bluuebunny/crossref_metadata_2025 huggingface dataset:
Calcium excitability and oscillations in suprachiasmatic nucleus neurons and glia in vitro
Authors: van den Pol AN , Finkbeiner SM , Cornell-Bell AH
Published: March 2018

Converging lines of evidence suggest that the hypothalamic suprachiasmatic nucleus (SCN) is the site of the endogenous biological clock controlling mammalian circadian rhythms. To study the calcium responses of the cellular components that make up the clock, computer- controlled digital video and confocal scanning laser microscopy were used with the Ca2+ indicator dye fluo-3 to examine dispersed SCN cells and SCN explants with repeated sampling over time. Ca2+ plays an important second messenger role in a wide variety of cellular mechanisms from gene regulation to electrical activity and neurotransmitter release, and may play a role in clock function and entrainment. SCN neurons and astrocytes showed an intracellular Ca2+ increase in response to glutamate and 5-HT, two major neurotransmitters in afferents to the SCN. Astrocytes showed a marked heterogeneity in their response to the serial perfusion of different transmitters; some responded to both 5-HT and glutamate, some to neither, and others to only one or the other. Under constant conditions, most neurons showed irregular temporal patterns of Ca2+ transients. Expression of regular neuronal oscillations could be blocked by the inhibitory transmitter GABA. Astrocytes, on the other hand, showed very regular rhythms of cytoplasmic Ca2+ concentrations with periods ranging from 7 to 20 sec. This periodic oscillation could be initiated by in vitro application of glutamate, the putative neurotransmitter conveying visual input to the SCN critical for clock entrainment. Long-distance communication between glial cells, seen as waves of fluorescence moving from cell to cell, probably through gap junctions, was induced by glutamate, 5-HT, and ATP. These waves increased the period length of cellular Ca2+ rises to 45–70 sec. Spontaneously oscillating cells were common in culture medium, serum, or rat cerebrospinal fluid, but rare in HEPES buffer. One source for cytoplasmic Ca2+ increases was an influx of extracellular Ca2+, as seen under depolarizing conditions in about 75% of the astroglia studied. All neurotransmitter-induced Ca2+ fluxes were not dependent on voltage changes, as Ca2+ oscillations could be initiated under both normal and depolarizing conditions. Since neurotransmitters could induce a Ca2+ rise in the absence of extracellular Ca2+, the mechanisms of ultradian oscillations appear to depend on cycles of intracellular Ca2+ fluxes from Ca(2+)-sequestering organelles into the cytoplasm, followed by a subsequent Ca2+ reduction. In the adult SCN, fewer astrocytes are found than neurons, yet astrocytes frequently surround glutamate-immunoreactive axons in synaptic contact with SCN dendrites, isolating neurons from each other while maintaining contact with other astrocytes by gap junctions.(ABSTRACT TRUNCATED AT 400 WORDS)
Tumor necrosis factor modulates Ca2+ currents in cultured sympathetic neurons
Authors: Soliven B , Albert J
Published: March 2018

The effect of recombinant human tumor necrosis factor-alpha (rhTNF) on calcium currents of cultured neurons from neonatal rat superior cervical ganglia (SCG) was studied using whole-cell patch-clamp technique. We found that rhTNF-treated SCG neurons exhibited increased calcium current density without significant alteration in the steady- state parameters of activation and availability. The fraction of the current sensitive to dihydropyridines and omega-conotoxin also remained unchanged. Recovery from slow inactivation of the current, but not recovery from fast inactivation, was prolonged in rhTNF-treated cells when compared to that of control cells. We conclude that immune peptides such as rhTNF can alter cellular functions of sympathetic neurons via modulating ionic conductances. However, these changes observed in calcium currents of SCG neurons cannot account for the effect of rhTNF on norepinephrine secretion observed in a previous study. It is proposed that rhTNF exerts an additional effect at a later event in the exocytotic process.
The period gene encodes a predominantly nuclear protein in adult Drosophila
Authors: Liu X , Zwiebel LJ , Hinton D , Benzer S , Hall JC , Rosbash M
Published: March 2018

The period gene of Drosophila melanogaster (per) is important for the generation and maintenance of biological rhythms. Previous light microscopic observations indicated that per is expressed in a variety of tissues and cell types and suggested that the per protein (PER) may be present in different subcellular compartments. To understand how PER influences circadian rhythms, it is important to define its subcellular location, especially in adult flies where inducible promoter experiments suggested that it is most relevant to circadian locomotor activity rhythms. To this end, we report the results of an immunoelectron microscopic analysis of wild-type flies and per-beta- galactosidase (beta-gal) fusion gene transgenics using a polyclonal anti-PER antibody or an anti-beta-gal antibody, respectively. Most of the PER antigen and the fusion gene product were located within nuclei, suggesting that PER acts in that subcellular compartment to affect circadian rhythms. The results are discussed in terms of per's possible biochemical functions.
Expression of the period clock gene within different cell types in the brain of Drosophila adults and mosaic analysis of these cells' influence on circadian behavioral rhythms
Authors: Ewer J , Frisch B , Hamblen-Coyle MJ , Rosbash M , Hall JC
Published: March 2018

The product of the period (per) gene of Drosophila melanogaster is continuously required for the functioning of the circadian pacemaker of locomotor activity. We have used internally marked mosaics to determine the anatomical locations at which per expression is required for adult rhythmicity, and thus where the fly's circadian pacemaker is likely located in this holometabolous insect. We first provide a detailed description of the distribution and nature of per-expressing cells in the fly's CNS. Using an antibody to the per gene product, or to that of a reporter of per expression, in conjunction with an antibody to the embryonic lethal-abnormal visual system (elav) gene product--which is used as a marker of neuronal identity--we have experimentally confirmed previously proposed assignments of per-expressing cells to the neuronal and glial classes. Thus, we found that per expression and elav immunoreactivity colocalized in large cells located in the lateral cortex of the central brain, as well as in more dorsally located cells in the posterior central brain. In contrast, we found that cells located at the margins of the cortex and the neuropil, and within the neuropil, as well as smaller cortical cells found throughout the brain's cortex, were elav negative, supporting the notion that they are glial in nature. Using internally marked mosaics, we find that the pacemaker is located in brain but is not exclusive to the eyes, the ocelli, or the optic lobes, which is consistent with previous reports obtained in this and other insects of this class. Although the pacemaker may be a paired structure, we show that the functioning of one of them is sufficient for rhythmicity. Finally, we report that glial expression is sufficient for some behavioral rhythmicity to be manifest. However, the rhythmicities of animals for which per expression was confined to glia were weak, suggesting that neuronal per expression as well may be required for normal pacemaker function.
Binding and retrograde transport of leukemia inhibitory factor by the sensory nervous system
Authors: Hendry IA , Murphy M , Hilton DJ , Nicola NA , Bartlett PF
Published: March 2018

Leukemia inhibitory factor (LIF), a peptide growth factor with multiple activities, has recently been shown to support the generation and survival of sensory neurons in cultures of mouse neural crest and dorsal root ganglia (DRG). We have conducted binding experiments with 125I-LIF on cultures of DRG to determine the receptor distribution for LIF on these cells and found that at least 60% of the sensory neurons in the cultures bound 125I-LIF, all of which could be eliminated by the addition of unlabeled LIF. The other cells in the culture, which morphologically appeared to be Schwann cells, did not bind appreciable quantities of 125I-LIF. In order to investigate whether LIF is retrogradely transported to sensory neurons in vivo, 125I-LIF was injected into the footpads and gastrocnemius muscles of newborn and adult mice, following sciatic nerve ligation. Radioactivity accumulated in the distal portion of the sciatic nerve, indicating retrograde transport of LIF. Subsequent experiments on mice with unligated sciatic nerves showed that 125I-LIF is specifically transported into the sensory neurons of the DRG. There was no apparent transport of 125I-LIF into motor neurons in the spinal cord. These experiments demonstrate that LIF can specifically bind to and be transported by sensory neurons and further support the idea that LIF acts as a target-derived neurotrophic factor, analogous to NGF.
The subcellular localization of the G-protein Gi alpha in the basal ganglia reveals its potential role in both signal transduction and vesicle trafficking
Authors: Aronin N , DiFiglia M
Published: March 2018

The G-protein Gi is known to mediate signal transduction in cells by coupling its 41 kDa alpha-subunit to plasma membrane-bound receptors and inhibiting adenylyl cyclase or affecting ion channel function. Although this G-protein has been functionally associated with D2/dopamine and mu-opioid receptors in striatal membranes, its localization to neurons of the neostriatum, a brain region rich in adenylyl cyclase activity, has not been established. Light and electron microscopic study of the basal ganglia was conducted using the immunoperoxidase method and an antiserum directed against the alpha- subunit of Gi. In the neostriatum, immunoreactivity was localized to medium-sized spiny and aspiny neurons and axon terminals that formed symmetric synapses. Some astrocytes and glial processes that encapsulated axospinous complexes were also labeled. Immunoreactive axon terminals were numerous in the globus pallidus and substantia nigra, where they exhibited a dense pattern of distribution characteristic of neostriatal spiny projection neurons. Gi alpha immunoreactivity was distributed to multiple subcellular compartments. In neostriatal somata and dendrites, labeling was present intermittently along plasma membranes, and on rough and smooth endoplasmic reticulum and microtubules. In axon terminals, reaction product appeared on plasma membranes and heavily labeled the membranes of synaptic vesicles. The presence of Gi alpha in axon terminals was confirmed in purified synaptosome preparations. G-proteins consistent with the masses of Go alpha and Gi alpha, respectively, were ADP- ribosylated in the presence of pertussis toxin in striatal synaptosomes. Western blot analysis in purified synaptosome preparations of the neostriatum, globus pallidus, and substantia nigra with the same antiserum used in the immunohistochemistry demonstrated a predominant 41 kDa protein corresponding to the molecular mass of Gi alpha. Immunohistochemical localization of Gi alpha with the immunogold method in a crude striatal synaptosome preparation showed gold particles associated with synaptic vesicles and plasma membranes. Results provide the first direct evidence that Gi alpha is localized to medium-sized neostriatal projection neurons and interneurons, where it is likely to function in membrane-bound signal transduction at the postsynaptic and presynaptic level. The presence of Gi alpha in synaptic vesicle membranes points to another potentially important role for this G-protein in vesicle trafficking, such as that recently shown for smaller-molecular-mass G-proteins.
Pharmacological characterization of GABAB-mediated responses in the CA1 region of the rat hippocampal slice
Authors: Solis JM , Nicoll RA
Published: March 2018

It is generally accepted that the bicuculline-resistant responses to GABA are mediated through the activation of GABAB receptors that mediate a slow IPSP. However, a number of reported observations are difficult to reconcile with this model. Specifically, GABAB antagonists only partially block bicuculline-resistant GABA responses, and both 4- aminopyridine (4-AP) and carbachol have been reported to block responses to the selective GABAB agonist baclofen, but not GABA itself. Thus, it has been argued that baclofen and GABA increase potassium conductance through separate receptor mechanisms. This suggestion is not easily reconcilable with the postulated physiological role of GABAB receptors in mediating the slow IPSP. We have addressed these discrepancies by using the new GABAB antagonists 2-hydroxy-saclofen (2- OH-SAC) and CGP 35348 in the presence of the GABA uptake inhibitor SKF 89976A. The weak antagonism of 2-OH-SAC against the bicuculline- resistant GABA response was improved when the GABA uptake was inhibited with SKF 89976A, allowing for the application of lower GABA concentrations. Under these circumstances, 2-OH-SAC and CGP 35348 strongly antagonized GABA and baclofen responses, but did not have any effect on outward currents evoked by 5-HT. The slow IPSP evoked in the presence of glutamate antagonists was reversibly inhibited by CGP 35348 (IC50 = 14 microM), without affecting the fast IPSP. Carbachol (0.3–20 microM) had no effect on outward currents evoked by either baclofen or GABA. 4-AP (5 microM to 1 mM), despite causing a large increase in cell excitability, did not change baclofen responses. Higher concentrations of 4-AP (5 mM) induced inward current, and reduced both baclofen and GABA outward currents to a similar extent.(ABSTRACT TRUNCATED AT 250 WORDS)
Growth of the rat somatic sensory cortex and its constituent parts during postnatal development
Authors: Riddle D , Richards A , Zsuppan F , Purves D
Published: March 2018

We have compared the size and arrangement of the primary somatic sensory cortex (SI) and its constituent parts in juvenile (1 week old) and mature (10–12 weeks old) rats using succinic dehydrogenase histochemistry and digital image analysis. Our goal was to determine whether some regions of the maturing cortex grow more than others. To this end, we examined (1) the growth of barrels and the surrounding (interbarrel) cortex, (2) the growth of the major somatic representations within SI, and (3) the overall growth of SI compared to the neocortex as a whole. With respect to the first of these issues, SI barrels and barrel-like structures grow more than the intervening cortex. The growth of these elements varies according to region: barrels in the head representation more than double in size, whereas the barrel-like structures in the paw representations increase by only about half this amount. The growth of the major somatic representations within SI is also heterogeneous, the representation of the head enlarging to a greater extent than the representations of the paws. Thus, the ratio of the total area of head representation to the combined paw representation is 15% greater in adults than in juveniles. Finally, the primary somatic sensory cortex grows to a somewhat greater extent than the neocortex as a whole. These observations demonstrate that postnatal cortical growth is not uniform; it varies among cortical barrels and the immediately surrounding (interbarrel) cortex, among the representations of different body parts, and between SI and the rest of the neocortex. As an explanation of this differential growth, we suggest that the neuropil of metabolically (and/or electrically) more active cortical regions grows to a greater extent during maturation than that of less active regions.
Modulation of peptide release from single identified Aplysia neurons in culture
Authors: Whim MD , Lloyd PE
Published: March 2018

Aplysia neurons B1 and B2 contain large amounts of the neuropeptides SCPA and SCPB. When grown in culture, individual B1 and B2 cells incorporate 35S-methionine into the SCPs, which can be released in a stimulus- and calcium-dependent fashion (Lloyd et al., 1986). We now show that single cells can be stimulated in a manner to evoke release of the SCPs that declines only slightly with repeated stimulation. This has allowed us to examine the ability of several physiologically relevant agonists to modulate the stimulus-evoked release of the SCPs. Bath application of either FMRFamide or 5-HT resulted in a significant decrease in the amount of SCPs released by intracellular stimulation of B1 or B2. The action of 5-HT was dose dependent with an inhibition of release of approximately 70% at a concentration of 100 microM. SCPA did not significantly affect release. The bath application of several compounds that are expected to elevate intracellular levels of cAMP were also found to depress release. To investigate the possibility that the agonists inhibited the release of the SCPs via a hyperpolarization of membrane potential (and perhaps a loss of spikes in the neurites), we examined the actions of 5-HT, FMRFamide, and SCPA on several electrophysiological parameters intended to monitor the level of cell excitability. Surprisingly, even though 5-HT depressed the release of the SCPs from both cells, it depolarized and increased the excitability of B1, and hyperpolarized and decreased the excitability of B2. Furthermore, in contrast to the effects seen in culture, 5-HT depolarized both B1 and B2 in situ.(ABSTRACT TRUNCATED AT 250 WORDS)
Basic fibroblast growth factor and local injury protect photoreceptors from light damage in the rat
Authors: Faktorovich EG , Steinberg RH , Yasumura D , Matthes MT , LaVail MM
Published: March 2018

Injection of basic fibroblast growth factor (bFGF) into the eye, intravitreally or subretinally, delays photoreceptor degeneration in inherited retinal dystrophy in the rat, as does local injury to the retina (Faktorovich et al., 1990). To determine whether this heparin- binding peptide or local injury is effective in any other form of photoreceptor degeneration, we examined their protective roles in light damage. Albino rats of the F344 strain were exposed to 1 or 2 weeks of constant fluorescent light (115–200 footcandles), either with or without 1 microliter of bFGF solution (1150 ng/microliters in PBS) injected intravitreally or subretinally 2 d before the start of light exposure. Uninjected and intravitreally PBS-injected controls showed the loss of a majority of photoreceptor nuclei and the loss of most inner and outer segments after 1 week of light exposure, while intravitreal injection of bFGF resulted in significant photoreceptor rescue. The outer nuclear layer in bFGF-injected eyes was two to three times thicker than in controls, and the inner and outer segments showed a much greater degree of integrity. Following recovery in cyclic light for 10 d after 1 week of constant light exposure, bFGF-injected eyes showed much greater regeneration of photoreceptor inner and outer segments than did the controls. bFGF also increased the incidence of presumptive macrophages, located predominantly in the inner retina, but the evidence suggests they are not directly involved in photoreceptor rescue. Subretinal injection of bFGF resulted in photoreceptor rescue throughout most of the superior hemisphere in which the injection was made, with rescue extending into the inferior hemisphere in many of the eyes. Remarkably, the insertion of a dry needle or injection of PBS into the subretinal space also resulted in widespread photoreceptor rescue, extending through 70% or more of the superior hemisphere, and sometimes into the inferior hemispheres. This implicates the release and widespread diffusion of some endogenous survival-promoting factor from the site of injury in the retina. Our findings indicate that the photoreceptor rescue activity of bFGF is not restricted to inherited retinal dystrophy in the rat, and that light damage is an excellent model for studying the cellular site(s), kinetics, and molecular mechanisms of both the normal function of bFGF and its survival- promoting activity. Moreover, the injury-related rescue suggests that survival-promoting factors are readily available to provide a protective role in case of injury to the retina, presumably comparable to those that mediate the “conditioning lesion” effect in other neuronal systems.
Partial dopamine depletion of the prefrontal cortex leads to enhanced mesolimbic dopamine release elicited by repeated exposure to naturally reinforcing stimuli
Authors: Mitchell JB , Gratton A
Published: March 2018

High-speed chronoamperometry was used to monitor the extracellular concentration of dopamine within the nucleus accumbens, a terminal field of the mesolimbic dopamine system, in freely behaving rats exposed daily, on 6 consecutive days, to one of two naturally reinforcing stimuli; a highly palatable food or sex-related olfactory cues. The animals either were intact or had previously received microinjections of 6-hydroxydopamine into prefrontal cortex to lesion dopamine terminals. Food reliably elicited increases in dopamine levels within the nucleus accumbens, and if prefrontal cortical dopamine had been depleted, the response to food increased with repeated testing. Animals exposed to the sexually relevant olfactory stimulus showed progressively enhanced dopamine release with repeated testing, and this enhancement was potentiated by prefrontal cortical dopamine depletion. These results indicate that repeated exposure to naturally reinforcing events can lead to a hyperresponsiveness of the mesolimbic dopamine system upon future activation, and suggest that the dopamine projection to prefrontal cortex exerts an indirect, inhibitory influence on mesolimbic dopamine neurotransmission.
Cell-cell interactions during the migration of an identified commissural growth cone in the embryonic grasshopper
Authors: Myers PZ , Bastiani MJ
Published: March 2018

One of the fascicles of the posterior commissure of the embryonic grasshopper is pioneered by an individually identifiable neuron named Q1. Q1 initially grows along a longitudinal pathway established by another pioneer neuron, MP1, and then crosses to the midline, where it meets and fasciculates with the axon of the contralateral Q1. The Q1 growth cone follows the contralateral Q1 axon to the contralateral longitudinal pathway, where it then fasciculates with axons of the MP1/dMP2 fascicle. In this work, we have identified a small set of early neurons that Q1 could use as guidance cues while negotiating its way along a specific and stereotyped pathway to the midline. Furthermore, we have observed characteristic morphological changes in the Q1 growth cone that could indicate responses to changing adhesivity in the substrates it contacts. We have also quantified the pattern of dye coupling between neurons in this system. Most of the neurons to which Q1 becomes coupled retain a strong, consistent pattern of dye coupling that shows no recognizable variation at times when growth cones are making pathway decisions. However, we have found one clear instance of transient, site-specific dye coupling between the Q1 growth cone and the ipsilateral MP1 soma. The timing and pattern of dye coupling in this system suggest that dye coupling may play a role in synchronizing the initiation of axon outgrowth among a small population of neurons. Although dye coupling may not play a direct role in neuronal pathfinding, it may exert a profound indirect influence on neuronal interactions by regulating the timing of axon outgrowth.
Development of the embryonic neuromuscular synapse of Drosophila melanogaster
Authors: Broadie KS , Bate M
Published: March 2018

We have examined the embryonic development of an identified neuromuscular junction (NMJ) of Drosophila melanogaster using whole- cell patch-clamp and a variety of physiological and morphological techniques. Synaptic current at the embryonic NMJ is carried through a large-conductance (200 pS) L-glutamate receptor. Early synaptic communication is characterized by frequent, brief (< 10 msec) currents carried through few (1–10) receptors and relatively rare, prolonged currents (up to seconds) of similar amplitude. The brief currents have a time course similar to the mature larval excitatory junction currents (EJCs), but the prolonged currents are restricted to early stages of synaptogenesis. The amplitude of EJCs rapidly increases, and the frequency of the prolonged currents decreases, after the initial stages of synaptogenesis. Early prolonged (seconds), nonspiking synaptic potentials are replaced with rapid (> 0.10 sec), spiking synaptic potentials later in development. The early synapse appears tenuous, easily fatiguable, and with inconsistent communication properties. Synaptogenesis can be divided into a sequence of progressive stages. (1) Motor axon filopodia begin neurotransmitter expression and concurrent exploration of the myotube surface. (2) Myotubes uncouple to form single-cell units soon after motor axon contact. (3) A small number of transmitter receptors are homogeneously displayed on the myotube surface immediately following myotube uncoupling. (4) Endogenous transmitter release from pioneering growth cones is detected; nerve stimulation elicits postsynaptic EJC response. (5) Motor axon filopodia and transmitter receptors are localized to the mature synaptic zone; filopodial localization is complete in advance of receptor localization. (6) A functional neuromuscular synapse is formed; endogenous muscular activity begins; nerve stimulation leads to muscle contraction. (7) Morphological presynaptic specializations develop; synapse develops mature morphology. (8) A second motor axon synapses on the myotube at the pre-established synaptic zone. (9) Vigorous neuromuscular activity, characteristic of larval locomotory movements, begins. (10) A second stage of receptor expression begins and continues through the end of embryogenesis. In general, Drosophila neuromuscular synaptogenesis appears similar to neuromuscular synaptogenesis in known vertebrate preparations. We suggest that this system provides a model for synaptogenesis in which investigation can be readily extended to a genetic and molecular level.
Rapid evolution of the visual system: a cellular assay of the retina and dorsal lateral geniculate nucleus of the Spanish wildcat and the domestic cat
Authors: Williams RW , Cavada C , Reinoso-Suarez F
Published: March 2018

The large Spanish wildcat, Felis silvestris tartessia, has retained features of the Pleistocene ancestor of the modern domestic cat, F. catus. To gauge the direction and magnitude of short-term evolutionary change in this lineage, we have compared the retina, the optic nerve, and the dorsal lateral geniculate nucleus (LGN) of Spanish wildcats and their domestic relatives. Retinas of the two species have the same area. However, densities of cone photoreceptors are higher in wildcat-- over 100% higher in the area centralis--whereas rod densities are as high, or higher, in the domestic lineage. Densities of retinal ganglion cells are typically 20–100% higher across the wildcat retina, and the total ganglion cell population is nearly 70% higher than in the domestic cat. These differences are confined to the populations of beta and gamma retinal ganglion cells. In contrast, the population of alpha cells is almost precisely the same in both species. The wildcat LGN is much larger than that of the domestic cat and contains approximately 50% more neurons. However, cell size does not differ appreciably in either the retina or LGN of these species. The differences in total numbers of ganglion cells and LGN neurons correspond neatly to the overall decline in brain size in the domestic lineage and to allometric predictions based on average species differences in body size. We suggest that an increase in the severity of naturally occurring cell death is the most plausible mechanism that can account for the rapid evolutionary reduction in cell populations in this feline lineage.
Time course of extracellular dopamine and behavioral sensitization to cocaine. I. Dopamine axon terminals
Authors: Kalivas PW , Duffy P
Published: March 2018

Repeated administration of cocaine to rodents produces a progressive augmentation in motor activity known as behavioral sensitization. By using microdialysis in the ventral striatum, some studies have found that the development of behavioral sensitization is associated with a similar augmentation in dopamine release, while others have not. It was postulated that differences in doses and withdrawal periods may account for the discrepancies between studies. Rats were behaviorally sensitized to daily peripheral injections using two cocaine treatment regimens (15 mg/kg, i.p. x 5 d or 30 mg/kg, i.p. x 5 d). Using in vivo microdialysis in the ventral striatum, the effect of acute cocaine (15 mg/kg, i.p.) on extracellular dopamine content and motor behavior was examined at various times after discontinuing daily treatments. Twenty- four hours after discontinuing the low dose of daily cocaine, the increase in motor activity and extracellular dopamine elicited by an acute cocaine challenge was significantly elevated. In contrast, following the higher daily treatment regimen there was a significant augmentation in motor activity, but the increase in extracellular dopamine produced by cocaine was significantly reduced. When rats were challenged 10-14 d after discontinuing either dosage regimen of daily cocaine, the increase in both motor activity and extracellular dopamine was augmented. In general, the increase in extracellular dopamine by an acute cocaine challenge increased over time when rats were challenged between 1 and 22 d after discontinuing daily cocaine. Basal concentrations of extracellular dopamine were determined by measuring the in vivo flux of dopamine across the dialysis membrane, and there was no significant difference at 24 hr or 2 weeks following the last daily injection of saline or cocaine. It is concluded that behavioral sensitization to cocaine is generally associated with an augmentation in extracellular dopamine in the ventral striatum, but that high doses of daily cocaine produce apparent tolerance to the augmentation in extracellular dopamine during the early withdrawal period.
Initial tract formation in the mouse brain
Authors: Easter SS , Ross LS , Frankfurter A
Published: March 2018

Mouse embryos from embryonic days 8.5-10.5 (E8.5-E10.5) were fixed and labeled with an antibody to neuron-specific class III beta-tubulin (Moody et al., 1987; Lee et al., 1990a,b) to reveal the first neurons, axons, and tracts in the brain. They were studied in whole-mounts and in light microscopic sections. Some conclusions were checked by labeling tracts in older embryos (E11.5 and E12.5) with the lipophilic dye 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine. The first immunoreactive cells appeared at E8.5, prior to neural tube closure, in the neural plate immediately caudal to the optic vesicle. Cells along the dorsal midline of the mesencephalon issued the first axons, on E9.0; the cells were the mesencephalic nucleus of the trigeminal nerve, and the axons formed its descending tract. The tract reached the level of the trigeminal ganglion by E10.0 but did not enter the ganglion until after E12.5. On E9.5, the number of labeled cells and axons in the alar plate of the presumptive diencephalon and mesencephalon had increased substantially, and many of the rostral ones coursed into the basal plate to enter longitudinal tracts there. Two tracts originated from cells in the basal plate: the tract of the postoptic commissure (from the base of the optic stalk to the level of the cephalic flexure) and the medial longitudinal fasciculus (from the level of the cephalic flexure caudally through the mid and hind-brains). By E10.0, a small mammillotegmental tract paralleled the tract of the postoptic commissure, but immunolabeling was so widespread that discrete tracts were impossible to discern in the presumptive diencephalon and mesencephalon. The more rostral regions remained lightly labeled. In the cerebral vesicle, the presumptive cerebral cortex, the first immunoreactive cells appeared at E10.0; they had multiple processes oriented parallel to the pia, and were identified as the Cajal-Retzius cells. By E10.5, no tracts had formed in the cerebral vesicle. All the tracts formed by E10.0 were superficial, in the subpial lamina. Those that can be identified in the adult brain are very deep structures. These results are compared with previous descriptions of the embryonic brains of amphibians, fish, birds, and other mammals, including humans.
Directed expression of an oncogene to the olfactory neuronal lineage in transgenic mice
Authors: Largent BL , Sosnowski RG , Reed RR
Published: March 2018

The mammalian olfactory system provides a useful model to understand the cellular and molecular mechanisms governing the development of the nervous system. The olfactory neuroepithelium undergoes continual turnover in the adult animal, resulting in a neural tissue containing cells at various stages of neurogenesis. We have generated a transgenic mouse line to examine the effects of directed expression of an oncogene within the olfactory neuronal lineage. A hybrid oncogene was constructed utilizing the regulatory elements for the olfactory marker protein gene to direct the olfactory neuronal-specific expression of simian virus 40 T-antigen, a potent oncogene. The resulting transgenic mouse line expressed T-antigen only in olfactory neurons. Ten-month-old transgenic mice displayed significant hypoplasia of the neuronal elements in the olfactory neuroepithelium. The transgenic mice developed neuroblastomas of olfactory neuronal origin at a low frequency. Distinct clonal lines were derived from the primary culture of the tumor. GAP-43, a growth-associated neuronal marker, was expressed by some of the cell lines. One of the cell lines, 2.2, appeared to be responsive to neurotrophic effects from the presumptive target tissue, the olfactory bulb.
The highly irregular firing of cortical cells is inconsistent with temporal integration of random EPSPs
Authors: Softky WR , Koch C
Published: March 2018

How random is the discharge pattern of cortical neurons? We examined recordings from primary visual cortex (V1; Knierim and Van Essen, 1992) and extrastriate cortex (MT; Newsome et al., 1989a) of awake, behaving macaque monkey and compared them to analytical predictions. For nonbursting cells firing at sustained rates up to 300 Hz, we evaluated two indices of firing variability: the ratio of the variance to the mean for the number of action potentials evoked by a constant stimulus, and the rate-normalized coefficient of variation (Cv) of the interspike interval distribution. Firing in virtually all V1 and MT neurons was nearly consistent with a completely random process (e.g., Cv approximately 1). We tried to model this high variability by small, independent, and random EPSPs converging onto a leaky integrate-and- fire neuron (Knight, 1972). Both this and related models predicted very low firing variability (Cv << 1) for realistic EPSP depolarizations and membrane time constants. We also simulated a biophysically very detailed compartmental model of an anatomically reconstructed and physiologically characterized layer V cat pyramidal cell (Douglas et al., 1991) with passive dendrites and active soma. If independent, excitatory synaptic input fired the model cell at the high rates observed in monkey, the Cv and the variability in the number of spikes were both very low, in agreement with the integrate-and-fire models but in strong disagreement with the majority of our monkey data. The simulated cell only produced highly variable firing when Hodgkin-Huxley- like currents (INa and very strong IDR) were placed on distal dendrites. Now the simulated neuron acted more as a millisecond- resolution detector of dendritic spike coincidences than as a temporal integrator. We argue that neurons that act as temporal integrators over many synaptic inputs must fire very regularly. Only in the presence of either fast and strong dendritic nonlinearities or strong synchronization among individual synaptic events will the degree of predicted variability approach that of real cortical neurons.
Characterization of the glutamate transporter in retinal cones of the tiger salamander
Authors: Eliasof S , Werblin F
Published: March 2018

L-Glutamate elicits an inwardly rectifying current at hyperpolarized potentials in isolated retinal cones of the tiger salamander, as measured under whole-cell patch clamp. Evidence presented in this article supports the notion that cones possess a high-affinity glutamate transporter. This glutamate-elicited current shows no desensitization over a period of several minutes, and has an affinity (Km) of 10 microM. The inward current is mimicked by the amino acids L- aspartate, D-aspartate, L-cysteate, and to a lesser extent D-glutamate. It is neither blocked by the glutamate receptor antagonists kynurenic acid (1 mM), 6-cyano-7-nitroquinoxaline-2,3-dione (100 microM), or 2- amino-5-phosphonovalerate (100 microM), nor elicited by the glutamate receptor agonists (100 microM each) kainate, quisqualate, NMDA, or 2- amino-4-phosphonobutyrate. The glutamate-elicited current was reduced by the glutamate transport blockers dihydrokainate (DHKA), DL-threo- beta-hydroxyaspartate (beta HA), and L-trans-pyrrolidine-2,4- dicarboxylic acid. When glutamate was present on both sides of the membrane, the blockers reduced both uptake and release; the blocker- sensitive current as a function of membrane potential represents the transport current-voltage relation (I-V), and the reversal potential of the I-V represents the transporter equilibrium potential. This potential was a function of the equilibrium potential for glutamate. DHKA and beta HA depolarized horizontal cells in a retinal slice, and abolished their light responses, suggesting that in the absence of glutamate transport, glutamate concentrations in the cleft rise to a level that saturates the postsynaptic receptors. The high capacity of the cone glutamate transporter is well suited for the rapid removal of glutamate from the synaptic cleft required for the signaling of a light onset to postsynaptic cells.
Intracellular calcium levels during the period of delayed excitotoxicity
Authors: Dubinsky JM
Published: March 2018

Intracellular calcium concentrations ([Ca2+]i) among cultured hippocampal neurons were monitored during and in the hours following an excitotoxic glutamate application to determine the time course of changes involved in delayed excitotoxicity. After a 5 min toxic insult, [Ca2+]i increased immediately and remained elevated for an hour. Subsequently, [Ca2+]i declined to normal resting levels and remained so up to 13 hr following insult. Only a few neurons displayed greatly elevated [Ca2+]i at these extended times. Survival experiments in sister cultures indicated that 85% of the neurons died after 24 hr. Therefore, intracellular calcium returned to baseline levels prior to neuronal death. Additionally, during this period when basal calcium levels had recovered, the majority of neurons responded to a second excitatory amino acid application with a second increase in [Ca2+]i.
Evidence for presynaptic inhibition of the olfactory commissural pathway by cholinergic agonists and stimulation of the nucleus of the diagonal band
Authors: Nickell WT , Shipley MT
Published: March 2018

We have investigated the role of the projection from the magnocellular basal forebrain to the olfactory bulb in regulating synaptic transmission in the commissural connection between the two olfactory bulbs. Commissural fibers arise in the contralateral anterior olfactory nucleus, travel in the anterior wing of the anterior commissure (AC), and terminate in the granule cell layer of the olfactory bulb. Electrical stimulation of the commissure causes synaptic activation of granule cells in the granule cell layer of the bulb; the resulting field potential is a reliable indicator of this synaptic current. Microinjections of cholinergic agonists, but not of identical, or larger, quantities of vehicle, reduced the amplitude of this AC field potential. Systemic injection of scopolamine reversed this depression and returned the AC response amplitude to control levels. Irreversible AChE inhibition also reduced the amplitude of the AC response, and muscarinic blockade reversed this effect. Cholinergic terminals in the olfactory bulb arise entirely from the axons of magnocellular basal forebrain neurons in the nucleus of the diagonal band (NDB). Electrical stimulation of NDB, which should release ACh, as well as other transmitters, depressed the AC response. Brief trains of NDB shocks caused a moderate decrease in the AC response that lasted 1–2 sec. Longer shock trains, which caused marked potentiation of the NDB field potential, caused a profound, prolonged (> 20 sec) inhibition of the AC response. Antidromic tests demonstrated that NDB stimulation significantly decreased the excitability of AC terminals. This and other characteristics of the inhibition strongly suggest that the decrease in amplitude of the field potential response to AC stimulation caused by cholinergic agonists and stimulation of NDB is due to presynaptic inhibition leading to reduced release of transmitter from AC terminals. These results suggest that one function of the basal forebrain projection to the olfactory bulb is inhibition of the commissural connection between the two olfactory bulbs. As NDB has been implicated in theta pacemaker input to the olfactory bulb, phasic NDB inhibition of centrifugal afferents to the bulb could function to coordinate signal processing temporally in the olfactory system. Temporal coordination may be particularly important to olfactory circuit function, as this system lacks the point-to-point topographical organization characteristic of other sensory systems.
Characterization and topography of high-affinity 125I-neurotrophin-3 binding to mammalian brain
Authors: Altar CA , Criden MR , Lindsay RM , DiStefano PS
Published: March 2018

The binding of biologically active and 125I-labeled neurotrophin-3 (NT- 3) was studied with both dry film and emulsion autoradiography to compare with NGF binding and discover areas where NT-3 may function in vivo. The equilibrium binding of 300 pM 125I-NT-3 to rat brain sections was reversible and inhibited by unlabeled NT-3 (IC50, 420 pM). 125I-NT- 3 bound in a saturable manner, with high affinity (Kd, 227-269 pM), and with a capacity (Bmax, 26 fmol/mg protein) that exceeded that of NGF by threefold. As with NGF, 125I-NT-3 also bound to a second population of sites with lower affinity (Kd, 2.8 nM) and higher capacity (Bmax, 170 fmol/mg protein). 125I-NT-3 binding was not blocked by NGF, or serum proteins, and brain-derived neurotrophic factor (BDNF) competed for it in a distinctly biphasic manner (IC50 values of 230 pM and 37 nM). Microdensitometry confirmed graphically and by Hill analysis the monophasic displacement of 125I-NT-3 and the biphasic displacement of 125I-NT-3 binding by BDNF in hippocampus, caudate-putamen, neocortex, and olfactory tubercle. In rat or cat, the topography of 125I-NT-3 binding differed from that reported for 125I-NGF binding or for the low- affinity NGF receptor. The highest binding densities were found in neocortical layers 1 and 2, the stratum oriens and radiatum of hippocampus, molecular layer of the dentate gyrus, nucleus of the lateral olfactory tract, entorhinal cortex, anterior olfactory nucleus, anteromedial thalamic nucleus, and amygdala. Moderate densities were found in neocortical layers 4–6, the neostriatum, amygdala, the dorsal root ganglia, and the central gray of spinal cord. Emulsion autoradiography also revealed binding in nerve terminal-rich regions of superficial neocortex and hippocampus but not on neural cell bodies. Binding was absent in many other brain regions, including cholinergic nuclei, and in all peripheral organs studied including liver, kidney, pancreas, heart, and skeletal muscle. 125I-NT-3 binding to sections of human basal ganglia resembled that seen in rat or cat, including high densities in the caudate, putamen, and superficial neocortex. The unique distribution and pharmacology of 125I-NT-3 binding to BDNF- sensitive and -insensitive sites in brain predict predominantly neuronal actions for these factors that are likely to be more widespread and distinct from those of NGF.
The striatal mosaic in primates: striosomes and matrix are differentially enriched in ionotropic glutamate receptor subunits
Authors: Martin LJ , Blackstone CD , Huganir RL , Price DL
Published: March 2018

The cellular and subcellular distributions of the ionotropic alpha- amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)-preferring glutamate receptor (GluR) in monkey striatum were demonstrated immunocytochemically using anti-peptide antibodies to individual subunits of the AMPA receptor. These antibodies specifically recognize GluR1, GluR4, and an epitope common to GluR2 and GluR3 (designated as GluR2/3). On immunoblots, the antibodies detect proteins ranging from 102 to 108 kDa in total homogenates of monkey striatum, hippocampus, and cerebellum. By immunoblotting, GluR1 and GluR2/3 are considerably more abundant than GluR4 in the caudate nucleus. Within the caudate nucleus, putamen, and nucleus accumbens, numerous neuronal perikarya, dendrites, and spines show GluR1 and GluR2/3 immunoreactivities. GluR1- and GluR2/3-enriched striatal neurons have the morphology, transmitter specificity, and distribution of medium-sized (10–20 microns) spiny neurons; large (20–60 microns) round neurons exhibit GluR4 immunoreactivity. GluR1 immunoreactivity, but not GluR2/3 or GluR4 immunoreactivity, is more intense in the ventral striatum (i.e., nucleus accumbens) than in the dorsal striatum, and GluR1 is enriched within dendritic spines in the neuropil of the nucleus accumbens and striosomes in the dorsal striatum. In the caudate nucleus, these patches of dense GluR1 immunoreactivity align with regions low in calcium binding protein immunoreactivity and high in substance P immunoreactivity. Within striosomes, GluR1 immunoreactivity is more abundant than GluR2/3 immunoreactivity; GluR4 immunoreactivity is sparse in striosomes, but the matrix contains large, GluR4-positive cholinergic neurons. This study demonstrates that, within monkey striatum, subunits of ionotropic AMPA GluR have differential distributions within striosomes and matrix. Furthermore, the results suggest that neurons within striatal striosomes and matrix may express different combinations of GluR subunits, thus forming receptors with different channel properties and having consequences that may be relevant physiologically and pathophysiologically. Neurons within these two striatal compartments may have different roles in the synaptic plasticity of motor systems.