifically associated with dopamine-innervated brain regions
Authors: Girault JA , Horiuchi A , Gustafson EL , Rosen NL , Greengard P
Published: March 2018

ARPP-16 and ARPP-19 are 2 cAMP-regulated phosphoproteins of Mr = 16,000 and 19,000, respectively, which are identical except for the presence of 15 additional amino acids on the NH2-terminus of ARPP-19. The phosphorylation of these 2 proteins is regulated by cAMP and vasoactive intestinal peptide in reaggregate striatal cultures (Girault et al., 1988). Using immunoblots and immunocytochemistry, we have compared the regional, subcellular, phylogenetic, and ontogenetic distributions of these 2 proteins. ARPP-19 was found in all vertebrate species studied and, at various levels, in all tissues of adult rat. ARPP-19 was also present at high levels in malignant cell lines. During development ARPP- 19 concentrations were highest in the embryo and decreased during the pre- and postnatal periods. In contrast, ARPP-16 was detected only in some specific neurons of the dopamine-innervated regions of the basal ganglia and the cerebral cortex, which are known to possess D1 dopamine receptors, in particular the striatonigral neurons. ARPP-16 is phylogenetically recent, being found only in birds and mammals, and appears late in ontogenesis, increasing during the postnatal period. These 2 proteins provide a unique model for studying the specificity of signal transduction and gene expression in dopaminoceptive neurons.
Emergence and refinement of clustered horizontal connections in cat striate cortex
Authors: Callaway EM , Katz LC
Published: March 2018

Pyramidal cells in layer 2/3 of adult cat striate cortex have long, intrinsic horizontal axon collaterals within both layer 2/3 and layer 5. These collaterals form periodic “clusters” of finer axon branches that link columns of similar orientation selectivity. We have investigated the sequence of events and possible mechanisms underlying the development of these clustered intrinsic horizontal connections using a combination of neuronal tracers and intracellular staining. Small injections of fluorescent latex microspheres made during the first postnatal week (at P4–6), when examined in tangential sections, produced an even, unclustered distribution of retrogradely labeled cells up to 2 mm from the injection site. At P8, retrograde labeling extended over a larger area and clustering was discernible, primarily among the most distant labeled cells. At both P6 and P8, labeling was similar in layers 2/3 and 5, indicating that the transition from clustered to unclustered connections occurred simultaneously for cells in superficial and deep laminae. By the end of the second postnatal week (P12–15), retrogradely labeled cells were far more clustered both within and beyond the extent of P6 label; the density of labeled cells was high throughout the labeled region, but much higher within clusters. The periodicity of these nascent clusters was similar to that in the adult. Despite obvious clustering, the pattern of retrograde label observed following injections at 2–3 weeks (P12–21) differed markedly from the adult, in that the regions between clusters contained many labeled cells. Over the next 3 weeks, the connections were refined, so that by the sixth postnatal week (P36–38), regions between clusters contained very few retrogradely labeled cells and the overall pattern of retrograde label was indistinguishable from that in adults. Despite differences in postmigratory ages of neurons from the superficial and deep laminae, clustering of retrogradely labeled cells from these 2 populations was similar at all ages. Experiments in which 2–3 weeks elapsed between the time microsphere injections were made and animals were killed demonstrated that neither the initial formation of crude clusters nor their refinement was due to cell death. Instead, cluster refinement resulted from specific process elimination. When a red microsphere injection at P15 was followed by a green microsphere injection at exactly the same location on P29, the earlier injection resulted in crude clustering, as expected. Virtually all of the cells double-labeled by the later injection were within the densest clusters of label from the early injection.(ABSTRACT TRUNCATED AT 400 WORDS)
Fluorescence measurement of changes in intracellular calcium induced by excitatory amino acids in cultured cortical astrocytes
Authors: Jensen AM , Chiu SY
Published: March 2018

Population response of [Ca2+]i in cultured cortical astrocytes to excitatory amino acids was measured at room temperature using the calcium-sensitive dye fura-2. Quisqualic acid (QA), glutamate (Glu), and kainic acid (KA) caused a peak increase in [Ca2+]i in the order QA greater than Glu greater than KA. No response to N-methyl-D-aspartic acid (NMDA) was observed whether or not Mg2+ was present externally. Both QA and Glu (100 microM) frequently elicited a decaying oscillatory [Ca2+]i response during sustained agonist application; the period of oscillations initially was 23.5 sec and increased as the response was damped. Comparatively, the [Ca2+]i response to KA was nonoscillatory. Both responses to Glu and KA were reduced slightly by antagonist gamma- D-glutamylaminomethyl-sulfonic acid (1 mM), but virtually were abolished by kynurenic acid (3 mM). Replacement of external Na+ by choline had no significant effect on the Glu response. Removal of external Ca2+ reduced the peak response to QA, Glu, and KA to 40, 34, and 18%, respectively; and markedly reduced the degree of QA- and Glu- induced [Ca2+]i oscillations. Pretreatment with phorbol esters, a potent activator of protein kinase C, blocked the [Ca2+]i response to Glu but not KA. It is concluded that cortical astrocytes express Glu receptors of the non-NMDA type in culture and that receptor activation leads to Ca2+ influx and release of internal Ca2+. Mobilization of Ca2+ apparently occurs via the known Glu-mediated hydrolysis of inositol lipids, which may come under negative-feed-back control by protein kinase C activation. Oscillatory [Ca2+]i signaling offers the possibility of a dynamic population response in an electrically coupled glial network.
Intrinsic mechanisms of antinociception in inflammation: local opioid receptors and beta-endorphin
Authors: Stein C , Gramsch C , Herz A
Published: March 2018

This study examined antinociception induced through the activation of local opioid receptors in inflammation by endogenous opioids. Rats developed a unilateral localized inflammation upon injection of Freund's adjuvant into one hindpaw. Four to 6 d later they were subjected to cold water swim (CWS), an environmental stimulus known to activate intrinsic opioid systems. Following CWS (1 min) the animals' withdrawal threshold to noxious pressure applied onto the paws increased significantly more on the inflamed paw than on the noninflamed paw. This unilateral antinociceptive effect in inflamed paws was dose-dependently and stereospecifically reversible by intraplantar (i.pl.) but not systemic (i.v. or s.c.) administration of the opioid antagonist naloxone (18 micrograms). This suggested that CWS- induced antinociception in inflamed tissue was brought about by the activation of local opioid receptors. Antiinflammatory or vasoconstrictive events, as measured by paw volume and temperature, did not contribute to this unilateral antinociception. Receptor-selective antagonists indicated the involvement of mu- and delta- but not kappa- receptors. Intravenous application of a universal antibody to endogenous opioid peptides (3-E7) and a specific antibody to beta- endorphin, but not antisera against metenkephalin or dynorphin, abolished the CWS effect. Finally, the i.pl. injection of synthetic beta-endorphin (1–31) produced an antinociceptive effect in inflamed paws which was reversible by i.pl. naloxone and selective mu- and delta- receptor antagonists. These findings suggest that antinociception in inflamed tissue can be induced through the activation of local opioid receptors by endogenous beta-endorphin released during CWS.
A voltage-clamp analysis of gene-dosage effects of the Shaker locus on larval muscle potassium currents in Drosophila
Authors: Haugland FN , Wu CF
Published: March 2018

Mutations of the Shaker (Sh) locus of Drosophila reduce, eliminate, or otherwise alter a transient potassium current, IA, in muscle. Recent molecular studies indicate that the Sh locus produces several proteins by alternative splicing, but the relationships of the variety of Sh gene products to IA channels in the various excitable membranes still remain to be determined. In Drosophila, many enzymes have been shown to exhibit gene-dosage effects; their amounts vary in direct proportion to the number of structural genes present. We describe a physiological isolation of IA in larval muscle which allowed precise quantification of gene-dosage effects on IA in Sh heterozygotes and aneuploids. We found that doubling the number of Sh genes in aneuploids increased IA to twice that of normal, consistent with the notion that the Sh locus encodes the entire IA channel in larval muscle. We further examined heterozygous combinations of different Sh mutations for evidence of interactions among Sh gene products within the IA channel, which may yield clues to the possible subunit composition of the channel. Combinations among 5 Sh mutations plus their normal counterpart followed a simple gene-dosage effect; in each case the resulting IA was about the average of the homozygous currents, compatible with the notion of additive contributions from 2 independent populations of IA channels. Two additional Sh mutations caused pronounced departures from the simple dosage effect; the amplitude of IA in heterozygotes was significantly smaller than that expected from gene dosage, a strong dominant effect attributable to interactions among protein subunits. These contrasting observations may be accounted for by certain hetero- or homo-multimeric arrangements of Sh products in the IA channel.
Increasing intracellular free calcium induces circumferential contractions in isolated cochlear outer hair cells
Authors: Dulon D , Zajic G , Schacht J
Published: March 2018

The relationship between intracellular free calcium and the motile responses of outer hair cells isolated from the guinea pig cochlea was examined. Calcium levels were modulated by the addition of the calcium ionophores ionomycin or A23187 to the incubation medium and monitored with the fluorescent calcium indicator fluo-3. In the presence of 1.25 mM external calcium, the application of either ionophore (10 microM) led to an increase in intracellular free calcium from 157 +/- 76 nM to 1200 +/- 500 nM within 30–60 sec. Concurrently, cells elongated by 1–2 microns, cell diameter decreased, and cell volume shrank by 269 +/- 220 microns 3 (5.0 +/- 4.1%). The reduction in diameter was most pronounced in the middle portion of the cell (4.4% +/- 4.2%), also evident in the apical region (3.1% +/- 4.8%) but not significant in the basal region near the nucleus. This response was observed in outer hair cells from basal and apical turns of the cochlea and was reversed when the cells were rinsed with calcium-free medium supplemented with 2 mM EGTA. Optical imaging of the cell membrane with the potentiometric dye 1-(3- sulfonatopropyl)-4-[beta] [2-(di-n-butylaminol)-6-naphthyl vinyl] pyridinium betaine during the elevation of intracellular calcium demonstrated features of contractility at the lateral cell membrane. A rise in intracellular calcium as well as the motile response was still observed after a 5-min exposure of the cells to a calcium-free solution (supplemented with 2 mM EGTA), indicating that the ionophore was also able to liberate calcium from intracellular sites. However, depletion of calcium stores through prolonged incubation of the cells in calcium- free medium (30–60 min) suppressed both the calcium signal and the cell response. The calmodulin inhibitors trifluoperazine and pimozide (30 microM) blocked the cell motility induced by ionomycin while they left the increase of intracellular calcium unaffected. These observations suggest that calcium-dependent circumferential contractions in outer hair cells are mediated by calmodulin. The application to the extracellular medium of putative neurotransmitters of the cochlear efferent system such as acetylcholine and GABA led to neither an increase in intracellular calcium nor a modification of cell shape. Therefore, these neurotransmitters may not be directly involved in calcium-induced contractions in outer hair cells. The circumferential contractions altered the stiffness of the plasma membrane and the turgor of the cell. Under normal conditions, changes in cell volume were inversely proportional to the osmotic pressure of the extracellular medium following van't Hoff's law.(ABSTRACT TRUNCATED AT 400 WORDS)
Repeated in vivo visualization of neuromuscular junctions in adult mouse lateral gastrocnemius
Authors: Wigston DJ
Published: March 2018

The structure of individually identified neuromuscular junctions (NMJs) in mouse lateral gastrocnemius (LG) muscles was studied on 2 or more occasions over 3–6 months. Presynaptic motor nerve terminals and their underlying acetylcholine receptors were stained in living animals with the fluorescent dye 4-(4-diethylaminostyryl)-N-methylpyridinium iodide) and tetramethylrhodamine isothiocyanate-conjugated alpha-bungarotoxin (R alpha BTX), respectively, and visualized by video-enhanced fluorescence microscopy. The overall shape of most NMJs changed very little over this time except for enlargement of some junctions attributable to growth of the animals. A few junctions did, however, change appreciably: over 3–6 months about 15% underwent modifications such as additions to, or losses from, their original configuration. The frequency and extent of changes in LG NMJs were substantially less than in a similar study of NMJs from mouse soleus (Wigston, 1989). These findings, together with those from other laboratories, indicate a correlation between the extent of NMJ remodeling and the fiber-type composition of a muscle: NMJs in muscles consisting of predominantly fast-twitch myofibers appear to undergo less remodeling than NMJs in muscles containing a substantial fraction of slow-twitch fibers. Since fast- and slow-twitch muscles and their motoneurons exhibit strikingly different patterns of electrical activity, these findings suggest that structural remodeling at mammalian NMJs may depend on the amount of impulse activity experienced by motoneurons, their target muscle, or individual synaptic terminals.
Alterations in activation gating of single Shaker A-type potassium channels by the Sh5 mutation
Authors: Zagotta WN , Aldrich RW
Published: March 2018

Alterations in voltage-dependent gating of A-type potassium channels by the Sh5 mutation were studied using macroscopic and single-channel recording techniques on cultured embryonic myotubes from Drosophila. The mutation increases the voltage required to activate and inactivate the A1 channel by approximately 20 mV and decreases the steepness of the voltage dependence of steady-state inactivation. Boltzmann fits to the prepulse inactivation curves have slopes of e-fold/3.5 mV for wild type and e-fold/5.0 mV for Sh5. A kinetic analysis of single Sh5 A1 channels revealed that the transition rates into and out of the open state are not significantly affected by the mutation. In contrast, the latencies until the channel opens following a voltage step are increased at low voltages. These alterations probably do not result from an offset of the transmembrane voltage sensed by the channel as might be expected for an alteration in the surface charge of the protein. By using a kinetic model developed for wild-type A1 channels, we show that an alteration in the amplitude and voltage dependence of the deactivation rate for each subunit (beta) can account for all of the alterations observed.
Distributed processing of sensory information in the leech. II. Identification of interneurons contributing to the local bending reflex
Authors: Lockery SR , Kristan WB
Published: March 2018

Isolated midbody ganglia of the leech Hirudo medicinalis were surveyed for interneurons contributing to the dorsal component of the local bending reflex, i.e., to the excitation of dorsal excitatory motor neurons that follows stimulation of dorsal mechanoreceptors responsive to pressure (P cells). Nine types of local bending interneuron could be distinguished on physiological and morphological grounds--8 paired and 1 unpaired cell per ganglion. Synaptic latencies from sensory neurons to interneurons were consistent with a direct or possibly disynaptic pathway. Connections between interneurons appeared to be rare and hyperpolarization of individual interneurons during local bending produced small but reliable decrements in motor neuron response, suggesting that multiple parallel pathways contribute to the behavior. Paradoxically, most interneurons received substantial inputs from ventral as well as dorsal mechanoreceptors, indicating that interneurons that were distinguished by their contribution to dorsal local bending were, in fact, active in ventral and lateral bends as well. The capacity to detect a particular stimulus and produce the appropriate response cannot be localized to particular types of interneuron; rather, it appears to be a distributed property of the entire local bending network.
Quantitative autoradiography reveals selective changes in cerebellar GABA receptors of the rat mutant dystonic
Authors: Beales M , Lorden JF , Walz E , Oltmans GA
Published: March 2018

In the rat mutant dystonic (dt), glutamic acid decarboxylase (GAD) activity in the deep cerebellar nuclei (DCN) is elevated compared to normal littermates. The distribution of this increase within the DCN, and the effect upon GABA receptor density, was assessed in 25-d-old animals. GAD activity was increased 45, 41, and 74% in the medial, interpositus, and lateral divisions of the DCN, respectively. Autoradiographic analysis of GABAA receptor density, using the ligand 3H-muscimol (MUSC), revealed a significant decrease in MUSC binding in the DCN of the mutant. No changes in the binding of the benzodiazepine ligand 3H-flunitrazepam (FLU) were found in the DCN. At 18 other sites, including motor areas in the brain stem, midbrain, and forebrain, no significant changes were found in either MUSC or FLU binding. There also was a failure to find any significant changes in dt animals in the binding of ligands which label the muscarinic cholinergic receptor, dopamine D2 receptor, or serotonin 5-HT2 receptor. The results support earlier findings that GABAergic activity is increased in Purkinje cell terminals of the dt mutant and suggest that in response to this enhanced activity, GABA receptors in the DCN are down-regulated. At other levels of the neuraxis no consistent changes were found in any of the variables studied, suggesting that cerebellar dysfunction may be a primary component of the dystonic syndrome.
The amygdala central nucleus and appetitive Pavlovian conditioning: lesions impair one class of conditioned behavior
Authors: Gallagher M , Graham PW , Holland PC
Published: March 2018

Rats with neurotoxic lesions of the amygdala central nucleus (CN) were trained using appetitive Pavlovian conditioning procedures. Conditioned responses (CRs) that are representative of 2 classes of behavior were monitored. One type of CR resembled the orienting responses that were elicited by the conditioned stimuli (CSs) prior to pairing with food reinforcement: the other type of CR resembled the behavior elicited by food reinforcement itself. Holland (1977, 1984) has referred to these as CS-generated and unconditioned stimulus (US)-generated CRs, respectively. During differential conditioning, some lesioned and unlesioned rats received reinforced presentations of a visual cue and nonreinforced presentations of an auditory cue, and the others in each lesion condition received reinforced auditory and nonreinforced visual cue presentations. Relative to the control group, the CN-lesioned rats were impaired in the acquisition of CS-generated CRs to both visual and auditory CSs. Orienting responses and habituation to the CSs were, however, comparable for the lesion and control groups. Moreover, rats with CN lesions readily acquired the US-generated CRs. Thus, a specific class of conditioned behavior was impaired by CN damage. Many studies using aversive Pavlovian procedures have suggested that CN lesions impair fear conditioning. The present results suggest another role for CN in the conditioning of orienting/attentional responses.
Basic FGF reverses chemical and morphological deficits in the nigrostriatal system of MPTP-treated mice
Authors: Otto D , Unsicker K
Published: March 2018

The specific mechanisms underlying the restorative effects of adrenal chromaffin grafts in experimental parkinsonism are still obscure. Recent findings indicated an involvement of graft-induced trophic interactions in the course of recovery-related events. Evidence that basic fibroblast growth factor (bFGF), a potent trophic protein for neurons, (1) is present in chromaffin cells (Blottner et al., 1989) and (2) exerts trophic activities on embryonic mesencephalic neurons in vitro (Ferrari et al., 1989) provided the rationale for administering bFGF in gel foam implants unilaterally to the striatum of 1-methyl-4- phenyl-1,2,3,6-tetrahydropyridine (MPTP) lesioned mice. Simultaneous bFGF/MPTP treatment diminished bilaterally the reduction of striatal dopamine (DA) levels observed in cytochrome c/MPTP-treated mice and led to an ipsilateral reappearance of tyrosine hydroxylase (TH)-like immunoreactive fibers, most notably adjacent to the implant, 2 weeks after the surgery. Determinations of TH activities and TH immunoblotting demonstrated that bFGF almost fully reversed the loss of TH activity on either side but restored TH protein more on the ipsilateral than on the contralateral side. Furthermore, differences in dihydroxyphenylacetic acid levels, which were about twice as high on the contralateral side yet still reduced with respect to untreated mice, supported our assumption that the molar TH activity was increased on the untreated side, possibly due to an intrinsic compensatory up- regulation. Delayed administration of bFGF starting 8 d after the MPTP treatment was equally effective with regard to morphological parameters. Our results suggest that bFGF partially prevents the deleterious chemical and morphological consequences of an MPTP-mediated nigrostriatal lesion. Thus, bFGF mimics at least the morphological effects of chromaffin cell grafts to the MPTP-lesioned brain.
Neurotropic properties of pseudorabies virus: uptake and transneuronal passage in the rat central nervous system
Authors: Card JP , Rinaman L , Schwaber JS , Miselis RR , Whealy ME , Robbins AK , Enquist LW
Published: March 2018

Uptake, replication, and transneuronal passage of a swine neurotropic herpesvirus (pseudorabies virus, PRV) was evaluated in the rat CNS. PRV was localized in neural circuits innervating the tongue, stomach, esophagus and eye with light microscopic immunohistochemistry. In each instance, the distribution of PRV-immunoreactive neurons was entirely consistent with that observed following injection of cholera toxin- horseradish peroxidase conjugate (CT-HRP). Injections of the tongue resulted in retrograde transport of PRV and CT-HRP to hypoglossal motor neurons, while preganglionic neurons in the dorsal motor vagal nucleus or somatic motor neurons in the nucleus ambiguus were labeled following injections of the stomach or esophagus, respectively. At longer times after infection, viral antigens were found in astrocytes adjacent to infected neurons and their efferent axons and second-order neuron labeling became apparent. The distribution of second-order neurons was also entirely dependent upon the site of PRV injection. Following tongue injection, second-order neurons were observed in the trigeminal complex, the brain-stem tegmentum and in monoaminergic cell groups. Injection of the stomach or esophagus led to second-order neuron labeling confined to distinct subdivisions of the neucleus of the solitary tract and monoaminergic cell groups. Comparative quantitative analysis of the number of PRV immunoreactive neurons present in the diencephalon and brain stem following injection of virus into both the eye and stomach musculature of the same animal demonstrated that retrograde transport of PRV from the viscera was more efficient and occurred at a much faster rate than anterograde transport of virus. These data demonstrate projection-specific transport of PRV in the nervous system and provide further insight into the means through which this neurotropic virus infects the nervous system.
Specific routing of retinal ganglion cell axons at the mammalian optic chiasm during embryonic development
Authors: Sretavan DW
Published: March 2018

During development of the mammalian CNS, axons encounter multiple pathway choices on their way to central target structures. A major pathway branch point in the visual system occurs at the optic chiasm, where retinal ganglion cell axons may either enter the ipsilateral or the contralateral optic tract. To investigate whether embryonic mouse retinal ganglion cell axons, upon reaching the optic chiasm, selectively grow into the correct pathway, developing retinal ganglion cells were retrogradely labeled using either 1,1′dioctadecyl- 3,3,3′,3′tetramethylindocarbocyanine perchlorate (Dil) or fluorescent microspheres placed into the optic tract on one side. The distribution of ipsilaterally and contralaterally projecting ganglion cells in the embryo was then examined and compared to that of the adult animal. Results show that axon routing at the chiasm is already extremely adult- like as early as embryonic day 15 (E15), shortly after retinal axons arrive at the chiasm. [Retinal ganglion cell neurogenesis = E11-E18 (Drager, 1985); birth = E21.] Throughout the development of this pathway, routing errors are infrequent and are on the order of only about 3–8/1000 retinal ganglion cells. Thus, embryonic retinal ganglion cell axons do not project randomly at the optic chiasm but instead appear to be highly specific in their choice of pathway. To learn how correct pathway choices are made, retinal axons were retrogradely labeled with Dil and their trajectories at the optic chiasm were reconstructed. Results show that ipsilaterally and contralaterally projecting axons are highly intermixed as they enter the chiasm region but selectively grow into the correct pathway. For example, a contralaterally projecting axon near the entrance of the ipsilateral optic tract will turn and bypass this pathway and grow towards the midline to head into the contralateral optic tract. Similarly, axons far away from the ipsilateral optic tract frequently turn abruptly at right angles to enter the ipsilateral tract, directly crossing over contralaterally projecting axons heading to the opposite side. The sorting out of intermixed ipsilaterally and contralaterally projecting retinal axons into the appropriate optic tracts strongly suggests the presence of specific guidance cues at the optic chiasm during embryonic development. Together, results from this study demonstrate that the pattern of axon projection at the adult mammalian optic chiasm is gradually built upon a highly specific pattern of axon routing laid down early during development.
Laminar organization and age-related loss of cholinergic receptors in temporal neocortex of rhesus monkey
Authors: Wagster MV , Whitehouse PJ , Walker LC , Kellar KJ , Price DL
Published: March 2018

Using in vitro receptor autoradiography, the distributions of cholinergic muscarinic [3H-N-methyl scopolamine (NMS), 3H-pirenzepine (PZ), and 3H-oxotremorine-M (OXO-M)] and nicotinic [3H-acetylcholine (ACh)] receptors were mapped in the temporal cortices of rhesus monkeys (Macaca mulatta) ranging from 2-22 years of age. Although high-affinity 3H-PZ, low-affinity 3H-NMS binding (M1 sites) and high-affinity 3H-OXO- M, high-affinity 3H-NMS binding (M2 sites) occurred across all layers of the temporal neocortex, the laminar distribution of M1 and M2 receptor binding sites was different. M1 muscarinic receptor binding was concentrated in layers II and III, whereas M2 muscarinic receptor binding was greatest in layers IV and V. The concentration of both muscarinic (M1 and M2) and nicotinic receptor binding sites declined with increasing age, and decrements were uniform across all cortical layers. This investigation provides evidence for a decrease in cholinergic receptor binding with age in temporal cortices of rhesus monkeys. Moreover, these changes appear to precede previously reported age-associated memory deficits and neuropathological changes that occur in this species.
Asynchronous appearance and topographic segregation of neuropeptide- containing cells in the developing rat adrenal medulla
Authors: Henion PD , Landis SC
Published: March 2018

The developmental expression of neuropeptide Y (NPY) and leucine- enkephalin (L-Enk) was examined in embryonic, early postnatal, and adult chromaffin cells with double- and triple-label immunocytochemical techniques and compared to the expression of immunoreactivity for tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT). In addition, the establishment of preganglionic innervation was assessed by labeling for choline acetyltransferase (ChAT) and L-Enk. NPY-IR was detectable on embryonic (E) day 15 in a clustered subpopulation of TH-IR cells. L-Enk and PNMT-IR cells were initially present on E16 in a separate nonclustered population of TH-IR cells. By late embryonic development, twice as many TH-IR cells expressed NPY and 4 times as many expressed L-Enk as in the adult. In contrast to early embryonic development, NPY-IR was evident in both the clustered and nonclustered subpopulation of TH-IR cells at this time. The proportion of NPY-IR chromaffin cells decreased to adult values during the first postnatal week at the time when obviously clustered TH-IR cells were no longer observed. The embryonic rise in the proportion of L-Enk-IR cells correlates with the developmental increase in glucocorticoid production, while the postnatal decrease corresponds to the appearance of ChAT-IR in the preganglionic innervation of the adrenal medulla. These results indicate that NPY and L-Enk are expressed at different times and in different subpopulations of cells in the embryonic adrenal. Further, the observation that peptide expression by chromaffin cells undergoes marked changes during development raises the possibility that a number of factors including developmental history, environmental signals and impulse activity play a role in the regulation of neuropeptide expression in sympathoadrenal derivatives of the neural crest.
Molecular and morphological changes in the cat lateral geniculate nucleus and visual cortex induced by visual deprivation are revealed by monoclonal antibodies Cat-304 and Cat-301
Authors: Guimaraes A , Zaremba S , Hockfield S
Published: March 2018

Monoclonal antibody Cat-301 recognizes a surface-associated proteoglycan on subsets of neurons in the mammalian CNS (Hockfield and McKay, 1983). The expression of Cat-301 immunoreactivity on Y cells in the cat LGN is sharply reduced by early visual deprivation (Sur et al., 1988). We employed an immunosuppression strategy (Hockfield, 1987) to further study alterations in the expression of experience-dependent molecules. Newborn BALB/c mice were injected with LGN from dark-reared cats to induce a suppression of the immune response to antigens expressed in visually deprived animals. These mice were then immunized with LGN from normal cats to elicit an immune response to antigens with an expression dependent on normal early visual experience. This strategy permitted the generation of monoclonal antibody Cat-304, which recognizes a surface-associated antigen on neuronal cell bodies and proximal dendrites, and which appears histologically identical to Cat- 301. Further analyses show that Cat-304 and Cat-301 recognize different epitopes on the same 680-kDa chondroitin sulfate proteoglycan. We examined the effects of early visual deprivation on Cat-304 immunoreactivity in the LGN and visual cortex of cats. In LGN from normal cats, Cat-304 labels neurons in layers A, A1, and C, in interlaminar zones, and in the medial interlaminar nucleus. In LGN from dark-reared cats, the number of antibody-positive neurons is markedly reduced, and the cross-sectional area of the remaining positive neurons is smaller than normal. In cortical area 17 of normally reared cats, Cat 304-positive neurons are densely distributed in 2 bands, in layers IV and V/VI. Labeled neurons are also present in layers II and III. In area 17 of dark-reared cats, the number of antibody-positive neurons is reduced. The reduction in the number of labeled neurons is most pronounced in layers II/III and V/VI. Antibody-positive neurons are smaller in all cortical layers of dark-reared cats. The changes in the expression of Cat-301 immunoreactivity in dark-reared visual cortex and LGN are identical to those of Cat-304. The laminar differences in the effect of dark rearing on Cat-301 and Cat-304 expression in the visual cortex provides support for the suggestion that layer IV of cortical area 17 may be less susceptible to prolongation of plasticity by dark rearing than layers II/III and V/VI. Further, the biochemical and histological studies reported provide evidence that early visual experience regulates protein expression in the cat LGN and visual cortex.
Neuron-specific expression of high-molecular-weight clathrin light chain
Authors: Wong DH , Ignatius MJ , Parosky G , Parham P , Trojanowski JQ , Brodsky FM
Published: March 2018

High-molecular-weight forms of clathrin light chains LCa and LCb contain inserted sequences and are expressed in brain tissue but have not been observed in peripheral tissues. Monoclonal antibodies specific for the high-molecular-weight form of LCb and all forms of LCa were used to analyze their expression in different species and different neuronal cell types. High-molecular-weight light chains were found in bovine, rat, mouse, chicken, and human brain, indicating a conserved pattern of expression. Neuron-specific expression of the high-molecular- weight light chains was suggested by analysis of human brain gray matter and white matter. The former contained a higher proportion of light chains with insertion sequences. Immunohistochemical analysis localized the high-molecular-weight form of LCb to synapses and neuronal perikarya, but not to glial cells. Immunofluorescent labeling of cultured chicken dorsal root ganglia confirmed expression in neurons but not Schwann cells. These results indicate that the high-molecular- weight forms of clathrin light chains are restricted in expression and found in neuronal cells.
Firing properties of hippocampal neurons in a visually symmetrical environment: contributions of multiple sensory cues and mnemonic processes
Authors: Sharp PE , Kubie JL , Muller RU
Published: March 2018

The location-specific firing of hippocampal place cells can easily be brought under the control of experimenter-defined cues. Nevertheless, there is evidence that these firing fields are not determined just by immediate sensory input, but also by earlier states of the nervous system (O'Keefe and Speakman, 1987). Here, we report further on the roles of multiple visual cues and mnemonic processes in determining the firing of place cells. Rats were trained to chase food pellets in a cylinder with homogeneous gray walls and 1 white cue card. After a cell's field was recorded in this “standard” condition, probe sessions were conducted in which a second card was placed 180 degrees away from the first. This configuration created a diametrically symmetrical environment in which pairs of locations 180 degrees apart were identical with respect to views of the wall and cards. If place cells are strongly controlled by these immediately available views, firing in the 2-card configuration should be diametrically symmetrical. Alternatively, because the rat moves freely in the cylinder, information is available that pairs of visually identical places are not truly the same. If some mnemonic process stores and updates information about the rat's paths during the session, it is possible that the firing pattern will be different in 2 such places, especially because the original training was conducted in the 1-card, asymmetrical environment. Thirteen of 18 cells had a single, asymmetric firing pattern after the second card was introduced; the field was the same size and shape as in the 1-card configuration and in the same spatial relation to 1 of the 2 cards. The field position during 2-card sessions could be rotated 180 degrees by starting the rat by one card or the other. In further probe sessions in which the cue cards, entry location, and background cues were, in various combinations, rotated in relation to each other, these cells always showed a single field, similar in size and shape to that in the standard, and in the same relationship as in the standard to as many cues as possible. The remaining 5 cells showed complex changes over repeated 2-card sessions, and 3 of these showed paired fields, 180 degrees apart for at least some of the sessions. In one case, the second field disappeared with repeated exposures to 2 cards; in another, the second field persisted when only 1 card was used. We conclude that place cells are influenced both by the immediate sensory configuration and by internal neural states related to earlier experience in the environment.