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Sample records for aii amacrine cell

  1. The AII amacrine cell connectome: a dense network hub

    PubMed Central

    Marc, Robert E.; Anderson, James R.; Jones, Bryan W.; Sigulinsky, Crystal L.; Lauritzen, James S.

    2014-01-01

    The mammalian AII retinal amacrine cell is a narrow-field, multistratified glycinergic neuron best known for its role in collecting scotopic signals from rod bipolar cells and distributing them to ON and OFF cone pathways in a crossover network via a combination of inhibitory synapses and heterocellular AII::ON cone bipolar cell gap junctions. Long considered a simple cell, a full connectomics analysis shows that AII cells possess the most complex interaction repertoire of any known vertebrate neuron, contacting at least 28 different cell classes, including every class of retinal bipolar cell. Beyond its basic role in distributing rod signals to cone pathways, the AII cell may also mediate narrow-field feedback and feedforward inhibition for the photopic OFF channel, photopic ON-OFF inhibitory crossover signaling, and serves as a nexus for a collection of inhibitory networks arising from cone pathways that likely negotiate fast switching between cone and rod vision. Further analysis of the complete synaptic counts for five AII cells shows that (1) synaptic sampling is normalized for anatomic target encounter rates; (2) qualitative targeting is specific and apparently errorless; and (3) that AII cells strongly differentiate partner cohorts by synaptic and/or coupling weights. The AII network is a dense hub connecting all primary retinal excitatory channels via precisely weighted drive and specific polarities. Homologs of AII amacrine cells have yet to be identified in non-mammalians, but we propose that such homologs should be narrow-field glycinergic amacrine cells driving photopic ON-OFF crossover via heterocellular coupling with ON cone bipolar cells and glycinergic synapses on OFF cone bipolar cells. The specific evolutionary event creating the mammalian AII scotopic-photopic hub would then simply be the emergence of large numbers of pure rod bipolar cells. PMID:25237297

  2. Intrinsic bursting of AII amacrine cells underlies oscillations in the rd1 mouse retina.

    PubMed

    Choi, Hannah; Zhang, Lei; Cembrowski, Mark S; Sabottke, Carl F; Markowitz, Alexander L; Butts, Daniel A; Kath, William L; Singer, Joshua H; Riecke, Hermann

    2014-09-15

    In many forms of retinal degeneration, photoreceptors die but inner retinal circuits remain intact. In the rd1 mouse, an established model for blinding retinal diseases, spontaneous activity in the coupled network of AII amacrine and ON cone bipolar cells leads to rhythmic bursting of ganglion cells. Since such activity could impair retinal and/or cortical responses to restored photoreceptor function, understanding its nature is important for developing treatments of retinal pathologies. Here we analyzed a compartmental model of the wild-type mouse AII amacrine cell to predict that the cell's intrinsic membrane properties, specifically, interacting fast Na and slow, M-type K conductances, would allow its membrane potential to oscillate when light-evoked excitatory synaptic inputs were withdrawn following photoreceptor degeneration. We tested and confirmed this hypothesis experimentally by recording from AIIs in a slice preparation of rd1 retina. Additionally, recordings from ganglion cells in a whole mount preparation of rd1 retina demonstrated that activity in AIIs was propagated unchanged to elicit bursts of action potentials in ganglion cells. We conclude that oscillations are not an emergent property of a degenerated retinal network. Rather, they arise largely from the intrinsic properties of a single retinal interneuron, the AII amacrine cell. PMID:25008417

  3. Intrinsic bursting of AII amacrine cells underlies oscillations in the rd1 mouse retina

    PubMed Central

    Choi, Hannah; Zhang, Lei; Cembrowski, Mark S.; Sabottke, Carl F.; Markowitz, Alexander L.; Butts, Daniel A.; Kath, William L.; Singer, Joshua H.

    2014-01-01

    In many forms of retinal degeneration, photoreceptors die but inner retinal circuits remain intact. In the rd1 mouse, an established model for blinding retinal diseases, spontaneous activity in the coupled network of AII amacrine and ON cone bipolar cells leads to rhythmic bursting of ganglion cells. Since such activity could impair retinal and/or cortical responses to restored photoreceptor function, understanding its nature is important for developing treatments of retinal pathologies. Here we analyzed a compartmental model of the wild-type mouse AII amacrine cell to predict that the cell's intrinsic membrane properties, specifically, interacting fast Na and slow, M-type K conductances, would allow its membrane potential to oscillate when light-evoked excitatory synaptic inputs were withdrawn following photoreceptor degeneration. We tested and confirmed this hypothesis experimentally by recording from AIIs in a slice preparation of rd1 retina. Additionally, recordings from ganglion cells in a whole mount preparation of rd1 retina demonstrated that activity in AIIs was propagated unchanged to elicit bursts of action potentials in ganglion cells. We conclude that oscillations are not an emergent property of a degenerated retinal network. Rather, they arise largely from the intrinsic properties of a single retinal interneuron, the AII amacrine cell. PMID:25008417

  4. Elucidating the Role of AII Amacrine Cells in Glutamatergic Retinal Waves

    PubMed Central

    Firl, Alana; Ke, Jiang-Bin; Zhang, Lei; Fuerst, Peter G.; Singer, Joshua H.

    2015-01-01

    Spontaneous retinal activity mediated by glutamatergic neurotransmission—so-called “Stage 3” retinal waves—drives anti-correlated spiking in ON and OFF RGCs during the second week of postnatal development of the mouse. In the mature retina, the activity of a retinal interneuron called the AII amacrine cell is responsible for anti-correlated spiking in ON and OFF α-RGCs. In mature AIIs, membrane hyperpolarization elicits bursting behavior. Here, we postulated that bursting in AIIs underlies the initiation of glutamatergic retinal waves. We tested this hypothesis by using two-photon calcium imaging of spontaneous activity in populations of retinal neurons and by making whole-cell recordings from individual AIIs and α-RGCs in in vitro preparations of mouse retina. We found that AIIs participated in retinal waves, and that their activity was correlated with that of ON α-RGCs and anti-correlated with that of OFF α-RGCs. Though immature AIIs lacked the complement of membrane conductances necessary to generate bursting, pharmacological activation of the M-current, a conductance that modulates bursting in mature AIIs, blocked retinal wave generation. Interestingly, blockade of the pacemaker conductance Ih, a conductance absent in AIIs but present in both ON and OFF cone bipolar cells, caused a dramatic loss of spatial coherence of spontaneous activity. We conclude that during glutamatergic waves, AIIs act to coordinate and propagate activity generated by BCs rather than to initiate spontaneous activity. PMID:25632142

  5. Increased phosphorylation of Cx36 gap junctions in the AII amacrine cells of RD retina

    PubMed Central

    Ivanova, Elena; Yee, Christopher W.; Sagdullaev, Botir T.

    2015-01-01

    Retinal degeneration (RD) encompasses a family of diseases that lead to photoreceptor death and visual impairment. Visual decline due to photoreceptor cell loss is further compromised by emerging spontaneous hyperactivity in inner retinal cells. This aberrant activity acts as a barrier to signals from the remaining photoreceptors, hindering therapeutic strategies to restore light sensitivity in RD. Gap junctions, particularly those expressed in AII amacrine cells, have been shown to be integral to the generation of aberrant activity. It is unclear whether gap junction expression and coupling are altered in RD. To test this, we evaluated the expression and phosphorylation state of connexin36 (Cx36), the gap junction subunit predominantly expressed in AII amacrine cells, in two mouse models of RD, rd10 (slow degeneration) and rd1 (fast degeneration). Using Ser293-P antibody, which recognizes a phosphorylated form of connexin36, we found that phosphorylation of connexin36 in both slow and fast RD models was significantly greater than in wildtype controls. This elevated phosphorylation may underlie the increased gap junction coupling of AII amacrine cells exhibited by RD retina. PMID:26483638

  6. Diabetic hyperglycemia reduces Ca2+ permeability of extrasynaptic AMPA receptors in AII amacrine cells.

    PubMed

    Castilho, Áurea; Madsen, Eirik; Ambrósio, António F; Veruki, Margaret L; Hartveit, Espen

    2015-09-01

    There is increasing evidence that diabetic retinopathy is a primary neuropathological disorder that precedes the microvascular pathology associated with later stages of the disease. Recently, we found evidence for altered functional properties of synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors in A17, but not AII, amacrine cells in the mammalian retina, and the observed changes were consistent with an upregulation of the GluA2 subunit, a key determinant of functional properties of AMPA receptors, including Ca(2+) permeability and current-voltage (I-V) rectification properties. Here, we have investigated functional changes of extrasynaptic AMPA receptors in AII amacrine cells evoked by diabetes. With patch-clamp recording of nucleated patches from retinal slices, we measured Ca(2+) permeability and I-V rectification in rats with ∼3 wk of streptozotocin-induced diabetes and age-matched, noninjected controls. Under bi-ionic conditions (extracellular Ca(2+) concentration = 30 mM, intracellular Cs(+) concentration = 171 mM), the reversal potential (Erev) of AMPA-evoked currents indicated a significant reduction of Ca(2+) permeability in diabetic animals [Erev = -17.7 mV, relative permeability of Ca(2+) compared with Cs(+) (PCa/PCs) = 1.39] compared with normal animals (Erev = -7.7 mV, PCa/PCs = 2.35). Insulin treatment prevented the reduction of Ca(2+) permeability. I-V rectification was examined by calculating a rectification index (RI) as the ratio of the AMPA-evoked conductance at +40 and -60 mV. The degree of inward rectification in patches from diabetic animals (RI = 0.48) was significantly reduced compared with that in normal animals (RI = 0.30). These results suggest that diabetes evokes a change in the functional properties of extrasynaptic AMPA receptors of AII amacrine cells. These changes could be representative for extrasynaptic AMPA receptors elsewhere in AII amacrine cells and suggest that synaptic and extrasynaptic AMPA

  7. Relative contributions of rod and cone bipolar cell inputs to AII amacrine cell light responses in the mouse retina

    PubMed Central

    Pang, Ji-Jie; Abd-El-Barr, Muhammad M; Gao, Fan; Bramblett, Debra E; Paul, David L; Wu, Samuel M

    2007-01-01

    AII amacrine cells (AIIACs) are crucial relay stations for rod-mediated signals in the mammalian retina and they receive synaptic inputs from depolarizing and hyperpolarizing bipolar cells (DBCs and HBCs) as well as from other amacrine cells. Using whole-cell voltage-clamp technique in conjunction with pharmacological tools, we found that the light-evoked current response of AIIACs in the mouse retina is almost completely mediated by two DBC synaptic inputs: a 6,7-dinitro-quinoxaline-2,3-dione (DNQX)-resistant component mediated by cone DBCs (DBCCs) through an electrical synapse, and a DNQX-sensitive component mediated by rod DBCs (DBCRs). This scheme is supported by AIIAC current responses recorded from two knockout mice. The dynamic range of the AIIAC light response in the Bhlhb4−/− mouse (which lacks DBCRs) resembles that of the DNQX-resistant component, and that of the connexin36 (Cx36)−/− mouse resembles the DNQX-sensitive component. By comparing the light responses of the DBCCs with the DNQX-resistant AIIAC component, and light responses of the DBCRs with the DNQX-sensitive AIIAC component, we obtained the input–output relations of the DBCC→AIIAC electrical synapse and the DBCR→AIIAC chemical synapse. Similar to other glutamatergic chemical synapses in the retina, the DBCR→AIIAC synapse is non-linear. Its highest voltage gain (approximately 5) is found near the dark membrane potential, and it saturates for presynaptic signals larger than 5.5 mV. The DBCC→AIIAC electrical synapse is approximately linear (voltage gain of 0.92), consistent with the linear junctional conductance found in retinal electrical synapses. Moreover, relative DBCR and DBCC contributions to the AIIAC response at various light intensity levels are determined. PMID:17255172

  8. Connexin30.2: In Vitro Interaction with Connexin36 in HeLa Cells and Expression in AII Amacrine Cells and Intrinsically Photosensitive Ganglion Cells in the Mouse Retina

    PubMed Central

    Meyer, Arndt; Tetenborg, Stephan; Greb, Helena; Segelken, Jasmin; Dorgau, Birthe; Weiler, Reto; Hormuzdi, Sheriar G.; Janssen-Bienhold, Ulrike; Dedek, Karin

    2016-01-01

    Electrical coupling via gap junctions is an abundant phenomenon in the mammalian retina and occurs in all major cell types. Gap junction channels are assembled from different connexin subunits, and the connexin composition of the channel confers specific properties to the electrical synapse. In the mouse retina, gap junctions were demonstrated between intrinsically photosensitive ganglion cells and displaced amacrine cells but the underlying connexin remained undetermined. In the primary rod pathway, gap junctions play a crucial role, coupling AII amacrine cells among each other and to ON cone bipolar cells. Although it has long been known that connexin36 and connexin45 are necessary for the proper functioning of this most sensitive rod pathway, differences between homocellular AII/AII gap junctions and AII/ON bipolar cell gap junctions suggested the presence of an additional connexin in AII amacrine cells. Here, we used a connexin30.2-lacZ mouse line to study the expression of connexin30.2 in the retina. We show that connexin30.2 is expressed in intrinsically photosensitive ganglion cells and AII amacrine cells. Moreover, we tested whether connexin30.2 and connexin36—both expressed in AII amacrine cells—are able to interact with each other and are deposited in the same gap junctional plaques. Using newly generated anti-connexin30.2 antibodies, we show in HeLa cells that both connexins are indeed able to interact and may form heteromeric channels: both connexins were co-immunoprecipitated from transiently transfected HeLa cells and connexin30.2 gap junction plaques became significantly larger when co-expressed with connexin36. These data suggest that connexin36 is able to form heteromeric gap junctions with another connexin. We hypothesize that co-expression of connexin30.2 and connexin36 may endow AII amacrine cells with the means to differentially regulate its electrical coupling to different synaptic partners. PMID:27303262

  9. Functional NMDA receptors are expressed by both AII and A17 amacrine cells in the rod pathway of the mammalian retina.

    PubMed

    Zhou, Yifan; Tencerová, Barbora; Hartveit, Espen; Veruki, Margaret L

    2016-01-01

    At many glutamatergic synapses, non-N-methyl-d-aspartate (NMDA) and NMDA receptors are coexpressed postsynaptically. In the mammalian retina, glutamatergic rod bipolar cells are presynaptic to two rod amacrine cells (AII and A17) that constitute dyad postsynaptic partners opposite each presynaptic active zone. Whereas there is strong evidence for expression of non-NMDA receptors by both AII and A17 amacrines, the expression of NMDA receptors by the pre- and postsynaptic neurons in this microcircuit has not been resolved. In this study, using patch-clamp recording from visually identified cells in rat retinal slices, we investigated the expression and functional properties of NMDA receptors in these cells with a combination of pharmacological and biophysical methods. Pressure application of NMDA did not evoke a response in rod bipolar cells, but for both AII and A17 amacrines, NMDA evoked responses that were blocked by a competitive antagonist (CPP) applied extracellularly and an open channel blocker (MK-801) applied intracellularly. NMDA-evoked responses also displayed strong Mg(2+)-dependent voltage block and were independent of gap junction coupling. With low-frequency application (60-s intervals), NMDA-evoked responses remained stable for up to 50 min, but with higher-frequency stimulation (10- to 20-s intervals), NMDA responses were strongly and reversibly suppressed. We observed strong potentiation when NMDA was applied in nominally Ca(2+)-free extracellular solution, potentially reflecting Ca(2+)-dependent NMDA receptor inactivation. These results indicate that expression of functional (i.e., conductance-increasing) NMDA receptors is common to both AII and A17 amacrine cells and suggest that these receptors could play an important role for synaptic signaling, integration, or plasticity in the rod pathway. PMID:26561610

  10. Tyrosine hydroxylase positive perisomatic rings are formed around various amacrine cell types in the mammalian retina.

    PubMed

    Debertin, Gábor; Kántor, Orsolya; Kovács-Öller, Tamás; Balogh, Lajos; Szabó-Meleg, Edina; Orbán, József; Nyitrai, Miklós; Völgyi, Béla

    2015-08-01

    Dopaminergic neurons of the central nervous system are mainly found in nuclei of the midbrain and the hypothalamus that provide subcortical and cortical targets with a rich and divergent innervation. Disturbance of signaling through this system underlies a variety of deteriorating conditions such as Parkinson's disease and schizophrenia. Although retinal dopaminergic signaling is largely independent of the above circuitry, malfunction of the retinal dopaminergic system has been associated with anomalies in visual adaptation and a number of retinal disorders. Dopamine (DA) is released mainly in a paracrine manner by a population of tyrosine hydroxylase expressing (TH(+) ) amacrine cells (AC) of the mammalian retina; thus DA reaches virtually all retinal cell types by diffusion. Despite this paracrine release, however, the so called AII ACs have been considered as the main targets of DA signaling owing to a characteristic and robust ring-like TH(+) innervation to the soma/dendritic-stalk area of AII cells. This apparent selectivity of TH(+) innervation seems to contradict the divergent DAergic signaling scheme of other brain loci. In this study, however, we show evidence for intimate proximity between TH(+) rings and somata of neurochemically identified non-AII cells. We also show that this phenomenon is not species specific, as we observe it in popular mammalian animal models including the rabbit, the rat, and the mouse. Finally, our dataset suggests the existence of further, yet unidentified post-synaptic targets of TH(+) dendritic rings. Therefore, we hypothesize that TH(+) ring-like structures target the majority of ACs non-selectively and that such contacts are wide-spread among mammals. Therefore, this new view of inner retinal TH(+) innervation resembles the divergent DAergic innervation of other brain areas through the mesolimbic, mesocortical, and mesostriatal signaling streams. AII amacrine cells have been considered as the main targets of dopamine

  11. Synaptic connections of amacrine cells containing vesicular glutamate transporter 3 in baboon retinas

    PubMed Central

    MARSHAK, DAVID W.; CHUANG, ALICE Z.; DOLINO, DREW M.; JACOBY, ROY A.; LIU, WEILEY S.; LONG, YE; SHERMAN, MICHAEL B.; SUH, JAE M.; VILA, ALEJANDRO; MILLS, STEPHEN L.

    2016-01-01

    The goals of these experiments were to describe the morphology and synaptic connections of amacrine cells in the baboon retina that contain immunoreactive vesicular glutamate transporter 3 (vGluT3). These amacrine cells had the morphology characteristic of knotty bistratified type 1 cells, and their dendrites formed two plexuses on either side of the center of the inner plexiform layer. The primary dendrites received large synapses from amacrine cells, and the higher-order dendrites were both pre- and postsynaptic to other amacrine cells. Based on light microscopic immunolabeling results, these include AII cells and starburst cells, but not the polyaxonal amacrine cells tracer-coupled to ON parasol ganglion cells. The vGluT3 cells received input from ON bipolar cells at ribbon synapses and made synapses onto OFF bipolar cells, including the diffuse DB3a type. Many synapses from vGluT3 cells onto retinal ganglion cells were observed in both plexuses. At synapses where vGluT3 cells were presynaptic, two types of postsynaptic densities were observed; there were relatively thin ones characteristic of inhibitory synapses and relatively thick ones characteristic of excitatory synapses. In the light microscopic experiments with Neurobiotin-injected ganglion cells, vGluT3 cells made contacts with midget and parasol ganglion cells, including both ON and OFF types. Puncta containing immunoreactive gephyrin, an inhibitory synapse marker, were found at appositions between vGluT3 cells and each of the four types of labeled ganglion cells. The vGluT3 cells did not have detectable levels of immunoreactive γ-aminobutyric acid (GABA) or immunoreactive glycine transporter 1. Thus, the vGluT3 cells would be expected to have ON responses to light and make synapses onto neurons in both the ON and the OFF pathways. Taken with previous results, these findings suggest that vGluT3 cells release glycine at some of their output synapses and glutamate at others. PMID:26241195

  12. Identification of AⅡ amacrine, displaced amacrine, and bistratified ganglion cell types in human retina with antibodies against calretinin.

    PubMed

    Lee, Sammy C S; Weltzien, Felix; Madigan, Michele C; Martin, Paul R; Grünert, Ulrike

    2016-01-01

    Antibodies against calretinin are markers for one type of rod pathway interneuron (AⅡ amacrine cell) in the retina of some but not all mammalian species. The AⅡ cells play a crucial role in night-time (scotopic) vision and have been proposed as a target for optogenetic restoration of vision in retinal disease. In the present study we aimed to characterize the AⅡ cells in human retina. Postmortem human donor eyes were obtained with ethical approval and processed for calretinin immunofluorescence. Calretinin-positive somas in the inner nuclear and the ganglion cell layer were filled with the lipophilic dye DiI. The large majority (over 80%) of calretinin-immunoreactive cells is located in the inner nuclear layer, is immunopositive for glycine transporter 1, and shows the typical morphology of AⅡ amacrine cells. In addition, a small proportion of calretinin-positive cells in the inner nuclear layer and in the ganglion cell layer is glutamic acid decarboxylase-positive and shows the morphology of widefield amacrine cells (stellate, semilunar, and thorny amacrine cells). About half of the calretinin cells in the ganglion cell layer are bistratified ganglion cells resembling the small bistratified (presumed blue-ON/yellow-OFF) and the G17 ganglion cell previously described in primates. We conclude that in human retina, antibodies against calretinin can be used to identify AⅡ amacrine cells in the inner nuclear layer as well as widefield amacrine and small bistratified ganglion cells in the ganglion cell layer. PMID:26053777

  13. A polyaxonal amacrine cell population in the primate retina.

    PubMed

    Greschner, Martin; Field, Greg D; Li, Peter H; Schiff, Max L; Gauthier, Jeffrey L; Ahn, Daniel; Sher, Alexander; Litke, Alan M; Chichilnisky, E J

    2014-03-01

    Amacrine cells are the most diverse and least understood cell class in the retina. Polyaxonal amacrine cells (PACs) are a unique subset identified by multiple long axonal processes. To explore their functional properties, populations of PACs were identified by their distinctive radially propagating spikes in large-scale high-density multielectrode recordings of isolated macaque retina. One group of PACs exhibited stereotyped functional properties and receptive field mosaic organization similar to that of parasol ganglion cells. These PACs had receptive fields coincident with their dendritic fields, but much larger axonal fields, and slow radial spike propagation. They also exhibited ON-OFF light responses, transient response kinetics, sparse and coordinated firing during image transitions, receptive fields with antagonistic surrounds and fine spatial structure, nonlinear spatial summation, and strong homotypic neighbor electrical coupling. These findings reveal the functional organization and collective visual signaling by a distinctive, high-density amacrine cell population. PMID:24599459

  14. Electroanatomy of a unique amacrine cell in the rabbit retina.

    PubMed Central

    Miller, R F; Bloomfield, S A

    1983-01-01

    Intracellular electrophysiological recordings were obtained from a specialized class of "starburst" amacrine cells by using an isolated superperfused retina-eyecup preparation of the rabbit. These cells were injected intracellularly with horseradish peroxidase and identified with light microscopy. A computer-controlled image-processing system was used to map and display the three-dimensional dendritic organization and provide information on length and sublaminar distribution of dendritic processes. Starburst amacrines show an unusual dendritic architecture that includes thin intermediate dendritic segments. Analysis with steady-state cable equations suggests that these thin segments may provide electrical isolation of distal processes, raising the possibility that a single dendrite, which lies beyond the thin segment, may constitute a functional subunit of the cell. Images PMID:6574470

  15. Effects of Histamine on Light Responses of Amacrine Cells in Tiger Salamander Retina

    PubMed Central

    Yu, Yongchun; Satoh, Hiromasa; Vila, Alejandro; Wu, Samuel M.; Marshak, David W.

    2011-01-01

    Using immunofluorescence, we showed that histamine receptor 1 is expressed by horizontal cell axons and a subset of amacrine cells in the tiger salamander retina. The effects of histamine on light responses of amacrine cells were studied in slice preparations. Histamine modulated the light responses of many salamander amacrine cells, depending upon the morphological type. The most pronounced effects of histamine were decreases in the light responses of broadly stratified amacrine cells, particularly those having medium-sized dendritic field diameters. To determine whether the effects of histamine were direct, Co++ was substituted for Ca++ in the extracellular medium to block synaptic transmission. Histamine still affected broadly stratified amacrine cells, but not narrowly stratified amacrine cells under these conditions. Taken together, these findings suggest that inhibitory interactions between strata of the IPL and within the classical receptive fields of the ganglion cells would be particularly sensitive to histamine released from retinopetal axons. PMID:20878231

  16. An amacrine cell circuit for signaling steady illumination in the retina

    PubMed Central

    Jacoby, Jason; Zhu, Yongling; DeVries, Steven H.; Schwartz, Gregory

    2015-01-01

    Summary Decades of research have focused on the circuit connectivity between retinal neurons, yet only a handful of amacrine cells have been described functionally and placed in the context of a specific retinal circuit. Here we identify a circuit where inhibition from a specific amacrine cell plays a vital role in shaping the feature selectivity of a postsynaptic ganglion cell. We record from transgenically labeled CRH-1 amacrine cells and identify a postsynaptic target for CRH-1 amacrine cell inhibition in an atypical retinal ganglion cell (RGC) in mouse retina, the Suppressed-by-Contrast (SbC) RGC. Unlike other RGC types, SbC RGCs spike tonically in steady illumination and are suppressed by both increases and decreases in illumination. Inhibition from GABAergic CRH-1 amacrine cells shapes this unique contrast response profile to positive contrast. We show the existence and impact of this circuit with both paired recordings and cell-type specific ablation. PMID:26711334

  17. Muscarinic signaling influences the patterning and phenotype of cholinergic amacrine cells in the developing chick retina

    PubMed Central

    Stanke, Jennifer J; Lehman, Bret; Fischer, Andy J

    2008-01-01

    Background Many studies in the vertebrate retina have characterized the differentiation of amacrine cells as a homogenous class of neurons, but little is known about the genes and factors that regulate the development of distinct types of amacrine cells. Accordingly, the purpose of this study was to characterize the development of the cholinergic amacrine cells and identify factors that influence their development. Cholinergic amacrine cells in the embryonic chick retina were identified by using antibodies to choline acetyltransferase (ChAT). Results We found that as ChAT-immunoreactive cells differentiate they expressed the homeodomain transcription factors Pax6 and Islet1, and the cell-cycle inhibitor p27kip1. As differentiation proceeds, type-II cholinergic cells, displaced to the ganglion cell layer, transiently expressed high levels of cellular retinoic acid binding protein (CRABP) and neurofilament, while type-I cells in the inner nuclear layer did not. Although there is a 1:1 ratio of type-I to type-II cells in vivo, in dissociated cell cultures the type-I cells (ChAT-positive and CRABP-negative) out-numbered the type-II cells (ChAT and CRABP-positive cells) by 2:1. The relative abundance of type-I to type-II cells was not influenced by Sonic Hedgehog (Shh), but was affected by compounds that act at muscarinic acetylcholine receptors. In addition, the abundance and mosaic patterning of type-II cholinergic amacrine cells is disrupted by interfering with muscarinic signaling. Conclusion We conclude that: (1) during development type-I and type-II cholinergic amacrine cells are not homotypic, (2) the phenotypic differences between these subtypes of cells is controlled by the local microenvironment, and (3) appropriate levels of muscarinic signaling between the cholinergic amacrine cells are required for proper mosaic patterning. PMID:18254959

  18. Lgr5+ amacrine cells possess regenerative potential in the retina of adult mice

    PubMed Central

    Chen, Mengfei; Tian, Shenghe; Glasgow, Nathan G; Gibson, Gregory; Yang, Xiaoling; Shiber, Christen E; Funderburgh, James; Watkins, Simon; Johnson, Jon W; Schuman, Joel S; Liu, Hongjun

    2015-01-01

    Current knowledge indicates that the adult mammalian retina lacks regenerative capacity. Here, we show that the adult stem cell marker, leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5), is expressed in the retina of adult mice. Lgr5+ cells are generated at late stages of retinal development and exhibit properties of differentiated amacrine interneurons (amacrine cells). Nevertheless, Lgr5+ amacrine cells contribute to regeneration of new retinal cells in the adult stage. The generation of new retinal cells, including retinal neurons and Müller glia from Lgr5+ amacrine cells, begins in early adulthood and continues as the animal ages. Together, these findings suggest that the mammalian retina is not devoid of regeneration as previously thought. It is rather dynamic, and Lgr5+ amacrine cells function as an endogenous regenerative source. The identification of such cells in the mammalian retina may provide new insights into neuronal regeneration and point to therapeutic opportunities for age-related retinal degenerative diseases. PMID:25990970

  19. Sox2 Regulates Cholinergic Amacrine Cell Positioning and Dendritic Stratification in the Retina

    PubMed Central

    Whitney, Irene E.; Keeley, Patrick W.; St. John, Ace J.; Kautzman, Amanda G.; Kay, Jeremy N.

    2014-01-01

    The retina contains two populations of cholinergic amacrine cells, one positioned in the ganglion cell layer (GCL) and the other in the inner nuclear layer (INL), that together comprise ∼1/2 of a percent of all retinal neurons. The present study examined the genetic control of cholinergic amacrine cell number and distribution between these two layers. The total number of cholinergic amacrine cells was quantified in the C57BL/6J and A/J inbred mouse strains, and in 25 recombinant inbred strains derived from them, and variations in their number and ratio (GCL/INL) across these strains were mapped to genomic loci. The total cholinergic amacrine cell number was found to vary across the strains, from 27,000 to 40,000 cells, despite little variation within individual strains. The number of cells was always lower within the GCL relative to the INL, and the sizes of the two populations were strongly correlated, yet there was variation in their ratio between the strains. Approximately 1/3 of that variation in cell ratio was mapped to a locus on chromosome 3, where Sex determining region Y box 2 (Sox2) was identified as a candidate gene due to the presence of a 6-nucleotide insertion in the protein-coding sequence in C57BL/6J and because of robust and selective expression in cholinergic amacrine cells. Conditionally deleting Sox2 from the population of nascent cholinergic amacrine cells perturbed the normal ratio of cells situated in the GCL versus the INL and induced a bistratifying morphology, with dendrites distributed to both ON and OFF strata within the inner plexiform layer. PMID:25057212

  20. The area centralis in the chicken retina contains efferent target amacrine cells

    PubMed Central

    Weller, Cynthia; Lindstrom, Sarah H.; De Grip, Willem J.; Wilson, Martin

    2012-01-01

    The retinas of birds receive a substantial efferent, or centrifugal, input from a midbrain nucleus. The function of this input is presently unclear but previous work in the pigeon has shown that efferent input is excluded from the area centralis, suggesting that the functions of the area centralis and the efferent system are incompatible. Using an antibody specific to rods, we have identified the area centralis in another species, the chicken, and mapped the distribution of the unique amacrine cells that are the postsynaptic partners of efferent fibers. Efferent target amacrine cells are found within the chicken area centralis and their density is continuous across the border of the area centralis. In contrast to the pigeon retina then, we conclude that the chicken area centralis receives efferent input. We suggest that the difference between the 2 species is attributable to the presence of a fovea within the area centralis of the pigeon and its absence from that of the chicken. PMID:19296862

  1. Nerve Growth Factor Receptor TrkA Is Expressed by Horizontal and Amacrine Cells During Chicken Retinal Development

    PubMed Central

    KARLSSON, MIRIAM; CLARY, DOUGLAS O.; LEFCORT, FRANCES B.; REICHARDT, LOUIS F.; KARTEN, HARVEY J.; HALLBÖÖK, FINN

    2009-01-01

    Nerve growth factor is known to stimulate neurite outgrowth and support neuronal survival during embryonic development. We have studied the expression of the nerve growth factor receptor, TrkA, at both mRNA and protein levels during the course of chicken retinal development. Furthermore, we have compared the expression of trkA mRNA with that of the 75-kD low-affinity neurotrophin receptor (p75NTR). RNase protection assay identified peak-levels of trkA mRNA in the late embryonic retina. Using in situ hybridization and immunohistochemistry, we found cells expressing TrkA in both the internal and the external part of the inner nuclear layer, corresponding to amacrine and horizontal cells, respectively. The TrkA-expressing amacrine cell has a unistratified dendritic arborization in the second sublamina of the inner plexiform layer, and may represent the stellate amacrine cell described by Cajal. The horizontal cells, possessing arciform dendrite processes in the outer plexiform layer, showed strong TrkA immunoreactivity in both dendrites and cell bodies. During the course of retinal development, the TrkA-expressing amacrine cells decreased in number, whereas the TrkA-expressing horizontal cells persisted. Because nerve growth factor was expressed where the horizontal cells, but not where the amacrine cells were located, these findings raise the question of whether nerve growth factor could locally support the survival of TrkA-expressing interneurons during retinal development. PMID:9779944

  2. Postsynaptic Plasticity Triggered by Ca²⁺-Permeable AMPA Receptor Activation in Retinal Amacrine Cells.

    PubMed

    Kim, Mean-Hwan; von Gersdorff, Henrique

    2016-02-01

    Amacrine cells are thought to be a major locus for mechanisms of light adaptation and contrast enhancement in the retina. However, the potential for plasticity in their AMPA receptor currents remains largely unknown. Using paired patch-clamp recordings between bipolar cell terminals and amacrine cells, we have simultaneously measured presynaptic membrane capacitance changes and EPSCs. Repetitive bipolar cell depolarizations, designed to maintain the same amount of exocytosis, nevertheless significantly potentiated evoked EPSCs in a subpopulation of amacrine cells. Likewise, repetitive iontophoresis (or puffs) of glutamate (or AMPA) onto the dendrites of amacrine cells also significantly potentiated evoked currents and [Ca(2+)]i rises. However, strong postsynaptic Ca(2+) buffering with BAPTA abolished the potentiation and selective antagonists of Ca(2+)-permeable AMPA receptors also blocked the potentiation of AMPA-mediated currents. Together these results suggest that Ca(2+) influx via Ca(2+)-permeable AMPA receptors can elicit a rapid form of postsynaptic plasticity in a subgroup of amacrine cell dendrites. PMID:26804991

  3. Response Properties of a Newly Identified Tristratified Narrow Field Amacrine Cell in the Mouse Retina.

    PubMed

    Newkirk, G S; Hoon, M; Wong, R O; Detwiler, P B

    2015-01-01

    Amacrine cells were targeted for whole cell recording using two-photon fluorescence microscopy in a transgenic mouse line in which the promoter for dopamine receptor 2 drove expression of green fluorescent protein in a narrow field tristratified amacrine cell (TNAC) that had not been studied previously. Light evoked a multiphasic response that was the sum of hyperpolarizing and depolarization synaptic inputs consistent with distinct dendritic ramifications in the off and on sublamina of the inner plexiform layer. The amplitude and waveform of the response, which consisted of an initial brief hyperpolarization at light onset followed by recovery to a plateau potential close to dark resting potential and a hyperpolarizing response at the light offset varied little over an intensity range from 0.4 to ~10^6 Rh*/rod/s. This suggests that the cell functions as a differentiator that generates an output signal (a transient reduction in inhibitory input to downstream retina neurons) that is proportional to the derivative of light input independent of its intensity. The underlying circuitry appears to consist of rod and cone driven on and off bipolar cells that provide direct excitatory input to the cell as well as to GABAergic amacrine cells that are synaptically coupled to TNAC. Canonical reagents that blocked excitatory (glutamatergic) and inhibitory (GABA and glycine) synaptic transmission had effects on responses to scotopic stimuli consistent with the rod driven component of the proposed circuit. However, responses evoked by photopic stimuli were paradoxical and could not be interpreted on the basis of conventional thinking about the neuropharmacology of synaptic interactions in the retina. PMID:26352594

  4. Response Properties of a Newly Identified Tristratified Narrow Field Amacrine Cell in the Mouse Retina

    PubMed Central

    Newkirk, G. S.; Hoon, M.; Wong, R. O.; Detwiler, P. B.

    2015-01-01

    Amacrine cells were targeted for whole cell recording using two-photon fluorescence microscopy in a transgenic mouse line in which the promoter for dopamine receptor 2 drove expression of green fluorescent protein in a narrow field tristratified amacrine cell (TNAC) that had not been studied previously. Light evoked a multiphasic response that was the sum of hyperpolarizing and depolarization synaptic inputs consistent with distinct dendritic ramifications in the off and on sublamina of the inner plexiform layer. The amplitude and waveform of the response, which consisted of an initial brief hyperpolarization at light onset followed by recovery to a plateau potential close to dark resting potential and a hyperpolarizing response at the light offset varied little over an intensity range from 0.4 to ~10^6 Rh*/rod/s. This suggests that the cell functions as a differentiator that generates an output signal (a transient reduction in inhibitory input to downstream retina neurons) that is proportional to the derivative of light input independent of its intensity. The underlying circuitry appears to consist of rod and cone driven on and off bipolar cells that provide direct excitatory input to the cell as well as to GABAergic amacrine cells that are synaptically coupled to TNAC. Canonical reagents that blocked excitatory (glutamatergic) and inhibitory (GABA and glycine) synaptic transmission had effects on responses to scotopic stimuli consistent with the rod driven component of the proposed circuit. However, responses evoked by photopic stimuli were paradoxical and could not be interpreted on the basis of conventional thinking about the neuropharmacology of synaptic interactions in the retina. PMID:26352594

  5. Light-evoked current responses in rod bipolar cells, cone depolarizing bipolar cells and all amacrine cells in dark-adapted mouse retina

    PubMed Central

    Pang, Ji-Jie; Gao, Fan; Wu, Samuel M

    2004-01-01

    Light-evoked excitatory cation current (ΔIC) and inhibitory chloride current (ΔICl) of rod and cone depolarizing bipolar cells (DBCRs and DBCCs) and AII amacrine cells (AIIACs) in dark-adapted mouse retinal slices were studied by whole-cell voltage-clamp recording techniques, and the cell morphology was revealed by Lucifer yellow fluorescence with a confocal microscope. ΔIC of all DBCRs exhibited similar high sensitivity to 500 nm light, but two patterns of ΔICl were observed in DBCRs with slightly different axon morphology. At least two types of DBCCs were identified: one with axon terminals ramified in 70–85% of the depth of the inner plexiform layer (IPL) and DBCR-like ΔIC sensitivity, whereas the other with axon terminals ramified in 55–75% of IPL depth and much lower ΔIC sensitivity. The relative rod/cone inputs to DBCs and AIIACs were analysed by comparing the ΔIC and ΔICl thresholds and dynamic ranges with the corresponding values of rods and cones. On average, the sensitivity of a DBCR to the 500 nm light is about 20 times higher than that of a rod. The sensitivity of an AIIAC is more than 1000 times higher than that of a rod, suggesting that AIIAC responses are pooled through a coupled network of about 40 AIIACs. Interactions of rod and cone signals in dark-adapted mouse retina appear asymmetrical: rod signals spread into the cone system more efficiently than cone signals into the rod system. The mouse synaptic circuitry allows small rod signals to be highly amplified, and effectively transmitted to the cone system via rod–cone and AIIAC–DBCC coupling. PMID:15181169

  6. Ganglion Cell and Displaced Amacrine Cell Density Distribution in the Retina of the Howler Monkey (Alouatta caraya)

    PubMed Central

    Muniz, José Augusto Pereira Carneiro; de Athaide, Luana Modesto; Gomes, Bruno Duarte; Finlay, Barbara L.; Silveira, Luiz Carlos de Lima

    2014-01-01

    Unlike all other New World (platyrrine) monkeys, both male and female howler monkeys (Alouatta sp.) are obligatory trichromats. In all other platyrrines, only females can be trichromats, while males are always dichromats, as determined by multiple behavioral, electrophysiological, and genetic studies. In addition to obligatory trichromacy, Alouatta has an unusual fovea, with substantially higher peak cone density in the foveal pit than every other diurnal anthropoid monkey (both platyrrhines and catarrhines) and great ape yet examined, including humans. In addition to documenting the general organization of the retinal ganglion cell layer in Alouatta, the distribution of cones is compared to retinal ganglion cells, to explore possible relationships between their atypical trichromacy and foveal specialization. The number and distribution of retinal ganglion cells and displaced amacrine cells were determined in six flat-mounted retinas from five Alouatta caraya. Ganglion cell density peaked at 0.5 mm between the fovea and optic nerve head, reaching 40,700–45,200 cells/mm2. Displaced amacrine cell density distribution peaked between 0.5–1.75 mm from the fovea, reaching mean values between 2,050–3,100 cells/mm2. The mean number of ganglion cells was 1,133,000±79,000 cells and the mean number of displaced amacrine cells was 537,000±61,800 cells, in retinas of mean area 641±62 mm2. Ganglion cell and displaced amacrine cell density distribution in the Alouatta retina was consistent with that observed among several species of diurnal Anthropoidea, both platyrrhines and catarrhines. The principal alteration in the Alouatta retina appears not to be in the number of any retinal cell class, but rather a marked gradient in cone density within the fovea, which could potentially support high chromatic acuity in a restricted central region. PMID:25546077

  7. Glycinergic input of widefield, displaced amacrine cells of the mouse retina.

    PubMed

    Majumdar, Sriparna; Weiss, Jan; Wässle, Heinz

    2009-08-01

    Glycine receptors (GlyRs) of displaced amacrine cells of the mouse retina were analysed using whole cell recordings and immunocytochemical staining with subunit-specific antibodies. During the recordings the cells were filled with a fluorescent tracer and 11 different morphological types could be identified. The studies were performed in wild-type mice and in mutant mice deficient in the GlyRalpha1 (Glra1(spd-ot), 'oscillator' mouse), the GlyRalpha2 (Glra2(-/-)) and the GlyRalpha3 subunit (Glra3(-/-)). Based on their responses to the application of exogenous glycine in the retinas of wild-type and mutant mice, the cells were grouped into three major classes: group I cells (comprising the morphological types MA-S5, MA-S1, MA-S1/S5, A17, PA-S1, PA-S5 and WA-S1), group II cells (comprising the morphological types PA-S4, WA-S3 and WA-multi) and ON-starburst cells. For further analysis, spontaneous inhibitory postsynaptic currents (sIPSCs) were measured both in wild-type and mutant mouse retinas. Glycinergic sIPSCs and glycine induced currents of group I cells remained unaltered across wild-type and the three mutant mice (mean decay time constant of sIPSCs, tau approximately 25 ms). Group II cells showed glycinergic sIPSCs and glycine induced currents in wild-type, Glra1(spd-ot) and Glra3(-/-) mice (tau approximately 25 ms); however, glycinergic currents were absent in group II cells of Glra2(-/-) mice. Glycine induced currents and sIPSCs recorded from ON-starburst amacrine cells did not differ significantly between wild-type and the mutant mouse retinas (tau approximately 50-70 ms). We propose that GlyRs of group II cells are dominated by the alpha2 subunit; GlyRs of ON-starburst amacrine cells appear to be dominated by the alpha4 subunit. PMID:19528249

  8. Islet-1 Controls the Differentiation of Retinal Bipolar and Cholinergic Amacrine Cells

    PubMed Central

    Elshatory, Yasser; Everhart, Drew; Deng, Min; Xie, Xiaoling; Barlow, Robert B.; Gan, Lin

    2010-01-01

    Whereas the mammalian retina possesses a repertoire of factors known to establish general retinal cell types, these factors alone cannot explain the vast diversity of neuronal subtypes. In other CNS regions, the differentiation of diverse neuronal pools is governed by coordinately acting LIM-homeodomain proteins including the Islet-class factor Islet-1 (Isl1). We report that deletion of Isl1 profoundly disrupts retinal function as assessed by electroretinograms and vision as assessed by optomotor behavior. These deficits are coupled with marked reductions in mature ON- and OFF-bipolar (>76%), cholinergic amacrine (93%), and ganglion (71%) cells. Mosaic deletion of Isl1 permitted a chimeric analysis of “wild-type” cells in a predominantly Isl1-null environment, demonstrating a cell-autonomous role for Isl1 in rod bipolar and cholinergic amacrine development. Furthermore, the effects on bipolar cell development appear to be dissociable from the preceding retinal ganglion cell loss, because Pou4f2-null mice are devoid of similar defects in bipolar cell marker expression. Expression of the ON- and OFF-bipolar cell differentiation factors Bhlhb4 and Vsx1, respectively, requires the presence of Isl1, whereas the early bipolar cell marker Prox1 initially did not. Thus, Isl1 is required for engaging bipolar differentiation pathways but not for general bipolar cell specification. Spatiotemporal expression analysis of additional LIM-homeobox genes identifies a LIM-homeobox gene network during bipolar cell development that includes Lhx3 and Lhx4. We conclude that Isl1 has an indispensable role in retinal neuron differentiation within restricted cell populations and this function may reflect a broader role for other LIM-homeobox genes in retinal development, and perhaps in establishing neuronal subtypes. PMID:18003851

  9. Light-evoked synaptic activity of retinal ganglion and amacrine cells is regulated in developing mouse retina

    PubMed Central

    He, Quanhua; Wang, Ping; Tian, Ning

    2010-01-01

    Recent studies have shown a continued maturation of visual responsiveness and synaptic activity of retina after eye opening, including the size of receptive fields of retinal ganglion cells (RGCs), light-evoked synaptic output of RGCs, bipolar cell spontaneous synaptic inputs to RGCs, and the synaptic connections between RGCs and ON and OFF bipolar cells. Light deprivation retarded some of these age-dependent changes. However, many other functional and morphological features of RGCs are not sensitive to visual experience. To determine whether light-evoked synaptic responses of RGCs undergo developmental change, we directly examined the light-evoked synaptic inputs from ON and OFF synaptic pathways to RGCs in developing retinas and found that both light-evoked excitatory and inhibitory synaptic currents decreased, but not increased, with age. We also examined the light-evoked synaptic inputs from ON and OFF synaptic pathways to amacrine cells in developing retinas and found that the light-evoked synaptic input of amacrine cells is also down-regulated in developing mouse retina. Different from the developmental changes of RGC spontaneous synaptic activity, dark rearing has little effect on the developmental changes of light-evoked synaptic activity of both RGCs and amacrine cells. Therefore, we concluded that the synaptic mechanisms mediating spontaneous and light-evoked synaptic activity of RGCs and amacrine cells are likely to be different. PMID:21091802

  10. Vesicular γ-Aminobutyric Acid Transporter Expression in Amacrine and Horizontal Cells

    PubMed Central

    Cueva, Juan G.; Haverkamp, Silke; Reimer, Richard J.; Edwards, Robert; Wässle, Heinz; Brecha, Nicholas C.

    2010-01-01

    The vesicular γ-aminobutyric acid (GABA) transporter (VGAT), which transports the inhibitory amino acid transmitters GABA and glycine, is localized to synaptic vesicles in axon terminals. The localization of VGAT immunoreactivity to mouse and rat retina was evaluated with light and electron microscopy by using well-characterized VGAT antibodies. Specific VGAT immunoreactivity was localized to numerous varicose processes in all laminae of the inner plexiform layer (IPL) and to the outer plexiform layer (OPL). Amacrine cell somata characterized by weak VGAT immunoreactivity in the cytoplasm were located in the ganglion cell layer and proximal inner nuclear layer (INL) adjacent to the IPL. In rat retina, VGAT-immunoreactive cell bodies also contained GABA, glycine, or parvalbumin (PV) immunoreactivity, suggesting vesicular uptake of GABA or glycine by these cells. A few varicose VGAT-immunoreactive processes entered the OPL from the IPL. VGAT immunoreactivity in the OPL was predominantly localized to horizontal cell processes. VGAT and calcium binding protein-28K immunoreactivities (CaBP; a marker for horizontal cells) were colocalized in processes and terminals distributed to the OPL. Furthermore, VGAT immunoreactivity overlapped or was immediately adjacent to postsynaptic density-95 (PSD-95) immunoreactivity, which is prominent in photoreceptor terminals. Preem-bedding immunoelectron microscopy of mouse and rat retinae showed that VGAT immunoreactivity was localized to horizontal cell processes and their terminals. Immunoreactivity was distributed throughout the cytoplasm of the horizontal cell processes. Taken together, these findings demonstrate VGAT immunoreactivity in both amacrine and horizontal cell processes, suggesting these cells contain vesicles that accumulate GABA and glycine, possibly for vesicular release. PMID:11920703

  11. Parallel Inhibition of Dopamine Amacrine Cells and Intrinsically Photosensitive Retinal Ganglion Cells in a Non-Image-Forming Visual Circuit of the Mouse Retina.

    PubMed

    Vuong, Helen E; Hardi, Claudia N; Barnes, Steven; Brecha, Nicholas C

    2015-12-01

    An inner retinal microcircuit composed of dopamine (DA)-containing amacrine cells and melanopsin-containing, intrinsically photosensitive retinal ganglion cells (M1 ipRGCs) process information about the duration and intensity of light exposures, mediating light adaptation, circadian entrainment, pupillary reflexes, and other aspects of non-image-forming vision. The neural interaction is reciprocal: M1 ipRGCs excite DA amacrine cells, and these, in turn, feed inhibition back onto M1 ipRGCs. We found that the neuropeptide somatostatin [somatotropin release inhibiting factor (SRIF)] also inhibits the intrinsic light response of M1 ipRGCs and postulated that, to tune the bidirectional interaction of M1 ipRGCs and DA amacrine cells, SRIF amacrine cells would provide inhibitory modulation to both cell types. SRIF amacrine cells, DA amacrine cells, and M1 ipRGCs form numerous contacts. DA amacrine cells and M1 ipRGCs express the SRIF receptor subtypes sst(2A) and sst4 respectively. SRIF modulation of the microcircuit was investigated with targeted patch-clamp recordings of DA amacrine cells in TH-RFP mice and M1 ipRGCs in OPN4-EGFP mice. SRIF increases K(+) currents, decreases Ca(2+) currents, and inhibits spike activity in both cell types, actions reproduced by the selective sst(2A) agonist L-054,264 (N-[(1R)-2-[[[(1S*,3R*)-3-(aminomethyl)cyclohexyl]methyl]amino]-1-(1H-indol-3-ylmethyl)-2-oxoethyl]spiro[1H-indene-1,4'-piperidine]-1'-carboxamide) in DA amacrine cells and the selective sst4 agonist L-803,087 (N(2)-[4-(5,7-difluoro-2-phenyl-1H-indol-3-yl)-1-oxobutyl]-L-arginine methyl ester trifluoroacetate) in M1 ipRGCs. These parallel actions of SRIF may serve to counteract the disinhibition of M1 ipRGCs caused by SRIF inhibition of DA amacrine cells. This allows the actions of SRIF on DA amacrine cells to proceed with adjusting retinal DA levels without destabilizing light responses by M1 ipRGCs, which project to non-image-forming targets in the brain. PMID:26631476

  12. Gestational lead exposure selectively decreases retinal dopamine amacrine cells and dopamine content in adult mice

    SciTech Connect

    Fox, Donald A.; Hamilton, W. Ryan; Johnson, Jerry E.; Xiao, Weimin; Chaney, Shawntay; Mukherjee, Shradha; Miller, Diane B.; O'Callaghan, James P.

    2011-11-15

    Gestational lead exposure (GLE) produces supernormal scotopic electroretinograms (ERG) in children, monkeys and rats, and a novel retinal phenotype characterized by an increased number of rod photoreceptors and bipolar cells in adult mice and rats. Since the loss of dopaminergic amacrine cells (DA ACs) in GLE monkeys and rats contributes to supernormal ERGs, the retinal DA system was analyzed in mice following GLE. C57BL/6 female mice were exposed to low (27 ppm), moderate (55 ppm) or high (109 ppm) lead throughout gestation and until postnatal day 10 (PN10). Blood [Pb] in control, low-, moderate- and high-dose GLE was {<=} 1, {<=} 10, {approx} 25 and {approx} 40 {mu}g/dL, respectively, on PN10 and by PN30 all were {<=} 1 {mu}g/dL. At PN60, confocal-stereology studies used vertical sections and wholemounts to characterize tyrosine hydroxylase (TH) expression and the number of DA and other ACs. GLE dose-dependently and selectively decreased the number of TH-immunoreactive (IR) DA ACs and their synaptic plexus without affecting GABAergic, glycinergic or cholinergic ACs. Immunoblots and confocal revealed dose-dependent decreases in retinal TH protein expression and content, although monoamine oxidase-A protein and gene expression were unchanged. High-pressure liquid chromatography showed that GLE dose-dependently decreased retinal DA content, its metabolites and DA utilization/release. The mechanism of DA selective vulnerability is unknown. However, a GLE-induced loss/dysfunction of DA ACs during development could increase the number of rods and bipolar cells since DA helps regulate neuronal proliferation, whereas during adulthood it could produce ERG supernormality as well as altered circadian rhythms, dark/light adaptation and spatial contrast sensitivity. -- Highlights: Black-Right-Pointing-Pointer Peak [BPb] in control, low-, moderate- and high-dose newborn mice with gestational lead exposure: {<=} 1, {<=} 10, 25 and 40 {mu}g/dL Black

  13. Multiple Genes on Chromosome 7 Regulate Dopaminergic Amacrine Cell Number in the Mouse Retina

    PubMed Central

    Whitney, Irene E.; Raven, Mary A.; Ciobanu, Daniel C.; Williams, Robert W.; Reese, Benjamin E.

    2009-01-01

    Purpose The size of neuronal populations is modulated by gene variants that influence cell production and survival, in turn influencing neuronal connectivity, function, and disease risk. The size of the dopaminergic amacrine (DA) cell population is a highly heritable trait exhibiting six-fold variation among inbred strains of mice, and is used here to identify genes that modulate the number of DA cells. Methods The entire population was counted in retinal wholemounts from 37 genetically defined lines of mice, including six standard inbred strains, 25 recombinant inbred strains (AXB/BXA), reciprocal F1 hybrids, a chromosome (Chr) 7 consomic line, and three additional genetically modified lines. Results We mapped much of this variation to a broad locus on Chr 7 (Dopaminergic amacrine cell number control, Chr 7). The Dacnc7 locus is flanked by two candidate genes known to modulate the number of other types of retinal neuron—the pro-apoptotic gene, Bax, and tyrosinase. The Tyr mutation was shown to modulate DA cell number modestly, although in the direction opposite that predicted. In contrast, Bax deficiency increased the population four-fold. Bax expression was significantly greater in the A/J strain relative to C57BL/6J, an effect that may be due to an SNP in a p53 consensus binding site known to modulate transcription. Finally, we note a strong candidate situated at the peak of the Dacnc7 locus, Lrrk1, a Parkinson’s disease gene exhibiting mis-sense mutations segregating within the AXB/BXA cross. Conclusions Multiple polymorphic genes on Chr. 7 modulate the size of the population of DA cells. PMID:19168892

  14. Visual stimulation switches the polarity of excitatory input to starburst amacrine cells

    PubMed Central

    Vlasits, Anna L.; Bos, Rémi; Morrie, Ryan D.; Fortuny, Cécile; Flannery, John G.; Feller, Marla B.; Rivlin-Etzion, Michal

    2014-01-01

    Summary Direction-selective ganglion cells (DSGCs) are tuned to motion in one direction. Starburst amacrine cells (SACs) are thought to mediate this direction selectivity through precise anatomical wiring to DSGCs. Nevertheless, we previously found that visual adaptation can reverse DSGCs’ directional tuning, overcoming the circuit anatomy. Here we explore the role of SACs in the generation and adaptation of direction selectivity. First, using pharmaco-genetics and two-photon calcium imaging, we validate that SACs are necessary for direction selectivity. Next, we demonstrate that exposure to an adaptive stimulus dramatically alters SACs’ synaptic inputs. Specifically, after visual adaptation, On-SACs lose their excitatory input during light onset but gain an excitatory input during light offset. Our data suggest that visual stimulation alters the interactions between rod and cone-mediated inputs that converge on the terminals of On cone BCs. These results demonstrate how the sensory environment can modify computations performed by anatomically-defined neuronal circuits. PMID:25155960

  15. Parallel Inhibition of Dopamine Amacrine Cells and Intrinsically Photosensitive Retinal Ganglion Cells in a Non-Image-Forming Visual Circuit of the Mouse Retina

    PubMed Central

    Vuong, Helen E.; Hardi, Claudia N.; Barnes, Steven

    2015-01-01

    An inner retinal microcircuit composed of dopamine (DA)-containing amacrine cells and melanopsin-containing, intrinsically photosensitive retinal ganglion cells (M1 ipRGCs) process information about the duration and intensity of light exposures, mediating light adaptation, circadian entrainment, pupillary reflexes, and other aspects of non-image-forming vision. The neural interaction is reciprocal: M1 ipRGCs excite DA amacrine cells, and these, in turn, feed inhibition back onto M1 ipRGCs. We found that the neuropeptide somatostatin [somatotropin release inhibiting factor (SRIF)] also inhibits the intrinsic light response of M1 ipRGCs and postulated that, to tune the bidirectional interaction of M1 ipRGCs and DA amacrine cells, SRIF amacrine cells would provide inhibitory modulation to both cell types. SRIF amacrine cells, DA amacrine cells, and M1 ipRGCs form numerous contacts. DA amacrine cells and M1 ipRGCs express the SRIF receptor subtypes sst2A and sst4 respectively. SRIF modulation of the microcircuit was investigated with targeted patch-clamp recordings of DA amacrine cells in TH–RFP mice and M1 ipRGCs in OPN4–EGFP mice. SRIF increases K+ currents, decreases Ca2+ currents, and inhibits spike activity in both cell types, actions reproduced by the selective sst2A agonist L-054,264 (N-[(1R)-2-[[[(1S*,3R*)-3-(aminomethyl)cyclohexyl]methyl]amino]-1-(1H-indol-3-ylmethyl)-2-oxoethyl]spiro[1H-indene-1,4′-piperidine]-1′-carboxamide) in DA amacrine cells and the selective sst4 agonist L-803,087 (N2-[4-(5,7-difluoro-2-phenyl-1H-indol-3-yl)-1-oxobutyl]-l-arginine methyl ester trifluoroacetate) in M1 ipRGCs. These parallel actions of SRIF may serve to counteract the disinhibition of M1 ipRGCs caused by SRIF inhibition of DA amacrine cells. This allows the actions of SRIF on DA amacrine cells to proceed with adjusting retinal DA levels without destabilizing light responses by M1 ipRGCs, which project to non-image-forming targets in the brain. SIGNIFICANCE

  16. Corelease of acetylcholine and GABA by an amacrine cell: Evidence for independent mechanisms

    SciTech Connect

    O'Mally, D.M.

    1989-01-01

    The spatial resolution of the cholinergic cells was measured by illuminating the retina with moving gratings composed of light and dark bars. Retinas that were labelled with {sup 3}H-choline released acetylcholine in response to moving gratings composed of bars as small as 50 {mu}m; 300 to 800 {mu}m wide bars yielded maximal responses. Responses were obtained to gratings moving at speeds from 50 to 6000 {mu}m/sec. Three groups recently reported that the cholinergic cells also contain GABA. To confirm these findings, retinas were double-labeled with {sup 3}H-GABA and DAPI, and processed for autoradiography. The cells that accumulate DAPI were heavily labelled with silver grains due to uptake of {sup 3}H-GABA. Incubation of retinas in the presence of elevated concentrations of K{sup +} caused them to release both acetylcholine and GABA, and autoradiography showed depletion of radioactive GABA, and autoradiography showed depletion of radioactive GABA from the cholinergic amacrine cells. Retinas were double-labeled with {sup 14}C-GABA and {sup 3}H-acetylcholine, allowing simultaneous measurement of their release. The release of {sup 14}C-GABA was independent of extracellular Ca{sup ++}. Radioactive GABA synthesized endogenously from {sup 14}C-glutamate behave the same as radioactive GABA accumulated from the medium. In the same experiments, the simultaneously measured release of {sup 3}H-acetylcholine was strongly Ca{sup ++}-dependent, indicating that acetylcholine and GABA are released by different mechanisms.

  17. Tetrodotoxin-Resistant Sodium Channels Contribute to Directional Responses in Starburst Amacrine Cells

    PubMed Central

    Oesch, Nicholas W.; Taylor, W. Rowland

    2010-01-01

    The biophysical mechanisms that give rise to direction selectivity in the retina remain uncertain. Current evidence suggests that the directional signal first arises within the dendrites of starburst amacrine cells (SBACs). Two models have been proposed to explain this phenomenon, one based on mutual inhibitory interactions between SBACs, and the other positing an intrinsic dendritic mechanism requiring a voltage-gradient depolarizing towards the dendritic tips. We tested these models by recording current and voltage responses to visual stimuli in SBACs. In agreement with previous work, we found that the excitatory currents in the SBACs were directional, and remained directional when GABA receptors were blocked. Contrary to the mutual-inhibitory model, stimuli that produce strong directional signals in ganglion cells failed to reveal a significant inhibitory input to SBACs. Suppression of the tonic excitatory conductance, proposed to generate the dendritic voltage-gradient required for the dendrite autonomous model, failed to eliminate the directional signal in SBACs. However, selective block of tetrodotoxin-resistant sodium channels did reduce the strength of the directional excitatory signal in the SBACs. These results indicate that current models of direction-selectivity in the SBACs are inadequate, and suggest that voltage-gated excitatory channels, specifically tetrodotoxin-resistant sodium channels, are important elements in directional signaling. This is the first physiological evidence that tetrodotoxin-resistant sodium channels play a role in retinal information processing. PMID:20805982

  18. Acetylcholine induces GABA release onto rod bipolar cells through heteromeric nicotinic receptors expressed in A17 amacrine cells

    PubMed Central

    Elgueta, Claudio; Vielma, Alex H.; Palacios, Adrian G.; Schmachtenberg, Oliver

    2015-01-01

    Acetylcholine (ACh) is a major retinal neurotransmitter that modulates visual processing through a large repertoire of cholinergic receptors expressed on different retinal cell types. ACh is released from starburst amacrine cells (SACs) under scotopic conditions, but its effects on cells of the rod pathway have not been investigated. Using whole-cell patch clamp recordings in slices of rat retina, we found that ACh application triggers GABA release onto rod bipolar (RB) cells. GABA was released from A17 amacrine cells and activated postsynaptic GABAA and GABAC receptors in RB cells. The sensitivity of ACh-induced currents to nicotinic ACh receptor (nAChR) antagonists (TMPH ~ mecamylamine > erysodine > DhβE > MLA) together with the differential potency of specific agonists to mimic ACh responses (cytisine >> RJR2403 ~ choline), suggest that A17 cells express heteromeric nAChRs containing the β4 subunit. Activation of nAChRs induced GABA release after Ca2+ accumulation in A17 cell dendrites and varicosities mediated by L-type voltage-gated calcium channels (VGCCs) and intracellular Ca2+ stores. Inhibition of acetylcholinesterase depolarized A17 cells and increased spontaneous inhibitory postsynaptic currents in RB cells, indicating that endogenous ACh enhances GABAergic inhibition of RB cells. Moreover, injection of neostigmine or cytisine reduced the b-wave of the scotopic flash electroretinogram (ERG), suggesting that cholinergic modulation of GABA release controls RB cell activity in vivo. These results describe a novel regulatory mechanism of RB cell inhibition and complement our understanding of the neuromodulatory control of retinal signal processing. PMID:25709566

  19. Inner plexiform layer of jack mackerel retina: participation of amacrine and ganglion cells in its spatial organization.

    PubMed

    Podugolnikova, T A

    1985-01-01

    In the jack mackerel retina (Trachurus mediterraneus ponticus) the inner plexiform layer demonstrates a very high degree of differentiation and contains not less than 25 sublayers. Investigation with Golgi method revealed many varieties of neurons, which are responsible for the structural organization of the inner plexiform layer. There are 8 types of bipolar cells, 24 types of amacrine cells and 7 types of ganglion cells with layered processes. The branching levels of the processes of these neurons were determined. Several varieties of neurons are described for the first time. PMID:3832609

  20. The Synaptic and Morphological Basis of Orientation Selectivity in a Polyaxonal Amacrine Cell of the Rabbit Retina

    PubMed Central

    Murphy-Baum, Benjamin L.

    2015-01-01

    Much of the computational power of the retina derives from the activity of amacrine cells, a large and diverse group of GABAergic and glycinergic inhibitory interneurons. Here, we identify an ON-type orientation-selective, wide-field, polyaxonal amacrine cell (PAC) in the rabbit retina and demonstrate how its orientation selectivity arises from the structure of the dendritic arbor and the pattern of excitatory and inhibitory inputs. Excitation from ON bipolar cells and inhibition arising from the OFF pathway converge to generate a quasi-linear integration of visual signals in the receptive field center. This serves to suppress responses to high spatial frequencies, thereby improving sensitivity to larger objects and enhancing orientation selectivity. Inhibition also regulates the magnitude and time course of excitatory inputs to this PAC through serial inhibitory connections onto the presynaptic terminals of ON bipolar cells. This presynaptic inhibition is driven by graded potentials within local microcircuits, similar in extent to the size of single bipolar cell receptive fields. Additional presynaptic inhibition is generated by spiking amacrine cells on a larger spatial scale covering several hundred microns. The orientation selectivity of this PAC may be a substrate for the inhibition that mediates orientation selectivity in some types of ganglion cells. SIGNIFICANCE STATEMENT The retina comprises numerous excitatory and inhibitory circuits that encode specific features in the visual scene, such as orientation, contrast, or motion. Here, we identify a wide-field inhibitory neuron that responds to visual stimuli of a particular orientation, a feature selectivity that is primarily due to the elongated shape of the dendritic arbor. Integration of convergent excitatory and inhibitory inputs from the ON and OFF visual pathways suppress responses to small objects and fine textures, thus enhancing selectivity for larger objects. Feedback inhibition regulates the

  1. The Synaptic and Morphological Basis of Orientation Selectivity in a Polyaxonal Amacrine Cell of the Rabbit Retina.

    PubMed

    Murphy-Baum, Benjamin L; Taylor, W Rowland

    2015-09-30

    Much of the computational power of the retina derives from the activity of amacrine cells, a large and diverse group of GABAergic and glycinergic inhibitory interneurons. Here, we identify an ON-type orientation-selective, wide-field, polyaxonal amacrine cell (PAC) in the rabbit retina and demonstrate how its orientation selectivity arises from the structure of the dendritic arbor and the pattern of excitatory and inhibitory inputs. Excitation from ON bipolar cells and inhibition arising from the OFF pathway converge to generate a quasi-linear integration of visual signals in the receptive field center. This serves to suppress responses to high spatial frequencies, thereby improving sensitivity to larger objects and enhancing orientation selectivity. Inhibition also regulates the magnitude and time course of excitatory inputs to this PAC through serial inhibitory connections onto the presynaptic terminals of ON bipolar cells. This presynaptic inhibition is driven by graded potentials within local microcircuits, similar in extent to the size of single bipolar cell receptive fields. Additional presynaptic inhibition is generated by spiking amacrine cells on a larger spatial scale covering several hundred microns. The orientation selectivity of this PAC may be a substrate for the inhibition that mediates orientation selectivity in some types of ganglion cells. Significance statement: The retina comprises numerous excitatory and inhibitory circuits that encode specific features in the visual scene, such as orientation, contrast, or motion. Here, we identify a wide-field inhibitory neuron that responds to visual stimuli of a particular orientation, a feature selectivity that is primarily due to the elongated shape of the dendritic arbor. Integration of convergent excitatory and inhibitory inputs from the ON and OFF visual pathways suppress responses to small objects and fine textures, thus enhancing selectivity for larger objects. Feedback inhibition regulates the

  2. Morphology and function of three VIP-expressing amacrine cell types in the mouse retina.

    PubMed

    Akrouh, Alejandro; Kerschensteiner, Daniel

    2015-10-01

    Amacrine cells (ACs) are the most diverse class of neurons in the retina. The variety of signals provided by ACs allows the retina to encode a wide range of visual features. Of the 30-50 AC types in mammalian species, few have been studied in detail. Here, we combine genetic and viral strategies to identify and to characterize morphologically three vasoactive intestinal polypeptide-expressing GABAergic AC types (VIP1-, VIP2-, and VIP3-ACs) in mice. Somata of VIP1- and VIP2-ACs reside in the inner nuclear layer and somata of VIP3-ACs in the ganglion cell layer, and they show asymmetric distributions along the dorsoventral axis of the retina. Neurite arbors of VIP-ACs differ in size (VIP1-ACs ≈ VIP3-ACs > VIP2-ACs) and stratify in distinct sublaminae of the inner plexiform layer. To analyze light responses and underlying synaptic inputs, we target VIP-ACs under 2-photon guidance for patch-clamp recordings. VIP1-ACs depolarize strongly to light increments (ON) over a wide range of stimulus sizes but show size-selective responses to light decrements (OFF), depolarizing to small and hyperpolarizing to large stimuli. The switch in polarity of OFF responses is caused by pre- and postsynaptic surround inhibition. VIP2- and VIP3-ACs both show small depolarizations to ON stimuli and large hyperpolarizations to OFF stimuli but differ in their spatial response profiles. Depolarizations are caused by ON excitation outweighing ON inhibition, whereas hyperpolarizations result from pre- and postsynaptic OFF-ON crossover inhibition. VIP1-, VIP2-, and VIP3-ACs thus differ in response polarity and spatial tuning and contribute to the diversity of inhibitory and neuromodulatory signals in the retina. PMID:26311183

  3. Cyan fluorescent protein expression in ganglion and amacrine cells in a thy1-CFP transgenic mouse retina

    PubMed Central

    Vila, Alejandro; Huynh, Uyen-Chi N.; Brecha, Nicholas C.

    2008-01-01

    Purpose To characterize cyan fluorescent protein (CFP) expression in the retina of the thy1-CFP (B6.Cg-Tg(Thy1-CFP)23Jrs/J) transgenic mouse line. Methods CFP expression was characterized using morphometric methods and immunohistochemistry with antibodies to neurofilament light (NF-L), neuronal nuclei (NeuN), POU-domain protein (Brn3a) and calretinin, which immunolabel ganglion cells, and syntaxin 1 (HPC-1), glutamate decarboxylase 67 (GAD67), GABA plasma membrane transporter-1 (GAT-1), and choline acetyltransferase (ChAT), which immunolabel amacrine cells. Results CFP was extensively expressed in the inner retina, primarily in the inner plexiform layer (IPL), ganglion cell layer (GCL), nerve fiber layer, and optic nerve. CFP fluorescent cell bodies were in all retinal regions and their processes ramified in all laminae of the IPL. Some small, weakly CFP fluorescent somata were in the inner nuclear layer (INL). CFP-containing somata in the GCL ranged from 6 to 20 μm in diameter, and they had a density of 2636±347 cells/mm2 at 1.5 mm from the optic nerve head. Immunohistochemical studies demonstrated colocalization of CFP with the ganglion cell markers NF-L, NeuN, Brn3a, and calretinin. Immunohistochemistry with antibodies to HPC-1, GAD67, GAT-1, and ChAT indicated that the small, weakly fluorescent CFP cells in the INL and GCL were cholinergic amacrine cells. Conclusions The total number and density of CFP-fluorescent cells in the GCL were within the range of previous estimates of the total number of ganglion cells in the C57BL/6J line. Together these findings suggest that most ganglion cells in the thy1-CFP mouse line 23 express CFP. In conclusion, the thy1-CFP mouse line is highly useful for studies requiring the identification of ganglion cells. PMID:18728756

  4. An excitatory amacrine cell detects object motion and provides feature-selective input to ganglion cells in the mouse retina

    PubMed Central

    Kim, Tahnbee; Soto, Florentina; Kerschensteiner, Daniel

    2015-01-01

    Retinal circuits detect salient features of the visual world and report them to the brain through spike trains of retinal ganglion cells. The most abundant ganglion cell type in mice, the so-called W3 ganglion cell, selectively responds to movements of small objects. Where and how object motion sensitivity arises in the retina is incompletely understood. In this study, we use 2-photon-guided patch-clamp recordings to characterize responses of vesicular glutamate transporter 3 (VGluT3)-expressing amacrine cells (ACs) to a broad set of visual stimuli. We find that these ACs are object motion sensitive and analyze the synaptic mechanisms underlying this computation. Anatomical circuit reconstructions suggest that VGluT3-expressing ACs form glutamatergic synapses with W3 ganglion cells, and targeted recordings show that the tuning of W3 ganglion cells' excitatory input matches that of VGluT3-expressing ACs' responses. Synaptic excitation of W3 ganglion cells is diminished, and responses to object motion are suppressed in mice lacking VGluT3. Object motion, thus, is first detected by VGluT3-expressing ACs, which provide feature-selective excitatory input to W3 ganglion cells. DOI: http://dx.doi.org/10.7554/eLife.08025.001 PMID:25988808

  5. Neural architecture of the "transient" ON directionally selective (class IIb1) ganglion cells in rabbit retina, partly co-stratified with starburst amacrine cells.

    PubMed

    Famiglietti, Edward V

    2016-01-01

    Recent physiological studies coupled with intracellular staining have subdivided ON directionally selective (DS) ganglion cells of rabbit retina into two types. One exhibits more "transient" and more "brisk" responses (ON DS-t), and the other has more "sustained' and more "sluggish" responses (ON DS-s), although both represent the same three preferred directions and show preference for low stimulus velocity, as reported in previous studies of ON DS ganglion cells in rabbit retina. ON DS-s cells have the morphology of ganglion cells previously shown to project to the medial terminal nucleus (MTN) of the accessory optic system, and the MTN-projecting, class IVus1 cells have been well-characterized previously in terms of their dendritic morphology, branching pattern, and stratification. ON DS-t ganglion cells have a distinctly different morphology and exhibit heterotypic coupling to amacrine cells, including axon-bearing amacrine cells, with accompanying synchronous firing, while ON DS-s cells are not coupled. The present study shows that ON DS-t cells are morphologically identical to the previously well-characterized, "orphan" class IIb1 ganglion cell, previously regarded as a member of the "brisk-concentric" category of ganglion cells. Its branching pattern, quantitatively analyzed, is similar to that of the morphological counterparts of X and Y cells, and very different from that of the ON DS-s ganglion cell. Close analysis of the dendritic stratification of class IIb1 ganglion cells together with fiducial cells indicates that they differ from that of the ON DS-s cells. In agreement with one of the three previous studies, class IIb1/ON DS-t cells, unlike class IVus1/ON DS-s ganglion cells, in the main do not co-stratify with starburst amacrine cells. As the present study shows, however, portions of their dendrites do deviate from the main substratum, coming within range of starburst boutons. Parsimony favors DS input from starburst amacrine cells both to ON DS

  6. COUP-TFI and -TFII nuclear receptors are expressed in amacrine cells and play roles in regulating the differentiation of retinal progenitor cells.

    PubMed

    Inoue, Mariko; Iida, Atsumi; Satoh, Shinya; Kodama, Tatsuhiko; Watanabe, Sumiko

    2010-01-01

    Chicken ovalbumin upstream promoter transcription factors (COUP-TFs) are members of the steroid/thyroid hormone receptor superfamily. We have shown that two homologous COUP-TF genes, COUP-TFI and COUP-TFII, are expressed in developing mouse retina with a unique gradient along the dorsal-ventral axis. In this work, we aimed to characterize the detailed expression patterns of COUP-TFs in mature retina. Their functions in retinal progenitor cell differentiation into subtypes of mature retinal cells were also examined. Immunostaining of frozen mouse retinal sections with antibodies against COUP-TFs and markers for retinal subtypes revealed that COUP-TFI and -TFII are expressed in amacrine cells, especially in a glycinergic subtype in mature mouse retina. Forced expression of COUP-TFI and -TFII in mouse retinal explant culture by retrovirus-mediated gene transfer promoted amacrine and cone photoreceptor cell differentiation, whereas that of rod photoreceptors decreased. Cell proliferation and apoptosis were not affected by the perturbation of COUP-TFI and -TFII expression levels. Using the Y79 retinoblastoma cell line, we observed that COUP-TFI and -TFII suppressed the transcriptional activation of the Nrl gene. We then analyzed one another member of COUP-TF transcription factors, COUP-TFgamma, whose structure is relatively distant from those of COUP-TFI and -TFII. It is expressed mainly in horizontal cells and has weak activity in inducing amacrine cells when COUP-TFgamma was ectopically expressed in retinal explants. In summary, we found that COUP-TFI and -TFII play roles in amacrine cell differentiation, and COUP-TFgamma has distinct expression pattern and roles during retinal development. PMID:19766631

  7. Ectopic Retinal ON Bipolar Cell Synapses in the OFF Inner Plexiform Layer: Contacts with Dopaminergic Amacrine Cells and Melanopsin Ganglion Cells

    PubMed Central

    Dumitrescu, Olivia N.; Pucci, Francesco G.; Wong, Kwoon Y.; Berson, David M.

    2010-01-01

    A key principle of retinal organization is that distinct ON and OFF channels are relayed by separate populations of bipolar cells to different sublaminae of the inner plexiform layer (IPL). ON bipolar cell axons have been thought to synapse exclusively in the inner IPL (the ON sublamina) onto dendrites of ON-type amacrine and ganglion cells. However, M1 melanopsin-expressing ganglion cells and dopaminergic amacrine (DA) cells apparently violate this dogma. Both are driven by ON bipolar cells, but their dendrites stratify in the outermost IPL, within the OFF sublamina. Here, in the mouse retina, we show that some ON cone bipolar cells make ribbon synapses in the outermost OFF sublayer, where they costratify with and contact the dendrites of M1 and DA cells. Whole-cell recording and dye filling in retinal slices indicate that Type 6 ON cone bipolars provide some of this ectopic ON channel input. Imaging studies in dissociated bipolar cells show that these ectopic ribbon synapses are capable of vesicular release. There is thus an accessory ON sublayer in the outer IPL. PMID:19731338

  8. Distinctive receptive field and physiological properties of a wide-field amacrine cell in the macaque monkey retina

    PubMed Central

    Puller, Christian; Rieke, Fred; Neitz, Jay; Neitz, Maureen

    2015-01-01

    At early stages of visual processing, receptive fields are typically described as subtending local regions of space and thus performing computations on a narrow spatial scale. Nevertheless, stimulation well outside of the classical receptive field can exert clear and significant effects on visual processing. Given the distances over which they occur, the retinal mechanisms responsible for these long-range effects would certainly require signal propagation via active membrane properties. Here the physiology of a wide-field amacrine cell—the wiry cell—in macaque monkey retina is explored, revealing receptive fields that represent a striking departure from the classic structure. A single wiry cell integrates signals over wide regions of retina, 5–10 times larger than the classic receptive fields of most retinal ganglion cells. Wiry cells integrate signals over space much more effectively than predicted from passive signal propagation, and spatial integration is strongly attenuated during blockade of NMDA spikes but integration is insensitive to blockade of NaV channels with TTX. Thus these cells appear well suited for contributing to the long-range interactions of visual signals that characterize many aspects of visual perception. PMID:26133804

  9. Target-Specific Glycinergic Transmission from VGluT3-Expressing Amacrine Cells Shapes Suppressive Contrast Responses in the Retina.

    PubMed

    Tien, Nai-Wen; Kim, Tahnbee; Kerschensteiner, Daniel

    2016-05-17

    Neurons that release more than one transmitter exist throughout the CNS. Yet, how these neurons deploy multiple transmitters and shape the function of specific circuits is not well understood. VGluT3-expressing amacrine cells (VG3-ACs) provide glutamatergic input to ganglion cells activated by contrast and motion. Using optogenetics, we find that VG3-ACs provide selective glycinergic input to a retinal ganglion cell type suppressed by contrast and motion (SbC-RGCs). Firing of SbC-RGCs is suppressed at light ON and OFF over a broad range of stimulus sizes. Anatomical circuit reconstructions reveal that VG3-ACs form inhibitory synapses preferentially on processes that link ON and OFF arbors of SbC-RGC dendrites. Removal of VG3-ACs from mature circuits reduces inhibition and attenuates spike suppression of SbC-RGCs in a contrast- and size-selective manner, illustrating the modularity of retinal circuits. VG3-ACs thus use dual transmitters in a target-specific manner and shape suppressive contrast responses in the retina by glycinergic transmission. PMID:27160915

  10. Adenosine A2A Receptor Up-Regulates Retinal Wave Frequency via Starburst Amacrine Cells in the Developing Rat Retina

    PubMed Central

    Huang, Pin-Chien; Hsiao, Yu-Tien; Kao, Shao-Yen; Chen, Ching-Feng; Chen, Yu-Chieh; Chiang, Chung-Wei; Lee, Chien-fei; Lu, Juu-Chin; Chern, Yijuang; Wang, Chih-Tien

    2014-01-01

    Background Developing retinas display retinal waves, the patterned spontaneous activity essential for circuit refinement. During the first postnatal week in rodents, retinal waves are mediated by synaptic transmission between starburst amacrine cells (SACs) and retinal ganglion cells (RGCs). The neuromodulator adenosine is essential for the generation of retinal waves. However, the cellular basis underlying adenosine's regulation of retinal waves remains elusive. Here, we investigated whether and how the adenosine A2A receptor (A2AR) regulates retinal waves and whether A2AR regulation of retinal waves acts via presynaptic SACs. Methodology/Principal Findings We showed that A2AR was expressed in the inner plexiform layer and ganglion cell layer of the developing rat retina. Knockdown of A2AR decreased the frequency of spontaneous Ca2+ transients, suggesting that endogenous A2AR may up-regulate wave frequency. To investigate whether A2AR acts via presynaptic SACs, we targeted gene expression to SACs by the metabotropic glutamate receptor type II promoter. Ca2+ transient frequency was increased by expressing wild-type A2AR (A2AR-WT) in SACs, suggesting that A2AR may up-regulate retinal waves via presynaptic SACs. Subsequent patch-clamp recordings on RGCs revealed that presynaptic A2AR-WT increased the frequency of wave-associated postsynaptic currents (PSCs) or depolarizations compared to the control, without changing the RGC's excitability, membrane potentials, or PSC charge. These findings suggest that presynaptic A2AR may not affect the membrane properties of postsynaptic RGCs. In contrast, by expressing the C-terminal truncated A2AR mutant (A2AR-ΔC) in SACs, the wave frequency was reduced compared to the A2AR-WT, but was similar to the control, suggesting that the full-length A2AR in SACs is required for A2AR up-regulation of retinal waves. Conclusions/Significance A2AR up-regulates the frequency of retinal waves via presynaptic SACs, requiring its full

  11. Prdm13 regulates subtype specification of retinal amacrine interneurons and modulates visual sensitivity.

    PubMed

    Watanabe, Satoshi; Sanuki, Rikako; Sugita, Yuko; Imai, Wataru; Yamazaki, Ryoji; Kozuka, Takashi; Ohsuga, Mizuki; Furukawa, Takahisa

    2015-05-20

    Amacrine interneurons, which are highly diversified in morphological, neurochemical, and physiological features, play crucial roles in visual information processing in the retina. However, the specification mechanisms and functions in vision for each amacrine subtype are not well understood. We found that the Prdm13 transcriptional regulator is specifically expressed in developing and mature amacrine cells in the mouse retina. Most Prdm13-positive amacrine cells are Calbindin- and Calretinin-positive GABAergic or glycinergic neurons. Absence of Prdm13 significantly reduces GABAergic and glycinergic amacrines, resulting in a specific defect of the S2/S3 border neurite bundle in the inner plexiform layer. Forced expression of Prdm13 distinctively induces GABAergic and glycinergic amacrine cells but not cholinergic amacrine cells, whereas Ptf1a, an upstream transcriptional regulator of Prdm13, induces all of these subtypes. Moreover, Prdm13-deficient mice showed abnormally elevated spatial, temporal, and contrast sensitivities in vision. Together, these results show that Prdm13 regulates development of a subset of amacrine cells, which newly defines an amacrine subtype to negatively modulate visual sensitivities. Our current study provides new insights into mechanisms of the diversification of amacrine cells and their function in vision. PMID:25995483

  12. Segregated Glycine-Glutamate Co-transmission from vGluT3 Amacrine Cells to Contrast-Suppressed and Contrast-Enhanced Retinal Circuits.

    PubMed

    Lee, Seunghoon; Zhang, Yi; Chen, Minggang; Zhou, Z Jimmy

    2016-04-01

    Since the introduction of Dale's principle of "one neuron releases one transmitter at all its synapses," a growing number of exceptions to this principle have been identified. While the concept of neurotransmitter co-release by a single neuron is now well accepted, the specific synaptic circuitry and functional advantage of co-neurotransmission remain poorly understood in general. Here we report Ca(2+)-dependent co-release of a new combination of inhibitory and excitatory neurotransmitters, namely, glycine and glutamate, by the vGluT3-expressing amacrine cell (GAC) in the mouse retina. GACs selectively make glycinergic synapses with uniformity detectors (UDs) and provide a major inhibitory drive that underlies the suppressed-by-contrast trigger feature of UDs. Meanwhile, GACs release glutamate to excite OFF alpha ganglion cells and a few other nonlinear, contrast-sensitive ganglion cells. This coordinated inhibition and excitation of two separate neuronal circuits by a single interneuron suggests a unique advantage in differential detection of visual field uniformity and contrast. PMID:26996083

  13. Conditional Knock-Out of Vesicular GABA Transporter Gene from Starburst Amacrine Cells Reveals the Contributions of Multiple Synaptic Mechanisms Underlying Direction Selectivity in the Retina

    PubMed Central

    Pei, Zhe; Chen, Qiang; Koren, David; Giammarinaro, Benno; Acaron Ledesma, Hector

    2015-01-01

    Direction selectivity of direction-selective ganglion cells (DSGCs) in the retina results from patterned excitatory and inhibitory inputs onto DSGCs during motion stimuli. The inhibitory inputs onto DSGCs are directionally tuned to the antipreferred (null) direction and therefore potently suppress spiking during motion in the null direction. However, whether direction-selective inhibition is indispensable for direction selectivity is unclear. Here, we selectively eliminated the directional tuning of inhibitory inputs onto DSGCs by disrupting GABA release from the presynaptic interneuron starburst amacrine cell in the mouse retina. We found that, even without directionally tuned inhibition, direction selectivity can still be implemented in a subset of On-Off DSGCs by direction-selective excitation and a temporal offset between excitation and isotropic inhibition. Our results therefore demonstrate the concerted action of multiple synaptic mechanisms for robust direction selectivity in the retina. SIGNIFICANCE STATEMENT The direction-selective circuit in the retina has been a classic model to study neural computations by the brain. An important but unresolved question is how direction selectivity is implemented by directionally tuned excitatory and inhibitory mechanisms. Here we specifically removed the direction tuning of inhibition from the circuit. We found that direction tuning of inhibition is important but not indispensable for direction selectivity of DSGCs' spiking activity, and that the residual direction selectivity is implemented by direction-selective excitation and temporal offset between excitation and inhibition. Our results highlight the concerted actions of synaptic excitation and inhibition required for robust direction selectivity in the retina and provide critical insights into how patterned excitation and inhibition collectively implement sensory processing. PMID:26400950

  14. AdcAII of Streptococcus pneumoniae Affects Pneumococcal Invasiveness

    PubMed Central

    Brown, Lindsey R.; Gunnell, Steven M.; Cassella, Adam N.; Keller, Lance E.; Scherkenbach, Lisa A.; Mann, Beth; Brown, Matthew W.; Hill, Rebecca; Fitzkee, Nicholas C.; Rosch, Jason W.; Tuomanen, Elaine I.; Thornton, Justin A.

    2016-01-01

    Across bacterial species, metal binding proteins can serve functions in pathogenesis in addition to regulating metal homeostasis. We have compared and contrasted the activities of zinc (Zn2+)-binding lipoproteins AdcA and AdcAII in the Streptococcus pneumoniae TIGR4 background. Exposure to Zn2+-limiting conditions resulted in delayed growth in a strain lacking AdcAII (ΔAdcAII) when compared to wild type bacteria or a mutant lacking AdcA (ΔAdcA). AdcAII failed to interact with the extracellular matrix protein laminin despite homology to laminin-binding proteins of related streptococci. Deletion of AdcA or AdcAII led to significantly increased invasion of A549 human lung epithelial cells and a trend toward increased invasion in vivo. Loss of AdcAII, but not AdcA, was shown to negatively impact early colonization of the nasopharynx. Our findings suggest that expression of AdcAII affects invasiveness of S. pneumoniae in response to available Zn2+ concentrations. PMID:26752283

  15. Differential effects of P2Y1 deletion on glial activation and survival of photoreceptors and amacrine cells in the ischemic mouse retina

    PubMed Central

    Pannicke, T; Frommherz, I; Biedermann, B; Wagner, L; Sauer, K; Ulbricht, E; Härtig, W; Krügel, U; Ueberham, U; Arendt, T; Illes, P; Bringmann, A; Reichenbach, A; Grosche, A

    2014-01-01

    Gliosis of retinal Müller glial cells may have both beneficial and detrimental effects on neurons. To investigate the role of purinergic signaling in ischemia-induced reactive gliosis, transient retinal ischemia was evoked by elevation of the intraocular pressure in wild-type (Wt) mice and in mice deficient in the glia-specific nucleotide receptor P2Y1 (P2Y1 receptor-deficient (P2Y1R-KO)). While control retinae of P2Y1R-KO mice displayed reduced cell numbers in the ganglion cell and inner nuclear layers, ischemia induced apoptotic death of cells in all retinal layers in both, Wt and P2Y1R-KO mice, but the damage especially on photoreceptors was more pronounced in retinae of P2Y1R-KO mice. In contrast, gene expression profiling and histological data suggest an increased survival of amacrine cells in the postischemic retina of P2Y1R-KO mice. Interestingly, measuring the ischemia-induced downregulation of inwardly rectifying potassium channel (Kir)-mediated K+ currents as an indicator, reactive Müller cell gliosis was found to be weaker in P2Y1R-KO (current amplitude decreased by 18%) than in Wt mice (decrease by 68%). The inner retina harbors those neurons generating action potentials, which strongly rely on an intact ion homeostasis. This may explain why especially these cells appear to benefit from the preserved Kir4.1 expression in Müller cells, which should allow them to keep up their function in the context of spatial buffering of potassium. Especially under ischemic conditions, maintenance of this Müller cell function may dampen cytotoxic neuronal hyperexcitation and subsequent neuronal cell loss. In sum, we found that purinergic signaling modulates the gliotic activation pattern of Müller glia and lack of P2Y1 has janus-faced effects. In the end, the differential effects of a disrupted P2Y1 signaling onto neuronal survival in the ischemic retina call the putative therapeutical use of P2Y1-antagonists into question. PMID:25077539

  16. Differential effects of P2Y1 deletion on glial activation and survival of photoreceptors and amacrine cells in the ischemic mouse retina.

    PubMed

    Pannicke, T; Frommherz, I; Biedermann, B; Wagner, L; Sauer, K; Ulbricht, E; Härtig, W; Krügel, U; Ueberham, U; Arendt, T; Illes, P; Bringmann, A; Reichenbach, A; Grosche, A

    2014-01-01

    Gliosis of retinal Müller glial cells may have both beneficial and detrimental effects on neurons. To investigate the role of purinergic signaling in ischemia-induced reactive gliosis, transient retinal ischemia was evoked by elevation of the intraocular pressure in wild-type (Wt) mice and in mice deficient in the glia-specific nucleotide receptor P2Y1 (P2Y1 receptor-deficient (P2Y1R-KO)). While control retinae of P2Y1R-KO mice displayed reduced cell numbers in the ganglion cell and inner nuclear layers, ischemia induced apoptotic death of cells in all retinal layers in both, Wt and P2Y1R-KO mice, but the damage especially on photoreceptors was more pronounced in retinae of P2Y1R-KO mice. In contrast, gene expression profiling and histological data suggest an increased survival of amacrine cells in the postischemic retina of P2Y1R-KO mice. Interestingly, measuring the ischemia-induced downregulation of inwardly rectifying potassium channel (Kir)-mediated K(+) currents as an indicator, reactive Müller cell gliosis was found to be weaker in P2Y1R-KO (current amplitude decreased by 18%) than in Wt mice (decrease by 68%). The inner retina harbors those neurons generating action potentials, which strongly rely on an intact ion homeostasis. This may explain why especially these cells appear to benefit from the preserved Kir4.1 expression in Müller cells, which should allow them to keep up their function in the context of spatial buffering of potassium. Especially under ischemic conditions, maintenance of this Müller cell function may dampen cytotoxic neuronal hyperexcitation and subsequent neuronal cell loss. In sum, we found that purinergic signaling modulates the gliotic activation pattern of Müller glia and lack of P2Y1 has janus-faced effects. In the end, the differential effects of a disrupted P2Y1 signaling onto neuronal survival in the ischemic retina call the putative therapeutical use of P2Y1-antagonists into question. PMID:25077539

  17. Apolipoprotein A-II Plus Lipid Emulsion Enhance Cell Growth via SR-B1 and Target Pancreatic Cancer In Vitro and In Vivo

    PubMed Central

    Thanh LE, Thao N.; Gill, Anthony J.; Bulanadi, Jerikho C.; Patel, Mili; Waddington, Lynne J.; Rye, Kerry-Anne; Moghaddam, Minoo J.; Smith, Ross C.

    2016-01-01

    Background Apolipoprotein A-II (ApoA-II) is down regulated in the sera of pancreatic ductal adenocarcinoma (PDAC) patients, which may be due to increase utilization of high density lipoprotein (HDL) lipid by pancreatic cancer tissue. This study examined the influence of exogenous ApoA-II on lipid uptake and cell growth in pancreatic cancer (PC) both in vitro and in vivo. Methods Cryo transmission electron microscopy (TEM) examined ApoA-II’s influence on morphology of SMOFLipid emulsion. The influence of ApoA-II on proliferation of cancer cell lines was determined by incubating them with lipid+/-ApoA-II and anti-SR-B1 antibody. Lipid was labeled with the fluorophore, DiD, to trace lipid uptake by cancer cells in vitro by confocal microscopy and in vivo in PDAC patient derived xenograft tumours (PDXT) by fluorescence imaging. Scavenger receptor class B type-1(SR-B1) expression in PDAC cell lines and in PDAC PDXT was measured by western blotting and immunohistochemistry, respectively. Results ApoA-II spontaneously converted lipid emulsion into very small unilamellar rHDL like vesicles (rHDL/A-II) and enhanced lipid uptake in PANC-1, CFPAC-1 and primary tumour cells as shown by confocal microscopy. SR-B1 expression was 13.2, 10.6, 3.1 and 2.3 fold higher in PANC-1, MIAPaCa-2, CFPAC-1 and BxPC3 cell lines than the normal pancreatic cell line (HPDE6) and 3.7 fold greater in PDAC tissue than in normal pancreas. ApoA-II plus lipid significantly increased the uptake of labeled lipid and promoted cell growth in PANC-1, MIAPaCa-2, CFPAC-1 and BxPC3 cells which was inhibited by anti SR-B1 antibody. Further, ApoA-II increased the uptake of lipid in xenografts by 3.4 fold. Conclusion Our data suggest that ApoA-II enhance targeting potential of lipid in pancreatic cancer which may have imaging and drug delivery potentialities. PMID:27002321

  18. Fibrates increase human apolipoprotein A-II expression through activation of the peroxisome proliferator-activated receptor.

    PubMed Central

    Vu-Dac, N; Schoonjans, K; Kosykh, V; Dallongeville, J; Fruchart, J C; Staels, B; Auwerx, J

    1995-01-01

    In view of the evidence linking plasma high density lipoprotein (HDL)-cholesterol levels to a protective effect against coronary artery disease and the widespread use of fibrates in the treatment of hyperlipidemia, the goal of this study was to analyze the influence of fibrates on the expression of apolipoprotein (apo) A-II, a major protein constituent of HDL. Administration of fenofibrate (300 mg/d) to 16 patients with coronary artery disease resulted in a marked increase in plasma apo A-II concentrations (0.34 +/- 0.11 to 0.45 +/- 0.17 grams/liter; P < 0.01). This increase in plasma apo A-II was due to a direct effect on hepatic apo A-II production, since fenofibric acid induced apo A-II mRNA levels to 450 and 250% of control levels in primary cultures of human hepatocytes and in human hepatoblastoma HepG2 cells respectively. The induction in apo A-II mRNA levels was followed by an increase in apo A-II secretion in both cell culture systems. Transient transfection experiments of a reporter construct driven by the human apo A-II gene promoter indicated that fenofibrate induced apo A-II gene expression at the transcriptional level. Furthermore, several other peroxisome proliferators, such as the fibrate, Wy-14643, and the fatty acid, eicosatetraynoic acid (ETYA), also induced apo A-II gene transcription. Unilateral deletions and site-directed mutagenesis identified a sequence element located in the J-site of the apo A-II promoter mediating the responsiveness to fibrates and fatty acids. This element contains two imperfect half sites spaced by 1 oligonucleotide similar to a peroxisome proliferator responsive element (PPRE). Cotransfection assays showed that the peroxisome proliferator activated receptor (PPAR) transactivates the apo A-II promoter through this AII-PPRE. Gel retardation assays demonstrated that PPAR binds to the AII-PPRE with an affinity comparable to its binding affinity to the acyl coA oxidase (ACO)-PPRE. In conclusion, in humans fibrates increase

  19. Human apolipoprotein A-II protects against diet-induced atherosclerosis in transgenic rabbits

    PubMed Central

    Wang, Yao; Niimi, Manabu; Nishijima, Kazutoshi; Waqar, Ahmed Bilal; Yu, Ying; Koike, Tomonari; Kitajima, Shuji; Liu, Enqi; Inoue, Tomohiro; Kohashi, Masayuki; Keyamura, Yuka; Yoshikawa, Tomohiro; Zhang, Jifeng; Ma, Loretta; Zha, Xiaohui; Watanabe, Teruo; Asada, Yujiro; Chen, Y. Eugene; Fan, Jianglin

    2013-01-01

    Objective Apolipoprotein A-II (apo A-II) is the second major apolipoprotein of HDLs, yet its pathophysiological roles in the development of atherosclerosis remain unknown. We aimed to examine whether apo A-II plays any role in atherogenesis and if so, to elucidate the mechanism involved. Methods and Results We compared the susceptibility of human apo A-II transgenic (Tg) rabbits to cholesterol diet-induced atherosclerosis with non-Tg littermate rabbits. Tg rabbits developed significantly less aortic and coronary atherosclerosis than their non-Tg littermates while total plasma cholesterol levels were similar. Atherosclerotic lesions of Tg rabbits were characterized by reduced macrophages and smooth muscle cells and apo A-II immunoreactive proteins were frequently detected in the lesions. Tg rabbits exhibited low levels of plasma CRP and blood leukocytes compared to non-Tg rabbits and HDLs of Tg rabbit plasma exerted stronger cholesterol efflux activity and inhibitory effects on the inflammatory cytokine expression by macrophages in vitro than HDLs isolated from non-Tg rabbits. In addition, β-VLDLs of Tg rabbits were less sensitive to copper-induced oxidation than β-VLDLs of non-Tg rabbits. Conclusions These results suggest that enrichment of apo A-II in HDL particles has atheroprotective effects and apo A-II may become a target for the treatment of atherosclerosis. PMID:23241412

  20. Adaptation to background light enables contrast coding at rod bipolar cell synapses

    PubMed Central

    Ke, Jiang-Bin; Wang, Yanbin V.; Borghuis, Bart G.; Cembrowski, Mark S.; Riecke, Hermann; Kath, William L.; Demb, Jonathan B.; Singer, Joshua H.

    2013-01-01

    SUMMARY Rod photoreceptors contribute to vision over a ~6 log-unit range of light intensities. The wide dynamic range of rod vision is thought to depend upon light intensity-dependent switching between two parallel pathways linking rods to ganglion cells: a rod→rod bipolar (RB) cell pathway that operates at dim backgrounds and a rod→cone→cone bipolar cell pathway that operates at brighter backgrounds. We evaluated this conventional model of rod vision by recording rod-mediated light responses from ganglion and AII amacrine cells and by recording RB-mediated synaptic currents from AII amacrine cells in mouse retina. Contrary to the conventional model, we found that the RB pathway functioned at backgrounds sufficient to activate the rod→cone pathway. As background light intensity increased, the RB’s role changed from encoding the absorption of single photons to encoding contrast modulations around mean luminance. This transition is explained by the intrinsic dynamics of transmission from RB synapses. PMID:24373883

  1. Identification of a second cellulose synthase gene (acsAII) in Acetobacter xylinum.

    PubMed Central

    Saxena, I M; Brown, R M

    1995-01-01

    A second cellulose synthase gene (acsAII) coding for a 175-kDa polypeptide that is similar in size and sequence to the acsAB gene product has been identified in Acetobacter xylinum AY201. Evidence for the presence of this gene was obtained during analysis of A. xylinum mutants in which the acsAB gene was disrupted (I.M. Saxena, K. Kudlicka, K. Okuda, and R.M. Brown, Jr., J. Bacteriol. 176:5735-5752, 1994). Although these mutants produced no detectable cellulose, they exhibited significant cellulose synthase activity in vitro. The acsAII gene was isolated by using an acsAB gene fragment as a probe. The acsAII gene coded for cellulose synthase activity as determined from sequence analysis and study of mutants in which this gene was disrupted. A mutant in which only the acsAII gene was disrupted showed no significant differences in either the in vivo cellulose production or the in vitro cellulose synthase activity compared with wild-type cells. Mutants in which both the acsAII and acsAB genes were disrupted produced no cellulose in vivo and exhibited negligible cellulose synthase activity in vitro, thus confirming that the cellulose synthase activity observed in the acsAB mutants was coded by the acsAII gene. These results establish the presence of an additional gene for cellulose synthase expressed in cells of A. xylinum, yet this gene is not required for cellulose production when cells are grown under laboratory conditions. PMID:7665515

  2. Dopamine D1 receptor expression is bipolar cell type-specific in the mouse retina.

    PubMed

    Farshi, Pershang; Fyk-Kolodziej, Bozena; Krolewski, David M; Walker, Paul D; Ichinose, Tomomi

    2016-07-01

    In the retina, dopamine is a key molecule for daytime vision. Dopamine is released by retinal dopaminergic amacrine cells and transmits signaling either by conventional synaptic or by volume transmission. By means of volume transmission, dopamine modulates all layers of retinal neurons; however, it is not well understood how dopamine modulates visual signaling pathways in bipolar cells. Here we analyzed Drd1a-tdTomato BAC transgenic mice and found that the dopamine D1 receptor (D1R) is expressed in retinal bipolar cells in a type-dependent manner. Strong tdTomato fluorescence was detected in the inner nuclear layer and localized to type 1, 3b, and 4 OFF bipolar cells and type 5-2, XBC, 6, and 7 ON bipolar cells. In contrast, type 2, 3a, 5-1, 9, and rod bipolar cells did not express Drd1a-tdTomato. Other interneurons were also found to express tdTomato including horizontal cells and a subset (25%) of AII amacrine cells. Diverse visual processing pathways, such as color or motion-coded pathways, are thought to be initiated in retinal bipolar cells. Our results indicate that dopamine sculpts bipolar cell performance in a type-dependent manner to facilitate daytime vision. J. Comp. Neurol. 524:2059-2079, 2016. © 2015 Wiley Periodicals, Inc. PMID:26587737

  3. Global Ca2+ Signaling Drives Ribbon-Independent Synaptic Transmission at Rod Bipolar Cell Synapses

    PubMed Central

    Mehta, Bhupesh; Ke, Jiang-Bin; Zhang, Lei; Baden, Alexander D.; Markowitz, Alexander L.; Nayak, Subhashree; Briggman, Kevin L.

    2014-01-01

    Ribbon-type presynaptic active zones are a hallmark of excitatory retinal synapses, and the ribbon organelle is thought to serve as the organizing point of the presynaptic active zone. Imaging of exocytosis from isolated retinal neurons, however, has revealed ectopic release (i.e., release away from ribbons) in significant quantities. Here, we demonstrate in an in vitro mouse retinal slice preparation that ribbon-independent release from rod bipolar cells activates postsynaptic AMPARs on AII amacrine cells. This form of release appears to draw on a unique, ribbon-independent, vesicle pool. Experimental, anatomical, and computational analyses indicate that it is elicited by a significant, global elevation of intraterminal [Ca2+] arising following local buffer saturation. Our observations support the conclusion that ribbon-independent release provides a read-out of the average behavior of all of the active zones in a rod bipolar cell's terminal. PMID:24790194

  4. Global Ca2+ signaling drives ribbon-independent synaptic transmission at rod bipolar cell synapses.

    PubMed

    Mehta, Bhupesh; Ke, Jiang-Bin; Zhang, Lei; Baden, Alexander D; Markowitz, Alexander L; Nayak, Subhashree; Briggman, Kevin L; Zenisek, David; Singer, Joshua H

    2014-04-30

    Ribbon-type presynaptic active zones are a hallmark of excitatory retinal synapses, and the ribbon organelle is thought to serve as the organizing point of the presynaptic active zone. Imaging of exocytosis from isolated retinal neurons, however, has revealed ectopic release (i.e., release away from ribbons) in significant quantities. Here, we demonstrate in an in vitro mouse retinal slice preparation that ribbon-independent release from rod bipolar cells activates postsynaptic AMPARs on AII amacrine cells. This form of release appears to draw on a unique, ribbon-independent, vesicle pool. Experimental, anatomical, and computational analyses indicate that it is elicited by a significant, global elevation of intraterminal [Ca(2+)] arising following local buffer saturation. Our observations support the conclusion that ribbon-independent release provides a read-out of the average behavior of all of the active zones in a rod bipolar cell's terminal. PMID:24790194

  5. Angiotensin II (AII) vascular smooth muscle (VSM) receptors in the domestic fowl: interaction with endothelium (ENDO)

    SciTech Connect

    Stallone, J.N.; Nishimura, H.; Nasjletti, A.; Khosla, M.C.

    1986-03-05

    Previously, they reported that in the fowl, AII causes a vasodepressor response in vivo and endothelium-dependent vaso-relaxation in vitro, and that specific, high affinity AII receptors exist in the membrane fraction of aortic VSM. A membrane fraction was prepared from ENDO-deleted (ENDO-) aortae of leghorn hens and incubation with /sup 125/I-(Val/sup 5/)AII (0.5 nM) in 50 mM tris (pH 7.2) with 10 mM Mg/sup 2 +/, 0.2% BSA, and inhibitors (bacitracin, EGTA, phenanthroline) for 90 min at 12/sup 0/C. Ligand binding to the preparation (30 ..mu..g protein) was competitively displaced by AII analogs at relative potencies of: (Asn/sup 1/, Val/sup 5/)AII < (Asp/sup 1/, Ile/sup 5/)AII > (Asp/sup 1/, Val/sup 5/)AII > (Val/sup 5/)AIII > (Sar/sup 1/, Ile/sup 8/)AII. Maximum saturation binding of ligand (fmol/mg protein) to the ENDO(-) preparation (2169) was markedly lower than in identically prepared ENDO-intact (ENDO+) preparations (3498). /sup 14/C-arachidonic acid (3.6 nM) was incubated with minced ENDO(-) or ENDO(+) aortae (250 mg) for 30 min at 37/sup 0/C in Krebs-Ringer solution. Thin layer chromatography of ENDO(-) and ENDO(+) media yielded products that cochromatographed with prostaglandins PGE/sub 2/ and PGF/sub 2..cap alpha../, but not with 6-keto-PGF/sub 1..cap alpha../. Coincubation of substrate with indomethacin (50 ..mu..M) completely abolished PG production by both tissues. These data suggest that in the fowl aorta: (1) highly specific AII receptors exist in VSM, (2) AII receptors may exist in ENDO, and (3) PGI/sub 2/ is not involved in AII-induced vasodilation.

  6. Two regulatory elements of similar structure and placed in tandem account for the repressive activity of the first intron of the human apolipoprotein A-II gene.

    PubMed Central

    Bossu, J P; Chartier, F L; Fruchart, J C; Auwerx, J; Staels, B; Laine, B

    1996-01-01

    Recent reports indicate that apolipoprotein (apo) A-II, the second most abundant protein of high-density lipoproteins, plays a crucial role in counteracting the beneficial effect of apo A-I against atherogenesis. Transcription of the human apo A-II gene is controlled by an enhancer comprising 14 regulatory elements located upstream of its promoter whereas the first intron of this gene behaves as a silencer. Here we show that two sequence elements account for the repressive activity of this intron and correspond to negative regulatory elements termed NRE I and NRE II. The activity of intron I and the nuclear proteins binding to NRE I and II are encountered in hepatic cells but not in non-hepatic cells studied here. Both NREs form nucleoprotein complexes of very similar physicochemical characteristics and bind the same or closely related proteins. Site-directed mutagenesis, transient transfection and gel-shift analysis experiments indicate that both NREs exhibit similar structures, being composed of two sites required for maximal activity and optimal binding of transcription factors. Therefore two negative regulatory elements of similar structure and function, placed in tandem, account for the repressive activity of the first intron of the human apo A-II gene. These NREs do not exhibit structural similarity with known NREs of other genes. PMID:8809045

  7. 78 FR 64396 - Mixed Straddles; Straddle-by-Straddle Identification Under Section 1092(b)(2)(A)(i)(I); Correction

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-29

    ... Under Section 1092(b)(2)(A)(i)(I); Correction AGENCY: Internal Revenue Service (IRS), Treasury. ACTION... (78 FR 46807). The temporary regulations applied to all identified mixed straddles established after... straddles; straddle-by-straddle identification under section 1092(b)(2)(A)(i)(I) (Temporary). * * * * *...

  8. 78 FR 46854 - Mixed Straddles; Straddle-by-Straddle Identification Under Section 1092(b)(2)(A)(i)(I)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-02

    ... Under Section 1092(b)(2)(A)(i)(I) AGENCY: Internal Revenue Service (IRS), Treasury. ACTION: Notice of... identification under section 1092(b)(2)(A)(i)(I). [The text of the proposed amendments to Sec. 1.1092(b)-3(b)(6... the IRS are issuing temporary regulations that explain how to account for unrealized gain or loss on...

  9. An apolipoprotein A-II polymorphism (-265T/C, rs5082), regulates postprandial response to a saturated fat overload in healthy men

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Apolipoprotein (Apo) A-II is an apolipoprotein with an unknown role in lipid metabolism. It has been suggested that the presence of the less frequent allele of a single nucleotide polymorphism (Apo A-II -265T/C, rs5082) reduces the transcription rate of Apo A-II and enhances VLDL postprandial cleara...

  10. 49 CFR Appendix A-Ii to Part 541 - Lines With Antitheft Devices Which Are Exempted in-Part From the Parts-Marking Requirements of...

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ...-Part From the Parts-Marking Requirements of This Standard Pursuant to 49 CFR Part 543 A Appendix A-II... STANDARD Pt. 541, App. A-II Appendix A-II to Part 541—Lines With Antitheft Devices Which Are Exempted in-Part From the Parts-Marking Requirements of This Standard Pursuant to 49 CFR Part 543...

  11. Aii20J, a wide-spectrum thermostable N-acylhomoserine lactonase from the marine bacterium Tenacibaculum sp. 20J, can quench AHL-mediated acid resistance in Escherichia coli.

    PubMed

    Mayer, C; Romero, M; Muras, A; Otero, A

    2015-11-01

    Acyl homoserine lactones (AHLs) are produced by many Gram-negative bacteria to coordinate gene expression in cellular density dependent mechanisms known as quorum sensing (QS). Since the disruption of the communication systems significantly reduces virulence, the inhibition of quorumsensing processes or quorum quenching (QQ) represents an interesting anti-pathogenic strategy to control bacterial infections. Escherichia coli does not produce AHLs but possesses an orphan AHL receptor, SdiA, which is thought to be able to sense the QS signals produced by other bacteria and controls important traits as the expression of glutamate-dependent acid resistance mechanism, therefore constituting a putative target for QQ. A novel AHL-lactonase, named Aii20J, has been identified, cloned and over expressed from the marine bacterium Tenacibaculum sp. strain 20 J presenting a wide-spectrum QQ activity. The enzyme, belonging to the metallo-β-lactamase family, shares less than 31 % identity with the lactonase AiiA from Bacillus spp. Aii20J presents a much higher specific activity than the Bacillus enzyme, maintains its activity after incubation at 100 ºC for 10 minutes, is resistant to protease K and α-chymotrypsin, and is unaffected by wide ranges of pH. The addition of Aii20J (20 μg/mL) to cultures of E. coli K-12 to which OC6-HSL was added resulted in a significant reduction in cell viability in comparison with the acidresistant cultures derived from the presence of the signal. Results confirm the interaction between AHLs and SdiA in E. coli for the expression of virulence-related genes and reveal the potential use of Aii20J as anti-virulence strategy against important bacterial pathogens and in other biotechnological applications. PMID:26092757

  12. Apolipoprotein A-II polymorphism: relationships to behavioural and hormonal mediators of obesity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: The interaction between apolipoprotein A-II (APOA2) m265 genotype and saturated fat for obesity traits has been more extensively demonstrated than for any other locus, but behavioural and hormonal mechanisms underlying this relationship are unexplored. In this study, we evaluated relatio...

  13. Francisella tularensis Subtype A.II Genomic Plasticity in Comparison with Subtype A.I

    PubMed Central

    Larson, Marilynn A.; Nalbantoglu, Ufuk; Sayood, Khalid; Zentz, Emily B.; Bartling, Amanda M.; Francesconi, Stephen C.; Fey, Paul D.; Dempsey, Michael P.; Hinrichs, Steven H.

    2015-01-01

    Although Francisella tularensis is considered a monomorphic intracellular pathogen, molecular genotyping and virulence studies have demonstrated important differences within the tularensis subspecies (type A). To evaluate genetic variation within type A strains, sequencing and assembly of a new subtype A.II genome was achieved for comparison to other completed F. tularensis type A genomes. In contrast with the F. tularensis A.I strains (SCHU S4, FSC198, NE061598, and TI0902), substantial genomic variation was observed between the newly sequenced F. tularensis A.II strain (WY-00W4114) and the only other publically available A.II strain (WY96-3418). Genome differences between WY-00W4114 and WY96-3418 included three major chromosomal translocations, 1580 indels, and 286 nucleotide substitutions of which 159 were observed in predicted open reading frames and 127 were located in intergenic regions. The majority of WY-00W4114 nucleotide deletions occurred in intergenic regions, whereas most of the insertions and substitutions occurred in predicted genes. Of the nucleotide substitutions, 48 (30%) were synonymous and 111 (70%) were nonsynonymous. WY-00W4114 and WY96-3418 nucleotide polymorphisms were predominantly G/C to A/T allelic mutations, with WY-00W4114 having more A+T enrichment. In addition, the A.II genomes contained a considerably higher number of intact genes and longer repetitive sequences, including transposon remnants than the A.I genomes. Together these findings support the premise that F. tularensis A.II may have a fitness advantage compared to the A.I subtype due to the higher abundance of functional genes and repeated chromosomal sequences. A better understanding of the selective forces driving F. tularensis genetic diversity and plasticity is needed. PMID:25918839

  14. Transport of Apolipoproteins A-I and A-II by Human Thoracic Duct Lymph

    PubMed Central

    Anderson, David W.; Schaefer, Ernst J.; Bronzert, Thomas J.; Lindgren, Frank T.; Forte, Trudy; Starzl, Thomas E.; Niblack, Gary D.; Zech, Loren A.; Brewer, H. Bryan

    1981-01-01

    The daily transport of human plasma apolipoproteins A-I and A-II, triglyceride, and total cholesterol from the thoracic duct lymph into plasma was measured in two subjects before and three subjects after renal transplantation. Lymph triglyceride transport was ∼83% of the daily ingested fat loads, whereas lymph cholesterol transport was consistently greater than the amount of daily ingested cholesterol. Lymph apolipoprotein transport significantly (P < 0.05) exceeded the predicted apolipoprotein synthesis rate by an average of 659±578 mg/d for apolipoprotein A-I and 109±59 mg/d for apolipoprotein A-II among the five subjects. It is estimated that 22-77% (apolipoprotein A-I) and 28-82% (apolipoprotein A-II) of daily total body apolipoprotein synthesis takes place in the intestine. Lymph high density lipoprotein particles are mostly high density lipoprotein2b and high density lipoprotein2a and have a greater overall relative triglyceride content and a smaller relative cholesteryl ester content when compared with homologous plasma high density lipoproteins. The major quantity of both lymph apolipoprotein A-I (81±8%) and apolipoprotein A-II (90±11%) was found within high density lipoproteins with almost all of the remainder found in chylomicrons and very low density lipoproteins. The combined results are consistent with a major contribution of the intestine to total body synthesis of apolipoprotein A-I and apolipoprotein A-II. An important role of lymph in returning filtered apolipoprotein to plasma in association with high density lipoproteins is proposed. Accompanying the return of filtered apolipoprotein to the plasma is a probable transformation, both in size and composition, of at least some of the lymph high density lipoprotein2b and high density lipoprotein2a particles into high density lipoprotein3. Images PMID:7204560

  15. Cytokinetic studies reveal etiology of cytogenetic genotoxicity produced by a series of angiotensin II (AII) receptor antagonists may be perturbed DNA synthesis

    SciTech Connect

    Selden, J.R.; Miller, J.E.; Dolbeare, F.; Galloway, S.M.; Nichols, W.W. Lawrence Livermore National Lab., CA )

    1993-01-01

    Six of 13 AII receptor antagonists produced chromosomal aberrations in CHO cells. In addition, these six compounds perturbed cellular kinetics (i.e., reduced mitotic indices and cell yields). It was hypothesized that the mechanism of clastogenesis was not due to a direct genotoxic effect, but may result from disruption of DNA replication. Flow cytokinetic studies, using the BrdUrd-FITC/propidium iodide technique, were performed on all six clastogenic compounds, and a seventh candidate from this group. All seven altered CHO cell kinetics as follows: (1) The amount of BrdUrd per S phase cell was reduced; (2) Cell movement within S phase was inhibited; and (3) Lowest doses perturbing CHO cell kinetics were below minimum concentrations producing aberrations. These data provide evidence that this cytogenetic damage is mediated by a mechanism which disrupts cellular DNA synthesis.

  16. Investigations of AII-BVI mixed crystals with the piezoelectric photothermal method

    NASA Astrophysics Data System (ADS)

    Maliński, M.; Bychto, L.; Zakrzewski, J.; Firszt, F.; Pawlak, M.; Binnebesel, P.

    2005-06-01

    In this paper the frequency phase characteristics of the photothermal piezoelectric signal of a few AII-BVI mixed crystals are presented and discussed. The following groups of crystals were analyzed: Zn{1-x}BexTe, Zn{1-x}BexSe, Cd{1-x}MgxSe and Cd{1-x}MnxTe. From the fitting procedure the concentration dependence of the thermal diffusivity of these mixed crystals was determined.

  17. Goldfish Leptin-AI and Leptin-AII: Function and Central Mechanism in Feeding Control

    PubMed Central

    Yan, Ai-Fen; Chen, Ting; Chen, Shuang; Ren, Chun-Hua; Hu, Chao-Qun; Cai, Yi-Ming; Liu, Fang; Tang, Dong-Sheng

    2016-01-01

    In mammals, leptin is a peripheral satiety factor that inhibits feeding by regulating a variety of appetite-related hormones in the brain. However, most of the previous studies examining leptin in fish feeding were performed with mammalian leptins, which share very low sequence homologies with fish leptins. To elucidate the function and mechanism of endogenous fish leptins in feeding regulation, recombinant goldfish leptin-AI and leptin-AII were expressed in methylotrophic yeast and purified by immobilized metal ion affinity chromatography (IMAC). By intraperitoneal (IP) injection, both leptin-AI and leptin-AII were shown to inhibit the feeding behavior and to reduce the food consumption of goldfish in 2 h. In addition, co-treatment of leptin-AI or leptin-AII could block the feeding behavior and reduce the food consumption induced by neuropeptide Y (NPY) injection. High levels of leptin receptor (lepR) mRNA were detected in the hypothalamus, telencephalon, optic tectum and cerebellum of the goldfish brain. The appetite inhibitory effects of leptins were mediated by downregulating the mRNA levels of orexigenic NPY, agouti-related peptide (AgRP) and orexin and upregulating the mRNA levels of anorexigenic cocaine-amphetamine-regulated transcript (CART), cholecystokinin (CCK), melanin-concentrating hormone (MCH) and proopiomelanocortin (POMC) in different areas of the goldfish brain. Our study, as a whole, provides new insights into the functions and mechanisms of leptins in appetite control in a fish model. PMID:27249000

  18. Goldfish Leptin-AI and Leptin-AII: Function and Central Mechanism in Feeding Control.

    PubMed

    Yan, Ai-Fen; Chen, Ting; Chen, Shuang; Ren, Chun-Hua; Hu, Chao-Qun; Cai, Yi-Ming; Liu, Fang; Tang, Dong-Sheng

    2016-01-01

    In mammals, leptin is a peripheral satiety factor that inhibits feeding by regulating a variety of appetite-related hormones in the brain. However, most of the previous studies examining leptin in fish feeding were performed with mammalian leptins, which share very low sequence homologies with fish leptins. To elucidate the function and mechanism of endogenous fish leptins in feeding regulation, recombinant goldfish leptin-AI and leptin-AII were expressed in methylotrophic yeast and purified by immobilized metal ion affinity chromatography (IMAC). By intraperitoneal (IP) injection, both leptin-AI and leptin-AII were shown to inhibit the feeding behavior and to reduce the food consumption of goldfish in 2 h. In addition, co-treatment of leptin-AI or leptin-AII could block the feeding behavior and reduce the food consumption induced by neuropeptide Y (NPY) injection. High levels of leptin receptor (lepR) mRNA were detected in the hypothalamus, telencephalon, optic tectum and cerebellum of the goldfish brain. The appetite inhibitory effects of leptins were mediated by downregulating the mRNA levels of orexigenic NPY, agouti-related peptide (AgRP) and orexin and upregulating the mRNA levels of anorexigenic cocaine-amphetamine-regulated transcript (CART), cholecystokinin (CCK), melanin-concentrating hormone (MCH) and proopiomelanocortin (POMC) in different areas of the goldfish brain. Our study, as a whole, provides new insights into the functions and mechanisms of leptins in appetite control in a fish model. PMID:27249000

  19. Disruption in dopaminergic innervation during photoreceptor degeneration.

    PubMed

    Ivanova, Elena; Yee, Christopher W; Sagdullaev, Botir T

    2016-04-15

    Dopaminergic amacrine cells (DACs) release dopamine in response to light-driven synaptic inputs, and are critical to retinal light adaptation. Retinal degeneration (RD) compromises the light responsiveness of the retina and, subsequently, dopamine metabolism is impaired. As RD progresses, retinal neurons exhibit aberrant activity, driven by AII amacrine cells, a primary target of the retinal dopaminergic network. Surprisingly, DACs are an exception to this physiological change; DACs exhibit rhythmic activity in healthy retina, but do not burst in RD. The underlying mechanism of this divergent behavior is not known. It is also unclear whether RD leads to structural changes in DACs, impairing functional regulation of AII amacrine cells. Here we examine the anatomical details of DACs in three mouse models of human RD to determine how changes to the dopaminergic network may underlie physiological changes in RD. By using rd10, rd1, and rd1/C57 mice we were able to dissect the impacts of genetic background and the degenerative process on DAC structure in RD retina. We found that DACs density, soma size, and primary dendrite length are all significantly reduced. Using a novel adeno-associated virus-mediated technique to label AII amacrine cells in mouse retina, we observed diminished dopaminergic contacts to AII amacrine cells in RD mice. This was accompanied by changes to the components responsible for dopamine synthesis and release. Together, these data suggest that structural alterations of the retinal dopaminergic network underlie physiological changes during RD. PMID:26356010

  20. Effects of an inducible aiiA gene on disease resistance in Eucalyptus urophylla × Eucalyptus grandis.

    PubMed

    Ouyang, L J; Li, L M

    2016-08-01

    N-acyl-homoserine lactones (AHLs) are metabolites of mostly gram-negative bacteria and are critical signaling molecules in bacterial quorum-sensing systems. At threshold concentrations, AHLs can activate the expression of pathogenic genes and induce diseases. Therefore, reducing AHL concentrations is a key point of disease control in plants. AHL-lactonase, which is expressed by aiiA, is widespread in Bacillus sp and can hydrolyze AHLs. In the present study, we cloned aiiA from Bacillus subtilis by PCR. A plant expression vector of aiiA was constructed and name Pcam-PPP3-aiiA, in which expression of aiiA was controlled by the pathogen-inducible plant promoter PPP3. The recombinant plasmid was transferred into Eucalyptus × urophylla × E. grandis by an Agrobacterium-mediated transformation. PCR and Southern blotting showed that aiiA was successfully integrated into the E. urophylla × E. grandis genome and its expression was induced by Ralstonia solanacearum 12 h after inoculation, as shown by reverse transcription-PCR. The transcription efficacy of aiiA increased 43.88-, 30.65-, and 18.95-fold after inoculation with R. solanacearum, Erwinia carotovora ssp. zeae (Sabet) and Cylindrocladium quinqueseptatum, respectively as shown by RT-real-time PCR. Transgenic E.urophylla × E.grandis expressing the AIIA protein exhibited significantly enhanced disease resistance compared to non-transgenic plants by delaying the onset of wilting and reducing the disease index. PMID:26905275

  1. [Expression of gene aiiA carrying the promoter of gene cry3Aa in Bacillus thuringiensis].

    PubMed

    Zhu, Chen-Guang; Sun, Ming; Yu, Zi-Niu

    2003-07-01

    N-acyl-homoserine lactones (AHLs), are widely conserved signal molecules present in quorum-sensing systems of many Gram-negative bacteria. AHLs molecules mediate the expression of virulence genes of a range of bacterial pathogens. Recently, it has been reported that AiiA protein, which widely exists in Bacillus species, can inactivate the AHLs by hydrolyzing the lactone bond of AHLs, thus attenuate the diseases caused by the expression of virulence genes of bacterial pathogens. Bacillus thuringiensis, a type of Gram-positive bacteria, has been used extensively as a microbial insecticide in the last few decades. However, most of important insecticidal B. thuringiensis strains have not been exploited for bacterial disease control because they usually do not produce antibiotics that are effective against bacteria and fungi. The discovery of AiiA protein in B. thuringiensis shows the application potential of B. thuringiensis on biocontrol against bacterial diseases. In this study, in order to construct the B. thuringiensis recombinant strain that has high expression of AiiA protein, the promoter of insecticidal crystal protein coding gene cry3Aa of B. thuringiensis was selected. The promoter of gene cry3Aa is a non-sporulation promoter, it promotes the transcription earlier and longer than the promoters of other cry genes. The promoter of AiiA protein coding gene aiiA was replaced with the promoter of gene cry3Aa by overlapping PCR, resulting fusion gene pro3A-aiiA. The gene pro3A-aiiA was inserted into shuttle vector pHT304 at site BamH I / Sph I , resulting recombinant plasmid pBMB686. The plasmid pBMB686 was introduced into B. thuringiensis acrystalliferous strain BMB171, the resulting strain BMB686 had a higher and more stable expression level of protein AiiA comparing with the parental strain BMB171. Furthermore, the strain BMB686 exhibited stronger ability of AHLs inactivation and much more effective restraint to the potato's soft rot disease caused by Erwinia

  2. Identification and analysis of the salt tolerant property of AHL lactonase (AiiATSAWB ) of Bacillus species.

    PubMed

    Easwaran, Nalini; Karthikeyan, Sivashanmugam; Sridharan, Balasundaram; Gothandam, Kodiveri Muthukaliannan

    2015-05-01

    Bacterial biofilms communicate by a process called Quorum Sensing. Gram negative bacterial pathogens specifically talk through the production, detection, and response to the signal or autoinducer called Acyl Homoserine Lactones. Bacterial lactonases are important AHL hydrolysing or quorum quenching enzymes. The present study deals with ten endospore forming gram positive isolates of the saltern soil. Preliminary screening for Quorum Quenching activity with the QS Inhibition indicator strain Chromobacterium violaceum ATCC 12472, showed positive activity in four isolates namely TS2, TS16, TSAWB, and TS53B. AHL lactonase (AiiA) specific primers amplified Acyl Homoserine Lactone lactonase gene in the TSAWB genome alone. Phylogenetic relationship of the identified AiiATSAWB confirmed its evolutionary relationship with bacterial AiiA like AHL lactonase of the metallo-beta-lactamase super family. Our in vitro AHL hydrolysis assay under wide percentage (0-5) of salt solutions with TSAWB isolate and also its intracellular soluble protein fraction showed halotolerant AHL hydrolysis ability of the AiiATSAWB enzyme. In silico determination of putative tertiary structure, the ESBRI derived conserved salt bridges, aminoacid residue characterization with high mole percent of acidic and hydrophobic residues reaffirmed the halotolerant ability of the enzyme. So we propound the future use of purified AiiATSAWB , as hypertonic suspension for inhalation to substitute the action of inactivated host's paraoxonase in treating Pseudomonas aeruginosa infection in cystic fibrosis patients. PMID:25041996

  3. Localization of Neuropeptide Y1 Receptor Immunoreactivity in the Rat Retina and the Synaptic Connectivity of Y1 Immunoreactive Cells

    PubMed Central

    D'Angelo, Iona; Oh, Su-Ja; Chun, Myung-Hoon; Brecha, Nicholas C.

    2010-01-01

    Neuropeptide Y (NPY), an inhibitory neuropeptide expressed by a moderately dense population of wide-field amacrine cells in the rat retina, acts through multiple (Y1–y6) G-protein–coupled receptors. This study determined the cellular localization of Y1 receptors and the synaptic connectivity of Y1 processes in the inner plexiform layer (IPL) of the rat retina. Specific Y1 immunoreactivity was localized to horizontal cell bodies in the distal inner nuclear layer and their processes in the outer plexiform layer. Immunoreactivity was also prominent in cell processes located in strata 2 and 4, and puncta in strata 4 and 5 of the IPL. Double-label immunohistochemical experiments with calbindin, a horizontal cell marker, confirmed Y1 immunostaining in all horizontal cells. Double-label immunohistochemical experiments, using antibodies to choline acetyltransferase and vesicular acetylcholine transporter to label cholinergic amacrine cell processes, demonstrated that Y1 immunoreactivity in strata 2 and 4 of the IPL was localized to cholinergic amacrine cell processes. Electron microscopic studies of the inner retina showed that Y1-immunostained amacrine cell processes and puncta received synaptic inputs from unlabeled amacrine cell processes (65.2%) and bipolar cell axon terminals (34.8%). Y1-immunoreactive amacrine cell processes most frequently formed synaptic outputs onto unlabeled amacrine cell processes (34.0%) and ganglion cell dendrites (54.1%). NPY immunoreactivity in the rat retina is distributed primarily to strata 1 and 5 of the IPL, and the present findings, thus, suggest that NPY acts in a paracrine manner on Y1 receptors to influence both horizontal and amacrine cells. PMID:12455004

  4. Multilayer models in the piezo-PAS analysis of AII-BVI compounds

    NASA Astrophysics Data System (ADS)

    Maliński, M.; Zakrzewski, J.

    2003-01-01

    This article presents the results of computations of the photoacoustic piezoelectric spectra, both amplitude and phase, of a series of AII-BVI compounds and their comparison with experimental results. For the interpretation of experimental spectra several multilayer models were developed and applied. Computer analysis showed that it was not possible to interpret the experimental results in the model of a single layer in the case of real samples. In this article three multilayer models are presented and illustrated by experimental results and numerical characteristics.

  5. 26 CFR 1.1092(b)-3T - Mixed straddles; straddle-by-straddle identification under section 1092(b)(2)(A)(i)(I) (temporary).

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... identification under section 1092(b)(2)(A)(i)(I) (temporary). 1.1092(b)-3T Section 1.1092(b)-3T Internal Revenue... section 1092(b)(2)(A)(i)(I) (temporary). (a) In general. Except as otherwise provided, a taxpayer shall... constitute independent verification: (i) Separate account. Placement of one or more positions of a......

  6. Transient release kinetics of rod bipolar cells revealed by capacitance measurement of exocytosis from axon terminals in rat retinal slices.

    PubMed

    Oltedal, Leif; Hartveit, Espen

    2010-05-01

    Presynaptic transmitter release has mostly been studied through measurements of postsynaptic responses, but a few synapses offer direct access to the presynaptic terminal, thereby allowing capacitance measurements of exocytosis. For mammalian rod bipolar cells, synaptic transmission has been investigated in great detail by recording postsynaptic currents in AII amacrine cells. Presynaptic measurements of the dynamics of vesicular cycling have so far been limited to isolated rod bipolar cells in dissociated preparations. Here, we first used computer simulations of compartmental models of morphologically reconstructed rod bipolar cells to adapt the 'Sine + DC' technique for capacitance measurements of exocytosis at axon terminals of intact rod bipolar cells in retinal slices. In subsequent physiological recordings, voltage pulses that triggered presynaptic Ca(2+) influx evoked capacitance increases that were proportional to the pulse duration. With pulse durations 100 ms, the increase saturated at 10 fF, corresponding to the size of a readily releasable pool of vesicles. Pulse durations 400 ms evoked additional capacitance increases, probably reflecting recruitment from additional pools of vesicles. By using Ca(2+) tail current stimuli, we separated Ca(2+) influx from Ca(2+) channel activation kinetics, allowing us to estimate the intrinsic release kinetics of the readily releasable pool, yielding a time constant of 1.1 ms and a maximum release rate of 2-3 vesicles (release site)(1) ms(1). Following exocytosis, we observed endocytosis with time constants ranging from 0.7 to 17 s. Under physiological conditions, it is likely that release will be transient, with the kinetics limited by the activation kinetics of the voltage-gated Ca(2+) channels. PMID:20211976

  7. Structure And Specificity of a Quorum-Quenching Lactonase (AiiB) From Agrobacterium Tumefaciens

    SciTech Connect

    Liu, D.; Thomas, P.W.; Momb, J.; Hoang, Q.Q.; Petsko, G.A.; Ringe, D.; Fast, W.

    2009-06-03

    N-Acyl-l-homoserine lactone (AHL) mediated quorum-sensing regulates virulence factor production in a variety of Gram-negative bacteria. Proteins capable of degrading these autoinducers have been called 'quorum-quenching' enzymes, can block many quorum-sensing dependent phenotypes, and represent potentially useful reagents for clinical, agricultural, and industrial applications. The most characterized quorum-quenching enzymes to date are the AHL lactonases, which are metalloproteins that belong to the metallo-beta-lactamase superfamily. Here, we report the cloning, heterologous expression, purification, metal content, substrate specificity, and three-dimensional structure of AiiB, an AHL lactonase from Agrobacterium tumefaciens. Much like a homologous AHL lactonase from Bacillus thuringiensis, AiiB appears to be a metal-dependent AHL lactonase with broad specificity. A phosphate dianion is bound to the dinuclear zinc site and the active-site structure suggests specific mechanistic roles for an active site tyrosine and aspartate. To our knowledge, this is the second representative structure of an AHL lactonase and the first of an AHL lactonase from a microorganism that also produces AHL autoinducers. This work should help elucidate the hydrolytic ring-opening mechanism of this family of enzymes and also facilitate the design of more effective quorum-quenching catalysts.

  8. Internalization and lysosomal association of (/sup 125/I)angiotensin II in norepinephrine-containing cells of the rat adrenal medulla

    SciTech Connect

    Bianchi, C.; Gutkowska, J.; Charbonneau, C.; Ballak, M.; Anand-Srivastava, M.B.; De Lean, A.; Genest, J.; Cantin, M.

    1986-10-01

    The morphological localization of (/sup 125/I)angiotensin II (AII) in the rat adrenal medulla (AM) was studied by light- and electron-microscopic radioautography in vivo. With light microscopy the presence of binding sites for AII in both norepinephrine-containing (NE) and epinephrine-containing (E) cells was confirmed. With electron microscopy, it was found that AII binds to the cell surface of NE cells, is progressively internalized, and is associated with lysosomes and Golgi complex within 20 min, whereas in E cells AII seems to be internalized earlier and recycled back to the cell surface within 5 min without any appreciable association with intracellular organelles. These results suggest different intracellular pathways for AII in NE and E cells of the rat AM.

  9. 78 FR 64430 - Mixed Straddles; Straddle-by-Straddle Identification Under Section 1092(b)(2)(A)(i)(I); Correction

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-29

    ... Under Section 1092(b)(2)(A)(i)(I); Correction AGENCY: Internal Revenue Service (IRS), Treasury. ACTION...), published in the Federal Register on Friday, August 2, 2013 (78 FR 46807), serves as the text of the... Friday, August 2, 2013 (78 FR 46854). The proposed regulations were proposed to apply to all...

  10. 78 FR 46807 - Mixed Straddles; Straddle-by-Straddle Identification Under Section 1092(b)(2)(A)(i)(I)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-02

    ... rulemaking by cross-reference to temporary regulations (50 FR 3351, January 24, 1985). Included in the... Internal Revenue Service 26 CFR Part 1 RIN 1545-BL04 Mixed Straddles; Straddle-by-Straddle Identification Under Section 1092(b)(2)(A)(i)(I) AGENCY: Internal Revenue Service (IRS), Treasury. ACTION:...

  11. Luminescence processes in A3IILaNb3O12 (AII = Ba, Sr) layered perovskites

    NASA Astrophysics Data System (ADS)

    Chukova, Oksana; Gomenyuk, Olga; Nedilko, Sergiy; Polubinskii, Vitaliy; Scherbatsky, Vasyl; Sheludko, Vadim; Titov, Yuriy

    2014-08-01

    The A3IILaNb3O12 (AII = Sr, Ba) layered perovskites were synthesized by co-precipitation method. Luminescence properties of these compositions have been investigated for the first time. Emission spectra consist of wide bands with maxima located near 430 and 500 nm. Luminescence processes are discussed in connection with features of cation compositions, crystal structure and distortions of the NbO67- groups’ symmetry. Possible scheme of electron states and the lowest allowed electron transitions has been built for the niobate groups of Oh and C3v symmetry. In this scheme, the 430 and 500 nm emission bands were assigned to T1 → A1 and E → A1 electron transitions in the perfect and distorted NbO67- oxyanions, respectively.

  12. Inhibition to retinal rod bipolar cells is regulated by light levels

    PubMed Central

    Mazade, Reece E.; Klein, Justin S.

    2013-01-01

    The retina responds to a wide range of light stimuli by adaptation of retinal signaling to background light intensity and the use of two different photoreceptors: rods that sense dim light and cones that sense bright light. Rods signal to rod bipolar cells that receive significant inhibition from amacrine cells in the dark, especially from a rod bipolar cell-activated GABAergic amacrine cell. This inhibition modulates the output of rod bipolar cells onto downstream neurons. However, it was not clear how the inhibition of rod bipolar cells changes when rod signaling is limited by an adapting background light and cone signaling becomes dominant. We found that both light-evoked and spontaneous rod bipolar cell inhibition significantly decrease with light adaptation. This suggests a global decrease in the activity of amacrine cells that provide input to rod bipolar cells with light adaptation. However, inhibition to rod bipolar cells is also limited by GABAergic connections between amacrine cells, which decrease GABAergic input to rod bipolar cells. When we removed this serial inhibition, the light-evoked inhibition to rod bipolar cells remained after light adaptation. These results suggest that decreased inhibition to rod bipolar cells after light adaptation is due to decreased rod pathway activity as well as an active increase in inhibition between amacrine cells. Together these serve to limit rod bipolar cell inhibition after light adaptation, when the rod pathway is inactive and modulation of the signal is not required. This suggests an efficiency mechanism in the retina to limit unnecessary signaling. PMID:23596335

  13. Nosocomial Infections after Severe Trauma Are Associated With Lower Apolipoproteins B and AII

    PubMed Central

    Femling, Jon K.; West, Sonlee D.; Hauswald, Erik K.; Gresham, Hattie D.; Hall, Pamela R.

    2014-01-01

    BACKGROUND Infection following severe trauma is a significant cause of morbidity and mortality days to weeks after the initial injury. Apolipoproteins play important roles in host defense and circulating concentrations are altered by the acute inflammatory response. The purpose of this study was to determine if patients who acquire infection following severe trauma have significantly lower apolipoprotein levels than trauma patients who do not become infected. METHODS We conducted a case-control study on a prospectively identified cohort of adult patients admitted to our intensive care unit after severe trauma (Injury Severity Score ≥ 16). We compared plasma apolipoprotein levels between patients who acquired an infection within 30 days after trauma (cases) and those that remained infection free (controls). RESULTS Of 40 patients experiencing severe trauma, we identified 22 cases that developed an infection within 30 days after injury. Cases had significantly lower post-trauma plasma levels of apolipoprotein-B (p=0.02) and apolipoprotein-AII (p=0.02) compared to controls. Consistent with previous studies, cases also received greater volumes of crystalloid infusions (p<0.01) and blood transfusions (p<0.01). Cases also had a more profound inflammatory response as measured by Interleukin-6 levels (p=0.02). CONCLUSIONS Infection after severe trauma is associated with decreased circulating apolipoproteins as compared to uninfected controls. Profoundly decreased plasma apolipoproteins B and AII could potentially contribute to the impaired immunity after severe trauma. Apolipoproteins are potential targets for identifying those patients at risk of infection after trauma and for interventions aimed at preventing nosocomial infections. PMID:23511146

  14. Detection of angiotensin II binding to single adrenal zona glomerulosa cells by confocal Raman microspectroscopy

    NASA Astrophysics Data System (ADS)

    McCoy, Michael J.; Habermann, Timothy J.; Hanke, Craig J.; Adar, Fran; Campbell, William B.; Nithipatikom, Kasem

    1999-04-01

    We developed a confocal Raman microspectroscopic technique to study ligand-receptor bindings in single cells using Raman-labeled ligands and surface-enhanced Raman scattering (SERS). The adrenal zona glomerulosa (ZG) cells were used as a model in this study. ZG cells have a high density of angiotensin II (AII) receptors on the cellular membrane. There are two identified subtypes of AII receptors,namely AT1 and AT2 receptors. AII is a peptidic hormone, which upon binding to its receptors, stimulates the release of aldosterone from ZG cells. The cellular localization of these receptors subtypes was detected in single ZG cells by using immunocomplexation of receptors with specific antibodies and confocal Raman microspectroscopy. In the binding study, we used biotin-labeled AII to bind to its receptors in ZG cells. Then, avidin and Raman-labeled AII. The binding was measure directly on the single ZG cells. The results showed that the binding was displaced with unlabeled AII and specific AII antagonists. This is a rapid and sensitive technique for detection of cellular ligand bindings as well as antagonists screening in drug discovery.

  15. N-ACYL HOMOSERINE LACTONe LACTONASE, AiiA, INACTIVATION OF QUORUM-SENSING AGONISTS PRODUCED BY CHLAMYDOMONAS REINHARDTII (CHLOROPHYTA) AND CHARACTERIZATION OF aiiA TRANSGENIC ALGAE(1).

    PubMed

    Rajamani, Sathish; Teplitski, Max; Kumar, Anil; Krediet, Cory J; Sayre, Richard T; Bauer, Wolfgang D

    2011-10-01

    Eukaryotes such as plants and the unicellular green alga Chlamydomonas reinhardtii P. A. Dang. produce and secrete compounds that mimic N-acyl homoserine lactone (AHL) bacterial quorum-sensing (QS) signals and alter QS-regulated gene expression in the associated bacteria. Here, we show that the set of C. reinhardtii signal-mimic compounds that activate the CepR AHL receptor of Burkholderia cepacia are susceptible to inactivation by AiiA, an AHL lactonase enzyme of Bacillus. Inactivation of these algal mimics by AiiA suggests that the CepR-stimulatory class of mimics produced by C. reinhardtii may have a conserved lactone ring structure in common with AHL QS signals. To examine the role of AHL mimic compounds in the interactions of C. reinhardtii with bacteria, the aiiA gene codon optimized for Chlamydomonas was generated for the expression of AiiA as a chimeric fusion with cyan fluorescent protein (AimC). Culture filtrates of transgenic strains expressing the fusion protein AimC had significantly reduced levels of CepR signal-mimic activities. When parental and transgenic algae were cultured with a natural pond water bacterial community, a morphologically distinct, AHL-producing isolate of Aeromonas veronii was observed to colonize the transgenic algal cultures and form biofilms more readily than the parental algal cultures, indicating that secretion of the CepR signal mimics by the alga can significantly affect its interactions with bacteria it encounters in natural environments. The parental alga was also able to sequester and/or destroy AHLs in its growth media to further disrupt or manipulate bacterial QS. PMID:27020200

  16. Specific action of the lipoxygenase pathway in mediating angiotensin II-induced aldosterone synthesis in isolated adrenal glomerulosa cells.

    PubMed Central

    Nadler, J L; Natarajan, R; Stern, N

    1987-01-01

    Angiotensin II (AII) in adrenal glomerulosa cells activates phospholipase C resulting in the formation of inositol phosphates and diacylglycerol rich in arachidonic acid (AA). Although glomerulosa cells can metabolize AA via cyclooxygenase (CO), this pathway plays little role in aldosterone synthesis. Recent evidence suggests that the lipoxygenase (LO) pathway may be important for hormonal secretion in endocrine tissues such as the islet of Langerhans. However, the capacity of the glomerulosa cell to synthesize LO products and their role in aldosterone secretion is not known. To study this, the effect of nonselective and selective LO inhibitors on AII, ACTH, and potassium-induced aldosterone secretion and LO product formation was evaluated in isolated rat glomerulosa cells. BW755c, a nonselective LO inhibitor dose dependently reduced the AII-stimulated level of aldosterone without altering AII binding (91 +/- 6 to 36 +/- 4 ng/10(6) cells/h 10(-4) M, P less than 0.001). The same effect was observed with another nonselective LO blocker, phenidone, and a more selective 12-LO inhibitor, Baicalein. In contrast U-60257, a selective 5-LO inhibitor did not change the AII-stimulated levels of aldosterone (208 +/- 11% control, AII 10(-9) M vs. 222 +/- 38%, AII + U-60257). The LO blockers action was specific for AII since neither BW755c nor phenidone altered ACTH or K+-induced aldosterone secretion. AII stimulated the formation of the 12-LO product 12-hydroxyeicosatetraenoic acid (12-HETE) as measured by ultraviolet detection and HPLC in AA loaded cells and by a specific RIA in unlabeled cells (501 +/- 50 to 990 +/- 10 pg/10(5) cells, P less than 0.02). BW755c prevented the AII-mediated rise in 12-HETE formation. In contrast, neither ACTH nor K+ increased 12-HETE levels. The addition of 12-HETE or its unstable precursor 12-HPETE (10(-9) or 10(-8) M) completely restored AII action during LO blockade. AII also produced an increase in 15-HETE formation, but the 15-LO products

  17. Expression and Purification of Recombinant Human Apolipoprotein A-II in Pichia pastoris

    PubMed Central

    Su, Manman; Qi, Yitian; Wang, Mingxing; Chang, Weiqin; Peng, Shuang; Wang, Dingding

    2013-01-01

    Abstract Apolipoprotein A-II (ApoA-II) is the second most abundant protein constituent of high-density lipoprotein (HDL). The physiologic role of ApoA-II is poorly defined. ApoA-II may inhibit lecithin:cholesterol acyltransferase and cholesteryl-ester-transfer protein activities, but may increase the hepatic lipase activity. ApoA-II may also inhibit the hepatic cholesteryl uptake from HDL probably through the scavenger receptor class B type I depending pathway. Interpretation of data from transgenic and knockout mice of genes involved in lipoprotein metabolism has been often complicated as clinical implications because of species difference. So it is important to obtain human ApoA-II for further studies about its functions. In our studies, Pichia pastoris expression system was first used to express a high-level secreted recombinant human ApoA-II (rhApoA-II). We have cloned the cDNA encoding human ApoA-II and achieved its high-level secreting expression with a yield of 65 mg/L of yeast culture and the purification process was effective and easy to handle. The purified rhApoA-II can be used to further study its biological activities. PMID:24116940

  18. Methylation of 23S rRNA Nucleotide G748 by RlmAII Methyltransferase Renders Streptococcus pneumoniae Telithromycin Susceptible

    PubMed Central

    Sato, Yoshiharu; Shoji, Tatsuma; Yamamoto, Tomoko

    2013-01-01

    Several posttranscriptional modifications of bacterial rRNAs are important in determining antibiotic resistance or sensitivity. In all Gram-positive bacteria, dimethylation of nucleotide A2058, located in domain V of 23S rRNA, by the dimethyltransferase Erm(B) results in low susceptibility and resistance to telithromycin (TEL). However, this is insufficient to produce high-level resistance to TEL in Streptococcus pneumoniae. Inactivation of the methyltransferase RlmAII, which methylates the N-1 position of nucleotide G748, located in hairpin 35 of domain II of 23S rRNA, results in increased resistance to TEL in erm(B)-carrying S. pneumoniae. Sixteen TEL-resistant mutants (MICs, 16 to 32 μg/ml) were obtained from a clinically isolated S. pneumoniae strain showing low TEL susceptibility (MIC, 2 μg/ml), with mutation resulting in constitutive dimethylation of A2058 because of nucleotide differences in the regulatory region of erm(B) mRNA. Primer extension analysis showed that the degree of methylation at G748 in all TEL-resistant mutants was significantly reduced by a mutation in the gene encoding RlmAII to create a stop codon or change an amino acid residue. Furthermore, RNA footprinting with dimethyl sulfate and a molecular modeling study suggested that methylation of G748 may contribute to the stable interaction of TEL with domain II of 23S rRNA, even after dimethylation of A2058 by Erm(B). This novel finding shows that methylation of G748 by RlmAII renders S. pneumoniae TEL susceptible. PMID:23716046

  19. Oxidation of high density lipoproteins. II. Evidence for direct reduction of lipid hydroperoxides by methionine residues of apolipoproteins AI and AII.

    PubMed

    Garner, B; Waldeck, A R; Witting, P K; Rye, K A; Stocker, R

    1998-03-13

    Human high density lipoproteins (HDL) can reduce cholesteryl ester hydroperoxides to the corresponding hydroxides (Sattler W., Christison J. K., and Stocker, R. (1995) Free Radical Biol. & Med. 18, 421-429). Here we demonstrate that this reducing activity extended to hydroperoxides of phosphatidylcholine, was similar in HDL2 and HDL3, was independent of arylesterase and lecithin:cholesteryl acyltransferase activity, was unaffected by sulfhydryl reagents, and was expressed by reconstituted particles containing apoAI or apoAII only, as well as isolated human apoAI. Concomitant with the reduction of lipid hydroperoxides specific oxidized forms of apoAI and apoAII formed in blood-derived and reconstituted HDL. Similarly, specific oxidized forms of apoAI accumulated upon treatment of isolated apoAI with authentic cholesteryl linoleate hydroperoxide. These specific oxidized forms of apoAI and apoAII have been shown previously to contain Met sulfoxide (Met(O)) at Met residues and are also formed when HDL is exposed to Cu2+ or soybean lipoxygenase. Lipid hydroperoxide reduction and the associated formation of specific oxidized forms of apoAI and apoAII were inhibited by solubilizing HDL with SDS or by pretreatment of HDL with chloramine T. The inhibitory effect of chloramine T was dose-dependent and accompanied by the conversion of specific Met residues of apoAI and apoAII into Met(O). Canine HDL, which contains apoAI as the predominant apolipoprotein and which lacks the oxidation-sensitive Met residues Met112 and Met148, showed much weaker lipid hydroperoxide reducing activity and lower extents of formation of oxidized forms of apoAI than human HDL. We conclude that the oxidation of specific Met residues of apoAI and apoAII to Met(O) plays a significant role in the 2-electron reduction of hydroperoxides of cholesteryl esters and phosphatidylcholine associated with human HDL. PMID:9497326

  20. Apolipoprotein A-II is a key regulatory factor of HDL metabolism as appears from studies with transgenic animals and clinical outcomes.

    PubMed

    Maïga, Sira Fatoumata; Kalopissis, Athina-Despina; Chabert, Michèle

    2014-01-01

    The structure and metabolism of HDL are linked to their major apolipoproteins (apo) A-I and A-II. HDL metabolism is very dynamic and depends on the constant remodeling by lipases, lipid transfer proteins and receptors. HDL exert several cardioprotective effects, through their antioxidant and antiinflammatory capacities and through the stimulation of reverse cholesterol transport from extrahepatic tissues to the liver for excretion into bile. HDL also serve as plasma reservoir for C and E apolipoproteins, as transport vehicles for a great variety of proteins, and may have more physiological functions than previously recognized. In this review we will develop several aspects of HDL metabolism with emphasis on the structure/function of apo A-I and apo A-II. An important contribution to our understanding of the respective roles of apo A-I and apo A-II comes from studies using transgenic animal models that highlighted the stabilizatory role of apo A-II on HDL through inhibition of their remodeling by lipases. Clinical studies coupled with proteomic analyses revealed the presence of dysfunctional HDL in patients with cardiovascular disease. Beyond HDL cholesterol, a new notion is the functionality of HDL particles. In spite of abundant literature on HDL metabolic properties, a major question remains unanswered: which HDL particle(s) confer(s) protection against cardiovascular risk? PMID:24012775

  1. 49 CFR Appendix A-Ii to Part 541 - Lines With Antitheft Devices Which Are Exempted In-Part From the Parts-Marking Requirements of...

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 6 2012-10-01 2012-10-01 false Lines With Antitheft Devices Which Are Exempted In-Part From the Parts-Marking Requirements of This Standard Pursuant to 49 CFR Part 543 A Appendix A-II to Part 541 Transportation Other Regulations Relating to Transportation (Continued) NATIONAL HIGHWAY TRAFFIC SAFETY ADMINISTRATION, DEPARTMENT...

  2. Interactions of Apolipoproteins AI, AII, B and HDL, LDL, VLDL with Polyurethane and Polyurethane-PEO Surfaces.

    PubMed

    Cornelius, R M; Macri, J; Cornelius, K M; Brash, J L

    2015-11-10

    The lipoproteins (HDL, LDL, VLDL) are important components of blood present in high concentration. Surprisingly, their role in blood-biomaterial interactions has been largely ignored. In previous work apolipoprotein AI (the main protein component of HDL) was identified as a major constituent of protein layers adsorbed from plasma to biomaterials having a wide range of surface properties, and quantitative data on the adsorption of apo AI to a biomedical grade polyurethane were reported. In the present communication quantitative data on the adsorption of apo AI, apo AII and apoB (the latter being a constituent of LDL and VLDL), as well as the lipoprotein particles themselves (HDL, LDL, VLDL), to a biomedical segmented polyurethane (PU) with and without an additive containing poly(ethylene oxide) (material referred to as PEO) are reported. Using radiolabeled apo AI, apo AII, and apoB, adsorption levels on PU from buffer at a protein concentration of 50 μg/mL were found to be 0.34, 0.40, and 0.14 μg/cm(2) (12, 23, and 0.25 nmol/cm(2)) respectively. Adsorption to the PEO surface was <0.02 μg/cm(2) for all three apolipoproteins demonstrating the strong protein resistance of this material. In contrast to the apolipoproteins, significant amounts of the lipoproteins were found to adsorb to the PEO as well as to the PU surface. X-ray photoelectron spectra, following exposure of the surfaces to the lipoproteins, showed a strong phosphorus signal, confirming that adsorption had occurred. It therefore appears that a PEO-containing surface that is resistant to apolipoproteins may be less resistant to the corresponding lipoproteins. PMID:26513526

  3. Altered retinal cell differentiation in the AP-3 delta mutant (Mocha) mouse.

    PubMed

    Baguma-Nibasheka, Mark; Kablar, Boris

    2009-11-01

    Adaptor-related protein complex 3 delta 1 (Ap3d1) encodes the delta 1 subunit of an adaptor protein regulating intracellular vesicle-mediated transport, and the Ap3d-deletion mutant (Mocha) mouse undergoes rapid photoreceptor degeneration leading to blindness soon after birth. Previous microarray analysis revealed Ap3d down-regulation in the retina of mouse embryos specifically lacking cholinergic amacrine cells as a result of the absence of skeletal musculature. To investigate the role of Ap3d in the determination of retinal cell fate, the present study examined retinal morphology in newborn Ap3d-/- mice. The Ap3d-/- retina showed a complete absence of cholinergic amacrine cells and a decrease in parvalbumin-expressing amacrine cells and syntaxin- and VC1.1-expressing amacrine precursor cells, but had a normal number of cell layers and number of cells in each layer with no detectable difference in cell proliferation or apoptosis. These findings indicate that, despite having no apparent effect on the basic spatial organization of the retina at this stage of development, Ap3d could be involved in the regulation of progenitor cell competence and the eventual ratio of resulting differentiated cells. Finding the mouse mutant which phenocopies the eye defect seen in fetuses with no striated muscle was accomplished by the Systematic Subtractive Microarray Analysis Approach (SSMAA), explained in the discussion section. PMID:19631730

  4. Regulation of ERK5 by insulin and angiotensin-II in vascular smooth muscle cells

    SciTech Connect

    Sharma, Girish; Goalstone, Marc Lee; E-mail: Marc.Goalstone@uchsc.edu

    2007-03-23

    ERK5 is involved in proliferation of vascular smooth muscle cells (VSMC). The proliferative actions of insulin and angiotensin-II (A-II) in VSMC are mediated in part by ERK1/2. We hypothesized that insulin and A-II also regulate ERK5 activity in VSMC. Acute treatment (<60 min) with insulin or A-II increased phosphorylation of ERK1/2 at 15 min and ERK5 at 5 min. Chronic treatment ({<=}8 h) with insulin increased ERK1/2 phosphorylation by 4 h and ERK5 by 8 h. A-II-stimulated phosphorylation of ERK1/2 by 8 h and ERK5 by 4 h. The EC{sub 50} for insulin treatment effecting ERK1/2 and ERK5 phosphorylation was 1.5 and 0.1 nM, whereas the EC{sub 50} for A-II was 2 nM, each. Insulin plus A-II induced an additive effect only on ERK5 phosphorylation. Inhibition of insulin- and A-II-stimulated phosphorylation of ERK5 and ERK1/2 by PD98059 and Wortmannin exhibited differential and time-dependent effects. Taken together, these data indicate that insulin and A-II regulate the activity of ERK5, but different from that seen for ERK1/2.

  5. Fat3 and Ena/VASP proteins influence the emergence of asymmetric cell morphology in the developing retina.

    PubMed

    Krol, Alexandra; Henle, Steven J; Goodrich, Lisa V

    2016-06-15

    Neurons exhibit asymmetric morphologies throughout development - from migration to the elaboration of axons and dendrites - that are correctly oriented for the flow of information. For instance, retinal amacrine cells migrate towards the inner plexiform layer (IPL) and then retract their trailing processes, thereby acquiring a unipolar morphology with a single dendritic arbor restricted to the IPL. Here, we provide evidence that the Fat-like cadherin Fat3 acts during multiple stages of amacrine cell development in mice to orient overall changes in cell shape towards the IPL. Using a time-lapse imaging assay, we found that developing amacrine cells are less directed towards the IPL in the absence of Fat3, during both migration and retraction. Consistent with its predicted role as a cell-surface receptor, Fat3 functions cell-autonomously and is able to influence the cytoskeleton directly through its intracellular domain, which can bind and localize Ena/VASP family actin regulators. Indeed, a change in Ena/VASP protein distribution is sufficient to recapitulate the Fat3 mutant amacrine cell phenotype. Thus, Fat-like proteins might control the polarized development of tissues by sculpting the cytoskeleton of individual cells. PMID:27122175

  6. RlmCD-mediated U747 methylation promotes efficient G748 methylation by methyltransferase RlmAII in 23S rRNA in Streptococcus pneumoniae; interplay between two rRNA methylations responsible for telithromycin susceptibility

    PubMed Central

    Shoji, Tatsuma; Takaya, Akiko; Sato, Yoshiharu; Kimura, Satoshi; Suzuki, Tsutomu; Yamamoto, Tomoko

    2015-01-01

    Adenine at position 752 in a loop of helix 35 from positions 745 to 752 in domain II of 23S rRNA is involved in binding to the ribosome of telithromycin (TEL), a member of ketolides. Methylation of guanine at position 748 by the intrinsic methyltransferase RlmAII enhances binding of telithromycin (TEL) to A752 in Streptococcus pneumoniae. We have found that another intrinsic methylation of the adjacent uridine at position 747 enhances G748 methylation by RlmAII, rendering TEL susceptibility. U747 and another nucleotide, U1939, were methylated by the dual-specific methyltransferase RlmCD encoded by SP_1029 in S. pneumoniae. Inactivation of RlmCD reduced N1-methylated level of G748 by RlmAII in vivo, leading to TEL resistance when the nucleotide A2058, located in domain V of 23S rRNA, was dimethylated by the dimethyltransferase Erm(B). In vitro methylation of rRNA showed that RlmAII activity was significantly enhanced by RlmCD-mediated pre-methylation of 23S rRNA. These results suggest that RlmCD-mediated U747 methylation promotes efficient G748 methylation by RlmAII, thereby facilitating TEL binding to the ribosome. PMID:26365244

  7. Bioinformatic Analysis of Plasma Apolipoproteins A-I and A-II Revealed Unique Features of A-I/A-II HDL Particles in Human Plasma

    PubMed Central

    Kido, Toshimi; Kurata, Hideaki; Kondo, Kazuo; Itakura, Hiroshige; Okazaki, Mitsuyo; Urata, Takeyoshi; Yokoyama, Shinji

    2016-01-01

    Plasma concentration of apoA-I, apoA-II and apoA-II-unassociated apoA-I was analyzed in 314 Japanese subjects (177 males and 137 females), including one (male) homozygote and 37 (20 males and 17 females) heterozygotes of genetic CETP deficiency. ApoA-I unassociated with apoA-II markedly and linearly increased with HDL-cholesterol, while apoA-II increased only very slightly and the ratio of apoA-II-associated apoA-I to apoA-II stayed constant at 2 in molar ratio throughout the increase of HDL-cholesterol, among the wild type and heterozygous CETP deficiency. Thus, overall HDL concentration almost exclusively depends on HDL with apoA-I without apoA-II (LpAI) while concentration of HDL containing apoA-I and apoA-II (LpAI:AII) is constant having a fixed molar ratio of 2 : 1 regardless of total HDL and apoA-I concentration. Distribution of apoA-I between LpAI and LpAI:AII is consistent with a model of statistical partitioning regardless of sex and CETP genotype. The analysis also indicated that LpA-I accommodates on average 4 apoA-I molecules and has a clearance rate indistinguishable from LpAI:AII. Independent evidence indicated LpAI:A-II has a diameter 20% smaller than LpAI, consistent with a model having two apoA-I and one apoA-II. The functional contribution of these particles is to be investigated. PMID:27526664

  8. Bioinformatic Analysis of Plasma Apolipoproteins A-I and A-II Revealed Unique Features of A-I/A-II HDL Particles in Human Plasma.

    PubMed

    Kido, Toshimi; Kurata, Hideaki; Kondo, Kazuo; Itakura, Hiroshige; Okazaki, Mitsuyo; Urata, Takeyoshi; Yokoyama, Shinji

    2016-01-01

    Plasma concentration of apoA-I, apoA-II and apoA-II-unassociated apoA-I was analyzed in 314 Japanese subjects (177 males and 137 females), including one (male) homozygote and 37 (20 males and 17 females) heterozygotes of genetic CETP deficiency. ApoA-I unassociated with apoA-II markedly and linearly increased with HDL-cholesterol, while apoA-II increased only very slightly and the ratio of apoA-II-associated apoA-I to apoA-II stayed constant at 2 in molar ratio throughout the increase of HDL-cholesterol, among the wild type and heterozygous CETP deficiency. Thus, overall HDL concentration almost exclusively depends on HDL with apoA-I without apoA-II (LpAI) while concentration of HDL containing apoA-I and apoA-II (LpAI:AII) is constant having a fixed molar ratio of 2 : 1 regardless of total HDL and apoA-I concentration. Distribution of apoA-I between LpAI and LpAI:AII is consistent with a model of statistical partitioning regardless of sex and CETP genotype. The analysis also indicated that LpA-I accommodates on average 4 apoA-I molecules and has a clearance rate indistinguishable from LpAI:AII. Independent evidence indicated LpAI:A-II has a diameter 20% smaller than LpAI, consistent with a model having two apoA-I and one apoA-II. The functional contribution of these particles is to be investigated. PMID:27526664

  9. Regulation of corticotropin receptor number and messenger RNA in cultured human adrenocortical cells by corticotropin and angiotensin II.

    PubMed Central

    Lebrethon, M C; Naville, D; Begeot, M; Saez, J M

    1994-01-01

    The regulation of ACTH receptor binding sites and mRNA by ACTH and angiotensin II (A-II) was studied using cultured human adrenal fasciculata reticularis cells (HAC). These cells expressed two major ACTH receptor transcripts of 1.8 and 3.4 kb and three minor ones of 4, 7, and 11 kb. ACTH increased the levels of all these transcripts in a time- and dose-dependent manner. At a maximal concentration of 10(-8) M, ACTH enhanced 21- and 4-fold the level of ACTH receptor mRNA and the number of receptors per cell, respectively. Pretreatment of HAC with A-II produced a dose-dependent enhancement of ACTH receptor mRNA that was associated with an increase of both ACTH receptor number and responsiveness to this hormone. The effects of A-II were completely blocked by an AT1 receptor subtype antagonist but not by an AT2 antagonist. The effects of ACTH together with A-II on ACTH receptor mRNA were greater than those induced by each hormone alone. These results show that ACTH receptor number and mRNA are positively regulated by the two main hormones (ACTH and A-II) which, in vivo, regulate adrenocortical functions. In addition, they also show that HAC are a target for A-II. Thus, regulation of ACTH receptors may be one mechanism by which ACTH and A-II regulate adrenocortical functions under both normal and pathological conditions. Images PMID:8163681

  10. Zac1 functions through TGFβII to negatively regulate cell number in the developing retina

    PubMed Central

    Ma, Lin; Cantrup, Robert; Varrault, Annie; Colak, Dilek; Klenin, Natalia; Götz, Magdalena; McFarlane, Sarah; Journot, Laurent; Schuurmans, Carol

    2007-01-01

    Background Organs are programmed to acquire a particular size during development, but the regulatory mechanisms that dictate when dividing progenitor cells should permanently exit the cell cycle and stop producing additional daughter cells are poorly understood. In differentiated tissues, tumor suppressor genes maintain a constant cell number and intact tissue architecture by controlling proliferation, apoptosis and cell dispersal. Here we report a similar role for two tumor suppressor genes, the Zac1 zinc finger transcription factor and that encoding the cytokine TGFβII, in the developing retina. Results Using loss and gain-of-function approaches, we show that Zac1 is an essential negative regulator of retinal size. Zac1 mutants develop hypercellular retinae due to increased progenitor cell proliferation and reduced apoptosis at late developmental stages. Consequently, supernumerary rod photoreceptors and amacrine cells are generated, the latter of which form an ectopic cellular layer, while other retinal cells are present in their normal number and location. Strikingly, Zac1 functions as a direct negative regulator of a rod fate, while acting cell non-autonomously to modulate amacrine cell number. We implicate TGFβII, another tumor suppressor and cytokine, as a Zac1-dependent amacrine cell negative feedback signal. TGFβII and phospho-Smad2/3, its downstream effector, are expressed at reduced levels in Zac1 mutant retinae, and exogenous TGFβII relieves the mutant amacrine cell phenotype. Moreover, treatment of wild-type retinae with a soluble TGFβ inhibitor and TGFβ receptor II (TGFβRII) conditional mutants generate excess amacrine cells, phenocopying the Zac1 mutant phenotype. Conclusion We show here that Zac1 has an essential role in cell number control during retinal development, akin to its role in tumor surveillance in mature tissues. Furthermore, we demonstrate that Zac1 employs a novel cell non-autonomous strategy to regulate amacrine cell number

  11. Identification and characterization of the products of six region III flagellar genes (flaAII.3 through flaQII) of Salmonella typhimurium.

    PubMed Central

    Homma, M; Iino, T; Macnab, R M

    1988-01-01

    A portion of flagellar region III of the Salmonella typhimurium genome has been cloned and shown to contain six genes: flaAII.3, flaAIII, flaS, flaR, flaQI, and flaQII. Of these, all but flaQI were known to exist from mutant studies; the former flaQ has been renamed flaQII. The genes were shown by minicell analysis to encode proteins with apparent molecular masses of 28, 48, 15, 46, 17, and 37 kilodaltons, respectively. The presence of a flagellar-gene-specific promoter in the vicinity of flaQI was established by testing expression of the plasmid-encoded tetracycline resistance gene in artificial constructions. In minicell preparations, the flaAII.3 and flaR products were found principally in the cytoplasmic fraction; the rest were found principally in the membrane fraction. A comparison between the homologous genes of S. typhimurium and Escherichia coli confirmed that their genomic organizations were similar and that their products had similar molecular masses and isoelectric points. Images PMID:2834334

  12. An experimental study on amelioration of dyslipidemia-induced atherosclesis by Clematichinenoside through regulating Peroxisome proliferator-activated receptor-α mediated apolipoprotein A-I, A-II and C-III.

    PubMed

    Liu, Chao; Guo, Qianqian; Lu, Mengchen; Li, Yunman

    2015-08-15

    Prevention or amelioration the prevalence of atherosclerosis has been an effective strategy in the management of cardiovascular diseases. The aim of the study was to scrutinize the effect of Clematichinenoside (AR) on dyslipidemia-induced atherosclerosis and explore its capability on expression of Peroxisome proliferator-activated receptor-α (PPAR-alpha), apolipoprotein A-I (APOA1) and A-II (APOA2), and suppression of apolipoprotein C-III (APOC3) genes and proteins. In the present study, we investigated atherosclerosis effect of AR using a combination of high-fat diet and balloon injury model in rabbits. The levels of biochemical indicators were evaluated in plasma, liver and HepG2 cells using immunoassay technology. In order to expose the underlying mechanism, we evaluated the regulation of PPAR-alpha, APOA1, APOA2 and APOC3 expressions by AR, and we further evaluated the interactions between them after transfection with shRNA (shPPAR-alpha) and, the action of PPAR-alpha in HepG2 cells. We could find that AR markedly promoted the PPAR-alpha transfer from cytoplasm to nucleus which resulted in the alteration of APOA1, APOA2 and APOC3 expressions in HepG2 cells. Moreover, AR significantly reduced total cholesterol, triglycerides and low-density lipoprotein cholesterol (LDL-C) levels, and elevated high-density lipoprotein cholesterol (HDL-C) level, which play an important role in dyslipidemia-induced atherosclerosis. In conclusion, AR ameliorated atherosclerosis via the regulation of hepatic lipid metabolism, and AR also contributed to the activation of PPAR-alpha, APOA1, APOA2 and APOC3. Therefore, AR could be a potential therapeutic agent in the treatment of atherosclerosis. PMID:25979856

  13. Exploring the retinal connectome

    PubMed Central

    Anderson, James R.; Jones, Bryan W.; Watt, Carl B.; Shaw, Margaret V.; Yang, Jia-Hui; DeMill, David; Lauritzen, James S.; Lin, Yanhua; Rapp, Kevin D.; Mastronarde, David; Koshevoy, Pavel; Grimm, Bradley; Tasdizen, Tolga; Whitaker, Ross

    2011-01-01

    Purpose A connectome is a comprehensive description of synaptic connectivity for a neural domain. Our goal was to produce a connectome data set for the inner plexiform layer of the mammalian retina. This paper describes our first retinal connectome, validates the method, and provides key initial findings. Methods We acquired and assembled a 16.5 terabyte connectome data set RC1 for the rabbit retina at ≈2 nm resolution using automated transmission electron microscope imaging, automated mosaicking, and automated volume registration. RC1 represents a column of tissue 0.25 mm in diameter, spanning the inner nuclear, inner plexiform, and ganglion cell layers. To enhance ultrastructural tracing, we included molecular markers for 4-aminobutyrate (GABA), glutamate, glycine, taurine, glutamine, and the in vivo activity marker, 1-amino-4-guanidobutane. This enabled us to distinguish GABAergic and glycinergic amacrine cells; to identify ON bipolar cells coupled to glycinergic cells; and to discriminate different kinds of bipolar, amacrine, and ganglion cells based on their molecular signatures and activity. The data set was explored and annotated with Viking, our multiuser navigation tool. Annotations were exported to additional applications to render cells, visualize network graphs, and query the database. Results Exploration of RC1 showed that the 2 nm resolution readily recapitulated well known connections and revealed several new features of retinal organization: (1) The well known AII amacrine cell pathway displayed more complexity than previously reported, with no less than 17 distinct signaling modes, including ribbon synapse inputs from OFF bipolar cells, wide-field ON cone bipolar cells and rod bipolar cells, and extensive input from cone-pathway amacrine cells. (2) The axons of most cone bipolar cells formed a distinct signal integration compartment, with ON cone bipolar cell axonal synapses targeting diverse cell types. Both ON and OFF bipolar cells receive

  14. The major cell populations of the mouse retina.

    PubMed

    Jeon, C J; Strettoi, E; Masland, R H

    1998-11-01

    We report a quantitative analysis of the major populations of cells present in the retina of the C57 mouse. Rod and cone photoreceptors were counted using differential interference contrast microscopy in retinal whole mounts. Horizontal, bipolar, amacrine, and Müller cells were identified in serial section electron micrographs assembled into serial montages. Ganglion cells and displaced amacrine cells were counted by subtracting the number of axons in the optic nerve, learned from electron microscopy, from the total neurons of the ganglion cell layer. The results provide a base of reference for future work on genetically altered animals and put into perspective certain recent studies. Comparable data are now available for the retinas of the rabbit and the monkey. With the exception of the monkey fovea, the inner nuclear layers of the three species contain populations of cells that are, overall, quite similar. This contradicts the previous belief that the retinas of lower mammals are "amacrine-dominated", and therefore more complex, than those of higher mammals. PMID:9786999

  15. The -256T>C Polymorphism in the Apolipoprotein A-II Gene Promoter Is Associated with Body Mass Index and Food Intake in the Genetics of Lipid Lowering Drugs and Diet Network Study

    Technology Transfer Automated Retrieval System (TEKTRAN)

    BACKGROUND: Apolipoprotein A-II (APOA2) plays an ambiguous role in lipid metabolism, obesity, and atherosclerosis. METHODS: We studied the association between a functional APOA2 promoter polymorphism (-265T>C) and plasma lipids (fasting and postprandial), anthropometric variables, and food intake in...

  16. Light adaptation alters the source of inhibition to the mouse retinal OFF pathway

    PubMed Central

    Mazade, Reece E.

    2013-01-01

    Sensory systems must avoid saturation to encode a wide range of stimulus intensities. One way the retina accomplishes this is by using both dim-light-sensing rod and bright-light-sensing cone photoreceptor circuits. OFF cone bipolar cells are a key point in this process, as they receive both excitatory input from cones and inhibitory input from AII amacrine cells via the rod pathway. However, in addition to AII amacrine cell input, other inhibitory inputs from cone pathways also modulate OFF cone bipolar cell light signals. It is unknown how these inhibitory inputs to OFF cone bipolar cells change when switching between rod and cone pathways or whether all OFF cone bipolar cells receive rod pathway input. We found that one group of OFF cone bipolar cells (types 1, 2, and 4) receive rod-mediated inhibitory inputs that likely come from the rod-AII amacrine cell pathway, while another group of OFF cone bipolar cells (type 3) do not. In both cases, dark-adapted rod-dominant light responses showed a significant contribution of glycinergic inhibition, which decreased with light adaptation and was, surprisingly, compensated by an increase in GABAergic inhibition. As GABAergic input has distinct timing and spatial spread from glycinergic input, a shift from glycinergic to GABAergic inhibition could significantly alter OFF cone bipolar cell signaling to downstream OFF ganglion cells. Larger GABAergic input could reflect an adjustment of OFF bipolar cell spatial inhibition, which may be one mechanism that contributes to retinal spatial sensitivity in the light. PMID:23926034

  17. A Novel Retinal Oscillation Mechanism in an Autosomal Dominant Photoreceptor Degeneration Mouse Model.

    PubMed

    Tu, Hung-Ya; Chen, Yu-Jiun; McQuiston, Adam R; Chiao, Chuan-Chin; Chen, Ching-Kang

    2015-01-01

    It has been shown in rd1 and rd10 models of photoreceptor degeneration (PD) that inner retinal neurons display spontaneous and rhythmic activities. Furthermore, the rhythmic activity has been shown to require the gap junction protein connexin 36, which is likely located in AII amacrine cells (AII-ACs). In the present study, an autosomal dominant PD model called rhoΔCTA, whose rods overexpress a C-terminally truncated mutant rhodopsin and degenerate with a rate similar to that of rd1, was used to investigate the generality and mechanisms of heightened inner retinal activity following PD. To fluorescently identify cholinergic starburst amacrine cells (SACs), the rhoΔCTA mouse was introduced into a combined ChAT-IRES-Cre and Ai9 background. In this mouse, we observed excitatory postsynaptic current (EPSC) oscillation and non-rhythmic inhibitory postsynaptic current (IPSC) in both ON- and OFF-SACs. The IPSCs were more noticeable in OFF- than in ON-SACs. Similar to reported retinal ganglion cell (RGC) oscillation in rd1 mice, EPSC oscillation was synaptically driven by glutamate and sensitive to blockade of NaV channels and gap junctions. These data suggest that akin to rd1 mice, AII-AC is a prominent oscillator in rhoΔCTA mice. Surprisingly, OFF-SAC but not ON-SAC EPSC oscillation could readily be enhanced by GABAergic blockade. More importantly, weakening the AII-AC gap junction network by activating retinal dopamine receptors abolished oscillations in ON-SACs but not in OFF-SACs. Furthermore, the latter persisted in the presence of flupirtine, an M-type potassium channel activator recently reported to dampen intrinsic AII-AC bursting. These data suggest the existence of a novel oscillation mechanism in mice with PD. PMID:26793064

  18. A Novel Retinal Oscillation Mechanism in an Autosomal Dominant Photoreceptor Degeneration Mouse Model

    PubMed Central

    Tu, Hung-Ya; Chen, Yu-Jiun; McQuiston, Adam R.; Chiao, Chuan-Chin; Chen, Ching-Kang

    2016-01-01

    It has been shown in rd1 and rd10 models of photoreceptor degeneration (PD) that inner retinal neurons display spontaneous and rhythmic activities. Furthermore, the rhythmic activity has been shown to require the gap junction protein connexin 36, which is likely located in AII amacrine cells (AII-ACs). In the present study, an autosomal dominant PD model called rhoΔCTA, whose rods overexpress a C-terminally truncated mutant rhodopsin and degenerate with a rate similar to that of rd1, was used to investigate the generality and mechanisms of heightened inner retinal activity following PD. To fluorescently identify cholinergic starburst amacrine cells (SACs), the rhoΔCTA mouse was introduced into a combined ChAT-IRES-Cre and Ai9 background. In this mouse, we observed excitatory postsynaptic current (EPSC) oscillation and non-rhythmic inhibitory postsynaptic current (IPSC) in both ON- and OFF-SACs. The IPSCs were more noticeable in OFF- than in ON-SACs. Similar to reported retinal ganglion cell (RGC) oscillation in rd1 mice, EPSC oscillation was synaptically driven by glutamate and sensitive to blockade of NaV channels and gap junctions. These data suggest that akin to rd1 mice, AII-AC is a prominent oscillator in rhoΔCTA mice. Surprisingly, OFF-SAC but not ON-SAC EPSC oscillation could readily be enhanced by GABAergic blockade. More importantly, weakening the AII-AC gap junction network by activating retinal dopamine receptors abolished oscillations in ON-SACs but not in OFF-SACs. Furthermore, the latter persisted in the presence of flupirtine, an M-type potassium channel activator recently reported to dampen intrinsic AII-AC bursting. These data suggest the existence of a novel oscillation mechanism in mice with PD. PMID:26793064

  19. Three Forms of Spatial Temporal Feedforward Inhibition Are Common to Different Ganglion Cell Types in Rabbit Retina

    PubMed Central

    Chen, Xin; Hsueh, Hain-Ann; Greenberg, Kenneth

    2010-01-01

    There exist more than 30 different morphological amacrine cell types, but there may be fewer physiological types. Here we studied the amacrine cell outputs by measuring the temporal and spatial properties of feedforward inhibition to four different types of ganglion cells. These ganglion cells, each with concentric receptive field organization, appear to receive a different relative contribution of the same three forms of feed-forward inhibition, namely: local glycinergic, local sustained GABAergic, and broad transient GABAergic inhibition. Two of these inhibitory components, local glycinergic inhibition and local sustained GABAergic inhibition were localized to narrow regions confined to the dendritic fields of the ganglion cells. The third, a broad transient GABAergic inhibition, was driven from regions peripheral to the dendritic area. Each inhibitory component is also correlated with characteristic kinetics expressed in all ganglion cells: broad transient GABAergic inhibition had the shortest latency, local glycinergic inhibition had an intermediate latency, and local sustained GABAergic inhibition had the longest latency. We suggest each of these three inhibitory components represents the output from a distinct class of amacrine cell, mediates a specific visual function, and each forms a basic functional component for the four ganglion cell types. Similar subunits likely exist in the circuits of other ganglion cell types as well. PMID:20220071

  20. Complexin 3 Increases the Fidelity of Signaling in a Retinal Circuit by Regulating Exocytosis at Ribbon Synapses.

    PubMed

    Mortensen, Lena S; Park, Silvia J H; Ke, Jiang-Bin; Cooper, Benjamin H; Zhang, Lei; Imig, Cordelia; Löwel, Siegrid; Reim, Kerstin; Brose, Nils; Demb, Jonathan B; Rhee, Jeong-Seop; Singer, Joshua H

    2016-06-01

    Complexin (Cplx) proteins modulate the core SNARE complex to regulate exocytosis. To understand the contributions of Cplx to signaling in a well-characterized neural circuit, we investigated how Cplx3, a retina-specific paralog, shapes transmission at rod bipolar (RB)→AII amacrine cell synapses in the mouse retina. Knockout of Cplx3 strongly attenuated fast, phasic Ca(2+)-dependent transmission, dependent on local [Ca(2+)] nanodomains, but enhanced slower Ca(2+)-dependent transmission, dependent on global intraterminal [Ca(2+)] ([Ca(2+)]I). Surprisingly, coordinated multivesicular release persisted at Cplx3(-/-) synapses, although its onset was slowed. Light-dependent signaling at Cplx3(-/-) RB→AII synapses was sluggish, owing largely to increased asynchronous release at light offset. Consequently, propagation of RB output to retinal ganglion cells was suppressed dramatically. Our study links Cplx3 expression with synapse and circuit function in a specific retinal pathway and reveals a role for asynchronous release in circuit gain control. PMID:27239031

  1. Melanopsin-expressing ganglion cells on macaque and human retinas form two morphologically distinct populations.

    PubMed

    Liao, Hsi-Wen; Ren, Xiaozhi; Peterson, Beth B; Marshak, David W; Yau, King-Wai; Gamlin, Paul D; Dacey, Dennis M

    2016-10-01

    The long-term goal of this research is to understand how retinal ganglion cells that express the photopigment melanopsin, also known as OPN4, contribute to vision in humans and other primates. Here we report the results of anatomical studies using our polyclonal antibody specifically against human melanopsin that confirm and extend previous descriptions of melanopsin cells in primates. In macaque and human retina, two distinct populations of melanopsin cells were identified based on dendritic stratification in either the inner or the outer portion of the inner plexiform layer (IPL). Variation in dendritic field size and cell density with eccentricity was confirmed, and dendritic spines, a new feature of melanopsin cells, were described. The spines were the sites of input from DB6 diffuse bipolar cell axon terminals to the inner stratifying type of melanopsin cells. The outer stratifying melanopsin type received inputs from DB6 bipolar cells via a sparse outer axonal arbor. Outer stratifying melanopsin cells also received inputs from axon terminals of dopaminergic amacrine cells. On the outer stratifying melanopsin cells, ribbon synapses from bipolar cells and conventional synapses from amacrine cells were identified in electron microscopic immunolabeling experiments. Both inner and outer stratifying melanopsin cell types were retrogradely labeled following tracer injection in the lateral geniculate nucleus (LGN). In addition, a method for targeting melanopsin cells for intracellular injection using their intrinsic fluorescence was developed. This technique was used to demonstrate that melanopsin cells were tracer coupled to amacrine cells and would be applicable to electrophysiological experiments in the future. J. Comp. Neurol. 524:2845-2872, 2016. © 2016 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc. PMID:26972791

  2. QTL mapping of genetic determinants of lipoprotein metabolism in mice: Mutations of the apolipoprotein A-II gene affecting lipoprotein turnover

    SciTech Connect

    Weinreb, A.; Purcell-Huynh, D.A.; Castellani, L.W.

    1994-09-01

    Cholesterol and lipoproteins represent important risk factors for atherosclerosis. In order to better understand the genes involved in determining lipoprotein levels, quantitative trait locus (QTL) mapping was performed using a cross between NZB and SM/J mice. Significant LOD scores for loci determining total cholesterol, HDL cholesterol, LDL and VLDL cholesterol, triglycerides, free fatty acids, and apolipoprotein A-II (apoA-II) were obtained. NZB mice have a 7-10 fold higher apoA-II level SM/J. LOD scores of 19.6 (chow) and 10.3 (high fat) were obtained at the apoA-II gene locus. Comparison of apoA-II levels by apoA-II genotype reveals that {approximately}30% of the variance in apoA-II levels can be accounted for by differences within the apoA-II gene. Northern analysis of mRNA from NZB and SM/J mice fed a high fat diet failed to show any significant differences in mRNA levels. The rates of apoA-II protein synthesis relative to total protein synthesis between the two strains were similar, with a rate of 0.16% for NZB and 0.18% for SM/J. Sequencing of NZB and SM/J apoA-II cDNAs revealed a pro5 to gln5 substitution in SM/J. Therefore, differences in the apoA-II levels between NZB and SM/J may be partly due to a structural difference in apoA-II resulting in an increased rate of apoA-II clearance in SM/J. A coincident QTL for HDL at the same chromosome 1 locus suggests that a structural difference in apoA-II may be affecting the rate of HDL clearance. It is of interest to note that the pro5 to gln5 substitution leads to apoA-II amyloid deposition in the SAM mouse.

  3. Comparison of High-Density Lipoprotein Cholesterol to Apolipoprotein A-I and A-II to Predict Coronary Calcium and the Effect of Insulin Resistance

    PubMed Central

    Martin, Seth S.; Qasim, Atif N.; Wolfe, Megan; Clair, Caitlin; Schwartz, Stanley; Iqbal, Nayyar; Schutta, Mark; Bagheri, Roshanak; Mehta, Nehal N.; Rader, Daniel J.; Reilly, Muredach P.

    2011-01-01

    High-density lipoprotein (HDL) cholesterol and its apolipoproteins each capture unique lipid and cardiometabolic information important to risk quantification. It was hypothesized that metabolic factors, including insulin resistance and type 2 diabetes, would confound the association of HDL cholesterol with coronary artery calcification (CAC) and that apolipoprotein A-I (apoA-I) and/or apolipoprotein A-II (apoA-II) would add to HDL cholesterol in predicting CAC. Two community-based cross-sectional studies of white subjects were analyzed: the Penn Diabetes Heart Study (PDHS; n = 611 subjects with type 2 diabetes, 71.4% men) and the Study of Inherited Risk of Coronary Atherosclerosis (SIRCA; n = 803 subjects without diabetes, 52.8% men) using multivariable analysis of apoA-I, apoA-II, and HDL cholesterol stratified by diabetes status. HDL cholesterol was inversely associated with CAC after adjusting for age and gender in whites with type 2 diabetes (tobit ratio for a 1-SD increase in HDL cholesterol 0.58, 95% confidence interval [CI] 0.44 to 0.77, p <0.001) as well as those without diabetes (tobit ratio 0.72, 95% CI 0.59 to 0.88, p = 0.001). In contrast, apoA-I was a weaker predictor in subjects with (tobit ratio 0.64, 95% CI 0.45 to 0.90, p = 0.010) and without (tobit ratio 0.79, 95% CI 0.66 to 0.94, p = 0.010) diabetes, while apoA-II had no association with CAC. Control for metabolic variables, including triglycerides, waist circumference, and homeostasis model assessment of insulin resistance, attenuated these relations, particularly in subjects without diabetes. In likelihood ratio test analyses, HDL cholesterol added to apoA-I, apoA-II, and atherogenic apolipoprotein B lipoproteins but improved CAC prediction over metabolic factors only in subjects with diabetes. In conclusion, HDL cholesterol outperformed apoA-I and apoA-II in CAC prediction, but its association with CAC was attenuated by measures of insulin resistance. PMID:21257004

  4. Comparison of high-density lipoprotein cholesterol to apolipoprotein A-I and A-II to predict coronary calcium and the effect of insulin resistance.

    PubMed

    Martin, Seth S; Qasim, Atif N; Wolfe, Megan; St Clair, Caitlin; Schwartz, Stanley; Iqbal, Nayyar; Schutta, Mark; Bagheri, Roshanak; Mehta, Nehal N; Rader, Daniel J; Reilly, Muredach P

    2011-02-01

    High-density lipoprotein (HDL) cholesterol and its apolipoproteins each capture unique lipid and cardiometabolic information important to risk quantification. It was hypothesized that metabolic factors, including insulin resistance and type 2 diabetes, would confound the association of HDL cholesterol with coronary artery calcification (CAC) and that apolipoprotein A-I (apoA-I) and/or apolipoprotein A-II (apoA-II) would add to HDL cholesterol in predicting CAC. Two community-based cross-sectional studies of white subjects were analyzed: the Penn Diabetes Heart Study (PDHS; n = 611 subjects with type 2 diabetes, 71.4% men) and the Study of Inherited Risk of Coronary Atherosclerosis (SIRCA; n = 803 subjects without diabetes, 52.8% men) using multivariable analysis of apoA-I, apoA-II, and HDL cholesterol stratified by diabetes status. HDL cholesterol was inversely associated with CAC after adjusting for age and gender in whites with type 2 diabetes (tobit ratio for a 1-SD increase in HDL cholesterol 0.58, 95% confidence interval [CI] 0.44 to 0.77, p <0.001) as well as those without diabetes (tobit ratio 0.72, 95% CI 0.59 to 0.88, p = 0.001). In contrast, apoA-I was a weaker predictor in subjects with (tobit ratio 0.64, 95% CI 0.45 to 0.90, p = 0.010) and without (tobit ratio 0.79, 95% CI 0.66 to 0.94, p = 0.010) diabetes, while apoA-II had no association with CAC. Control for metabolic variables, including triglycerides, waist circumference, and homeostasis model assessment of insulin resistance, attenuated these relations, particularly in subjects without diabetes. In likelihood ratio test analyses, HDL cholesterol added to apoA-I, apoA-II, and atherogenic apolipoprotein B lipoproteins but improved CAC prediction over metabolic factors only in subjects with diabetes. In conclusion, HDL cholesterol outperformed apoA-I and apoA-II in CAC prediction, but its association with CAC was attenuated by measures of insulin resistance. PMID:21257004

  5. Structure, infrared and Raman spectroscopic studies of newly synthetic AII(SbV0.50FeIII0.50)(PO4)2 (Adbnd Ba, Sr, Pb) phosphates with yavapaiite structure

    NASA Astrophysics Data System (ADS)

    Aatiq, Abderrahim; Tigha, My Rachid; Fakhreddine, Rachid; Bregiroux, Damien; Wallez, Gilles

    2016-08-01

    The synthesis and structural study of three new AII(SbV0.5FeIII0.5)(PO4)2 (Adbnd Ba, Sr, Pb) phosphates belonging to the Asbnd Sbsbnd Fesbnd Psbnd O system were reported here for the first time. Structures of [Ba], [Sr] and [Pb] compounds, obtained by solid state reaction in air atmosphere, were determined at room temperature from X-ray powder diffraction using the Rietveld method. BaII(SbV0.5FeIII0.5)(PO4)2 features the yavapaiite-type structure, with space group C2/m, Z = 2 and a = 8.1568(4) Å; b = 5.1996(3) Å c = 7.8290(4) Å; β = 94.53(1)°. AII(SbV0.5FeIII0.5)(PO4)2 (Adbnd Sr, Pb) compounds have a distorted yavapaiite structure with space group C2/c, Z = 4 and a = 16.5215(2) Å; b = 5.1891(1) Å c = 8.0489(1) Å; β = 115.70(1)° for [Sr]; a = 16.6925(2) Å; b = 5.1832(1) Å c = 8.1215(1) Å; β = 115.03(1)° for [Pb]. Raman and Infrared spectroscopic study was used to obtain further structural information about the nature of bonding in selected compositions.

  6. Expression Profiling of Developing Zebrafish Retinal Cells.

    PubMed

    Mullally, Madelyn; Albrecht, Caitlin; Horton, Mary; Laboissonniere, Lauren A; Goetz, Jillian J; Chowdhury, Rebecca; Manning, Alicia; Wester, Andrea K; Bose, Quinton; Trimarchi, Jeffrey M

    2016-08-01

    During retinal development, a variety of different types of neurons are produced. Understanding how each of these types of retinal nerve cells is generated is important from a developmental biology perspective. It is equally important if one is interested in how to regenerate cells after an injury or a disease. To gain more insight into how retinal neurons develop in the zebrafish, we performed single-cell mRNA profiling and in situ hybridizations (ISHs) on retinal sections and whole-mount zebrafish. Through the series of ISHs, designed and performed solely by undergraduate students in the laboratory, we were able to retrospectively identify our single-cell mRNA profiles as most likely coming from developing amacrine cells. Further analysis of these profiles will reveal genes that can be mutated using genome editing techniques. Together these studies increase our knowledge of the genes driving development of different cell types in the zebrafish retina. PMID:26982811

  7. Developmental changes in the expression level of connexin36 in the rat retina.

    PubMed

    Kovács-Öller, Tamás; Raics, Katalin; Orbán, József; Nyitrai, Miklós; Völgyi, Béla

    2014-11-01

    Connexin36 (Cx36) is the major gap junction forming protein in the brain and the retina; thus, alterations in its expression indicate changes in the corresponding circuitry. Many structural changes occur in the early postnatal retina before functional neuronal circuits are finalized, including those that incorporate gap junctions. To reveal the time-lapse formation of inner retinal gap junctions, we examine the developing postnatal rat retina from birth (P0) to young adult age (P20) and follow the expression of Cx36 in the mRNA and protein levels. We found a continuous elevation in the expression of both the Cx36 transcript and protein between P0 and P20 and a somewhat delayed Cx36 plaque formation throughout the inner plexiform layer (IPL) starting at P10. By using tristratificated calretinin positive (CaR(+)) fibers in the IPL as a guide, we detected a clear preference of Cx36 plaques for the ON sublamina from the earliest time of detection. This distributional preference became more pronounced at P15 and P20 due to the emergence and widespread expression of large (>0.1 μm(2)) Cx36 plaques in the ON sublamina. Finally, we showed that parvalbumin-positive (PV(+)) AII amacrine cell dendrites colocalize with Cx36 plaques as early as P10 in strata 3 and 4, whereas colocalizations in stratum 5 became characteristic only around P20. We conclude that Cx36 expression in the rat IPL displays a characteristic succession of changes during retinogenesis reflecting the formation of the underlying electrical synaptic circuitry. In particular, AII cell gap junctions, first formed with ON cone bipolar cells and later with other AII amacrine cells, accounted for the observed Cx36 expressional changes. PMID:25110193

  8. Complex inhibitory microcircuitry regulates retinal signaling near visual threshold.

    PubMed

    Grimes, William N; Zhang, Jun; Tian, Hua; Graydon, Cole W; Hoon, Mrinalini; Rieke, Fred; Diamond, Jeffrey S

    2015-07-01

    Neuronal microcircuits, small, localized signaling motifs involving two or more neurons, underlie signal processing and computation in the brain. Compartmentalized signaling within a neuron may enable it to participate in multiple, independent microcircuits. Each A17 amacrine cell in the mammalian retina contains within its dendrites hundreds of synaptic feedback microcircuits that operate independently to modulate feedforward signaling in the inner retina. Each of these microcircuits comprises a small (<1 μm) synaptic varicosity that typically receives one excitatory synapse from a presynaptic rod bipolar cell (RBC) and returns two reciprocal inhibitory synapses back onto the same RBC terminal. Feedback inhibition from the A17 sculpts the feedforward signal from the RBC to the AII, a critical component of the circuitry mediating night vision. Here, we show that the two inhibitory synapses from the A17 to the RBC express kinetically distinct populations of GABA receptors: rapidly activating GABA(A)Rs are enriched at one synapse while more slowly activating GABA(C)Rs are enriched at the other. Anatomical and electrophysiological data suggest that macromolecular complexes of voltage-gated (Cav) channels and Ca(2+)-activated K(+) channels help to regulate GABA release from A17 varicosities and limit GABA(C)R activation under certain conditions. Finally, we find that selective elimination of A17-mediated feedback inhibition reduces the signal to noise ratio of responses to dim flashes recorded in the feedforward pathway (i.e., the AII amacrine cell). We conclude that A17-mediated feedback inhibition improves the signal to noise ratio of RBC-AII transmission near visual threshold, thereby improving visual sensitivity at night. PMID:25972578

  9. Characterization of insulin-like growth factor I and insulin receptors on cultured bovine adrenal fasciculata cells. Role of these peptides on adrenal cell function

    SciTech Connect

    Penhoat, A.; Chatelain, P.G.; Jaillard, C.; Saez, J.M.

    1988-06-01

    We have characterized insulin-like growth factor I (IGF-I) and insulin receptors in cultured bovine adrenal cells by binding and cross-linking affinity experiments. At equilibrium the dissociation constant and the number of binding sites per cell for IGF-I were 1.4 +/- (SE) 0.3 x 10(-9) M and 19,200 +/- 2,100, respectively. Under reduction conditions, disuccinimidyl suberate cross-linked (/sup 125/I)iodo-IGF-I to one receptor complex with an Mr of 125,000. Adrenal cells also contain specific insulin receptors with an apparent dissociation constant (Kd) of 10(-9) M. Under reduction conditions (/sup 125/I)iodo-insulin binds to one band with an approximate Mr of 125,000. IGF-I and insulin at micromolar concentrations, but not at nanomolar concentrations, slightly stimulated DNA synthesis, but markedly potentiated the mitogenic action of fibroblast growth factor. Adrenal cells cultured in a serum-free medium containing transferrin, ascorbic acid, and insulin (5 micrograms/ml) maintained fairly constant angiotensin-II (A-II) receptor concentration per cell and increased cAMP release on response to ACTH and their steroidogenic response to both ACTH and A-II. When the cells were cultured in the same medium without insulin, the number of A-II receptors significantly decreased to 65% and the increased responsiveness was blunted. Treatment of such cells for 3 days with increasing concentrations of IGF-I (1-100 ng/ml) produced a 2- to 3-fold increase in A-II receptors and enhanced the cAMP response (3- to 4-fold) to ACTH and the steroidogenic response (4- to 6-fold) to ACTH and A-II. These effects were time and dose dependent (ED50 approximately equal to 10(-9) M). Insulin at micromolar concentrations produced an effect similar to that of IGF-I, but at nanomolar concentrations the effect was far less.

  10. Therapeutic enzyme deimmunization by combinatorial T-cell epitope removal using neutral drift

    PubMed Central

    Cantor, Jason R.; Yoo, Tae Hyeon; Dixit, Aakanksha; Iverson, Brent L.; Forsthuber, Thomas G.; Georgiou, George

    2011-01-01

    A number of heterologous enzymes have been investigated for cancer treatment and other therapeutic applications; however, immunogenicity issues have limited their clinical utility. Here, a new approach has been created for heterologous enzyme deimmunization whereby combinatorial saturation mutagenesis is coupled with a screening strategy that capitalizes on the evolutionary biology concept of neutral drift, and combined with iterative computational prediction of T-cell epitopes to achieve extensive reengineering of a protein sequence for reduced MHC-II binding propensity without affecting catalytic and pharmacological properties. Escherichia coli L-asparaginase II (EcAII), the only nonhuman enzyme approved for repeated administration, is critical in treatment of childhood acute lymphoblastic leukemia (ALL), but elicits adverse antibody responses in a significant fraction of patients. The neutral drift screening of combinatorial saturation mutagenesis libraries at a total of 12 positions was used to isolate an EcAII variant containing eight amino acid substitutions within computationally predicted T-cell epitopes—of which four were nonconservative—while still exhibiting kcat/KM = 106 M-1 s-1 for L-Asn hydrolysis. Further, immunization of HLA-transgenic mice expressing the ALL-associated DRB1*0401 allele with the engineered variant resulted in significantly reduced T-cell responses and a 10-fold reduction in anti-EcAII IgG titers relative to the existing therapeutic. This significant reduction in the immunogenicity of EcAII may be clinically relevant for ALL treatment and illustrates the potential of employing neutral drift screens to achieve large jumps in sequence space as may be required for the deimmunization of heterologous proteins. PMID:21209329

  11. Nitric Oxide Modulates the Temporal Properties of the Glutamate Response in Type 4 OFF Bipolar Cells

    PubMed Central

    Vielma, Alex H.; Agurto, Adolfo; Valdés, Joaquín; Palacios, Adrián G.; Schmachtenberg, Oliver

    2014-01-01

    Nitric oxide (NO) is involved in retinal signal processing, but its cellular actions are only partly understood. An established source of retinal NO are NOACs, a group of nNOS-expressing amacrine cells which signal onto bipolar, other amacrine and ganglion cells in the inner plexiform layer. Here, we report that NO regulates glutamate responses in morphologically and electrophysiologically identified type 4 OFF cone bipolar cells through activation of the soluble guanylyl cyclase-cGMP-PKG pathway. The glutamate response of these cells consists of two components, a fast phasic current sensitive to kainate receptor agonists, and a secondary component with slow kinetics, inhibited by AMPA receptor antagonists. NO shortened the duration of the AMPA receptor-dependent component of the glutamate response, while the kainate receptor-dependent component remained unchanged. Application of 8-Br-cGMP mimicked this effect, while inhibition of soluble guanylate cyclase or protein kinase G prevented it, supporting a mechanism involving a cGMP signaling pathway. Notably, perfusion with a NOS-inhibitor prolonged the duration of the glutamate response, while the NO precursor L-arginine shortened it, in agreement with a modulation by endogenous NO. Furthermore, NO accelerated the response recovery during repeated stimulation of type 4 cone bipolar cells, suggesting that the temporal response properties of this OFF bipolar cell type are regulated by NO. These results reveal a novel cellular mechanism of NO signaling in the retina, and represent the first functional evidence of NO modulating OFF cone bipolar cells. PMID:25463389

  12. Meis1 regulates Foxn4 expression during retinal progenitor cell differentiation

    PubMed Central

    Islam, Mohammed M.; Li, Ying; Luo, Huijun; Xiang, Mengqing; Cai, Li

    2013-01-01

    Summary The transcription factor forkhead box N4 (Foxn4) is a key regulator in a variety of biological processes during development. In particular, Foxn4 plays an essential role in the genesis of horizontal and amacrine neurons from neural progenitors in the vertebrate retina. Although the functions of Foxn4 have been well established, the transcriptional regulation of Foxn4 expression during progenitor cell differentiation remains unclear. Here, we report that an evolutionarily conserved 129 bp noncoding DNA fragment (Foxn4CR4.2 or CR4.2), located ∼26 kb upstream of Foxn4 transcription start site, functions as a cis-element for Foxn4 regulation. CR4.2 directs gene expression in Foxn4-positive cells, primarily in progenitors, differentiating horizontal and amacrine cells. We further determined that the gene regulatory activity of CR4.2 is modulated by Meis1 binding motif, which is bound and activated by Meis1 transcription factor. Deletion of the Meis1 binding motif or knockdown of Meis1 expression abolishes the gene regulatory activity of CR4.2. In addition, knockdown of Meis1 expression diminishes the endogenous Foxn4 expression and affects cell lineage development. Together, we demonstrate that CR4.2 and its interacting Meis1 transcription factor play important roles in regulating Foxn4 expression during early retinogenesis. These findings provide new insights into molecular mechanisms that govern gene regulation in retinal progenitors and specific cell lineage development. PMID:24244849

  13. Lmo4 and Other LIM domain only factors are necessary and sufficient for multiple retinal cell type development.

    PubMed

    Jin, Kangxin; Xiao, Dongchang; Andersen, Bogi; Xiang, Mengqing

    2016-08-01

    Understanding the molecular basis by which distinct cell types are specified is a central issue in retinogenesis and retinal disease development. Here we examined the role of LIM domain only 4 (Lmo4) in retinal development using both gain-of-function and loss-of-function approaches. By immunostaining, Lmo4 was found to be expressed in mouse retina from E10.5 to mature stages. Retroviral delivery of Lmo4 into retinal progenitor cells could promote the amacrine, bipolar and Müller cell fates at the expense of photoreceptors. It also inhibited the fate of early-born retinal ganglion cells. Using a dominant-negative form of Lmo4 which suppresses transcriptional activities of all LIM domain only factors, we demonstrated that LIM domain only factors are both necessary and sufficient for promoting amacrine and bipolar cell development, but not for the differentiation of ganglion, horizontal, Müller, or photoreceptor cells. Taken together, our study uncovers multiple roles of Lmo4 during retinal development and demonstrates the importance of LIM domain only factors in ensuring proper retinal cell specification and differentiation. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 900-915, 2016. PMID:26579872

  14. Interstitial deletion of chromosome 1q [del(1)(q24q25.3)] identified by fluorescence in situ hybridization and gene dosage analysis of apolipoprotein A-II, coagulation factor V, and antithrombin III

    SciTech Connect

    Takano, Takako; Yamanouchi, Yasuko; Mori, Yosuke

    1997-01-20

    We report on a 12-month-old Japanese boy with an interstitial deletion of the long-arm of chromosome 1 and meningomyelocele, hydrocephalus, anal atresia, atrial septal defect, left renal agenesis, bilateral cryptorchidism, talipes equinovarus, low birth weight, growth/developmental retardation, and many minor anomalies. By conventional GTG-banding, his karyotype was first interpreted as 46,XY,de1(1)(q23q24), but it was corrected as 46,XY.ish del(1)(q24q25.3) by fluorescence in situ hybridization using 11 known cosmid clones as probes. His serum levels of apolipoprotein A-II (gene symbol: APOA2, previously assigned to 1q21-q23) and coagulation factor V (F5, 1q21-q25) were normal, while serum concentration and activity of antithrombin III (AT3, 1q23-q25.1) was low. The results indicated that localization of APOA2 and F5 are proximal to the deleted region and AT3 is located within the deletion extent in the patient. 16 refs., 4 figs.

  15. Sip1 regulates the generation of the inner nuclear layer retinal cell lineages in mammals.

    PubMed

    Menuchin-Lasowski, Yotam; Oren-Giladi, Pazit; Xie, Qing; Ezra-Elia, Raaya; Ofri, Ron; Peled-Hajaj, Shany; Farhy, Chen; Higashi, Yujiro; Van de Putte, Tom; Kondoh, Hisato; Huylebroeck, Danny; Cvekl, Ales; Ashery-Padan, Ruth

    2016-08-01

    The transcription factor Sip1 (Zeb2) plays multiple roles during CNS development from early acquisition of neural fate to cortical neurogenesis and gliogenesis. In humans, SIP1 (ZEB2) haploinsufficiency leads to Mowat-Wilson syndrome, a complex congenital anomaly including intellectual disability, epilepsy and Hirschsprung disease. Here we uncover the role of Sip1 in retinogenesis. Somatic deletion of Sip1 from mouse retinal progenitors primarily affects the generation of inner nuclear layer cell types, resulting in complete loss of horizontal cells and reduced numbers of amacrine and bipolar cells, while the number of Muller glia is increased. Molecular analysis places Sip1 downstream of the eye field transcription factor Pax6 and upstream of Ptf1a in the gene network required for generating the horizontal and amacrine lineages. Intriguingly, characterization of differentiation dynamics reveals that Sip1 has a role in promoting the timely differentiation of retinal interneurons, assuring generation of the proper number of the diverse neuronal and glial cell subtypes that constitute the functional retina in mammals. PMID:27385012

  16. Reprogramming chick RPE progeny cells to differentiate towards retinal neurons by ash1

    PubMed Central

    Mao, Weiming; Yan, Run-Tao

    2008-01-01

    Purpose Harnessing a cell culture of retinal pigment epithelium (RPE) to give rise to retinal neurons may offer a source of developing neurons for cell-replacement studies. This study explores the possibility of reprogramming RPE progeny cells to differentiate toward retinal neurons with achaete-scute homolog 1 (ash1), a proneural gene that is expressed in progenitor cells in the developing retina and promotes amacrine cell production when overexpressed in the chick retina. Methods Replication Competent Avian Splice (RCAS) retrovirus was used to drive the ectopic expression of ash1 in cell cultures of dissociated RPE isolated from day 6 chick embryos. RCAS expressing green fluorescent protein (RCAS-GFP) was used as control. The cultures were examined for de novo generation of neuron-like cells by molecular, cellular, and physiologic criteria. Results In control cultures infected with RCAS-GFP, RPE cells appeared cobblestone-like and often darkly pigmented. In cultures infected with RCAS-ash1, however, cells remained de-pigmented and frequently formed clusters. Further examination at the morphological and molecular levels showed the development of elaborate processes characteristic of neurons and the expression of genes/markers that identify different types of retinal neurons. The most prevalently expressed neural marker was calretinin, which in the chick retina identifies amacrine, ganglion, and horizontal cells. As an assay for functional maturation, the reprogrammed cells were analyzed for the presence of functional, ionotropic glutamate receptors that lead to a rise in the cytosolic free calcium (Ca2+) concentration. Calcium imaging showed that reprogrammed cells responded to glutamate and N-methyl-D-aspartate (NMDA) by increasing their Ca2+ concentrations, which, after reaching a peak level, returned to the basal level. The response curves of reprogrammed cells resembled those of cultured retinal neurons. Conclusions These results suggest that RPE progeny cells

  17. Nicotinic Antagonists Enhance Process Outgrowth by Rat Retinal Ganglion Cells in Culture

    NASA Astrophysics Data System (ADS)

    Lipton, Stuart A.; Frosch, Matthew P.; Phillips, Micheal D.; Tauck, David L.; Aizenman, Elias

    1988-03-01

    Functional nicotinic cholinergic receptors are found on mammalian retinal ganglion cell neurons in culture. The neurotransmitter acetylcholine (ACh) can be detected in the medium of many of these retinal cultures, after release presumably from the choline acetyltransferase-positive amacrine cells. The postsynaptic effect of endogenous or applied ACh on the ganglion cells can be blocked with specific nicotinic antagonists. Here it is shown that within 24 hours of producing such a pharmacologic blockade, the retinal ganglion cells begin to sprout or regenerate neuronal processes. Thus, the growth-enhancing effect of nicotinic antagonists may be due to the removal of inhibition to growth by tonic levels of ACh present in the culture medium. Since there is a spontaneous leak of ACh in the intact retina, the effects of nicotinic cholinergic drugs on process outgrowth in culture may reflect a normal control mechanism for growth or regeneration of retinal ganglion cell processes that is exerted by ACh in vivo.

  18. Synaptic Input of ON-Bipolar Cells onto the Dopaminergic Neurons of the Mouse Retina

    PubMed Central

    Contini, Massimo; Lin, Bin; Kobayashi, Kazuto; Okano, Hideyuki; Masland, Richard H.; Raviola, Elio

    2010-01-01

    In the retina, dopamine fulfills a crucial role in neural adaptation to photopic illumination, but the pathway that carries cone signals to the dopaminergic amacrine (DA) cells was not known. We identified the site of ON-cone bipolar input onto DA cells in transgenic mice in which both types of catecholaminergic amacrine (CA) cells were labeled with green fluorescent protein or human placental alkaline phosphatase (PLAP). In confocal Z series of retinal whole mounts stained with antibodies to tyrosine hydroxylase (TH), DA cells gave rise to varicose processes that descended obliquely through the scleral half of the inner plexiform layer (IPL) and formed a loose, tangential plexus in the middle of this layer. Comparison with the distribution of the dendrites of type 2 CA cells and examination of neurobiotin-injected DA cells proved that their vitreal processes were situated in stratum S3 of the IPL. Electron microscope demonstration of PLAP activity showed that bipolar cell endings in S3 established ribbon synapses onto a postsynaptic dyad in which one or both processes were labeled by a precipitate of lead phosphate and therefore belonged to DA cells. In places, the postsynaptic DA cell processes returned a reciprocal synapse onto the bipolar endings. Confocal images of sections stained with antibodies to TH, kinesin Kif3a, which labels synaptic ribbons, and glutamate or GABAA receptors, confirmed that ribbon-containing endings made glutamatergic synapses onto DA cells processes in S3 and received from them GABAergic synapses. The presynaptic ON-bipolar cells most likely belonged to the CB3 (type 5) variety. PMID:20394057

  19. Characterisation of light responses in the retina of mice lacking principle components of rod, cone and melanopsin phototransduction signalling pathways.

    PubMed

    Hughes, Steven; Rodgers, Jessica; Hickey, Doron; Foster, Russell G; Peirson, Stuart N; Hankins, Mark W

    2016-01-01

    Gnat(-/-), Cnga3(-/-), Opn4(-/-) triple knockout (TKO) mice lack essential components of phototransduction signalling pathways present in rods, cones and photosensitive retinal ganglion cells (pRGCs), and are therefore expected to lack all sensitivity to light. However, a number of studies have shown that light responses persist in these mice. In this study we use multielectrode array (MEA) recordings and light-induced c-fos expression to further characterise the light responses of the TKO retina. Small, but robust electroretinogram type responses are routinely detected during MEA recordings, with properties consistent with rod driven responses. Furthermore, a distinctive pattern of light-induced c-fos expression is evident in the TKO retina, with c-fos expression largely restricted to a small subset of amacrine cells that express disabled-1 (Dab1) but lack expression of glycine transporter-1 (GlyT-1). Collectively these data are consistent with the persistence of a novel light sensing pathway in the TKO retina that originates in rod photoreceptors, potentially a rare subset of rods with distinct functional properties, and which is propagated to an atypical subtype of AII amacrine cells. Furthermore, the minimal responses observed following UV light stimulation suggest only a limited role for the non-visual opsin OPN5 in driving excitatory light responses within the mouse retina. PMID:27301998

  20. Characterisation of light responses in the retina of mice lacking principle components of rod, cone and melanopsin phototransduction signalling pathways

    PubMed Central

    Hughes, Steven; Rodgers, Jessica; Hickey, Doron; Foster, Russell G.; Peirson, Stuart N.; Hankins, Mark W.

    2016-01-01

    Gnat−/−, Cnga3−/−, Opn4−/− triple knockout (TKO) mice lack essential components of phototransduction signalling pathways present in rods, cones and photosensitive retinal ganglion cells (pRGCs), and are therefore expected to lack all sensitivity to light. However, a number of studies have shown that light responses persist in these mice. In this study we use multielectrode array (MEA) recordings and light-induced c-fos expression to further characterise the light responses of the TKO retina. Small, but robust electroretinogram type responses are routinely detected during MEA recordings, with properties consistent with rod driven responses. Furthermore, a distinctive pattern of light-induced c-fos expression is evident in the TKO retina, with c-fos expression largely restricted to a small subset of amacrine cells that express disabled-1 (Dab1) but lack expression of glycine transporter-1 (GlyT-1). Collectively these data are consistent with the persistence of a novel light sensing pathway in the TKO retina that originates in rod photoreceptors, potentially a rare subset of rods with distinct functional properties, and which is propagated to an atypical subtype of AII amacrine cells. Furthermore, the minimal responses observed following UV light stimulation suggest only a limited role for the non-visual opsin OPN5 in driving excitatory light responses within the mouse retina. PMID:27301998

  1. Characterization of inhibitory postsynaptic currents in rod bipolar cells of the mouse retina.

    PubMed

    Frech, Moritz J; Backus, Kurt H

    2004-01-01

    The synaptic terminals of mammalian rod bipolar cells are the targets of multiple presynaptic inhibitory inputs arriving from glycinergic and GABAergic amacrine cells. To investigate the contribution of these different inhibitory receptor types, we have applied the patch-clamp technique in acutely isolated slices of the adult mouse retina. By using the whole-cell configuration, we measured and analyzed the spontaneous postsynaptic currents (PSCs) in rod bipolar cells. The spontaneous synaptic activity of rod bipolar cells was very low. However, when amacrine cells were depolarized by AMPA or kainate, the PSC frequency in rod bipolar cells increased significantly. These PSCs comprised several types that could be distinguished by pharmacological and kinetic criteria. Strychnine-sensitive, glycinergic PSCs were characterized by a mean peak amplitude of -43.5 pA and a weighted decay time constant (tauw) of 10.9 ms. PSCs that persisted in the presence of strychnine, but were completely inhibited by bicuculline, were mediated by GABAARs. They had a mean peak amplitude of -20.0 pA and a significantly faster tauw of 5.8 ms. Few PSCs remained in the presence of strychnine and bicuculline, suggesting that they were mediated by GABACRs. These PSCs were characterized by much smaller amplitudes (-6.2 pA) and a significantly slower decay kinetics (tauw=51.0 ms). We conclude that rod bipolar cells express at least three types of functionally different inhibitory receptors, namely GABAARs, GABACRs, and GlyRs that may ultimately regulate the Ca2+ influx into rod bipolar cell terminals, thereby modulating their glutamate release. PMID:15579227

  2. A Thy1-CFP DBA/2J mouse line with cyan fluorescent protein expression in retinal ganglion cells

    PubMed Central

    RAYMOND, IONA D.; POOL, ANGELA L.; VILA, ALEJANDRO; BRECHA, NICHOLAS C.

    2013-01-01

    A DBA/2J (D2) transgenic mouse line with cyan fluorescent protein (CFP) reporter expression in ganglion cells was developed for the analysis of ganglion cells during progressive glaucoma. The Thy1-CFP D2 (CFP-D2) line was created by congenically breeding the D2 line, which develops pigmentary glaucoma, and the Thy1-CFP line, which expresses CFP in ganglion cells. Microsatellite marker analysis of CFP-D2 progeny verified the genetic inclusion of the D2 isa and ipd loci. Specific mutations within these loci lead to dysfunctional melanosomal proteins and glaucomatous phenotype in D2 mice. Polymerase chain reaction analysis confirmed the inclusion of the Thy1-CFP transgene. CFP-fluorescent ganglion cells, 6–20 μm in diameter, were distributed in all retinal regions, CFP processes were throughout the inner plexiform layer, and CFP-fluorescent axons were in the fiber layer and optic nerve head. Immunohistochemistry with antibodies to ganglion cell markers NF-L, NeuN, Brn3a, and SMI32 was used to confirm CFP expression in ganglion cells. Immunohistochemistry with antibodies to amacrine cell markers HPC-1 and ChAT was used to confirm weak CFP expression in cholinergic amacrine cells. CFP-D2 mice developed a glaucomatous phenotype, including iris disease, ganglion cell loss, attrition of the fiber layer, and elevated intraocular pressure. A CFP-D2 transgenic line with CFP-expressing ganglion cells was developed, which has (1) a predominantly D2 genetic background, (2) CFP-expressing ganglion cells, and (3) age-related progressive glaucoma. This line will be of value for experimental studies investigating ganglion cells and their axons in vivo and in vitro during the progressive development of glaucoma. PMID:19930759

  3. Synthesis of hydroxyeicosatetraenoic acids (HETE's) by adrenal glomerulosa cells and incorporation into cellular lipids

    SciTech Connect

    Campbell, W.B.; Richards, C.F.; Brady, M.T.; Falck, J.R.

    1986-03-05

    The role of lipoxygenase metabolites of arachidonic acid (AA) in the regulation of aldosterone secretion was studied in isolated rat adrenal glomerulosa cells. Cells were incubated with /sup 14/C-AA in the presence of angiotensin (AII). The media was extracted, metabolites isolated by HPLC, and structures of the metabolites determined by UV absorbance and mass spectrometry. The major products were 12- and 15-HETE with lesser amounts of 11- and 5-HETE. When adrenal cells were incubated with 15-, 12- or 5-HPETE or their respective HETE's (0.03-300nM), there was no significant change in basal or AII-stimulated aldosterone release. Cells were incubated with (/sup 3/H)-AA, -5-HETE, -15-HETE, -12-HETE or -LTB. The cellular lipids were extracted and analyzed by TLC. AA was incorporated into phospholipids (22%), cholesterol esters (50%) and triglycerides (21%). Neither the HETE's or LTB/sub 4/ were incorporated into phospholipids. 5-HETE was taken up into di- and mono-glycerides. The rates of incorporation of AA and 5-HETE were similar (+ 1/2 = 10 min). The incorporation of 5-HETE into glycerol esters did not modify the release of aldosterone by the cells. Thus, while adrenal cells synthesize HETE's, these eicosanoids do not appear to alter the synthesis of aldosterone.

  4. Heterogeneous Expression of the Core Circadian Clock Proteins among Neuronal Cell Types in Mouse Retina

    PubMed Central

    Liu, Xiaoqin; Zhang, Zhijing; Ribelayga, Christophe P.

    2012-01-01

    Circadian rhythms in metabolism, physiology, and behavior originate from cell-autonomous circadian clocks located in many organs and structures throughout the body and that share a common molecular mechanism based on the clock genes and their protein products. In the mammalian neural retina, despite evidence supporting the presence of several circadian clocks regulating many facets of retinal physiology and function, the exact cellular location and genetic signature of the retinal clock cells remain largely unknown. Here we examined the expression of the core circadian clock proteins CLOCK, BMAL1, NPAS2, PERIOD 1(PER1), PERIOD 2 (PER2), and CRYPTOCHROME2 (CRY2) in identified neurons of the mouse retina during daily and circadian cycles. We found concurrent clock protein expression in most retinal neurons, including cone photoreceptors, dopaminergic amacrine cells, and melanopsin-expressing intrinsically photosensitive ganglion cells. Remarkably, diurnal and circadian rhythms of expression of all clock proteins were observed in the cones whereas only CRY2 expression was found to be rhythmic in the dopaminergic amacrine cells. Only a low level of expression of the clock proteins was detected in the rods at any time of the daily or circadian cycle. Our observations provide evidence that cones and not rods are cell-autonomous circadian clocks and reveal an important disparity in the expression of the core clock components among neuronal cell types. We propose that the overall temporal architecture of the mammalian retina does not result from the synchronous activity of pervasive identical clocks but rather reflects the cellular and regional heterogeneity in clock function within retinal tissue. PMID:23189207

  5. Dopamine release from interplexiform cells in the retina: effects of GnRH, FMRFamide, bicuculline, and enkephalin on horizontal cell activity.

    PubMed

    Umino, O; Dowling, J E

    1991-10-01

    In teleost fish, dopaminergic interplexiform cells provide an intraretinal centrifugal pathway from the inner to the outer plexiform layer, where they make abundant synapses on cone-related horizontal cells. The interplexiform cells receive all their input in the inner plexiform layer from centrifugal fibers and amacrine cells. In fish, centrifugal fibers contain gonadotropin hormone-releasing hormone (GnRH)-like and FMRFamide-like peptides (Munz et al., 1982; Stell et al., 1984), whereas amacrine cells contain a variety of neuroactive substances, including a number of peptides. In this study, we examined the effects of GnRH, FMRFamide, bicuculline, and enkephalin on horizontal cell activity in the white perch retina in an attempt to understand the synaptic inputs to the interplexiform cells. When the retina was superfused with Ringer's solution containing GnRH, horizontal cells depolarized (approximately 10 mV), and their responses to small spots increased, whereas their responses to full-field lights decreased. Thus, GnRH closely mimicked the effects of dopamine on horizontal cells. The GnRH antagonist [D-Phe2, Pro3, D-Phe6]-GnRH blocked the effects of GnRH, as did haloperidol. GnRH also had no effect on horizontal cells in retinas treated with 6-hydroxydopamine. The results indicate that GnRH acts by stimulating the release of dopamine from interplexiform cells. FMRFamide alone produced no changes on either the membrane potential or light responses of horizontal cells, but it did suppress the effects of GnRH on horizontal cells in some experiments. FRMFamide also reversed the effects of prolonged darkness on horizontal cell responses. When bicuculline was applied to the retina, horizontal cells also depolarized (approximately 10 mV), responses to full-field illumination decreased, and responses to small spots increased. Most of the effects of bicuculline were suppressed by haloperidol, indicating that bicuculline also stimulates the release of dopamine from

  6. Receptive field properties of bipolar cell axon terminals in direction-selective sublaminas of the mouse retina

    PubMed Central

    Chen, Minggang; Lee, Seunghoon; Park, Silvia J. H.; Looger, Loren L.

    2014-01-01

    Retinal bipolar cells (BCs) transmit visual signals in parallel channels from the outer to the inner retina, where they provide glutamatergic inputs to specific networks of amacrine and ganglion cells. Intricate network computation at BC axon terminals has been proposed as a mechanism for complex network computation, such as direction selectivity, but direct knowledge of the receptive field property and the synaptic connectivity of the axon terminals of various BC types is required in order to understand the role of axonal computation by BCs. The present study tested the essential assumptions of the presynaptic model of direction selectivity at axon terminals of three functionally distinct BC types that ramify in the direction-selective strata of the mouse retina. Results from two-photon Ca2+ imaging, optogenetic stimulation, and dual patch-clamp recording demonstrated that 1) CB5 cells do not receive fast GABAergic synaptic feedback from starburst amacrine cells (SACs); 2) light-evoked and spontaneous Ca2+ responses are well coordinated among various local regions of CB5 axon terminals; 3) CB5 axon terminals are not directionally selective; 4) CB5 cells consist of two novel functional subtypes with distinct receptive field structures; 5) CB7 cells provide direct excitatory synaptic inputs to, but receive no direct GABAergic synaptic feedback from, SACs; and 6) CB7 axon terminals are not directionally selective, either. These findings help to simplify models of direction selectivity by ruling out complex computation at BC terminals. They also show that CB5 comprises two functional subclasses of BCs. PMID:25031256

  7. Receptive field properties of bipolar cell axon terminals in direction-selective sublaminas of the mouse retina.

    PubMed

    Chen, Minggang; Lee, Seunghoon; Park, Silvia J H; Looger, Loren L; Zhou, Z Jimmy

    2014-10-15

    Retinal bipolar cells (BCs) transmit visual signals in parallel channels from the outer to the inner retina, where they provide glutamatergic inputs to specific networks of amacrine and ganglion cells. Intricate network computation at BC axon terminals has been proposed as a mechanism for complex network computation, such as direction selectivity, but direct knowledge of the receptive field property and the synaptic connectivity of the axon terminals of various BC types is required in order to understand the role of axonal computation by BCs. The present study tested the essential assumptions of the presynaptic model of direction selectivity at axon terminals of three functionally distinct BC types that ramify in the direction-selective strata of the mouse retina. Results from two-photon Ca(2+) imaging, optogenetic stimulation, and dual patch-clamp recording demonstrated that 1) CB5 cells do not receive fast GABAergic synaptic feedback from starburst amacrine cells (SACs); 2) light-evoked and spontaneous Ca(2+) responses are well coordinated among various local regions of CB5 axon terminals; 3) CB5 axon terminals are not directionally selective; 4) CB5 cells consist of two novel functional subtypes with distinct receptive field structures; 5) CB7 cells provide direct excitatory synaptic inputs to, but receive no direct GABAergic synaptic feedback from, SACs; and 6) CB7 axon terminals are not directionally selective, either. These findings help to simplify models of direction selectivity by ruling out complex computation at BC terminals. They also show that CB5 comprises two functional subclasses of BCs. PMID:25031256

  8. Efficient generation of retinal progenitor cells from human embryonic stem cells

    PubMed Central

    Lamba, Deepak A.; Karl, Mike O.; Ware, Carol B.; Reh, Thomas A.

    2006-01-01

    The retina is subject to degenerative conditions, leading to blindness. Although retinal regeneration is robust in lower vertebrates, regeneration does not occur in the adult mammalian retina. Thus, we have developed efficient methods for deriving retinal neurons from human embryonic stem (hES) cells. Under appropriate culture conditions, up to 80% of the H1 line can be directed to the retinal progenitor fate, and express a gene expression profile similar to progenitors derived from human fetal retina. The hES cell-derived progenitors differentiate primarily into inner retinal neurons (ganglion and amacrine cells), with functional glutamate receptors. Upon coculture with retinas derived from a mouse model of retinal degeneration, the hES cell derived retinal progenitors integrate with the degenerated mouse retina and increase in their expression of photoreceptor-specific markers. These results demonstrate that human ES cells can be selectively directed to a neural retinal cell fate and thus may be useful in the treatment of retinal degenerations. PMID:16908856

  9. Embryonic Stem Cell-Derived Microvesicles Induce Gene Expression Changes in Müller Cells of the Retina

    PubMed Central

    Katsman, Diana; Stackpole, Emma J.; Domin, Daniel R.; Farber, Debora B.

    2012-01-01

    Cell-derived microvesicles (MVs), recognized as important components of cell-cell communication, contain mRNAs, miRNAs, proteins and lipids and transfer their bioactive contents from parent cells to cells of other origins. We have studied the effect that MVs released from embryonic stem cells (ESMVs) have on retinal progenitor Müller cells. Cultured human Müller cells were exposed to mouse ESMVs every 48 hours for a total of 9 treatments. Morphological changes were observed by light microscopy in the treated cells, which grew as individual heterogeneous cells, compared to the uniform, spindle-like adherent cellular sheets of untreated cells. ESMVs transferred to Müller cells embryonic stem cell (ESC) mRNAs involved in the maintenance of pluripotency, including Oct4 and Sox2, and the miRNAs of the 290 cluster, important regulators of the ESC-specific cell cycle. Moreover, ESMV exposure induced up-regulation of the basal levels of endogenous human Oct4 mRNA in Müller cells. mRNA and miRNA microarrays of ESMV-treated vs. untreated Müller cells revealed the up-regulation of genes and miRNAs involved in the induction of pluripotency, cellular proliferation, early ocular genes and genes important for retinal protection and remodeling, as well as the down-regulation of inhibitory and scar-related genes and miRNAs involved in differentiation and cell cycle arrest. To further characterize the heterogeneous cell population of ESMV-treated Müller cells, their expression of retinal cell markers was compared to that in untreated control cells by immunocytochemistry. Markers for amacrine, ganglion and rod photoreceptors were present in treated but not in control Müller cells. Together, our findings indicate that ESMs induce de-differentiation and pluripotency in their target Müller cells, which may turn on an early retinogenic program of differentiation. PMID:23226281

  10. Short-wavelength cone-opponent retinal ganglion cells in mammals

    PubMed Central

    MARSHAK, DAVID W.; MILLS, STEPHEN L.

    2014-01-01

    In all of the mammalian species studied to date, the short-wavelength-sensitive (S) cones and the S-cone bipolar cells that receive their input are very similar, but the retinal ganglion cells that receive synapses from the S-cone bipolar cells appear to be quite different. Here, we review the literature on mammalian retinal ganglion cells that respond selectively to stimulation of S-cones and respond with opposite polarity to longer wavelength stimuli. There are at least three basic mechanisms to generate these color-opponent responses, including: (1) opponency is generated in the outer plexiform layer by horizontal cells and is conveyed to the ganglion cells via S-cone bipolar cells, (2) inputs from bipolar cells with different cone inputs and opposite response polarity converge directly on the ganglion cells, and (3) inputs from S-cone bipolar cells are inverted by S-cone amacrine cells. These are not mutually exclusive; some mammalian ganglion cells that respond selectively to S-cone stimulation seem to utilize at least two of them. Based on these findings, we suggest that the small bistratified ganglion cells described in primates are not the ancestral type, as proposed previously. Instead, the known types of ganglion cells in this pathway evolved from monostratified ancestral types and became bistratified in some mammalian lineages. PMID:24759445

  11. Short-wavelength cone-opponent retinal ganglion cells in mammals.

    PubMed

    Marshak, David W; Mills, Stephen L

    2014-03-01

    In all of the mammalian species studied to date, the short-wavelength-sensitive (S) cones and the S-cone bipolar cells that receive their input are very similar, but the retinal ganglion cells that receive synapses from the S-cone bipolar cells appear to be quite different. Here, we review the literature on mammalian retinal ganglion cells that respond selectively to stimulation of S-cones and respond with opposite polarity to longer wavelength stimuli. There are at least three basic mechanisms to generate these color-opponent responses, including: (1) opponency is generated in the outer plexiform layer by horizontal cells and is conveyed to the ganglion cells via S-cone bipolar cells, (2) inputs from bipolar cells with different cone inputs and opposite response polarity converge directly on the ganglion cells, and (3) inputs from S-cone bipolar cells are inverted by S-cone amacrine cells. These are not mutually exclusive; some mammalian ganglion cells that respond selectively to S-cone stimulation seem to utilize at least two of them. Based on these findings, we suggest that the small bistratified ganglion cells described in primates are not the ancestral type, as proposed previously. Instead, the known types of ganglion cells in this pathway evolved from monostratified ancestral types and became bistratified in some mammalian lineages. PMID:24759445

  12. Broad Thorny Ganglion Cells: A Candidate for Visual Pursuit Error Signaling in the Primate Retina

    PubMed Central

    Manookin, Michael B.; Neitz, Jay; Rieke, Fred

    2015-01-01

    Functional analyses exist only for a few of the morphologically described primate ganglion cell types, and their correlates in other mammalian species remain elusive. Here, we recorded light responses of broad thorny cells in the whole-mounted macaque retina. They showed ON-OFF-center light responses that were strongly suppressed by stimulation of the receptive field surround. Spike responses were delayed compared with parasol ganglion cells and other ON-OFF cells, including recursive bistratified ganglion cells and A1 amacrine cells. The receptive field structure was shaped by direct excitatory synaptic input and strong presynaptic and postsynaptic inhibition in both ON and OFF pathways. The cells responded strongly to dark or bright stimuli moving either in or out of the receptive field, independent of the direction of motion. However, they did not show a maintained spike response either to a uniform background or to a drifting plaid pattern. These properties could be ideally suited for guiding movements involved in visual pursuit. The functional characteristics reported here permit the first direct cross-species comparison of putative homologous ganglion cell types. Based on morphological similarities, broad thorny ganglion cells have been proposed to be homologs of rabbit local edge detector ganglion cells, but we now show that the two cells have quite distinct physiological properties. Thus, our data argue against broad thorny cells as the homologs of local edge detector cells. PMID:25834063

  13. Mesenchymal Stromal Cells Prevent Renal Fibrosis in a Rat Model of Unilateral Ureteral Obstruction by Suppressing the Renin-Angiotensin System via HuR

    PubMed Central

    Gregorini, Marilena; Corradetti, Valeria; Rocca, Chiara; Pattonieri, Eleonora Francesca; Valsania, Teresa; Milanesi, Samantha; Serpieri, Nicoletta; Bedino, Giulia; Esposito, Pasquale; Libetta, Carmelo; Avanzini, Maria Antonietta; Mantelli, Melissa; Ingo, Daniela; Peressini, Sabrina; Albertini, Riccardo; Dal Canton, Antonio; Rampino, Teresa

    2016-01-01

    We studied Mesenchymal Stromal Cells (MSC) effects in experimental Unilateral Ureteral Obstruction (UUO), a fibrogenic renal disease. Rats were divided in 5 groups: sham, UUO, MSC treated-UUO, ACEi treated-UUO, MSC+ACEi treated- UUO. Data were collected at 1, 7, 21 days. UUO induced monocyte renal infiltration, tubular cell apoptosis, tubular atrophy, interstitial fibrosis and overexpression of TGFβ, Renin mRNA (RENmRNA), increase of Renin, Angiotensin II (AII) and aldosterone serum levels. Both lisinopril (ACEi) and MSC treatment prevented monocyte infiltration, reduced tubular cell apoptosis, renal fibrosis and TGFβ expression. Combined therapy provided a further suppression of monocyte infiltration and tubular injury. Lisinopril alone caused a rebound activation of Renin-Angiotensin System (RAS), while MSC suppressed RENmRNA and Renin synthesis and induced a decrease of AII and aldosterone serum levels. Furthermore, in in-vitro and in-vivo experiments, MSC inhibit Human antigen R (HuR) trascription, an enhancer of RENmRNA stability by IL10 release. In conclusion, we demonstrate that in UUO MSC prevent fibrosis, by decreasing HuR-dependent RENmRNA stability. Our findings give a clue to understand the molecular mechanism through which MSC may prevent fibrosis in a wide and heterogeneous number of diseases that share RAS activation as common upstream pathogenic mechanism. PMID:26866372

  14. Calcium buffer proteins are specific markers of human retinal neurons.

    PubMed

    Kántor, Orsolya; Mezey, Szilvia; Adeghate, Jennifer; Naumann, Angela; Nitschke, Roland; Énzsöly, Anna; Szabó, Arnold; Lukáts, Ákos; Németh, János; Somogyvári, Zoltán; Völgyi, Béla

    2016-07-01

    Ca(2+)-buffer proteins (CaBPs) modulate the temporal and spatial characteristics of transient intracellular Ca(2+)-concentration changes in neurons in order to fine-tune the strength and duration of the output signal. CaBPs have been used as neurochemical markers to identify and trace neurons of several brain loci including the mammalian retina. The CaBP content of retinal neurons, however, varies between species and, thus, the results inferred from animal models cannot be utilised directly by clinical ophthalmologists. Moreover, the shortage of well-preserved human samples greatly impedes human retina studies at the cellular and network level. Our purpose has therefore been to examine the distribution of major CaBPs, including calretinin, calbindin-D28, parvalbumin and the recently discovered secretagogin in exceptionally well-preserved human retinal samples. Based on a combination of immunohistochemistry, Neurolucida tracing and Lucifer yellow injections, we have established a database in which the CaBP marker composition can be defined for morphologically identified cell types of the human retina. Hence, we describe the full CaBP make-up for a number of human retinal neurons, including HII horizontal cells, AII amacrine cells, type-1 tyrosine-hydroxylase-expressing amacrine cells and other lesser known neurons. We have also found a number of unidentified cells whose morphology remains to be characterised. We present several examples of the colocalisation of two or three CaBPs with slightly different subcellular distributions in the same cell strongly suggesting a compartment-specific division of labour of Ca(2+)-buffering by CaBPs. Our work thus provides a neurochemical framework for future ophthalmological studies and renders new information concerning the cellular and subcellular distribution of CaBPs for experimental neuroscience. PMID:26899253

  15. Quorum quenching activity in cell-free lysate of endophytic bacteria isolated from Pterocarpus santalinus Linn., and its effect on quorum sensing regulated biofilm in Pseudomonas aeruginosa PAO1.

    PubMed

    Rajesh, P S; Ravishankar Rai, V

    2014-01-01

    Quorum sensing mechanism allows the microorganisms to resist the antibiotic treatment by forming biofilms. Quorum quenching is one of the mechanisms to control the development of drug resistance in microbes. Endophyte bacteria are beneficial to plant growth as they support the immune system against the pathogen attack. The endophytic bacteria present in Pterocarpus santalinus were screened for the presence of N-acyl homoserine lactones (AHLs) degrading bacteria using biosensor strains and further confirmed by quantifying the violacein production. Cell-free lysate of endophytic bacteria, Bacillus firmus PT18 and Enterobacter asburiae PT39 exhibited potent AHL degrading ability by inhibiting about 80% violacein production in biosensor strain. Furthermore, when the cell-free lysate was applied to Pseudomonas aeruginosa PAO1 and PAO1-JP2 biofilm it resulted in significant (p<0.01) inhibition of biofilm formation. The biofilm inhibition was confirmed by visualization of biofilm slides under fluorescence microscopy, which showed decrease in total biomass formation in treated slides. Isolation and amplification of the gene (aiiA) indicated that the presence of AHL lactonase in cell-free lysate and sequence alignment indicated that AiiA contains a "HXHXDH" zinc-binding motif that is being conserved in several groups of metallohydrolases. Therefore, the study shows the potential of AHLs degradation by AHL lactonase present in cell-free lysate of isolated endophytic bacteria and inhibition of quorum sensing regulated biofilm formation in P. aeruginosa PAO1. PMID:24268182

  16. Two polysialic acid synthases, mouse ST8Sia II and IV, synthesize different degrees of polysialic acids on different substrate glycoproteins in mouse neuroblastoma Neuro2a cells.

    PubMed

    Kojima, N; Tachida, Y; Tsuji, S

    1997-12-01

    We previously cloned cDNAs encoding two different polysialic acid (PSA) synthases, ST8Sia II and IV, from mouse, and showed that both mouse ST8Sia II and IV can synthesize PSA on the neural cell adhesion molecule (NCAM) as well as other glycoproteins such as fetuin, at least in vitro (Kojima, N., Tachida, Y., Yoshida, Y., and Tsuji, S. (1996) J. Biol. Chem. 271, 19457-19463]. In the present study, to clarify how the two PSA synthases act differently in vivo, we first cloned PSA-expressing cell lines (N2a-II and N2a-IV) by stable transfection of the cDNA encoding either mST8Sia II or IV into mouse neuroblastoma Neuro2a cells, which do not express PSA but express NCAM, then compared the expression of the PSA and NCAM isoforms and de novo synthesis of PSA between N2a-II and N2a-IV. Western blotting with an anti-NCAM polyclonal antibody showed that NCAM was expressed as the polysialylated form in both ST8Sia II cDNA-transfected and ST8Sia IV cDNA-transfected Neuro2a cells, but that the polysialylated NCAMs expressed in ST8Sia IV cDNA-transfected clones migrated much slower on SDS-PAGE than those expressed in ST8Sia II cDNA-transfected clones. The slower migration of polysialylated NCAM of the ST8Sia IV cDNA-transfected clone (N2a-IV) than that of the ST8Sia II cDNA-transfected clone (N2a-II) was also observed when cells were metabolically labeled with [3H]glucosamine or pulse-chase labeled with [35S] methionine followed by immunoprecipitation with anti-PSA antibody or anti-NCAM monoclonal antibody. In addition, polysialylated N-glycans of PSA-carrying glycoproteins prepared from [3H] glucosamine-labeled N2a-IV by immunoprecipitation with anti-PSA monoclonal antibody were eluted at a much higher salt concentration than those from [3H] glucosamine-labeled N2a-II on an anion-exchange column. These results indicated that the degree of de novo polysialylation of NCAM by mST8Sia IV was much higher than that by mST8Sia II. In N2a-IV, NCAM-120, -140, and -180 were expressed as

  17. Evaluation of the percentage of ganglion cells in the ganglion cell layer of the rodent retina

    PubMed Central

    Schlamp, Cassandra L.; Montgomery, Angela D.; Mac Nair, Caitlin E.; Schuart, Claudia; Willmer, Daniel J.

    2013-01-01

    Purpose Retinal ganglion cells comprise a percentage of the neurons actually residing in the ganglion cell layer (GCL) of the rodent retina. This estimate is useful to extrapolate ganglion cell loss in models of optic nerve disease, but the values reported in the literature are highly variable depending on the methods used to obtain them. Methods We tested three retrograde labeling methods and two immunostaining methods to calculate ganglion cell number in the mouse retina (C57BL/6). Additionally, a double-stain retrograde staining method was used to label rats (Long-Evans). The number of total neurons was estimated using a nuclear stain and selecting for nuclei that met specific criteria. Cholinergic amacrine cells were identified using transgenic mice expressing Tomato fluorescent protein. Total neurons and total ganglion cell numbers were measured in microscopic fields of 104 µm2 to determine the percentage of neurons comprising ganglion cells in each field. Results Historical estimates of the percentage of ganglion cells in the mouse GCL range from 36.1% to 67.5% depending on the method used. Experimentally, retrograde labeling methods yielded a combined estimate of 50.3% in mice. A retrograde method also yielded a value of 50.21% for rat retinas. Immunolabeling estimates were higher at 64.8%. Immunolabeling may introduce overestimates, however, with non-specific labeling effects, or ectopic expression of antigens in neurons other than ganglion cells. Conclusions Since immunolabeling methods may overestimate ganglion cell numbers, we conclude that 50%, which is consistently derived from retrograde labeling methods, is a reliable estimate of the ganglion cells in the neuronal population of the GCL. PMID:23825918

  18. DSCAM Promotes Refinement in the Mouse Retina through Cell Death and Restriction of Exploring Dendrites

    PubMed Central

    Li, Shuai; Sukeena, Joshua M.; Simmons, Aaron B.; Hansen, Ethan J.; Nuhn, Renee E.; Samuels, Ivy S.

    2015-01-01

    In this study we develop and use a gain-of-function mouse allele of the Down syndrome cell adhesion molecule (Dscam) to complement loss-of-function models. We assay the role of Dscam in promoting cell death, spacing, and laminar targeting of neurons in the developing mouse retina. We find that ectopic or overexpression of Dscam is sufficient to drive cell death. Gain-of-function studies indicate that Dscam is not sufficient to increase spatial organization, prevent cell-to-cell pairing, or promote active avoidance in the mouse retina, despite the similarity of the Dscam loss-of-function phenotype in the mouse retina to phenotypes observed in Drosophila Dscam1 mutants. Both gain- and loss-of-function studies support a role for Dscam in targeting neurites; DSCAM is necessary for precise dendrite lamination, and is sufficient to retarget neurites of outer retinal cells after ectopic expression. We further demonstrate that DSCAM guides dendrite targeting in type 2 dopaminergic amacrine cells, by restricting the stratum in which exploring retinal dendrites stabilize, in a Dscam dosage-dependent manner. Based on these results we propose a single model to account for the numerous Dscam gain- and loss-of-function phenotypes reported in the mouse retina whereby DSCAM eliminates inappropriately placed cells and connections. PMID:25855178

  19. Synaptotagmin-7 Is Essential for Ca2+-Triggered Delayed Asynchronous Release But Not for Ca2+-Dependent Vesicle Priming in Retinal Ribbon Synapses

    PubMed Central

    Bacaj, Taulant

    2015-01-01

    Most synapses release neurotransmitters in two phases: (1) a fast synchronous phase lasting a few milliseconds; and (2) a delayed “asynchronous” phase lasting hundreds of milliseconds. Ca2+ triggers fast synchronous neurotransmitter release by binding to synaptotagmin-1, synaptotagmin-2, or synaptotagmin-9, but how Ca2+ triggers delayed asynchronous release has long remained enigmatic. Recent results suggested that consistent with the Ca2+-sensor function of synaptotagmin-7 in neuroendocrine exocytosis, synaptotagmin-7 also functions as a Ca2+ sensor for synaptic vesicle exocytosis but operates during delayed asynchronous release. Puzzlingly, a subsequent study postulated that synaptotagmin-7 is not a Ca2+ sensor for release but mediates Ca2+-dependent vesicle repriming after intense stimulation. To address these issues, we here analyzed synaptic transmission at rod bipolar neuron–AII amacrine cell synapses in acute mouse retina slices as a model system. Using paired recordings, we show that knock-out of synaptotagmin-7 selectively impairs delayed asynchronous release but not fast synchronous release. Delayed asynchronous release was blocked in wild-type synapses by intracellular addition of high concentrations of the slow Ca2+-chelator EGTA, but EGTA had no effect in synaptotagmin-7 knock-out neurons because delayed asynchronous release was already impaired. Moreover, direct measurements of vesicle repriming failed to uncover an effect of the synaptotagmin-7 knock-out on vesicle repriming. Our data demonstrate that synaptotagmin-7 is selectively essential for Ca2+-dependent delayed asynchronous release in retinal rod bipolar cell synapses, that its function can be blocked by simply introducing a slow Ca2+ buffer into the cells, and that synaptotagmin-7 is not required for normal vesicle repriming. SIGNIFICANCE STATEMENT How Ca2+ triggers delayed asynchronous release has long remained enigmatic. Synaptotagmin-7 has been implicated recently as Ca2+ sensor in

  20. A synaptic mechanism for retinal adaptation to luminance and contrast.

    PubMed

    Jarsky, Tim; Cembrowski, Mark; Logan, Stephen M; Kath, William L; Riecke, Hermann; Demb, Jonathan B; Singer, Joshua H

    2011-07-27

    The gain of signaling in primary sensory circuits is matched to the stimulus intensity by the process of adaptation. Retinal neural circuits adapt to visual scene statistics, including the mean (background adaptation) and the temporal variance (contrast adaptation) of the light stimulus. The intrinsic properties of retinal bipolar cells and synapses contribute to background and contrast adaptation, but it is unclear whether both forms of adaptation depend on the same cellular mechanisms. Studies of bipolar cell synapses identified synaptic mechanisms of gain control, but the relevance of these mechanisms to visual processing is uncertain because of the historical focus on fast, phasic transmission rather than the tonic transmission evoked by ambient light. Here, we studied use-dependent regulation of bipolar cell synaptic transmission evoked by small, ongoing modulations of membrane potential (V(M)) in the physiological range. We made paired whole-cell recordings from rod bipolar (RB) and AII amacrine cells in a mouse retinal slice preparation. Quasi-white noise voltage commands modulated RB V(M) and evoked EPSCs in the AII. We mimicked changes in background luminance or contrast, respectively, by depolarizing the V(M) or increasing its variance. A linear systems analysis of synaptic transmission showed that increasing either the mean or the variance of the presynaptic V(M) reduced gain. Further electrophysiological and computational analyses demonstrated that adaptation to mean potential resulted from both Ca channel inactivation and vesicle depletion, whereas adaptation to variance resulted from vesicle depletion alone. Thus, background and contrast adaptation apparently depend in part on a common synaptic mechanism. PMID:21795549

  1. Cellular Distribution and Subcellular Localization of Molecular Components of Vesicular Transmitter Release in Horizontal Cells of Rabbit Retina

    PubMed Central

    HIRANO, ARLENE A.; BRANDSTÄTTER, JOHANN H.; BRECHA, NICHOLAS C.

    2010-01-01

    The mechanism underlying transmitter release from retinal horizontal cells is poorly understood. We investigated the possibility of vesicular transmitter release from mammalian horizontal cells by examining the expression of synaptic proteins that participate in vesicular transmitter release at chemical synapses. Using immunocytochemistry, we evaluated the cellular and subcellular distribution of complexin I/II, syntaxin-1, and synapsin I in rabbit retina. Strong labeling for complexin I/II, proteins that regulate a late step in vesicular transmitter release, was found in both synaptic layers of the retina, and in somata of A- and B-type horizontal cells, of γ-aminobutyric acid (GABA)- and glycinergic amacrine cells, and of ganglion cells. Immunoelectron microscopy demonstrated the presence of complexin I/II in horizontal cell processes postsynaptic to rod and cone ribbon synapses. Syntaxin-1, a core protein of the soluble N-ethylmaleimide-sensitive-factor attachment protein receptor (SNARE) complex known to bind to complexin, and synapsin I, a synaptic vesicle-associated protein involved in the Ca2+-dependent recruitment of synaptic vesicles for transmitter release, were also present in the horizontal cells and their processes at photoreceptor synapses. Photoreceptors and bipolar cells did not express any of these proteins at their axon terminals. The presence of complexin I/II, syntaxin-1, and synapsin I in rabbit horizontal cell processes and tips suggests that a vesicular mechanism may underlie transmitter release from mammalian horizontal cells. PMID:15912504

  2. Transition of differential histone H3 methylation in photoreceptors and other retinal cells during retinal differentiation

    PubMed Central

    Ueno, Kazuko; Iwagawa, Toshiro; Kuribayashi, Hiroshi; Baba, Yukihiro; Nakauchi, Hiromitsu; Murakami, Akira; Nagasaki, Masao; Suzuki, Yutaka; Watanabe, Sumiko

    2016-01-01

    To analyze cell lineage-specific transitions in global transcriptional and epigenetic changes during retinogenesis, we purified retinal cells from normal mice during postnatal development into two fractions, namely, photoreceptors and other retinal cells, based on Cd73 expression, and performed RNA sequencing and ChIP sequencing of H3K27me3 and H3K4me3. Genes expressed in the photoreceptor lineage were marked with H3K4me3 in the Cd73-positive cell fraction; however, the level of H3K27me3 was very low in both Cd73-positive and -negative populations. H3K27me3 may be involved in spatio-temporal onset of a subset of bipolar-related genes. Subsets of genes expressed in amacrine and retinal ganglion cells, which are early-born retinal cell types, were suggested to be maintained in a silent state by H3K27me3 during late-stage retinogenesis. In the outer nuclear layer, upregulation of Rho and rod-related genes were observed in Ezh2-ablated retina, suggesting a role for H3K27me3 in the maintenance of proper expression levels. Taken together, our data on the transition of lineage-specific molecular signatures during development suggest that histone methylation is involved in retinal differentiation and maintenance through cell lineage-specific mechanisms. PMID:27377164

  3. Transition of differential histone H3 methylation in photoreceptors and other retinal cells during retinal differentiation.

    PubMed

    Ueno, Kazuko; Iwagawa, Toshiro; Kuribayashi, Hiroshi; Baba, Yukihiro; Nakauchi, Hiromitsu; Murakami, Akira; Nagasaki, Masao; Suzuki, Yutaka; Watanabe, Sumiko

    2016-01-01

    To analyze cell lineage-specific transitions in global transcriptional and epigenetic changes during retinogenesis, we purified retinal cells from normal mice during postnatal development into two fractions, namely, photoreceptors and other retinal cells, based on Cd73 expression, and performed RNA sequencing and ChIP sequencing of H3K27me3 and H3K4me3. Genes expressed in the photoreceptor lineage were marked with H3K4me3 in the Cd73-positive cell fraction; however, the level of H3K27me3 was very low in both Cd73-positive and -negative populations. H3K27me3 may be involved in spatio-temporal onset of a subset of bipolar-related genes. Subsets of genes expressed in amacrine and retinal ganglion cells, which are early-born retinal cell types, were suggested to be maintained in a silent state by H3K27me3 during late-stage retinogenesis. In the outer nuclear layer, upregulation of Rho and rod-related genes were observed in Ezh2-ablated retina, suggesting a role for H3K27me3 in the maintenance of proper expression levels. Taken together, our data on the transition of lineage-specific molecular signatures during development suggest that histone methylation is involved in retinal differentiation and maintenance through cell lineage-specific mechanisms. PMID:27377164

  4. Developmental mechanisms that regulate retinal ganglion cell dendritic morphology

    PubMed Central

    Tian, Ning

    2011-01-01

    One of the fundamental features of retinal ganglion cells (RGCs) is that dendrites of individual RGCs are confined to one or a few narrow strata within the inner plexiform layer (IPL), and each RGC synapses only with a small group of presynaptic bipolar and amacrine cells with axons/dendrites ramified in the same strata to process distinct visual features. The underlying mechanisms which control the development of this laminar-restricted distribution pattern of RGC dendrites have been extensively studied, and it is still an open question whether the dendritic pattern of RGCs is determined by molecular cues or by activity-dependent refinement. Accumulating evidence suggests that both molecular cues and activity-dependent refinement might regulate RGC dendrites in a cell subtype-specific manner. However, identification of morphological subtypes of RGCs before they have achieved their mature dendritic pattern is a major challenge in the study of RGC dendritic development. This problem is now being circumvented through the use of molecular markers in genetically engineered mouse lines to identify RGC subsets early during development. Another unanswered fundamental question in the study of activity-dependent refinement of RGC dendrites is how changes in synaptic activity lead to the changes in dendritic morphology. Recent studies have started to shed light on the molecular basis of activity-dependent dendritic refinement of RGCs by showing that some molecular cascades control the cytoskeleton reorganization of RGCs. PMID:21542137

  5. Ionotropic glutamate receptors mediate OFF responses in light-adapted ON bipolar cells

    PubMed Central

    Pang, Ji-Jie; Gao, Fan; Wu, Samuel M.

    2013-01-01

    Previous studies have suggested that photoreceptor synaptic inputs to depolarizing bipolar cells (DBCs or ON bipolar cells) are mediated by mGluR6 receptors and those to hyperpolarizing bipolar cells (HBCs or OFF bipolar cells) are mediated by AMPA/kainate receptors. Here we show that in addition to mGluR6 receptors which mediate the sign-inverting, depolarizing light responses, subpopulations of cone-dominated and rod/cone mixed DBCs use GluR4 AMPA receptors to generate a transient sign-preserving OFF response under light adapted conditions. These AMPA receptors are located at the basal junctions postsynaptic to rods and they are silent under dark-adapted conditions, as tonic glutamate release in darkness desensitizes these receptors. Light adaptation enhances rod-cone coupling and thus allows cone photocurrents with an abrupt OFF depolarization to enter the rods. The abrupt rod depolarization triggers glutamate activation of unoccupied AMPA receptors, resulting in a transient OFF response in DBCs. It has been widely accepted that the DNQX-sensitive, OFF transient responses in retinal amacrine cells and ganglion cells are mediated exclusively by HBCs. Our results suggests that this view needs revision as AMPA receptors in subpopulations of DBCs are likely to significantly contribute to the DNQX-sensitive OFF transient responses in light-adapted third- and higher-order visual neurons. PMID:22842089

  6. Inner retinal inhibition shapes the receptive field of retinal ganglion cells in primate

    PubMed Central

    Protti, D A; Di Marco, S; Huang, J Y; Vonhoff, C R; Nguyen, V; Solomon, S G

    2014-01-01

    Abstract The centre–surround organisation of receptive fields is a feature of most retinal ganglion cells (RGCs) and is critical for spatial discrimination and contrast detection. Although lateral inhibitory processes are known to be important in generating the receptive field surround, the contribution of each of the two synaptic layers in the primate retina remains unclear. Here we studied the spatial organisation of excitatory and inhibitory synaptic inputs onto ON and OFF ganglion cells in the primate retina. All RGCs showed an increase in excitation in response to stimulus of preferred polarity. Inhibition onto RGCs comprised two types of responses to preferred polarity: some RGCs showed an increase in inhibition whilst others showed removal of tonic inhibition. Excitatory inputs were strongly spatially tuned but inhibitory inputs showed more variable organisation: in some neurons they were as strongly tuned as excitation, and in others inhibitory inputs showed no spatial tuning. We targeted one source of inner retinal inhibition by functionally ablating spiking amacrine cells with bath application of tetrodotoxin (TTX). TTX significantly reduced the spatial tuning of excitatory inputs. In addition, TTX reduced inhibition onto those RGCs where a stimulus of preferred polarity increased inhibition. Reconstruction of the spatial tuning properties by somatic injection of excitatory and inhibitory synaptic conductances verified that TTX-mediated inhibition onto bipolar cells increases the strength of the surround in RGC spiking output. These results indicate that in the primate retina inhibitory mechanisms in the inner plexiform layer sharpen the spatial tuning of ganglion cells. PMID:24042496

  7. Receptive fields of retinal bipolar cells are mediated by heterogeneous synaptic circuitry

    PubMed Central

    Zhang, Ai-Jun; Wu, Samuel M.

    2009-01-01

    Center-surround antagonistic receptive field (CSARF) organization is the basic synaptic circuit that serves as elementary building blocks for spatial information processing in the visual system. Cells with such receptive fields converge into higher-order visual neurons to form more complex receptive fields. Retinal bipolar cells (BCs) are the first neurons along the visual pathway that exhibit CSARF organization. BCs have been classified according to their response polarities and rod/cone inputs, and they project signals to target cells at different sublaminae of the inner plexiform layer. On the other hand, CSARFs of various types of BCs have been assumed be organized the same way. Here we examined center and surround responses of over 250 salamander BCs, and demonstrated that different types of BCs exhibit different patterns of dye coupling, receptive field center size, surround response strength, and conductance changes associated with center and surround responses. We show that BC receptive field center sizes varied with the degree of BC-BC coupling, and that surround responses of different BCs are mediated by different combinations of five lateral synaptic pathways mediated by the horizontal cells and amacrine cells. The finding of heterogeneous receptive field circuitry fundamentally challenges the common assumption that CSARFs of different subtypes of visual neurons are mediated by the same synaptic pathways. BCs carrying different visual signals use different synaptic circuits to process spatial information, allowing shape and contrast computation be differentially modulated by various lighting and adaptation conditions. PMID:19158304

  8. Identification of Protein Interaction Partners in Mammalian Cells Using SILAC-immunoprecipitation Quantitative Proteomics

    PubMed Central

    Emmott, Edward; Goodfellow, Ian

    2014-01-01

    Quantitative proteomics combined with immuno-affinity purification, SILAC immunoprecipitation, represent a powerful means for the discovery of novel protein:protein interactions. By allowing the accurate relative quantification of protein abundance in both control and test samples, true interactions may be easily distinguished from experimental contaminants. Low affinity interactions can be preserved through the use of less-stringent buffer conditions and remain readily identifiable. This protocol discusses the labeling of tissue culture cells with stable isotope labeled amino acids, transfection and immunoprecipitation of an affinity tagged protein of interest, followed by the preparation for submission to a mass spectrometry facility. This protocol then discusses how to analyze and interpret the data returned from the mass spectrometer in order to identify cellular partners interacting with a protein of interest. As an example this technique is applied to identify proteins binding to the eukaryotic translation initiation factors: eIF4AI and eIF4AII. PMID:25046639

  9. Purification of an angiotensin II binding protein by using antibodies to a peptide encoded by angiotensin II complementary RNA

    SciTech Connect

    Elton, T.S.; Dion, L.D.; Bost, K.L.; Oparil, S.; Blalock, J.E.

    1988-04-01

    The authors have generated a monospecific antibody to a synthetic peptide encoded by an RNA complementary to the mRNA for angiotensin II (AII) and determined whether this antibody recognizes the AII receptor. They demonstrate that the antibody competes specifically with /sup 125/I-labeled AII for the same binding site on rat adrenal membranes. Furthermore, they show this antibody inhibits the secretion of aldosterone from cultured rat adrenal cells, suggesting that the antibody recognizes the biologically relevant AII receptor. Finally, they demonstrate that antibody to the complementary peptide can be used to immunoaffinity-purify a protein of M/sub r/ 66,000 that specifically binds radiolabeled AII.

  10. A high frequency resonance in the responses of retinal ganglion cells to rapidly modulated stimuli: a computer model.

    PubMed

    Miller, J A; Denning, K S; George, J S; Marshak, D W; Kenyon, G T

    2006-01-01

    Brisk Y-type ganglion cells in the cat retina exhibit a high frequency resonance (HFR) in their responses to large, rapidly modulated stimuli. We used a computer model to test whether negative feedback mediated by axon-bearing amacrine cells onto ganglion cells could account for the experimentally observed properties of HFRs. Temporal modulation transfer functions (tMTFs) recorded from model ganglion cells exhibited HFR peaks whose amplitude, width, and locations were qualitatively consistent with experimental data. Moreover, the wide spatial distribution of axon-mediated feedback accounted for the observed increase in HFR amplitude with stimulus size. Model phase plots were qualitatively similar to those recorded from Y ganglion cells, including an anomalous phase advance that in our model coincided with the amplification of low-order harmonics that overlapped the HFR peak. When axon-mediated feedback in the model was directed primarily to bipolar cells, whose synaptic output was graded, or else when the model was replaced with a simple cascade of linear filters, it was possible to produce large HFR peaks but the region of anomalous phase advance was always eliminated, suggesting the critical involvement of strongly non-linear feedback loops. To investigate whether HFRs might contribute to visual processing, we simulated high frequency ocular tremor by rapidly modulating a naturalistic image. Visual signals riding on top of the imposed jitter conveyed an enhanced representation of large objects. We conclude that by amplifying responses to ocular tremor, HFRs may selectively enhance the processing of large image features. PMID:17020633

  11. Multiple Components of Ganglion Cell Desensitization in Response to Prosthetic Stimulation

    PubMed Central

    Freeman, Daniel K; Fried, Shelley I

    2011-01-01

    Retinal prostheses aim to restore functional vision to those blinded by outer retinal diseases using electric stimulation of surviving neurons. Previous work indicates that repetitive stimulation with stimuli that activate the synaptic network reduces the sensitivity of retinal neurons to further stimulation. Such desensitization may contribute to the fading of visual percepts over time reported by human subjects. Here, we show that desensitization may be more complex than previously considered. We recorded spike trains from rabbit retinal ganglion cells and found that desensitization persists in the presence of inhibitory blockers (strychnine and picrotoxin), indicating amacrine cell inhibition is not solely responsible for reducing sensitivity in response to electric stimulation. The threshold for direct activation of the ganglion cell changes little during the simultaneous desensitization of the synaptically mediated response, indicating that desensitization likely occurs upstream of the spike generator. In addition to the rapid desensitization acting over hundreds of milliseconds (τ = 176.4 ± 8.8ms), we report the presence of a slow acting desensitization with a time course of seconds (τ = 14.0 ± 1.1sec). The time course of the two components of desensitization that we found are similar to the two phases of brightness fading seen in human subjects. This suggests that the reduction in ganglion cell firing due to desensitization may be responsible for the fading of visual percepts over time in response to prosthetic stimulation. PMID:21248379

  12. Retinal Connectomics: Towards Complete, Accurate Networks

    PubMed Central

    Marc, Robert E.; Jones, Bryan W.; Watt, Carl B.; Anderson, James R.; Sigulinsky, Crystal; Lauritzen, Scott

    2013-01-01

    Connectomics is a strategy for mapping complex neural networks based on high-speed automated electron optical imaging, computational assembly of neural data volumes, web-based navigational tools to explore 1012–1015 byte (terabyte to petabyte) image volumes, and annotation and markup tools to convert images into rich networks with cellular metadata. These collections of network data and associated metadata, analyzed using tools from graph theory and classification theory, can be merged with classical systems theory, giving a more completely parameterized view of how biologic information processing systems are implemented in retina and brain. Networks have two separable features: topology and connection attributes. The first findings from connectomics strongly validate the idea that the topologies complete retinal networks are far more complex than the simple schematics that emerged from classical anatomy. In particular, connectomics has permitted an aggressive refactoring of the retinal inner plexiform layer, demonstrating that network function cannot be simply inferred from stratification; exposing the complex geometric rules for inserting different cells into a shared network; revealing unexpected bidirectional signaling pathways between mammalian rod and cone systems; documenting selective feedforward systems, novel candidate signaling architectures, new coupling motifs, and the highly complex architecture of the mammalian AII amacrine cell. This is but the beginning, as the underlying principles of connectomics are readily transferrable to non-neural cell complexes and provide new contexts for assessing intercellular communication. PMID:24016532

  13. Aberrant Activity in Degenerated Retinas Revealed by Electrical Imaging

    PubMed Central

    Zeck, Günther

    2016-01-01

    In this review, I present and discuss the current understanding of aberrant electrical activity found in the ganglion cell layer (GCL) of rod-degenerated (rd) mouse retinas. The reported electrophysiological properties revealed by electrical imaging using high-density microelectrode arrays can be subdivided between spiking activity originating from retinal ganglion cells (RGCs) and local field potentials (LFPs) reflecting strong trans-membrane currents within the GCL. RGCs in rd retinas show increased and rhythmic spiking compared to age-matched wild-type retinas. Fundamental spiking frequencies range from 5 to 15 Hz in various mouse models. The rhythmic RGC spiking is driven by a presynaptic network comprising AII amacrine and bipolar cells. In the healthy retina this rhythm-generating circuit is inhibited by photoreceptor input. A unique physiological feature of rd retinas is rhythmic LFP manifested as spatially-restricted low-frequency (5–15 Hz) voltage changes. Their spatiotemporal characterization revealed propagation and correlation with RGC spiking. LFPs rely on gap-junctional coupling and are shaped by glycinergic and by GABAergic transmission. The aberrant RGC spiking and LFPs provide a simple readout of the functionality of the remaining retinal circuitry which can be used in the development of improved vision restoration strategies. PMID:26903810

  14. Retinal connectomics: towards complete, accurate networks.

    PubMed

    Marc, Robert E; Jones, Bryan W; Watt, Carl B; Anderson, James R; Sigulinsky, Crystal; Lauritzen, Scott

    2013-11-01

    Connectomics is a strategy for mapping complex neural networks based on high-speed automated electron optical imaging, computational assembly of neural data volumes, web-based navigational tools to explore 10(12)-10(15) byte (terabyte to petabyte) image volumes, and annotation and markup tools to convert images into rich networks with cellular metadata. These collections of network data and associated metadata, analyzed using tools from graph theory and classification theory, can be merged with classical systems theory, giving a more completely parameterized view of how biologic information processing systems are implemented in retina and brain. Networks have two separable features: topology and connection attributes. The first findings from connectomics strongly validate the idea that the topologies of complete retinal networks are far more complex than the simple schematics that emerged from classical anatomy. In particular, connectomics has permitted an aggressive refactoring of the retinal inner plexiform layer, demonstrating that network function cannot be simply inferred from stratification; exposing the complex geometric rules for inserting different cells into a shared network; revealing unexpected bidirectional signaling pathways between mammalian rod and cone systems; documenting selective feedforward systems, novel candidate signaling architectures, new coupling motifs, and the highly complex architecture of the mammalian AII amacrine cell. This is but the beginning, as the underlying principles of connectomics are readily transferrable to non-neural cell complexes and provide new contexts for assessing intercellular communication. PMID:24016532

  15. Neets'aii Gwiindaii: Living in the Chandalar Country.

    ERIC Educational Resources Information Center

    Peter, Katherine

    The book is Katherine Peter's own account, in her native Gwich'in Athabaskan language, of life in 1936-47 in Arctic Village, Alaska. An introductory section offers background information on the author's life and the origins of this work. The text that follows is in Gwich'in Athabaskan on the left page, and in English on the right page. It tells of…

  16. Photoresponse diversity among the five types of intrinsically photosensitive retinal ganglion cells.

    PubMed

    Zhao, Xiwu; Stafford, Ben K; Godin, Ashley L; King, W Michael; Wong, Kwoon Y

    2014-04-01

    Intrinsically photosensitive retinal ganglion cells (ipRGCs) mediate non-image-forming visual responses, including pupillary constriction, circadian photoentrainment and suppression of pineal melatonin secretion. Five morphological types of ipRGCs, M1-M5, have been identified in mice. In order to understand their functions better, we studied the photoresponses of all five cell types, by whole-cell recording from fluorescently labelled ipRGCs visualized using multiphoton microscopy. All ipRGC types generated melanopsin-based ('intrinsic') as well as synaptically driven ('extrinsic') light responses. The intrinsic photoresponses of M1 cells were lower threshold, higher amplitude and faster than those of M2-M5. The peak amplitudes of extrinsic light responses differed among the ipRGC types; however, the responses of all cell types had comparable thresholds, kinetics and waveforms, and all cells received rod input. While all five types exhibited inhibitory amacrine-cell and excitatory bipolar-cell inputs from the 'on' channel, M1 and M3 received additional 'off'-channel inhibition, possibly through their 'off'-sublamina dendrites. The M2-M5 ipRGCs had centre-surround-organized receptive fields, implicating a capacity to detect spatial contrast. In contrast, the receptive fields of M1 cells lacked surround antagonism, which might be caused by the surround of the inhibitory input nullifying the surround of the excitatory input. All ipRGCs responded robustly to a wide range of motion speeds, and M1-M4 cells appeared tuned to different speeds, suggesting that they might analyse the speed of motion. Retrograde labelling revealed that M1-M4 cells project to the superior colliculus, suggesting that the contrast and motion information signalled by these cells could be used by this sensorimotor area to detect novel objects and motion in the visual field. PMID:24396062

  17. Photoresponse diversity among the five types of intrinsically photosensitive retinal ganglion cells

    PubMed Central

    Zhao, Xiwu; Stafford, Ben K; Godin, Ashley L; King, W Michael; Wong, Kwoon Y

    2014-01-01

    Intrinsically photosensitive retinal ganglion cells (ipRGCs) mediate non-image-forming visual responses, including pupillary constriction, circadian photoentrainment and suppression of pineal melatonin secretion. Five morphological types of ipRGCs, M1–M5, have been identified in mice. In order to understand their functions better, we studied the photoresponses of all five cell types, by whole-cell recording from fluorescently labelled ipRGCs visualized using multiphoton microscopy. All ipRGC types generated melanopsin-based (‘intrinsic’) as well as synaptically driven (‘extrinsic’) light responses. The intrinsic photoresponses of M1 cells were lower threshold, higher amplitude and faster than those of M2–M5. The peak amplitudes of extrinsic light responses differed among the ipRGC types; however, the responses of all cell types had comparable thresholds, kinetics and waveforms, and all cells received rod input. While all five types exhibited inhibitory amacrine-cell and excitatory bipolar-cell inputs from the ‘on’ channel, M1 and M3 received additional ‘off’-channel inhibition, possibly through their ‘off’-sublamina dendrites. The M2–M5 ipRGCs had centre–surround-organized receptive fields, implicating a capacity to detect spatial contrast. In contrast, the receptive fields of M1 cells lacked surround antagonism, which might be caused by the surround of the inhibitory input nullifying the surround of the excitatory input. All ipRGCs responded robustly to a wide range of motion speeds, and M1–M4 cells appeared tuned to different speeds, suggesting that they might analyse the speed of motion. Retrograde labelling revealed that M1–M4 cells project to the superior colliculus, suggesting that the contrast and motion information signalled by these cells could be used by this sensorimotor area to detect novel objects and motion in the visual field. PMID:24396062

  18. Automated computation of arbor densities: a step toward identifying neuronal cell types

    PubMed Central

    Sümbül, Uygar; Zlateski, Aleksandar; Vishwanathan, Ashwin; Masland, Richard H.; Seung, H. Sebastian

    2014-01-01

    The shape and position of a neuron convey information regarding its molecular and functional identity. The identification of cell types from structure, a classic method, relies on the time-consuming step of arbor tracing. However, as genetic tools and imaging methods make data-driven approaches to neuronal circuit analysis feasible, the need for automated processing increases. Here, we first establish that mouse retinal ganglion cell types can be as precise about distributing their arbor volumes across the inner plexiform layer as they are about distributing the skeletons of the arbors. Then, we describe an automated approach to computing the spatial distribution of the dendritic arbors, or arbor density, with respect to a global depth coordinate based on this observation. Our method involves three-dimensional reconstruction of neuronal arbors by a supervised machine learning algorithm, post-processing of the enhanced stacks to remove somata and isolate the neuron of interest, and registration of neurons to each other using automatically detected arbors of the starburst amacrine interneurons as fiducial markers. In principle, this method could be generalizable to other structures of the CNS, provided that they allow sparse labeling of the cells and contain a reliable axis of spatial reference. PMID:25505389

  19. Muscarinic acetylcholine receptor-mediated stimulation of retinal ganglion cell photoreceptors.

    PubMed

    Sodhi, Puneet; Hartwick, Andrew T E

    2016-09-01

    Melanopsin-dependent phototransduction in intrinsically photosensitive retinal ganglion cells (ipRGCs) involves a Gq-coupled phospholipase C (PLC) signaling cascade. Acetylcholine, released in the mammalian retina by starburst amacrine cells, can also activate Gq-PLC pathways through certain muscarinic acetylcholine receptors (mAChRs). Using multielectrode array recordings of rat retinas, we demonstrate that robust spiking responses can be evoked in neonatal and adult ipRGCs after bath application of the muscarinic agonist carbachol. The stimulatory action of carbachol on ipRGCs was a direct effect, as confirmed through calcium imaging experiments on isolated ipRGCs in purified cultures. Using flickering (6 Hz) yellow light stimuli at irradiances below the threshold for melanopsin activation, spiking responses could be elicited in ipRGCs that were suppressed by mAChR antagonism. Therefore, this work identified a novel melanopsin-independent pathway for stimulating sustained spiking in ganglion cell photoreceptors. This mAChR-mediated pathway could enhance ipRGC spiking responses in conditions known to evoke retinal acetylcholine release, such as those involving flickering or moving visual stimuli. Furthermore, this work identifies a pharmacological approach for light-independent ipRGC stimulation that could be targeted by mAChR agonists. PMID:27055770

  20. Teneurin-3 Specifies Morphological and Functional Connectivity of Retinal Ganglion Cells in the Vertebrate Visual System

    PubMed Central

    Antinucci, Paride; Nikolaou, Nikolas; Meyer, Martin P.; Hindges, Robert

    2013-01-01

    Summary A striking feature of the CNS is the precise wiring of its neuronal connections. During vertebrate visual system development, different subtypes of retinal ganglion cells (RGCs) form specific connections with their corresponding synaptic partners. However, the underlying molecular mechanisms remain to be fully elucidated. Here, we report that the cell-adhesive transmembrane protein Teneurin-3 (Tenm3) is required by zebrafish RGCs for acquisition of their correct morphological and functional connectivity in vivo. Teneurin-3 is expressed by RGCs and their presynaptic amacrine and postsynaptic tectal cell targets. Knockdown of Teneurin-3 leads to RGC dendrite stratification defects within the inner plexiform layer, as well as mistargeting of dendritic processes into outer portions of the retina. Moreover, a subset of RGC axons exhibits tectal laminar arborization errors. Finally, functional analysis of RGCs targeting the tectum reveals a selective deficit in the development of orientation selectivity after Teneurin-3 knockdown. These results suggest that Teneurin-3 plays an instructive role in the functional wiring of the vertebrate visual system. PMID:24183672

  1. Pharmacological Analysis of Intrinsic Neuronal Oscillations in rd10 Retina

    PubMed Central

    Biswas, Sonia; Haselier, Christine; Mataruga, Anja; Thumann, Gabriele; Walter, Peter; Müller, Frank

    2014-01-01

    In the widely used mouse model of retinal degeneration, rd1, the loss of photoreceptors leads to rhythmic electrical activity of around 10–16 Hz in the remaining retinal network. Recent studies suggest that this oscillation is formed within the electrically coupled network of AII amacrine cells and ON-bipolar cells. A second mouse model, rd10, displays a delayed onset and slower progression of degeneration, making this mouse strain a better model for human retinitis pigmentosa. In rd10, oscillations occur at a frequency of 3–7 Hz, raising the question whether oscillations have the same origin in the two mouse models. As rd10 is increasingly being used as a model to develop experimental therapies, it is important to understand the mechanisms underlying the spontaneous rhythmic activity. To study the properties of oscillations in rd10 retina we combined multi electrode recordings with pharmacological manipulation of the retinal network. Oscillations were abolished by blockers for ionotropic glutamate receptors and gap junctions. Frequency and amplitude of oscillations were modulated strongly by blockers of inhibitory receptors and to a lesser extent by blockers of HCN channels. In summary, although we found certain differences in the pharmacological modulation of rhythmic activity in rd10 compared to rd1, the overall pattern looked similar. This suggests that the generation of rhythmic activity may underlie similar mechanisms in rd1 and rd10 retina. PMID:24918437

  2. Mechanisms of allele-selective down-regulation of HLA class I in Burkitt's lymphoma.

    PubMed

    Imreh, M P; Zhang, Q J; de Campos-Lima, P O; Imreh, S; Krausa, P; Browning, M; Klein, G; Masucci, M G

    1995-07-01

    Burkitt lymphomas (BL) that arise in HLA-AII-positive individuals are characterized by selective loss/down-regulation of the HLA AII polypeptide. We have investigated the molecular basis of such down-regulation by comparing 5 pairs of BL lines and Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (LCL) derived from the normal B cells of the same individuals. The presence of apparently intact HLA AII genes was confirmed in all 5 BL/LCL pairs by polymerase chain reaction (PCR) typing and by Southern-blot hybridization with HLA A locus-specific probes. Northern-blot analysis with locus- and allele-specific probes revealed a significantly lower expression or absence of AII-specific mRNA in all 5 BL lines compared to the corresponding LCLs. Up-regulation of AII-specific mRNA was achieved by IFN alpha treatment of 2 BL lines with low HLA AII expression (BL-28 and BL-72) while the treatment had no effect in 3 BL lines (WWI-BL, WW2-BL and BL41) that did not express the endogenous gene. HLA AII expression was restored by transfection of the gene in WWI-BL whereas transfectants of BL-41 remained AII-negative. An HLA-AII-promoter-driven chloramphenicol acetyl transferase reporter gene (pAIICAT) was active in WWI-BL but not in BL-41. HLA-AII was expressed in hybrids of BL-41 with an AII-positive LCL, while expression of the endogenous HLA AII gene could not be restored by fusion of BL-41 with an AII-negative LCL, although an adequate set of transcription factors was present in the hybrid. Our results suggest that genetic defects and lack of transcription factors may contribute to the selective down-regulation of HLA AII in BL cells. PMID:7601573

  3. The RNA binding protein RBPMS is a selective marker of ganglion cells in the mammalian retina

    PubMed Central

    Rodriguez, Allen R.; de Sevilla Müller, Luis Pérez; Brecha, Nicholas C.

    2014-01-01

    There are few neurochemical markers that reliably identify retinal ganglion cells (RGCs), which are a heterogeneous population of cells that integrate and transmit the visual signal from the retina to the central visual nuclei. We have developed and characterized a new set of affinity purified guinea pig and rabbit antibodies against RNA-binding protein with multiple splicing (RBPMS). On Western blots these antibodies recognize a single band at ~24 kDa, corresponding to RBPMS, and they strongly label RGC and displaced RGC (dRGC) somata in mouse, rat, guinea pig, rabbit and monkey retina. RBPMS immunoreactive cells and RGCs identified by other techniques have a similar range of somal diameters and areas. The density of RBPMS cells in mouse and rat retina is comparable to earlier semi-quantitative estimates of RGCs. RBPMS is mainly expressed in medium and large DAPI-, DRAQ5-, NeuroTrace- and NeuN-stained cells in the ganglion cell layer (GCL), and RBPMS is not expressed in syntaxin (HPC-1) immunoreactive cells in the inner nuclear layer (INL) and GCL, consistent with their identity as RGCs, and not displaced amacrine cells. In mouse and rat retina, most RBPMS cells are lost following optic nerve crush or transection at three weeks, and all Brn3a, SMI-32 and melanopsin immunoreactive RGCs also express RBPMS immunoreactivity. RBPMS immunoreactivity is localized to CFP-fluorescent RGCs in the B6.Cg-Tg(Thy1-CFP)23Jrs/J mouse line. These findings show that antibodies against RBPMS are robust reagents that exclusively identify RGCs and dRGCs in multiple mammalian species, and they will be especially useful for quantification of RGCs. PMID:24318667

  4. Angiotensin II receptors in the gonads

    SciTech Connect

    Aguilera, G.; Millan, M.A.; Harwood, J.P.

    1989-05-01

    The presence of components of the renin-angiotensin system in ovaries and testes suggests that angiotensin II (AII) is involved in gonadal function, and thus we sought to characterize receptors for AII in rat and primate gonads. In the testes, autoradiographic studies showed receptors in the interstitium in all species. In rat interstitial cells fractionated by Percoll gradient, AII receptors coincided with hCG receptors indicating that AII receptors are located on the Leydig cells. In Leydig cells and membranes from rat and rhesus monkey prepuberal testes, AII receptors were specific for AII analogues and of high affinity (Kd=nM). During development, AII receptor content in rat testes decreases with age parallel to a fall in the ratio of interstitial to tubular tissue. In the ovary, the distribution of AII receptors was dependent on the stage of development, being high in the germinal epithelium and stromal tissue between five and 15 days, and becoming localized in secondary follicles in 20-and 40-day-old rats. No binding was found in primordial or primary follicles. In rhesus monkey ovary, AII receptors were higher in stromal tissue and lower in granulosa and luteal cells of the follicles. Characterization of the binding in rat and monkey ovarian membranes showed a single class of sites with a Kd in the nmol/L range and specificity similar to that of the adrenal glomerulosa and testicular AII receptors. Receptors for AII were also present in membrane fractions from PMSG/hCG primed rat ovaries. Infusion of AII (25 ng/min) or captopril (1.4 micrograms/min) during the PMSG/hCG induction period had no effect on ovarian weight or AII receptor concentration in the ovaries.

  5. ECIL guidelines for the diagnosis of Pneumocystis jirovecii pneumonia in patients with haematological malignancies and stem cell transplant recipients.

    PubMed

    Alanio, Alexandre; Hauser, Philippe M; Lagrou, Katrien; Melchers, Willem J G; Helweg-Larsen, Jannik; Matos, Olga; Cesaro, Simone; Maschmeyer, Georg; Einsele, Hermann; Donnelly, J Peter; Cordonnier, Catherine; Maertens, Johan; Bretagne, Stéphane

    2016-09-01

    The Fifth European Conference on Infections in Leukaemia (ECIL-5) convened a meeting to establish evidence-based recommendations for using tests to diagnose Pneumocystis jirovecii pneumonia (PCP) in adult patients with haematological malignancies. Immunofluorescence assays are recommended as the most sensitive microscopic method (recommendation A-II: ). Real-time PCR is recommended for the routine diagnosis of PCP ( A-II: ). Bronchoalveolar lavage (BAL) fluid is recommended as the best specimen as it yields good negative predictive value ( A-II: ). Non-invasive specimens can be suitable alternatives ( B-II: ), acknowledging that PCP cannot be ruled out in case of a negative PCR result ( A-II: ). Detecting β-d-glucan in serum can contribute to the diagnosis but not the follow-up of PCP ( A-II: ). A negative serum β-d-glucan result can exclude PCP in a patient at risk ( A-II: ), whereas a positive test result may indicate other fungal infections. Genotyping using multilocus sequence markers can be used to investigate suspected outbreaks ( A-II: ). The routine detection of dihydropteroate synthase mutations in cases of treatment failure is not recommended ( B-II: ) since these mutations do not affect response to high-dose co-trimoxazole. The clinical utility of these diagnostic tests for the early management of PCP should be further assessed in prospective, randomized interventional studies. PMID:27550991

  6. Functional inhibition in direction-selective retinal ganglion cells: spatiotemporal extent and intralaminar interactions.

    PubMed

    Stasheff, Steven F; Masland, Richard H

    2002-08-01

    We recorded from ON-OFF direction-selective ganglion cells (DS cells) in the rabbit retina to investigate in detail the inhibition that contributes to direction selectivity in these cells. Using paired stimuli moving sequentially across the cells' receptive fields in the preferred direction, we directly confirmed the prediction of that a wave of inhibition accompanies any moving excitatory stimulus on its null side, at a fixed spatial offset. Varying the interstimulus distance, stimulus size, luminance, and speed yielded a spatiotemporal map of the strength of inhibition within this region. This "null" inhibition was maximal at an intermediate distance behind a moving stimulus: 1/2 to 11/2 times the width of the receptive field. The strength of inhibition depended more on the distance behind the stimulus than on stimulus speed, and the inhibition often lasted 1-2 s. These spatial and temporal parameters appear to account for the known spatial frequency and velocity tuning of ON-OFF DS cells to drifting contrast gratings. Stimuli that elicit distinct ON and OFF responses to leading and trailing edges revealed that an excitatory response of either polarity could inhibit a subsequent response of either polarity. For example, an OFF response inhibited either an ON or OFF response of a subsequent stimulus. This inhibition apparently is conferred by a neural element or network spanning the ON and OFF sublayers of the inner plexiform layer, such as a multistratified amacrine cell. Trials using a stationary flashing spot as a probe demonstrated that the total amount of inhibition conferred on the DS cell was equivalent for stimuli moving in either the null or preferred direction. Apparently the cell does not act as a classic "integrate and fire" neuron, summing all inputs at the soma. Rather, computation of stimulus direction likely involves interactions between excitatory and inhibitory inputs in local regions of the dendrites. PMID:12163551

  7. Characterization of Three-Dimensional Retinal Tissue Derived from Human Embryonic Stem Cells in Adherent Monolayer Cultures.

    PubMed

    Singh, Ratnesh K; Mallela, Ramya K; Cornuet, Pamela K; Reifler, Aaron N; Chervenak, Andrew P; West, Michael D; Wong, Kwoon Y; Nasonkin, Igor O

    2015-12-01

    Stem cell-based therapy of retinal degenerative conditions is a promising modality to treat blindness, but requires new strategies to improve the number of functionally integrating cells. Grafting semidifferentiated retinal tissue rather than progenitors allows preservation of tissue structure and connectivity in retinal grafts, mandatory for vision restoration. Using human embryonic stem cells (hESCs), we derived retinal tissue growing in adherent conditions consisting of conjoined neural retina and retinal pigment epithelial (RPE) cells and evaluated cell fate determination and maturation in this tissue. We found that deriving such tissue in adherent conditions robustly induces all eye field genes (RX, PAX6, LHX2, SIX3, SIX6) and produces four layers of pure populations of retinal cells: RPE (expressing NHERF1, EZRIN, RPE65, DCT, TYR, TYRP, MITF, PMEL), early photoreceptors (PRs) (coexpressing CRX and RCVRN), inner nuclear layer neurons (expressing CALB2), and retinal ganglion cells [RGCs, expressing BRN3B and Neurofilament (NF) 200]. Furthermore, we found that retinal progenitors divide at the apical side of the hESC-derived retinal tissue (next to the RPE layer) and then migrate toward the basal side, similar to that found during embryonic retinogenesis. We detected synaptogenesis in hESC-derived retinal tissue, and found neurons containing many synaptophysin-positive boutons within the RGC and PR layers. We also observed long NF200-positive axons projected by RGCs toward the apical side. Whole-cell recordings demonstrated that putative amacrine and/or ganglion cells exhibited electrophysiological responses reminiscent of those in normal retinal neurons. These responses included voltage-gated Na(+) and K(+) currents, depolarization-induced spiking, and responses to neurotransmitter receptor agonists. Differentiation in adherent conditions allows generation of long and flexible pieces of 3D retinal tissue suitable for isolating transplantable slices of tissue

  8. Muller glia, vision-guided ocular growth, retinal stem cells, and a little serendipity: the Cogan lecture.

    PubMed

    Fischer, Andy J

    2011-09-01

    Hypothesis-driven science is expected to result in a continuum of studies and findings along a discrete path. By comparison, serendipity can lead to new directions that branch into different paths. Herein, I describe a diverse series of findings that were motivated by hypotheses, but driven by serendipity. I summarize how investigations into vision-guided ocular growth in the chick eye led to the identification of glucagonergic amacrine cells as key regulators of ocular elongation. Studies designed to assess the impact of the ablation of different types of neurons on vision-guided ocular growth led to the finding of numerous proliferating cells within damaged retinas. These proliferating cells were Müller glia-derived retinal progenitors with a capacity to produce new neurons. Studies designed to investigate Müller glia-derived progenitors led to the identification of a domain of neural stem cells that form a circumferential marginal zone (CMZ) that lines the periphery of the retina. Accelerated ocular growth, caused by visual deprivation, stimulated the proliferation of CMZ progenitors. We formulated a hypothesis that growth-regulating glucagonergic cells may regulate both overall eye size (scleral growth) and the growth of the retina (proliferation of CMZ cells). Subsequent studies identified unusual types of glucagonergic neurons with terminals that ramify within the CMZ; these cells use visual cues to control equatorial ocular growth and the proliferation of CMZ cells. Finally, while studying the signaling pathways that stimulate CMZ and Müller glia-derived progenitors, serendipity led to the discovery of a novel type of glial cell that is scattered across the inner retinal layers. PMID:21960640

  9. Relative contribution of rod and cone inputs to bipolar cells and ganglion cells in the tiger salamander retina.

    PubMed

    Hensley, S H; Yang, X L; Wu, S M

    1993-06-01

    1. The relative contribution of rod and cone inputs to bipolar and ganglion cells were studied by comparing the response-irradiance relations, spectral sensitivities, and response waveforms of these neurons recorded from the isolated, flat-mounted tiger salamander retina under dark-adapted conditions. 2. Bipolar cells could be differentiated both on the basis of the polarity of the light response and on their relative rod/cone input. Thus some depolarizing bipolar cells appeared more strongly influenced by rod input (DBCR), whereas others were more influenced by cone input (DBCC). Similarly, hyperpolarizing bipolar cells could be divided into those that received rod-dominant input (HBCR) or cone-dominant input (HBCC). 3. The light onset response of sustained-ON ganglion cells reflected both rod-dominant input from DBCRs and cone-dominant input from DBCCs. 4. OFF ganglion cells displayed both a rod-dominant sustained light offset response and a cone-dominant transient light offset response, suggesting input from both HBCRs and HBCCs. 5. In ON-OFF ganglion cells, the light onset response was strongly rod dominated and was presumably mediated by DBCRs, whereas the light offset response displayed both rod and cone influence, suggesting input from HBCRs and HBCCs. The contribution of cones to the light onset response of ON-OFF ganglion cells was only observed in the presence of a rod-adapting background light. 6. A suppression of the light offset responses of OFF and ON-OFF ganglion cells was observed, which was dependent both on the wavelength and irradiance of the light stimulus. 7. These results indicate that the photoreceptor inputs to bipolar cells in the tiger salamander retina are segregated such that they form separate rod-dominant and cone-dominant pathways. Thus the response properties of the different types of ganglion cells are influenced not only by the excitatory and inhibitory inputs they receive from the bipolar and amacrine cells but also whether these

  10. Involvement of ER stress in retinal cell death

    PubMed Central

    Shimazawa, Masamitsu; Inokuchi, Yuta; Ito, Yasushi; Murata, Hiroshi; Aihara, Makoto; Miura, Masayuki; Araie, Makoto

    2007-01-01

    amacrine cells at 12 and 24 h, respectively, and to microglial cells at 72 h. BiP and CHOP were increased at 12 h after NMDA injection, and the increases persisted for the remainder of the 72 h observation period. Conclusions These data indicate that ER-stress may play a pivotal role in RGC death, whether induced by NMDA or IOP elevation. PMID:17438523

  11. Mechanisms that limit the light stimulus frequency following through the APB sensitive and insensitive rod Off-pathways

    PubMed Central

    Bai, Xia; Zhu, Junling; Yang, Jinnan; Savoie, Brian T.; Wang, Guo-Yong

    2009-01-01

    In the retina, rod signal pathways process scotopic visual information. Light decrements are mediated by two distinct groups of rod pathways in the dark adapted retina that can be differentiated on the basis of their sensitivity to the glutamate agonist DL-2-amino-4-phosphonobutyric acid (APB). We have found that the APB sensitive and insensitive rod Off-pathways signal different light decrement information: the APB sensitive rod Off-pathway conveys slow and low frequency light signals, whereas the APB insensitive rod Off-pathways mediate fast and high frequency light signals (Wang, 2006). However, the mechanisms which limit the frequency following through the APB sensitive and insensitive rod Off-pathways remain unknown. In the current study, whole-cell patch-clamp recordings were made from ganglion cells in dark and light adapted mouse retina to identify the mechanisms that limit the frequency following through the APB sensitive and insensitive rod Off-pathways. The results showed that the sites from AII amacrine cells to Off cone bipolar cells are the major mechanisms that limit the frequency following through the APB sensitive rod Off-pathway. In the APB insensitive rod Off-pathways, rods themselves limited the frequency following through these pathways. Moreover, ganglion cells were able to follow higher frequencies under photopic conditions than under scotopic conditions. The Off responses followed lower frequencies than On responses under photopic conditions. This finding was observed in cells that yielded On or Off responses only as well as in On-Off cells. PMID:19406212

  12. Physiological and morphological characterization of ganglion cells in the salamander retina.

    PubMed

    Wang, Jing; Jacoby, Roy; Wu, Samuel M

    2016-02-01

    Retinal ganglion cells (RGCs) integrate visual information from the retina and transmit collective signals to the brain. A systematic investigation of functional and morphological characteristics of various types of RGCs is important to comprehensively understand how the visual system encodes and transmits information via various RGC pathways. This study evaluated both physiological and morphological properties of 67 RGCs in dark-adapted flat-mounted salamander retina by examining light-evoked cation and chloride current responses via voltage-clamp recordings and visualizing morphology by Lucifer yellow fluorescence with a confocal microscope. Six groups of RGCs were described: asymmetrical ON-OFF RGCs, symmetrical ON RGCs, OFF RGCs, and narrow-, medium- and wide-field ON-OFF RGCs. Dendritic field diameters of RGCs ranged 102-490 μm: narrow field (<200 μm, 31% of RGCs), medium field (200-300 μm, 45%) and wide field (>300 μm, 24%). Dendritic ramification patterns of RGCs agree with the sublamina A/B rule. 34% of RGCs were monostratified, 24% bistratified and 42% diffusely stratified. 70% of ON RGCs and OFF RGCs were monostratified. Wide-field RGCs were diffusely stratified. 82% of RGCs generated light-evoked ON-OFF responses, while 11% generated ON responses and 7% OFF responses. Response sensitivity analysis suggested that some RGCs obtained separated rod/cone bipolar cell inputs whereas others obtained mixed bipolar cell inputs. 25% of neurons in the RGC layer were displaced amacrine cells. Although more types may be defined by more refined classification criteria, this report is to incorporate more physiological properties into RGC classification. PMID:26731645

  13. Absence of phosphoglucose isomerase-1 in retinal photoreceptor, pigment epithelium and Muller cells.

    PubMed

    Archer, Simon N; Ahuja, Poonam; Caffé, Romeo; Mikol, Catherine; Foster, Russell G; van Veen, Theo; von Schantz, Malcolm

    2004-06-01

    Macroarray analysis was used to compare equal amounts of cDNA from wild-type and rd/rd (retinal degeneration) mice, collected at P90 when photoreceptor degeneration is virtually complete. A stronger signal for the glycolytic enzyme phosphoglucose isomerase (Gpi1) was observed in the rd/rd sample. Extracellularly, Gpi1 may act as a cytokine, independently described as neuroleukin and autocrine motility factor. Retinal Gpi1 expression was investigated by Northern and Western blot analysis and immunohistochemistry. Double-labelling was performed with antibodies against Gpi1 and calbindin-D, glutamine synthetase, RPE65, calretinin and ultraviolet opsin in order to provide positive cell type identification. Northern and Western blots showed double expression levels per microgram of RNA and protein, respectively, in the rd/rd retina compared with wild-type. However, the total amount of Gpi1 protein per retina was indistinguishable. Gpi1 immunoreactivity was found in ganglion, amacrine, horizontal and bipolar cells, but not in rods, cones, pigment epithelium and Muller cells. This distribution explains why the absolute amounts of Gpi1 protein were not appreciably different between wild-type and the rd/rd phenotype, where rods and cones are absent, whilst the relative contribution of Gpi1 to the total protein and RNA pools differed. Some extracellular immunoreactivity was observed in the photoreceptor matrix around cones in freshly fixed tissue only, which could possibly reflect a role as a cytokine. We propose that glycolysis in Gpi1-negative cells proceeds entirely through the pentose phosphate pathway, creating NADPH at the cost of organic carbon. We hypothesize that the unique metabolic needs of photoreceptors justify this trade-off. PMID:15182299

  14. APP involvement in retinogenesis of mice.

    PubMed

    Dinet, Virginie; An, Na; Ciccotosto, Giuseppe D; Bruban, Julien; Maoui, Agathe; Bellingham, Shayne A; Hill, Andrew F; Andersen, Olav M; Nykjaer, Anders; Jonet, Laurent; Cappai, Roberto; Mascarelli, Frédéric

    2011-03-01

    Very few studies have examined expression and function of amyloid precursor protein (APP) in the retina. We showed that APP mRNA and protein are expressed according to the different waves of retinal differentiation. Depletion of App led to an absence of amacrine cells, a 50% increase in the number of horizontal cells and alteration of the synapses. The retinas of adult APP(-/-) mice showed only half as many glycinergic amacrine cells as wild-type retinas. We identified Ptf1a, which plays a role in controlling both amacrine and horizontal cell fates, as a downstream effector of APP. The observation of a similar phenotype in sorLA knockout mice, a major regulator of APP processing, suggests that regulation of APP functions via sorLA controls the determination of amacrine and horizontal cell fate. These findings provide novel insights that indicate that APP plays an important role in retinal differentiation. PMID:20978902

  15. Elevated intraocular pressure decreases response sensitivity of inner retinal neurons in experimental glaucoma mice

    PubMed Central

    Pang, Ji-Jie; Frankfort, Benjamin J.; Gross, Ronald L.; Wu, Samuel M.

    2015-01-01

    Glaucoma is the second leading cause of blindness in the United States and the world, characterized by progressive degeneration of the optic nerve and retinal ganglion cells (RGCs). Glaucoma patients exhibit an early diffuse loss of retinal sensitivity followed by focal loss of RGCs in sectored patterns. Recent evidence has suggested that this early sensitivity loss may be associated with dysfunctions in the inner retina, but detailed cellular and synaptic mechanisms underlying such sensitivity changes are largely unknown. In this study, we use whole-cell voltage-clamp techniques to analyze light responses of individual bipolar cells (BCs), AII amacrine cells (AIIACs), and ON and sustained OFF alpha-ganglion cells (ONαGCs and sOFFαGCs) in dark-adapted mouse retinas with elevated intraocular pressure (IOP). We present evidence showing that elevated IOP suppresses the rod ON BC inputs to AIIACs, resulting in less sensitive AIIACs, which alter AIIAC inputs to ONαGCs via the AIIAC→cone ON BC→ONαGC pathway, resulting in lower ONαGC sensitivity. The altered AIIAC response also reduces sOFFαGC sensitivity via the AIIAC→sOFFαGC chemical synapses. These sensitivity decreases in αGCs and AIIACs were found in mice with elevated IOP for 3–7 wk, a stage when little RGC or optic nerve degeneration was observed. Our finding that elevated IOP alters neuronal function in the inner retina before irreversible structural damage occurs provides useful information for developing new diagnostic tools and treatments for glaucoma in human patients. PMID:25675503

  16. Autocrine protective mechanisms of human granulocyte colony-stimulating factor (G-CSF) on retinal ganglion cells after optic nerve crush.

    PubMed

    Huang, Shun-Ping; Fang, Kan-Tang; Chang, Chung-Hsing; Huang, Tzu-Lun; Wen, Yao-Tseng; Tsai, Rong-Kung

    2016-02-01

    This study investigated the role of autocrine mechanisms in the anti-apoptotic effects of human granulocyte colony-stimulating factor (G-CSF) on retinal ganglion cells (RGCs) after optic nerve (ON) crush. We observed that both G-CSF and G-CSF receptor (G-CSFR) are expressed in normal rat retina. Further dual immunofluorescence staining showed G-CSFR immunoreactive cells were colocalized with RGCs, Müller cells, horizontal and amacrine cells. These results confirm that G-CSF is an endogenous ligand in the retina. The semi-quantitative RT-PCR finding demonstrated the transcription levels of G-CSF and G-CSFR were up-regulated after ON crush injury. G-CSF treatment further increased and prolonged the expression level of G-CSFR in the retina. G-CSF has been shown to enhance transdifferentiation of the mobilized hematopoietic stem cells into tissue to repair central nervous system injury. We test the hypothesis that the hematopoietic stem cells recruited by G-CSF treatment can transdifferentiate into RGCs after ON crush by performing sublethal irradiation of the rats 5 days before ON crush. The flow cytometric analysis showed the number of CD34 positive cells in the peripheral blood is significantly lower in the irradiated, crushed and G-CSF-treated group than the sham control group or crush and G-CSF treated group. Nevertheless, the G-CSF treatment enhances the RGC survival after sublethal irradiation and ON crush injury. These data indicate that G-CSF seems unlikely to induce hematopoietic stem cell transdifferentiation into RGCs after ON crush injury. In conclusion, G-CSF may serve an endogenous protective signaling in the retina through direct activation of intrinsic G-CSF receptors and downstream signaling pathways to rescue RGCs after ON crush injury, exogenous G-CSF administration can enhance the anti-apoptotic effects on RGCs. PMID:26518178

  17. Angiotensin II receptor heterogeneity

    SciTech Connect

    Herblin, W.F.; Chiu, A.T.; McCall, D.E.; Ardecky, R.J.; Carini, D.J.; Duncia, J.V.; Pease, L.J.; Wong, P.C.; Wexler, R.R.; Johnson, A.L. )

    1991-04-01

    The possibility of receptor heterogeneity in the angiotensin II (AII) system has been suggested previously, based on differences in Kd values or sensitivity to thiol reagents. One of the authors earliest indications was the frequent observation of incomplete inhibition of the binding of AII to adrenal cortical membranes. Autoradiographic studies demonstrated that all of the labeling of the rat adrenal was blocked by unlabeled AII or saralasin, but not by DuP 753. The predominant receptor in the rat adrenal cortex (80%) is sensitive to dithiothreitol (DTT) and DuP 753, and is designated AII-1. The residual sites in the adrenal cortex and almost all of the sites in the rat adrenal medulla are insensitive to both DTT and DuP 753, but were blocked by EXP655. These sites have been confirmed by ligand binding studies and are designated AII-2. The rabbit adrenal cortex is unique in yielding a nonuniform distribution of AII-2 sites around the outer layer of glomerulosa cells. In the rabbit kidney, the sites on the glomeruli are AII-1, but the sites on the kidney capsule are AII-2. Angiotensin III appears to have a higher affinity for AII-2 sites since it inhibits the binding to the rabbit kidney capsule but not the glomeruli. Elucidation of the distribution and function of these diverse sites should permit the development of more selective and specific therapeutic strategies.

  18. Proteomic identification of altered apolipoprotein patterns in pulmonary hypertension and vasculopathy of sickle cell disease

    PubMed Central

    Yuditskaya, Susan; Tumblin, Ashaunta; Hoehn, Gerard T.; Wang, Guanghui; Drake, Steven K.; Xu, Xiuli; Ying, Saixia; Chi, Amy H.; Remaley, Alan T.; Shen, Rong-Fong; Munson, Peter J.; Suffredini, Anthony F.

    2009-01-01

    Pulmonary arterial hypertension (PAH) is emerging as a major complication and independent risk factor for death among adults with sickle cell disease (SCD). Using surface-enhanced laser desorption/ionization time of flight mass spectrometry (SELDI-TOF MS), we searched for biomarkers of PAH in plasma specimens from 27 homozygous sickle cell anemia (HbSS) patients with PAH and 28 without PAH. In PAH patients, analysis consistently showed lower abundance of a 28.1-kDa peak (P < .001), identified by high-resolution mass spectrometry as the oxidant-scavenging protein apolipoprotein A-I (apoA-I), which correlated with clinical assays of apoA-I (r = .58, P < .001) and high-density lipoprotein (HDL) levels (r = .50, P = .001). Consistent with endothelial dysfunction that may mediate this effect in PAH, HbSS patients with lower apoA-I levels also displayed impaired vasodilatory responses to acetylcholine (mean ± SEM, 189% ± 34% [n = 13] vs 339% ± 51% [n = 13], P < .001). As a group, patients with SCD demonstrated significantly lower apoA-I levels than African-American control subjects. The PAH cohort was further characterized by high levels of apolipoproteins A-II and B and serum amyloid A, and low levels of haptoglobin dimers and plasminogen. These results imply a relationship of apolipoproteins to the development of PAH vasculopathy in SCD, potentially involving an unexpected mechanistic parallel to atherosclerosis, another proliferative vasculopathy. PMID:19023114

  19. Mutant WDR36 directly affects axon growth of retinal ganglion cells leading to progressive retinal degeneration in mice

    PubMed Central

    Chi, Zai-Long; Yasumoto, Fumie; Sergeev, Yuri; Minami, Masayoshi; Obazawa, Minoru; Kimura, Itaru; Takada, Yuichiro; Iwata, Takeshi

    2010-01-01

    Primary open-angle glaucoma (POAG) is one of the three principal subtypes of glaucoma and among the leading cause of blindness worldwide. POAG is defined by cell death of the retinal ganglion cells (RGCs) and surrounding neuronal cells at higher or normal intraocular pressure (IOP). Coded by one of the three genes responsible for POAG, WD repeat-containing protein 36 (WDR36) has two domains with a similar folding. To address whether WDR36 is functionally important in the retina, we developed four transgenic mice strains overexpressing a wild-type (Wt) and three mutant variants of D606G, deletion of amino acids at positions 605–607 (Del605–607) and at 601–640 (Del601–640) equivalent to the location of the D658G mutation observed in POAG patients. A triple amino acid deletion of mouse Wdr36 at positions 605–607 corresponding to the deletion at positions 657–659 in humans developed progressive retinal degeneration at the peripheral retina with normal IOP. RGCs and connecting amacrine cell synapses were affected at the peripheral retina. Axon outgrowth rate of cultured RGC directly isolated from transgenic animal was significantly reduced by the Wdr36 mutation compared with Wt. Molecular modeling of wild and mutant mouse Wdr36 revealed that deletion at positions 605–607 removed three residues and a hydrogen bond, required to stabilize anti-parallel β-sheet of the 6th β-propeller in the second domain. We concluded that WDR36 plays an important functional role in the retina homeostasis and mutation to this gene can cause devastating retinal damage. These data will improve understanding of the functional property of WDR36 in the retina and provide a new animal model for glaucoma therapeutics. PMID:20631153

  20. Ability of retinal Müller glial cells to protect neurons against excitotoxicity in vitro depends upon maturation and neuron-glial interactions.

    PubMed

    Heidinger, V; Hicks, D; Sahel, J; Dreyfus, H

    1999-02-01

    Glutamate is the most abundant excitatory amino acid in the central nervous system. It has also been described as a potent toxin when present in high concentrations because excessive stimulation of its receptors leads to neuronal death. Glial influence on neuronal survival has already been shown in the central nervous system, but the mechanisms underlying glial neuroprotection are only partly known. When cells isolated from newborn rat retina were maintained in culture as enriched neuronal populations, 80% of the cells were destroyed by application of excitotoxic concentrations of glutamate. Massive neuronal death was also observed in newborn retinal cultures containing large numbers of glia, or when neurons were seeded onto feeder layers of purified cells prepared from immature (postnatal 8 day) rat retina. When newborn retinal neurons were seeded onto feeder layers of purified glial cells prepared from adult retinas, application of excitotoxic amino acids no longer led to neuronal death. Furthermore, neuronal death was not observed in mixed neuron/glial cultures prepared from adult retina. However, in all cases (newborn and adult) application of kainate led to amacrine cell-specific death. Activity of glutamine synthetase, a key glial enzyme involved in glutamate detoxification, was assayed in these cultures in the presence or absence of exogenous glutamate. Whereas pure glial cultures alone (from young or adult retina) showed low activity that was not stimulated by glutamate addition, mixed or co-cultured neurons and adult glia exhibited up to threefold higher levels of activity following glutamate treatment. These data indicate that two conditions must be satisfied to observe glial neuroprotection: maturation of glutamine synthetase expression, and neuron-glial signalling through glutamate-elicited responses. PMID:9932869

  1. Angiotensin II receptors in testes

    SciTech Connect

    Millan, M.A.; Aguilera, G.

    1988-05-01

    Receptors for angiotensin II (AII) were identified and characterized in testes of rats and several primate species. Autoradiographic analysis of the binding of 125I-labeled (Sar1,Ile8)AII to rat, rhesus monkey, cebus monkey, and human testicular slide-mounted frozen sections indicated specific binding to Leydig cells in the interstitium. In rat collagenase-dispersed interstitial cells fractionated by Percoll gradient, AII receptor content was parallel to that of hCG receptors, confirming that the AII receptors are in the Leydig cells. In rat dispersed Leydig cells, binding was specific for AII and its analogs and of high affinity (Kd, 4.8 nM), with a receptor concentration of 15 fmol/10(6) cells. Studies of AII receptors in rat testes during development reveals the presence of high receptor density in newborn rats which decreases toward the adult age (4934 +/- 309, 1460 +/- 228, 772 +/- 169, and 82 +/- 12 fmol/mg protein at 5, 15, 20, and 30 days of age, respectively) with no change in affinity. At all ages receptors were located in the interstitium, and the decrease in binding was parallel to the decrease in the interstitial to tubular ratio observed with age. AII receptor properties in membrane-rich fractions from prepuberal testes were similar in the rat and rhesus monkey. Binding was time and temperature dependent, reaching a plateau at 60 min at 37 C, and was increased by divalent cations, EGTA, and dithiothreitol up to 0.5 mM. In membranes from prepuberal monkey testes, AII receptors were specific for AII analogs and of high affinity (Kd, 4.2 nM) with a receptor concentration of 7599 +/- 1342 fmol/mg protein. The presence of AII receptors in Leydig cells in rat and primate testes in conjunction with reports of the presence of other components of the renin-angiotensin system in the testes suggests that the peptide has a physiological role in testicular function.

  2. In Vitro Cytotoxic Effects of Gold Nanoparticles Coated with Functional Acyl Homoserine Lactone Lactonase Protein from Bacillus licheniformis and Their Antibiofilm Activity against Proteus Species

    PubMed Central

    Vinoj, Gopalakrishnan; Pati, Rashmirekha; Sonawane, Avinash

    2014-01-01

    N-acylated homoserine lactonases are known to inhibit the signaling molecules of the biofilm-forming pathogens. In this study, gold nanoparticles were coated with N-acylated homoserine lactonase proteins (AiiA AuNPs) purified from Bacillus licheniformis. The AiiA AuNPs were characterized by UV-visible spectra, Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), and X-ray diffraction (XRD). The synthesized AiiA AuNPs were found to be spherical in shape and 10 to 30 nm in size. Treatment with AiiA protein-coated AuNPs showed maximum reduction in exopolysaccharide production, metabolic activities, and cell surface hydrophobicity and potent antibiofilm activity against multidrug-resistant Proteus species compared to treatment with AiiA protein alone. AiiA AuNPs exhibited potent antibiofilm activity at 2 to 8 μM concentrations without being harmful to the macrophages. We conclude that at a specific dose, AuNPs coated with AiiA can kill bacteria without harming the host cells, thus representing a potential template for the design of novel antibiofilm and antibacterial protein drugs to decrease bacterial colonization and to overcome the problem of drug resistance. In summary, our data suggest that the combined effect of the lactonase and the gold nanoparticles of the AiiA AuNPs has promising antibiofilm activity against biofilm-forming and multidrug-resistant Proteus species. PMID:25403677

  3. mTor signaling is required for the formation of proliferating Müller glia-derived progenitor cells in the chick retina.

    PubMed

    Zelinka, Christopher P; Volkov, Leo; Goodman, Zachary A; Todd, Levi; Palazzo, Isabella; Bishop, William A; Fischer, Andy J

    2016-06-01

    We investigate the roles of mTor signaling in the formation of Müller glia-derived progenitor cells (MGPCs) in the chick retina. During embryonic development, pS6 (a readout of active mTor signaling) is present in early-stage retinal progenitors, differentiating amacrine and ganglion cells, and late-stage progenitors or maturing Müller glia. By contrast, pS6 is present at low levels in a few scattered cell types in mature, healthy retina. Following retinal damage, in which MGPCs are known to form, mTor signaling is rapidly activated in Müller glia. Inhibition of mTor in damaged retinas prevented the accumulation of pS6 in Müller glia and reduced numbers of proliferating MGPCs. Inhibition of mTor had no effect on MAPK signaling or on upregulation of the stem cell factor Klf4, whereas Pax6 upregulation was significantly reduced. Inhibition of mTor potently blocked the MGPC-promoting effects of Hedgehog, Wnt and glucocorticoid signaling in damaged retinas. In the absence of retinal damage, insulin, IGF1 and FGF2 induced pS6 in Müller glia, and this was blocked by mTor inhibitor. In FGF2-treated retinas, in which MGPCs are known to form, inhibition of mTor blocked the accumulation of pS6, the upregulation of Pax6 and the formation of proliferating MGPCs. We conclude that mTor signaling is required, but not sufficient, to stimulate Müller glia to give rise to proliferating progenitors, and the network of signaling pathways that drive the formation of MGPCs requires activation of mTor. PMID:27068108

  4. Neuroprotective effect of memantine on the retinal ganglion cells of APPswe/PS1ΔE9 mice and its immunomodulatory mechanisms.

    PubMed

    Gao, Lixiong; Chen, Xi; Tang, Yongping; Zhao, Jinghui; Li, Qiyou; Fan, Xiaotang; Xu, Haiwei; Yin, Zheng Qin

    2015-06-01

    Besides the cognitive impairment and degeneration in the brain, vision dysfunction and retina damage are always prevalent in patients with Alzheimer's disease (AD). The uncompetitive antagonist of the N-methyl-d-aspartate receptor, memantine (MEM), has been proven to improve the cognition of patients with AD. However, limited information exists regarding the mechanism of neurodegeneration and the possible neuroprotective mechanisms of MEM on the retinas of patients with AD. In the present study, by using APPswe/PS1ΔE9 double transgenic (dtg) mice, we found that MEM rescued the loss of retinal ganglion cells (RGCs), as well as improved visual impairments, including improving the P50 component in pattern electroretinograms and the latency delay of the P2 component in flash visual evoked potentials of APPswe/PS1ΔE9 dtg mice. The activated microglia in the retinas of APPswe/PS1ΔE9 dtg mice were also inhibited by MEM. Additionally, the level of glutamine synthetase expressed by Müller cells within the RGC layer was upregulated in APPswe/PS1ΔE9 dtg mice, which was inhibited by MEM. Simultaneously, MEM also reduced the apoptosis of choline acetyl transferase-immunoreactive cholinergic amacrine cells within the RGC layer of AD mice. Moreover, the phosphorylation level of extracellular regulated protein kinases 1 and 2 was increased in APPswe/PS1ΔE9 dtg mice, which was blocked by MEM treatment. These findings suggest that MEM protects RGCs in the retinas of APPswe/PS1ΔE9 dtg mice by modulating the immune response of microglia and the adapted response of Müller cells, making MEM a potential ophthalmic treatment alternative in patients with AD. PMID:25912193

  5. The role of potassium and other ions in the control of aldosterone synthesis

    SciTech Connect

    Kenyon, C.J.; Shepherd, R.M.; Fraser, R.; Pediani, J.D.; Elder, H.Y. )

    1991-01-01

    Fast and slow K+ efflux components, independently regulated by angiotensin II (AII), have been identified in bovine adrenocortical cells. The authors have further investigated the role of potassium in the control of aldosterone synthesis in two ways. Firstly, isotopic tracers, in conjunction with channel modulators, have been used to study the interrelationship of K+ and Ca2+ in the control of AII-stimulated aldosterone synthesis. Secondly, electron probe X-ray microanalysis (EPXMA) was used to quantify potassium, sodium, chlorine and phosphorous in control and AII-stimulated cells. The effects of verapamil on 43K efflux were measured at two stages during AII stimulation. During the first ten minutes of treatment, when efflux via the fast component predominates, AII and verapamil both slowed efflux and their effects were additive. If verapamil was added later, at the time when efflux by the fast component appeared exhausted and the stimulatory effect of AII on the slow efflux component was apparent, it again slowed efflux. These data suggest that verapamil prevents calcium-gated K+ channels from opening by blocking Ca2+ channels. However, verapamil had no effect on AII-stimulated calcium efflux. In addition to blocking Ca2+ channels, verapamil may directly inhibit potassium efflux. EPXMA showed a bimodal distribution of potassium concentrations in control cells. However, in cells stimulated with AII for five minutes, the mean potassium content was less than in controls and was not bimodally distributed. Sodium content was increased by AII-treatment, chlorine was lowered and phosphorus remained unchanged. The data confirm previous observations that AII inhibits Na+/K+ ATPase activity.

  6. Dopamine modulation of rod pathway signaling by suppression of GABAC feedback to rod-driven depolarizing bipolar cells.

    PubMed

    Smith, Benjamin J; Côté, Patrice D; Tremblay, François

    2015-09-01

    Reducing signal gain in the highly sensitive rod pathway prevents saturation as background light levels increase, allowing the dark-adapted retina to encode stimuli over a range of background luminances. Dopamine release is increased during light adaptation and is generally accepted to suppress rod signaling in light-adapted retinas. However, recent research has suggested that dopamine, acting through D1 receptors, could additionally produce a sensitization of the rod pathway in dim light conditions via gamma-aminobutyric acid (GABA) type C receptors. Here, we evaluated the overall activity of the depolarizing bipolar cell (DBC) population in vivo to ensure the integrity of long-distance network interactions by quantifying the b-wave of the electroretinogram in mice. We showed that dopamine, acting through D1 receptors, reduced the amplitude and sensitivity of rod-driven DBCs during light adaptation by suppressing GABA type A receptor-mediated serial inhibition onto rod DBC GABA type C receptors. Block of D1 receptors did not suppress rod-driven DBC sensitivity when GABAA -mediated serial inhibition was blocked by gabazine, suggesting that the reduction in rod-driven DBC sensitivity in the absence of D1 receptors was due to disinhibition of serial inhibitory GABAergic circuitry rather than a direct facilitatory effect on GABA release onto rod-driven DBC GABA type C receptors. Finally, the large population of GABAergic A17 wide-field amacrine cells known to maintain reciprocal inhibition with rod DBCs could be excluded from the proposed disinhibitory circuit after treatment with 5,7-dihydroxytryptamine. PMID:26080286

  7. Light-evoked lateral GABAergic inhibition at single bipolar cell synaptic terminals is driven by distinct retinal microcircuits

    PubMed Central

    Vigh, Jozsef; Vickers, Evan; von Gersdorff, Henrique

    2011-01-01

    Inhibitory amacrine cells (ACs) filter visual signals crossing the retina by modulating the excitatory, glutamatergic output of bipolar cells (BCs) on multiple temporal and spatial scales. Reciprocal feedback from ACs provides focal inhibition that is temporally locked to the activity of presynaptic BC activity, whereas lateral feedback originates from ACs excited by distant BCs. These distinct feedback mechanisms permit temporal and spatial computation at BC terminals. Here, we used a unique preparation to study light-evoked inhibitory postsynaptic currents (IPSCs) recorded from axotomized terminals of ON-type mixed rod/cone BCs (Mb) in goldfish retinal slices. In this preparation, light-evoked IPSCs could only reach axotomized BC terminals via the lateral feedback pathway, allowing us to study lateral feedback in the absence of overlapping reciprocal feedback components. We found that light evokes ON and OFF lateral IPSCs (L-IPSCs) in Mb terminals having different temporal patterns and conveyed via distinct retinal pathways. The relative contribution of rods versus cones to ON and OFF L-IPSCs was light intensity dependent. ACs presynaptic to Mb BC terminals received inputs via AMPA/KA and NMDA type receptors in both the ON and OFF pathways, and employed TTX-sensitive sodium channels to boost signal transfer along their processes. ON and OFF L-IPSCs, like reciprocal feedback IPSCs, were mediated by both GABAA and GABAC receptors. However, our results suggest that lateral and reciprocal feedback do not cross-depress each other, and are therefore mediated by distinct populations of ACs. These findings demonstrate that retinal inhibitory circuits are highly specialized to modulate BC output at different light intensities. PMID:22049431

  8. A web-based archive for topographic maps of retinal cell distribution in vertebrates.

    PubMed

    Collin, Shaun P

    2008-01-01

    Clinical and Experimental Optometry, in conjunction with Optometrists Association Australia and Professor Shaun P Collin of the University of Queensland, announce the launch of a web-based archive of previously published topographic maps of retinal cell distribution in vertebrates. At present, the archive boasts more than 770 different maps of the distribution of retinal neurons (for example, photoreceptors, bipolar cells, amacrine cells, horizontal cells and ganglion cells) in nearly 200 species within all vertebrate classes (Cephalospidomorpha, Actinopterygii, Sarcopterygii, Amphibia, Reptilia, Aves and Mammalia). The distribution of retinal neurons has been studied for more than 100 years and has become a powerful means of predicting the spatial resolving power of the eye and the retinal regions containing specialisations, such as areae centrales, horizontal streaks and foveae, where increased densities of neurons define the way in which a species visually samples its environment. The location of these retinal specialisations thereby identifies the part(s) of the visual field of critical importance for localising food and mates and for predator surveillance. The distribution of sampling elements even reflects the symmetry of a species' ecological habitat. The archive is a unique collection of most of the currently available retinal maps, which also presents relevant information, where known, about eye size, retinal cell density, retinal orientation, cell number, spatial resolving power and the type of specialisation, in addition to basic physical parameters of each species (body size, weight, sex and developmental stage). The archive is accessible at http://www.optometrists.asn.au/ceo/retinalsearch and will be updated regularly. The powerful database is interactive and freely available, providing the opportunity to upload both published and unpublished topographic maps. Following a review process, previously unpublished maps will be 'published' and available

  9. Abnormal Retinal Development in the Btrc Null Mouse

    PubMed Central

    Baguma-Nibasheka, Mark; Kablar, Boris

    2016-01-01

    Previous microarray analysis revealed beta-transducin repeat containing (Btrc) down-regulation in the retina of mouse embryos specifically lacking cholinergic amacrine cells (CACs) as a result of the absence of skeletal musculature and fetal ocular movements. To investigate the role of Btrc in the determination of retinal cell fate, the present study examined retinal morphology in Btrc−/− mouse fetuses. The Btrc−/− retina showed a normal number of cell layers and number of cells per layer with normal cell proliferation and apoptosis. However, there was a complete absence of CACs and a decrease in tyrosine hydroxylase-expressing amacrine cells. The population of other amacrine cell subtypes was normal, whereas that of the precursor cells was decreased. There was also a reduction in the number of retinal ganglion cells, whereas their progenitors were increased. These findings suggest a role for Btrc in regulating the eventual ratio of resulting differentiated retinal cell types. PMID:19705444

  10. Identification of a Retinal Circuit for Recurrent Suppression Using Indirect Electrical Imaging.

    PubMed

    Greschner, Martin; Heitman, Alexander K; Field, Greg D; Li, Peter H; Ahn, Daniel; Sher, Alexander; Litke, Alan M; Chichilnisky, E J

    2016-08-01

    Understanding the function of modulatory interneuron networks is a major challenge, because such networks typically operate over long spatial scales and involve many neurons of different types. Here, we use an indirect electrical imaging method to reveal the function of a spatially extended, recurrent retinal circuit composed of two cell types. This recurrent circuit produces peripheral response suppression of early visual signals in the primate magnocellular visual pathway. We identify a type of polyaxonal amacrine cell physiologically via its distinctive electrical signature, revealed by electrical coupling with ON parasol retinal ganglion cells recorded using a large-scale multi-electrode array. Coupling causes the amacrine cells to fire spikes that propagate radially over long distances, producing GABA-ergic inhibition of other ON parasol cells recorded near the amacrine cell axonal projections. We propose and test a model for the function of this amacrine cell type, in which the extra-classical receptive field of ON parasol cells is formed by reciprocal inhibition from other ON parasol cells in the periphery, via the electrically coupled amacrine cell network. PMID:27397894

  11. Comparative analysis of three purification protocols for retinal ganglion cells from rat

    PubMed Central

    Gao, Fengjuan; Li, Tingting; Hu, Jianyan; Zhou, Xujiao; Wu, Jihong

    2016-01-01

    Purpose To make comparative analyses of the common three purification protocols for retinal ganglion cells (RGCs), providing a solid practical basis for selecting the method for purifying RGCs for use in subsequent experiments. Methods Rat RGCs were isolated and purified using three methods, including two-step immunopanning (TIP) separation, two-step immunopanning-magnetic (TIPM) separation, and flow cytometric (FC) separation. Immunocytochemical staining, quantitative real-time PCR, flow cytometry, electrophysiology, and Cell Counting Kit-8 (CCK-8) analyses were performed to compare the purity, yield, and viability of the RGCs. Results The RGC yields from the TIP, TIPM, and FC methods were 24.60±15.98 × 104, 5.28±4.42 × 104, and 5.4±2.7 × 103 per retina, respectively. We easily controlled the relative purity of the RGCs with the FC method and even reached 100% of the maximum expected purity. However, the RGC purity was only 80.97±5.45% and 95.41±3.23% using the TIP and TIPM methods, respectively. The contaminant cells were mainly large, star-shaped, glial fibrillary acidic protein (GFAP)-positive astrocytes and small, round, syntaxin 1-positive amacrine cells with multiple short neurites. The RGCs purified with FC could not be cultured successively in our study; however, the TIP-RGCs survived more than 20 days with good viability, while the TIPM-RGCs survived less than 9 days. Conclusions The three protocols for purifying the RGCs each had its own pros and cons. The RGCs isolated by the TIP method exhibited the highest viability and yield but had low purity. The purity of the RGCs isolated with the FC method could reach approximately 100% but had a low yield and cell viability. The TIPM method was reliable and produced RGCs with considerable purity, yield, and viability. This study provides a solid practical basis for selecting the method for purifying RGCs for use in subsequent experiments. PMID:27122968

  12. Impaired Glutathione Redox System Paradoxically Suppresses Angiotensin II-Induced Vascular Remodeling

    PubMed Central

    Izawa, Kazuma; Okada, Motoi; Sumitomo, Kazuhiro; Nakagawa, Naoki; Aizawa, Yoshiaki; Kawabe, Junichi; Kikuchi, Kenjiro; Hasebe, Naoyuki

    2014-01-01

    Background Angiotensin II (AII) plays a central role in vascular remodeling via oxidative stress. However, the interaction between AII and reduced glutathione (GSH) redox status in cardiovascular remodeling remains unknown. Methods In vivo: The cuff-induced vascular injury model was applied to Sprague Dawley rats. Then we administered saline or a GSH inhibitor, buthionine sulfoximine (BSO, 30 mmol/L in drinking water) for a week, subsequently administered 4 more weeks by osmotic pump with saline or AII (200 ng/kg/minute) to the rats. In vitro: Incorporation of bromodeoxyuridine (BrdU) was measured to determine DNA synthesis in cultured rat vascular smooth muscle cells (VSMCs). Results BSO reduced whole blood GSH levels. Systolic blood pressure was increased up to 215±4 mmHg by AII at 4 weeks (p<0.01), which was not affected by BSO. Superoxide production in vascular wall was increased by AII and BSO alone, and was markedly enhanced by AII+BSO. The left ventricular weight to body weight ratio was significantly increased in AII and AII+BSO as compared to controls (2.52±0.08, 2.50±0.09 and 2.10±0.07 mg/g respectively, p<0.05). Surprisingly, the co-treatment of BSO totally abolished these morphological changes. Although the vascular circumferential wall stress was well compensated in AII, significantly increased in AII+BSO. The anti-single-stranded DNA staining revealed increasing apoptotic cells in the neointima of injured arteries in BSO groups. BrdU incorporation in cultured VSMCs with AII was increased dose-dependently. Furthermore it was totally abolished by BSO and was reversed by GSH monoethyl ester. Conclusions We demonstrated that a vast oxidative stress in impaired GSH redox system totally abolished AII-induced vascular, not cardiac remodeling via enhancement of apoptosis in the neointima and suppression of cell growth in the media. The drastic suppression of remodeling may result in fragile vasculature intolerable to mechanical stress by AII. PMID

  13. Co-release of acetylcholine and gamma-aminobutyric acid by a retinal neuron

    SciTech Connect

    O'Malley, D.M.; Masland, R.H.

    1989-05-01

    Rabbit retinas were vitally stained with 4',6-diamidino-2-phenylindole (DAPI), a fluorescent compound that selectively accumulates within the cholinergic amacrine cells. The retinas were then incubated in vitro in the presence of radioactive gamma-aminobutyric acid (GABA) and autoradiographed. The cells that accumulated DAPI were found to accumulate GABA, confirming immunohistochemical evidence that the cholinergic amacrine cells contain GABA. Incubation of retinas in the presence of elevated concentrations of K+ caused them to release acetylcholine and GABA, and autoradiography showed depletion of radioactive GABA from the cholinergic amacrine cells. This indicates that the cholinergic amacrine cells can secrete acetylcholine and GABA. Retinas were double-labeled with (14C)GABA and (3H)acetylcholine, allowing simultaneous measurement of their release. The release of (14C)GABA was found to be independent of extracellular Ca2+. Radioactive GABA synthesized endogenously from (14C)glutamate behaved the same way as radioactive GABA accumulated from the medium. In the same experiments the simultaneously measured release of (3H)acetylcholine was strongly Ca2+-dependent, indicating that the releases of acetylcholine and GABA are controlled by different mechanisms. Synaptic vesicles immunologically isolated from double-labeled retinas contained much (3H)acetylcholine and little or no (14C)GABA. These results suggest that the cholinergic amacrine cells release acetylcholine primarily by vesicle exocytosis and release GABA primarily by means of a carrier.

  14. Synaptic circuitry mediating light-evoked signals in dark-adapted mouse retina.

    PubMed

    Wu, Samuel M; Gao, Fan; Pang, Ji-Jie

    2004-12-01

    Light-evoked excitatory cation current (DeltaIC) and inhibitory chloride current (DeltaICl) of rod and cone bipolar cells and AII amacrine cells (AIIACs) were recorded from slices of dark-adapted mouse retinas, and alpha ganglion cells were recorded from flatmounts of dark-adapted mouse retinas. The cell morphology was revealed by Lucifer yellow fluorescence with a confocal microscope. DeltaIC of all rod depolarizing bipolar cells (DBCRs) exhibited similar high sensitivity to 500 nm light, but two patterns of DeltaICl were observed with slightly different axon morphologies. At least two types of cone depolarizing bipolar cells (DBCCs) were identified: one with axon terminals ramified in 70-85% of IPL depth and DBCR-like DeltaIC sensitivity, and the other with axon terminals ramified in 55-75% of IPL depth and much lower DeltaIC sensitivity. The relative rod/cone inputs to DBCs and AIIACs were analyzed by comparing the DeltaIC and DeltaICl thresholds and dynamic ranges with the corresponding values of rods and cones. On average, the sensitivity of a DBCR to the 500 nm light is about 20 times higher than that of a rod. The sensitivity of an AIIAC is more than 1000 times higher than that of a rod, suggesting that AIIAC responses are pooled through a coupled network of about 40 AIIACs. Interactions of rod and cone signals in dark-adapted mouse retinas appear asymmetrical: rod signals spread into the cone system more efficiently than cone signals into the rod system. The mouse synaptic circuitry allows small rod signals to be highly amplified and effectively transmitted to the cone system via rod/cone and AIIAC/DBCC coupling. Three types of alpha ganglion cells (alphaGCs) were identified. (1) ONGCs exhibits no spike activity in darkness, increased spikes in light, sustained inward DeltaIC, sustained outward DeltaICl of varying amplitude, and large soma (20-25 microm in diameter) with an alpha-cell-like dendritic field about 180-350 microm stratifying near 70% of the IPL

  15. Retinoids increase human apolipoprotein A-11 expression through activation of the retinoid X receptor but not the retinoic acid receptor.

    PubMed Central

    Vu-Dac, N; Schoonjans, K; Kosykh, V; Dallongeville, J; Heyman, R A; Staels, B; Auwerx, J

    1996-01-01

    Considering the link between plasma high-density lipoprotein (HDL) cholesterol levels and a protective effect against coronary artery disease as well as the suggested beneficial effects of retinoids on the production of the major HDL apolipoprotein (apo), apo A-I, the goal of this study was to analyze the influence of retinoids on the expression of apo A-II, the other major HDL protein. Retinoic acid (RA) derivatives have a direct effect on hepatic apo A-II production, since all-trans (at) RA induces apo A-II mRNA levels and apo A-II secretion in primary cultures of human hepatocytes. In the HepG2 human hepatoblastoma cell line, both at-RA and 9-cis RA as well as the retinoid X receptor (RXR)-specific agonist LGD 1069, but not the RA receptor (RAR) agonist ethyl-p-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-l-pro penyl]-benzoic acid (TTNPB), induce apo A-II mRNA levels. Transient-transfection experiments with a reporter construct driven by the human apo A-II gene promoter indicated that 9-cis RA and at-RA, as well as the RXR agonists LGD 1069 and LG 100268, induced apo A-II gene expression at the transcriptional level. Only minimal effects of the RAR agonist TTNPB were observed on the apo A-II promoter reporter construct. Unilateral deletions and site-directed mutagenesis identified the J site of the apo A-II promoter mediating the responsiveness to RA. This element contains two imperfect half-sites spaced by 1 oligonucleotide. Cotransfection assays in combination with the use of RXR or RAR agonists showed that RXR but not RAR transactivates the apo A-II promoter through this element. By contrast, RAR inhibits the inductive effects of RXR on the apo A-II J site in a dose-dependent fashion. Gel retardation assays demonstrated that RXR homodimers bind, although with a lower affinity than RAR-RXR heterodimers, to the AH-RXR response element. In conclusion, retinoids induce hepatic apo A-II production at the transcriptional level via the interaction of

  16. Predominance of Giardia duodenalis Assemblage AII in Fresh Leafy Vegetables from a Market in Southern Brazil.

    PubMed

    Tiyo, Rogerio; de Souza, Carla Zangari; Arruda Piovesani, Ana Flávia; Tiyo, Bruna Tiaki; Colli, Cristiane Maria; Marchioro, Ariella Andrade; Gomes, Monica Lucia; Falavigna-Guilherme, Ana Lucia

    2016-06-01

    We investigated the presence of Giardia duodenalis cysts and its genotypes in raw leafy vegetables sold in a Brazilian market. These products are different from those sold in most street markets because the producers themselves display and sell their products and rely on specialized technical and sanitary assistance. Vegetable and water samples were collected from 14 (80%) producers who cultivated vegetables that are typically consumed raw for sale at the market, obtained at the market and farms, respectively. A total of 128 samples of leafy greens (chives, parsley, cabbage, arugula, watercress, and chicory) and 14 water samples were analyzed by direct immunofluorescence and PCR techniques. The positive samples were genotyped (GHD gene) using PCR and restriction fragment length polymorphism. The analyses indicated that 16 (12.5%) of 128 samples were positive by PCR, while 1 (0.8%) of 128 samples were positive by immunofluorescence. Giardia cysts were not detected in the water samples obtained at the farms. The molecular technique revealed a genotype with zoonotic potential, which underscores the challenge in the control of giardiasis dissemination via the consumption of raw vegetables. PMID:27296610

  17. Apolipoprotein A-II polymorphism: relationships to behavioural and hormonal mediators of obesity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    New obesity loci continue to be identified through genomewide association studies in populations of increasing size and ethnic diversity but understanding of the mechanisms by which known genetic variants contribute to obesity remains limited. Several well-established obesity candidates encode prote...

  18. Angiotensinergic involvement in olfactory function

    SciTech Connect

    Speth, R.C.; Parker, J.L.; Wright, J.W.; Harding, J.W.

    1986-03-05

    The olfactory bulbs (OB) from Sprague-Dawley and Wistar-Kyoto rats were frozen and sectioned in a sagittal plane, 20 ..mu.. thick. Sections incubated with /sup 125/-Sar/sup 1/, Ile/sup 8/-AII indicated a high density of AII receptor binding sites in the external layers of the OB. Since the primary olfactory neurons synapse with the mitral cells in these layers, this suggests that AII may affect olfactory input to the OB. To test this hypothesis, male Sprague-Dawley rats, 9-12 weeks of age, n = 8, were administered 0.2 ml of 0.17 M ZnSO/sub 4/ into each nostril to lesion the primary olfactory neurons and their axon terminals in the OB. Rats treated with ZnSO/sub 4/ showed an impairment in their ability to find a buried food pellet, P = 0.041, Mann-Whitney test. Nine days post-treatment, the rats were sacrificed and AII receptors binding in homogenates of the OB was determined. There was a 23% increase (P < 0.05) in AII receptor density in the ZnSO/sub 4/ treated rat OB; it was correlated with the extent of the olfactory deficit, r/sub s/ = .91, Spearman Rank Order Test, P < .01. However, there was a 24% decrease in OB weight in the ZnSO/sub 4/ group, so the number of AII receptors per OB was unchanged. These data suggest that AII plays a role in olfaction. Localizing AII receptor changes within the OB by quantitative autoradiography will characterize the changes in AII receptor density caused by ZnSO/sub 4/.

  19. A Role for Synaptic Input Distribution in a Dendritic Computation of Motion Direction in the Retina.

    PubMed

    Vlasits, Anna L; Morrie, Ryan D; Tran-Van-Minh, Alexandra; Bleckert, Adam; Gainer, Christian F; DiGregorio, David A; Feller, Marla B

    2016-03-16

    The starburst amacrine cell in the mouse retina presents an opportunity to examine the precise role of sensory input location on neuronal computations. Using visual receptive field mapping, glutamate uncaging, two-photon Ca(2+) imaging, and genetic labeling of putative synapses, we identify a unique arrangement of excitatory inputs and neurotransmitter release sites on starburst amacrine cell dendrites: the excitatory input distribution is skewed away from the release sites. By comparing computational simulations with Ca(2+) transients recorded near release sites, we show that this anatomical arrangement of inputs and outputs supports a dendritic mechanism for computing motion direction. Direction-selective Ca(2+) transients persist in the presence of a GABA-A receptor antagonist, though the directional tuning is reduced. These results indicate a synergistic interaction between dendritic and circuit mechanisms for generating direction selectivity in the starburst amacrine cell. PMID:26985724

  20. Reciprocal regulation between sirtuin-1 and angiotensin-II in the substantia nigra: implications for aging and neurodegeneration

    PubMed Central

    Diaz-Ruiz, Carmen; Rodriguez-Perez, Ana I.; Beiroa, Daniel; Rodriguez-Pallares, Jannette; Labandeira-Garcia, Jose L.

    2015-01-01

    Local angiotensin II (AII) and sirtuin 1 (SIRT1) play a major role in the modulation of neuroinflammation, oxidative stress and aging-related dopaminergic vulnerability to damage. However, it is not known whether the modulation is related to reciprocal regulation between SIRT1 and AII. In the present study, a single intraventricular injection of AII increased nigral SIRT1 levels in young adult rats. Although AII activity is known to be increased in aged rats, levels of SIRT1 were significantly lower than in young controls. Treatment with the SIRT1-activating compound resveratrol increased nigral SIRT1 levels in aged rats. Levels of SIRT1 were significantly higher in aged wild type mice than in AII type-1 receptor (AT1) deficient mice. In cell culture studies, treatment with AII also induced a transitory increase in levels of SIRT1 in the MES 23.5 dopaminergic neuron and the N9 microglial cell lines. In aged rats, treatment with resveratrol induced a significant decrease in the expression of AT1 receptors and markers of NADPH-oxidase activation (p47phox). In aged transgenic mice over-expressing SIRT1, levels of AT1 and p47phox were lower than in aged wild type controls. In vitro, the inhibitory effects of resveratrol on AII/AT1/NADPH-oxidase activity were confirmed in primary mesencephalic cultures, the N9 microglial cell line, and the dopaminergic neuron cell line MES 23.5, and they were blocked by the SIRT1 specific inhibitor EX527. The present findings show that SIRT1 and the axis AII/AT1/NADPH-oxidase regulate each other. This is impaired in aged animals and may be mitigated with sirtuin-activating compounds. PMID:26384348

  1. Glycinergic pathways in the goldfish retina

    SciTech Connect

    Marc, R.E.; Lam, D.M.

    1981-02-01

    Autoradiographic localization of high affinity (3H)glycine uptake in the retina of the goldfish has been used to study some anatomical and physiological properties of potentially glycinergic neurons. There are two classes of retinal cells exhibiting high affinity glycine uptake: Aa amacrine cells and I2 interplexiform cells. Aa amacrine cells constitute about 20% of the somas in the amacrine cell layer and send their dendrites to the middle of the inner plexiform layer. There they are both pre- and postsynaptic primarily to other amacrine cells. Photic modulation of glycine uptake indicates that they are probably red-hyperpolarizing/green-depolarizing neurons. I2 interplexiform cells are a newly discovered type of interplexiform cell; in the outer plexiform layer, they receive direct synaptic input from the somas of red-dominated GABAergic H1 horizontal cells and are apparently presynaptic to dendrites of unidentified types of horizontal cells. The connections of I2 interplexiform cells have not been successfully characterized in the inner plexiform layer. These findings extend our knowledge of neurochemically specific pathways in the cyprinid retina and indicate that glycine, like GABA, is a neurotransmitter primarily involved with circuits coding ''red'' information.

  2. Cell Structure

    MedlinePlus

    ... Cells, Tissues, & Membranes Cell Structure & Function Cell Structure Cell Function Body Tissues Epithelial Tissue Connective Tissue Muscle Tissue ... apparatus , and lysosomes . « Previous (Cell Structure & Function) Next (Cell Function) » Contact Us | Privacy Policy | Accessibility | FOIA | File Formats ...

  3. Stem Cells

    MedlinePlus

    Stem cells are cells with the potential to develop into many different types of cells in the body. ... the body. There are two main types of stem cells: embryonic stem cells and adult stem cells. Stem ...

  4. Stem Cells

    MedlinePlus

    Stem cells are cells with the potential to develop into many different types of cells in the body. They serve as a repair ... body. There are two main types of stem cells: embryonic stem cells and adult stem cells. Stem ...

  5. Decreased Expression of DREAM Promotes the Degeneration of Retinal Neurons

    PubMed Central

    Chintala, Shravan; Cheng, Mei; Zhang, Xiao

    2015-01-01

    The intrinsic mechanisms that promote the degeneration of retinal ganglion cells (RGCs) following the activation of N-Methyl-D-aspartic acid-type glutamate receptors (NMDARs) are unclear. In this study, we have investigated the role of downstream regulatory element antagonist modulator (DREAM) in NMDA-mediated degeneration of the retina. NMDA, phosphate-buffered saline (PBS), and MK801 were injected into the vitreous humor of C57BL/6 mice. At 12, 24, and 48 hours after injection, expression of DREAM in the retina was determined by immunohistochemistry, western blot analysis, and electrophoretic mobility-shift assay (EMSA). Apoptotic death of cells in the retina was determined by terminal deoxynucleotidyl transferace dUTP nick end labeling (TUNEL) assays. Degeneration of RGCs in cross sections and in whole mount retinas was determined by using antibodies against Tuj1 and Brn3a respectively. Degeneration of amacrine cells and bipolar cells was determined by using antibodies against calretinin and protein kinase C (PKC)-alpha respectively. DREAM was expressed constitutively in RGCs, amacrine cells, bipolar cells, as well as in the inner plexiform layer (IPL). NMDA promoted a progressive decrease in DREAM levels in all three cell types over time, and at 48 h after NMDA-treatment very low DREAM levels were evident in the IPL only. DREAM expression in retinal nuclear proteins was decreased progressively after NMDA-treatment, and correlated with its decreased binding to the c-fos-DRE oligonucleotides. A decrease in DREAM expression correlated significantly with apoptotic death of RGCs, amacrine cells and bipolar cells. Treatment of eyes with NMDA antagonist MK801, restored DREAM expression to almost normal levels in the retina, and significantly decreased NMDA-mediated apoptotic death of RGCs, amacrine cells, and bipolar cells. Results presented in this study show for the first time that down-regulation of DREAM promotes the degeneration of RGCs, amacrine cells, and

  6. Renal sympathetic baroreflex effects of angiotensin II infusions into the rostral ventrolateral medulla of the rabbit.

    PubMed

    Saigusa, T; Head, G A

    1993-05-01

    1. The effects of local infusion of angiotensin II (AII) into the rostral ventrolateral medulla (RVLM) pressor area on the renal sympathetic baroreflex were compared with the excitatory amino acid glutamate in urethane anaesthetized rabbits with chronically implanted renal nerve electrodes. Baroreflex blood pressure-renal nerve activity curves were obtained by intravenous infusion of phenylephrine and nitroprusside before and after treatments. 2. Infusion of 4 pmol/min of AII into the RVLM increased blood pressure by 12 +/- 2 mmHg and transiently increased resting sympathetic nerve activity. The renal sympathetic baroreflex curves were shifted to the right. The upper plateau of the sympathetic reflex increased by 29 +/- 8% (n = 6, P < 0.025). 3. Infusions of glutamate into the RVLM, at a dose which was equipressor to that of AII, also increased resting renal sympathetic nerve activity. In contrast to AII, this increase was maintained throughout the infusion. Glutamate shifted the reflex curve to the right and increased the upper plateau of the sympathetic reflex by 44 +/- 5% without affecting the lower plateau. 4. These results support the suggestion that AII can act at the level of the RVLM pressor area to facilitate baroreflex control of renal sympathetic activity in a similar fashion to that produced by fourth ventricular administration. 5. Thus the RVLM is a likely candidate site for modulation of the renal sympathetic baroreflex. The similarity of the actions of AII to those of glutamate suggest that it may directly excite sympathetic vasomotor cells in this region. PMID:8100748

  7. T Cells

    MedlinePlus

    ... or turn off the immune response. Cytotoxic or “killer” T cells directly attack and destroy cells bearing ... involve selective activation of helper T cells and killer T cells, with a corresponding decrease in regulatory ...

  8. Cell division

    MedlinePlus Videos and Cool Tools

    ... hours after conception, the fertilized egg cell remains a single cell. After approximately 30 hours, it divides ... 3 days, the fertilized egg cell has become a berry-like structure made up of 16 cells. ...

  9. The erythropoietin receptor is not required for the development, function, and aging of rods and cells in the retinal periphery

    PubMed Central

    Caprara, Christian; Britschgi, Corinne; Samardzija, Marijana

    2014-01-01

    . Analysis of the retinal morphology in the two knockdown lines did not reveal any developmental defects or signs of accelerated degeneration in the senescent tissue. Similarly, retinal function was not altered under scotopic and photopic conditions. In addition, EpoR knockdown had no influence on cell viability under acute hypoxic conditions. Retinal angiogenesis and vasculature were normal in the absence of EPOR. However, expression of some EPOR-signaling target genes was significantly altered in the retinas of the EpoRflox/flox;α-Cre mice. Conclusions Our data suggest that expression of EPOR in rod photoreceptors, Müller cells, and amacrine, horizontal, and ganglion cells of the peripheral retina is not required for the maturation, function, and survival of these cells in aging tissue. Based on the expression of the EPOR-signaling target genes, we postulate that expression of soluble EPOR in the retina may modulate endogenous EPO-EPOR signaling. PMID:24644405

  10. Cell counting.

    PubMed

    Phelan, M C; Lawler, G

    2001-05-01

    This unit presents protocols for counting cells using either a hemacytometer or electronically using a Coulter counter. Cell counting with a hemacytometer permits effective discrimination of live from dead cells using trypan blue exclusion. In addition, the procedure is less subject to errors arising from cell clumping or size heterogeneity. Counting cells is more quickly and easily performed using an electronic counter, but live-dead discrimination is unreliable. Cell populations containing large numbers of dead cells and/or cell clumps are difficult to count accurately. In addition, electronic counting requires resetting of the instrument for cell populations of different sizes; heterogeneous populations can give rise to inaccurate counts, and resting and activated cells may require counting at separate settings. In general, electronic cell counting is best performed on fresh peripheral blood cells. PMID:18770655

  11. 75 FR 20645 - Order Instituting Administrative Proceedings Pursuant to Section 15E(a)(2)(A)(ii) of the...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-20

    ... Credit Rating Co., Ltd. April 14, 2010. Dagong Global Credit Rating Co., Ltd. (``Dagong''), a credit rating agency based in Beijing, China, submitted an application with the Securities and Exchange Commission (``Commission'') for registration as a nationally recognized statistical rating...

  12. Determination of a quantum efficiency of A^II B^VI compounds on the basis of photoacoustic spectra

    NASA Astrophysics Data System (ADS)

    Maliński, M.; Bychto, L.; Zakrzewski, J.

    2005-10-01

    The method of determination of the quantum efficiency η of irradiative relaxation processes in semiconductors is introduced and discussed. The correlation of the amplitude photoacoustic spectra in the high absorption region with the annealing process is used to estimate the values of η. The values of η, both for as grown and annealed crystals were determined and have been discussed. The results are presented for microphone and piezoelectric detection methods.

  13. 76 FR 53312 - Airworthiness Directives; Agusta S.p.A. Model A109A and A109AII Helicopters

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-08-26

    ... Privacy Act Statement in the Federal Register published on April 11, 2000 (65 FR 19477-78). Regulatory... Regulatory Policies and Procedures (44 FR 11034, February 26, 1979); 3. Will not affect intrastate aviation... action requires a one-time inspection to determine the tightening torque value of the hub plug,...

  14. Mechanism of the Quorum-Quenching Lactonase (AiiA) from Bacillus thuringiensis. 2. Substrate Modeling and Active Site Mutations†

    PubMed Central

    2008-01-01

    The N-acyl-l-homoserine lactone hydrolases (AHL lactonases) have attracted considerable attention because of their ability to quench AHL-mediated quorum-sensing pathways in Gram-negative bacteria and because of their relation to other enzymes in the metallo-β-lactamase superfamily. To elucidate the detailed catalytic mechanism of AHL lactonase, mutations are made on residues that presumably contribute to substrate binding and catalysis. Steady-state kinetic studies are carried out on both the wild-type and mutant enzymes using a spectrum of substrates. Two mutations, Y194F and D108N, present significant effects on the overall catalysis. On the basis of a high-resolution structural model of the enzyme−product complex, a hybrid quantum mechanical/molecular mechanical method is used to model the substrate binding orientation and to probe the effect of the Y194F mutation. Combining all experimental and computational results, we propose a detailed mechanism for the ring-opening hydrolysis of AHL substrates as catalyzed by the AHL lactonase from Bacillus thuringiensis. Several features of the mechanism that are also found in related enzymes are discussed and may help to define an evolutionary thread that connects the hydrolytic enzymes of this mechanistically diverse superfamily. PMID:18627130

  15. Pharmacological reactivity of neoplastic and non-neoplastic associated neovasculature to vasoconstrictors.

    PubMed

    Andrade, S P; Beraldo, W T

    1998-12-01

    Angiogenesis and the pharmacological responses of the tumour and non-tumour associated neovasculature have been investigated. Cannulated sponge discs in mice were used to host the angiogenic stimulators, while 133Xe washout was employed to assess local blood flow. Enhancement of blood flow was detected in implants bearing B16 cells, 3T3 cells and angiotensin II (AII)-treated at day 7. The responses of non-neoplastic associated neovasculature at day 14 post sponge implantation to the vasoconstrictors used endothelin-1 (Et-1), AII, platelet activating factor (PAF) and 5-hydroxytryptamine (5-HT) were dose-dependent. By contrast, the newly formed blood vessels induced by tumour cells were markedly insensitive to the vasoconstrictors agonists Et-1 and AII, while fully responsive to PAF and 5-HT. The vessels resulting from neoplastic stimulus exhibited altered pharmacological reactivity, suggesting that the characteristics of the neovasculature are dependent on the nature of the angiogenic stimuli. PMID:10319023

  16. Galvanic Cells

    ERIC Educational Resources Information Center

    Young, I. G.

    1973-01-01

    Many standard physical chemistry textbooks contain ambiguities which lead to confusion about standard electrode potentials, calculating cell voltages, and writing reactions for galvanic cells. This article shows how standard electrode potentials can be used to calculate cell voltages and deduce cell reactions. (Author/RH)

  17. Cell Biochips

    NASA Astrophysics Data System (ADS)

    Pioufle, B. Le; Picollet-D'Hahan, N.

    A cell biochip is a microsystem, equipped with electronic and microfluidic functions, designed to manipulate or analyse living cells. The first publications in this emerging area of research appeared toward the end of the 1980s. In 1989 Washizu described a biochip designed to fuse two cells by electropermeabilisation of the cytoplasmic membrane [1]. Research centers have devised a whole range of cell chip structures, for simultaneous or sequential analysis of single cells, cell groups, or cell tissues reconstituted on the chip. The cells are arranged in a square array on a parallel cell chip for parallel analysis, while they are examined and processed one by one in a microchannel in the case of a series cell chip. In contrast to these biochips for high-throughput analysis of a large number of cells, single-cell chips focus on the analysis of a single isolated cell. As in DNA microarrays, where a large number of oligonucleotides are ordered in a matrix array, parallel cell chips order living cells in a similar way. At each point of the array, the cells can be isolated, provided that the cell type allows this, e.g., blood cells, or cultivated in groups (most adhesion cells can only survive in groups). The aim is to allow massively parallel analysis or processing. Le Pioufle et al. describe a microdevice for the culture of single cells or small groups of cells in a micropit array [2]. Each pit is equipped to stimulate the cell or group of cells either electrically or fluidically. Among the applications envisaged are gene transfer, cell sorting, and screening in pharmacology. A complementary approach, combining the DNA microarray and cell biochip ideas, has been put forward by Bailey et al. [3]. Genes previously arrayed on the chip transfect the cultured cells on the substrate depending on their position in the array (see Fig. 19.1). This way of achieving differential lipofection on a chip was then taken up again by Yoshikawa et al. [4] with primary cells, more

  18. Synaptic mechanisms of adaptation and sensitization in the retina

    PubMed Central

    Nikolaev, Anton; Leung, Kin-Mei; Odermatt, Benjamin; Lagnado, Leon

    2014-01-01

    Sensory systems continually adjust the way stimuli are processed. What are the circuit mechanisms underlying this plasticity? We investigated how synapses in the retina of zebrafish adjust to changes in the temporal contrast of a visual stimulus by imaging activity in vivo. Following an increase in contrast, bipolar cell synapses with strong initial responses depressed, whereas synapses with weak initial responses facilitated. Depression and facilitation predominated in different strata of the inner retina, where bipolar cell output was anticorrelated with the activity of amacrine cell synapses providing inhibitory feedback. Pharmacological block of GABAergic feedback converted facilitating bipolar cell synapses into depressing ones. These results indicate that depression intrinsic to bipolar cell synapses causes adaptation of the ganglion cell response to contrast, whereas depression in amacrine cell synapses causes sensitization. Distinct microcircuits segregating to different layers of the retina can cause simultaneous increases or decreases in the gain of neural responses. PMID:23685718

  19. Cell division

    MedlinePlus Videos and Cool Tools

    ... structure made up of 16 cells. This structure is called a morula, which is Latin for mulberry. The cells continue to divide ... days following conception into a blastocyst. Although it is only the size of a pinhead, the blastocyst ...

  20. Immunohistochemical and Calcium Imaging Methods in Wholemount Rat Retina

    PubMed Central

    Sargoy, Allison; Barnes, Steven; Brecha, Nicholas C.; De Sevilla Müller, Luis Pérez

    2015-01-01

    In this paper we describe the tools, reagents, and the practical steps that are needed for: 1) successful preparation of wholemount retinas for immunohistochemistry and, 2) calcium imaging for the study of voltage gated calcium channel (VGCC) mediated calcium signaling in retinal ganglion cells. The calcium imaging method we describe circumvents issues concerning non-specific loading of displaced amacrine cells in the ganglion cell layer. PMID:25349920

  1. Solar cells

    NASA Astrophysics Data System (ADS)

    Cuquel, A.; Roussel, M.

    The physical and electronic characteristics of solar cells are discussed in terms of space applications. The principles underlying the photovoltaic effect are reviewed, including an analytic model for predicting the performance of individual cells and arrays of cells. Attention is given to the effects of electromagnetic and ionizing radiation, micrometeors, thermal and mechanical stresses, pollution and degassing encountered in space. The responses of different types of solar cells to the various performance-degrading agents are examined, with emphasis on techniques for quality assurance in the manufacture and mounting of Si cells.

  2. mglur6b:EGFP Transgenic zebrafish suggest novel functions of metabotropic glutamate signaling in retina and other brain regions.

    PubMed

    Glasauer, Stella M K; Wäger, Robert; Gesemann, Matthias; Neuhauss, Stephan C F

    2016-08-15

    Metabotropic glutamate receptors (mGluRs) are mainly known for regulating excitability of neurons. However, mGluR6 at the photoreceptor-ON bipolar cell synapse mediates sign inversion through glutamatergic inhibition. Although this is currently the only confirmed function of mGluR6, other functions have been suggested. Here we present Tg(mglur6b:EGFP)zh1, a new transgenic zebrafish line recapitulating endogenous expression of one of the two mglur6 paralogs in zebrafish. Investigating transgene as well as endogenous mglur6b expression within the zebrafish retina indicates that EGFP and mglur6b mRNA are not only expressed in bipolar cells, but also in a subset of ganglion and amacrine cells. The amacrine cells labeled in Tg(mglur6b:EGFP)zh1 constitute a novel cholinergic, non-GABAergic, non-starburst amacrine cell type described for the first time in teleost fishes. Apart from the retina, we found transgene expression in subsets of periventricular neurons of the hypothalamus, Purkinje cells of the cerebellum, various cell types of the optic tectum, and mitral/ruffed cells of the olfactory bulb. These findings suggest novel functions of mGluR6 besides sign inversion at ON bipolar cell dendrites, opening up the possibility that inhibitory glutamatergic signaling may be more prevalent than currently thought. J. Comp. Neurol. 524:2363-2378, 2016. © 2016 Wiley Periodicals, Inc. PMID:27121676

  3. Types of Stem Cells

    MedlinePlus

    ... PDF) Download an introduction to stem cells and stem cell research. Stem Cell Glossary Stem cell terms to know. ... stem cells blog from the International Society for Stem Cell Research. Learn About Stem Cells From Lab to You ...

  4. Long-Term Plasticity Mediated by mGluR1 at a Retinal Reciprocal Synapse

    PubMed Central

    Vigh, Jozsef; Li, Geng-Lin; Hull, Court; von Gersdorff, Henrique

    2013-01-01

    Summary The flow of information across the retina is controlled by reciprocal synapses between bipolar cell terminals and amacrine cells. However, the synaptic delays and properties of plasticity at these synapses are not known. Here we report that glutamate release from goldfish Mb-type bipolar cell terminals can trigger fast (delay of 2–3 ms) and transient GABAA IPSCs and a much slower and more sustained GABAC feedback. Synaptically released glutamate activated mGluR1 receptors on amacrine cells and, depending on the strength of presynaptic activity, potentiated subsequent feedback. This poststimulus enhancement of GABAergic feedback lasted for up to 10 min. This form of mGluR1-mediated long-term synaptic plasticity may provide retinal reciprocal synapses with adaptive capabilities. PMID:15882646

  5. Electrolytic cell

    NASA Astrophysics Data System (ADS)

    Bullock, J. S.; Hale, B. D.

    1984-09-01

    An apparatus is described for the separation of the anolyte and the catholyte during electrolysis. The electrolyte flows through an electrolytic cell between the oppositely charged electrodes. The cell is equipped with a wedge-shaped device, the tapered end is located between the electrodes on the effluent side of the cell. The wedge diverts the flow of the electrolyte to either side of the wedge, substantially separating the anolyte and the catholyte.

  6. Glutamatergic Monopolar Interneurons Provide a Novel Pathway of Excitation in the Mouse Retina.

    PubMed

    Della Santina, Luca; Kuo, Sidney P; Yoshimatsu, Takeshi; Okawa, Haruhisa; Suzuki, Sachihiro C; Hoon, Mrinalini; Tsuboyama, Kotaro; Rieke, Fred; Wong, Rachel O L

    2016-08-01

    Excitatory and inhibitory neurons in the CNS are distinguished by several features, including morphology, transmitter content, and synapse architecture [1]. Such distinctions are exemplified in the vertebrate retina. Retinal bipolar cells are polarized glutamatergic neurons receiving direct photoreceptor input, whereas amacrine cells are usually monopolar inhibitory interneurons with synapses almost exclusively in the inner retina [2]. Bipolar but not amacrine cell synapses have presynaptic ribbon-like structures at their transmitter release sites. We identified a monopolar interneuron in the mouse retina that resembles amacrine cells morphologically but is glutamatergic and, unexpectedly, makes ribbon synapses. These glutamatergic monopolar interneurons (GluMIs) do not receive direct photoreceptor input, and their light responses are strongly shaped by both ON and OFF pathway-derived inhibitory input. GluMIs contact and make almost as many synapses as type 2 OFF bipolar cells onto OFF-sustained A-type (AOFF-S) retinal ganglion cells (RGCs). However, GluMIs and type 2 OFF bipolar cells possess functionally distinct light-driven responses and may therefore mediate separate components of the excitatory synaptic input to AOFF-S RGCs. The identification of GluMIs thus unveils a novel cellular component of excitatory circuits in the vertebrate retina, underscoring the complexity in defining cell types even in this well-characterized region of the CNS. PMID:27426514

  7. Characterization of glucagon-expressing neurons in the chicken retina

    PubMed Central

    Fischer, Andy J.; Skorupa, Dana; Schonberg, David L.; Walton, Nathaniel A.

    2008-01-01

    We have recently identified large glucagon-expressing neurons that densely ramify neurites in the peripheral edge of the retina and regulate the proliferation of progenitors in the circumferential marginal zone (CMZ) of the postnatal chicken eye (Fischer et al., 2005). However, nothing is known about the transmitters and proteins that are expressed by the glucagon-expressing neurons in the avian retina. We used antibodies to cell-distinguishing markers to better characterize the different types of glucagon-expressing neurons. We found that the large glucagon-expressing neurons were immunoreactive for substance P, neurofilament, Pax6, AP2α, HuD, calretinin, trkB and trkC. Colocalization of glucagon and substance P in the large glucagon-expressing neurons indicates that these cells are the “bullwhip cells” that have been briefly described by Ehrlich, Keyser and Karten (1987). Similar to the bullwhip cells, the conventional glucagon-expressing amacrine cells were immunoreactive for calretinin, HuD, Pax6, and AP2α. Unlike bullwhip cells, the conventional glucagon-expressing amacrine cells were immunoreactive for GABA. While glucagon-immunoreactive amacrine cells were negative for substance P in central regions of the retina, a subset of this type of amacrine cell was immunoreactive for substance P in far peripheral regions of the retina. An additional type of glucagon/substance P-expressing neuron, resembling the bullwhip cells, was found in far peripheral and dorsal regions of the retina. Based on morphology, distribution within the retina, and histological markers, we conclude that there may be 4 different types of glucagon-expressing neurons in the avian retina. PMID:16572462

  8. Cell Migration

    PubMed Central

    Trepat, Xavier; Chen, Zaozao; Jacobson, Ken

    2015-01-01

    Cell migration is fundamental to establishing and maintaining the proper organization of multicellular organisms. Morphogenesis can be viewed as a consequence, in part, of cell locomotion, from large-scale migrations of epithelial sheets during gastrulation, to the movement of individual cells during development of the nervous system. In an adult organism, cell migration is essential for proper immune response, wound repair, and tissue homeostasis, while aberrant cell migration is found in various pathologies. Indeed, as our knowledge of migration increases, we can look forward to, for example, abating the spread of highly malignant cancer cells, retarding the invasion of white cells in the inflammatory process, or enhancing the healing of wounds. This article is organized in two main sections. The first section is devoted to the single-cell migrating in isolation such as occurs when leukocytes migrate during the immune response or when fibroblasts squeeze through connective tissue. The second section is devoted to cells collectively migrating as part of multicellular clusters or sheets. This second type of migration is prevalent in development, wound healing, and in some forms of cancer metastasis. PMID:23720251

  9. Cell Chauvinism

    ERIC Educational Resources Information Center

    Keller, Dolores Elaine

    1972-01-01

    Indicates that biological terminology, such as mother cell'' and labels of sex factors in bacteria, reflect discrimination against females by reinforcing perpetuation of stereotyped gender roles. (AL)

  10. Cell migration.

    PubMed

    Trepat, Xavier; Chen, Zaozao; Jacobson, Ken

    2012-10-01

    Cell migration is fundamental to establishing and maintaining the proper organization of multicellular organisms. Morphogenesis can be viewed as a consequence, in part, of cell locomotion, from large-scale migrations of epithelial sheets during gastrulation, to the movement of individual cells during development of the nervous system. In an adult organism, cell migration is essential for proper immune response, wound repair, and tissue homeostasis, while aberrant cell migration is found in various pathologies. Indeed, as our knowledge of migration increases, we can look forward to, for example, abating the spread of highly malignant cancer cells, retarding the invasion of white cells in the inflammatory process, or enhancing the healing of wounds. This article is organized in two main sections. The first section is devoted to the single-cell migrating in isolation such as occurs when leukocytes migrate during the immune response or when fibroblasts squeeze through connective tissue. The second section is devoted to cells collectively migrating as part of multicellular clusters or sheets. This second type of migration is prevalent in development, wound healing, and in some forms of cancer metastasis. PMID:23720251