Ooto, Sotaro; Akagi, Tadamichi; Kageyama, Ryoichiro; Akita, Joe; Mandai, Michiko; Honda, Yoshihito; Takahashi, Masayo
It has long been believed that the retina of mature mammals is incapable of regeneration. In this study, using the N-methyl-D-aspartate neurotoxicity model of adult rat retina, we observed that some Müller glial cells were stimulated to proliferate in response to a toxic injury and produce bipolar cells and rod photoreceptors. Although these newly produced neurons were limited in number, retinoic acid treatment promoted the number of regenerated bipolar cells. Moreover, misexpression of basic helix-loop-helix and homeobox genes promoted the induction of amacrine, horizontal, and rod photoreceptor specific phenotypes. These findings demonstrated that retinal neurons regenerated even in adult mammalian retina after toxic injury. Furthermore, we could partially control the fate of the regenerated neurons with extrinsic factors or intrinsic genes. The Müller glial cells constitute a potential source for the regeneration of adult mammalian retina and can be a target for drug delivery and gene therapy in retinal degenerative diseases.
Ooto, Sotaro; Akagi, Tadamichi; Kageyama, Ryoichiro; Akita, Joe; Mandai, Michiko; Honda, Yoshihito; Takahashi, Masayo
It has long been believed that the retina of mature mammals is incapable of regeneration. In this study, using the N-methyl-d-aspartate neurotoxicity model of adult rat retina, we observed that some Müller glial cells were stimulated to proliferate in response to a toxic injury and produce bipolar cells and rod photoreceptors. Although these newly produced neurons were limited in number, retinoic acid treatment promoted the number of regenerated bipolar cells. Moreover, misexpression of basic helix–loop–helix and homeobox genes promoted the induction of amacrine, horizontal, and rod photoreceptor specific phenotypes. These findings demonstrated that retinal neurons regenerated even in adult mammalian retina after toxic injury. Furthermore, we could partially control the fate of the regenerated neurons with extrinsic factors or intrinsic genes. The Müller glial cells constitute a potential source for the regeneration of adult mammalian retina and can be a target for drug delivery and gene therapy in retinal degenerative diseases. PMID:15353594
1983) Neuronal subpopulations in cat retina which accumulate the GABA agonist , ( H)muscimol: A combined Golgi and autoradiographic study, J. Comp...chromatography.Preliminary findings indicate that acetylcholine produces no significant changes in the release of gamma-aminobutyric acid or taurine but causes a...gamma-aminobutyric acid, glycine, and taurine . Concentrations of ACh less than 1 mmol were ineffective in 6 causing amino acid release. After exposure
Tosini, Gianluca; Fukuhara, Chiaki
Many physiological, cellular, and biochemical parameters in the retina of vertebrates show daily rhythms that, in many cases, also persist under constant conditions. This demonstrates that they are driven by a circadian pacemaker. The presence of an autonomous circadian clock in the retina of vertebrates was first demonstrated in Xenopus laevis and then, several years later, in mammals. In X. laevis and in chicken, the retinal circadian pacemaker has been localized in the photoreceptor layer, whereas in mammals, such information is not yet available. Recent advances in molecular techniques have led to the identification of a group of genes that are believed to constitute the molecular core of the circadian clock. These genes are expressed in the retina, although with a slightly different 24-h profile from that observed in the central circadian pacemaker. This result suggests that some difference (at the molecular level) may exist between the retinal clock and the clock located in the suprachiasmatic nuclei of hypothalamus. The present review will focus on the current knowledge of the retinal rhythmicity and the mechanisms responsible for its control.
Matsuoka, Ryota L; Nguyen-Ba-Charvet, Kim T; Parray, Aijaz; Badea, Tudor C; Chédotal, Alain; Kolodkin, Alex L
In the vertebrate retina, establishment of precise synaptic connections among distinct retinal neuron cell types is critical for processing visual information and for accurate visual perception. Retinal ganglion cells (RGCs), amacrine cells and bipolar cells establish stereotypic neurite arborization patterns to form functional neural circuits in the inner plexiform layer (IPL), a laminar region that is conventionally divided into five major parallel sublaminae. However, the molecular mechanisms governing distinct retinal subtype targeting to specific sublaminae within the IPL remain to be elucidated. Here we show that the transmembrane semaphorin Sema6A signals through its receptor PlexinA4 (PlexA4) to control lamina-specific neuronal stratification in the mouse retina. Expression analyses demonstrate that Sema6A and PlexA4 proteins are expressed in a complementary fashion in the developing retina: Sema6A in most ON sublaminae and PlexA4 in OFF sublaminae of the IPL. Mice with null mutations in PlexA4 or Sema6A exhibit severe defects in stereotypic lamina-specific neurite arborization of tyrosine hydroxylase (TH)-expressing dopaminergic amacrine cells, intrinsically photosensitive RGCs (ipRGCs) and calbindin-positive cells in the IPL. Sema6A and PlexA4 genetically interact in vivo for the regulation of dopaminergic amacrine cell laminar targeting. Therefore, neuronal targeting to subdivisions of the IPL in the mammalian retina is directed by repulsive transmembrane guidance cues present on neuronal processes.
Matsuoka, Ryota L.; Nguyen-Ba-Charvet, Kim T.; Parray, Aijaz; Badea, Tudor C.; Chédotal, Alain; Kolodkin, Alex L.
In the vertebrate retina, establishment of precise synaptic connections among distinct retinal neuron cell types is critical for processing visual information and for accurate visual perception. Retinal ganglion cells (RGCs), amacrine cells, and bipolar cells establish stereotypic neurite arborization patterns to form functional neural circuits in the inner plexiform layer (IPL)1–3: a laminar region that is conventionally divided into five major parallel sublaminae1,2. However, the molecular mechanisms governing distinct retinal subtype targeting to specific sublaminae within the IPL remain to be elucidated. Here, we show that the transmembrane semaphorin Sema6A signals through its receptor PlexinA4 (PlexA4) to control lamina-specific neuronal stratification in the mouse retina. Expression analyses demonstrate that Sema6A and PlexA4 proteins are expressed in a complementary fashion in the developing retina: Sema6A in most ON sublaminae and PlexA4 in OFF sublaminae of the IPL. Mice with null mutations in PlexA4 or Sema6A exhibit severe defects in stereotypic lamina-specific neurite arborization of tyrosine hydroxylase (TH)-expressing dopaminergic amacrine cells, intrinsically photosensitive RGCs (ipRGCs), and calbindin-positive cells in the IPL. Sema6A and PlexA4 genetically interact in vivo with respect to the regulation of dopaminergic amacrine cell laminar targeting. Therefore, neuronal targeting to subdivisions of the IPL in the mammalian retina is directed by repulsive transmembrane guidance cues present on neuronal processes. PMID:21270798
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
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.
Dakubo, Gabriel D; Wallace, Valerie A
The mature vertebrate retina develops from a population of multipotential neural progenitor cells that give rise to all of the retinal neurons and one glial cell type. Retinal histogenesis is regulated, in part, by cell extrinsic cues. A growing number of studies now implicate signaling by members of the Hedgehog (Hh) family of morphogens in vertebrate retinal development. In this review we will discuss the role of Hh signals from retinal ganglion cells (RGCs), the projection neurons of the retina, on proliferation, differentiation and lamination in the neural retina.
Novikova, Iu P; Aleĭnikova, K S; Krasnov, M S; Poplinskaia, V A; Grigorian, E N
Adult rat and newt retinas were studied during long organotypic 3D cultivation. A high proliferation level was discovered in the region of growth by applying DNA synthesis markers and in vitro mitosis registration in newt retina. Aggregates were formed in the retina spheroid cavity because dedifferentiated cells migrated into this region. Small cell populations in nuclear layers also had dividing and migration capacity. Rosette formation has been shown in newt retina. It is a characteristic of fetal retinal development under pathological conditions. The antiG FAP antibody dye demonstrated an increase in the parent M@uller cell population and generation of a small cell pool with short GFAP-extensions de novo. Recoverin expression studies detected its translocation from photoreceptor extensions to the cell bodies. Moreover, protein was presented in some cells inside the spheroid. It has been shown for the first time that cell proliferation occurred in the developing adult rat retinal spheroid in vitro; BrdU-positive cells and multiple mitoses were revealed in this zone. However, the source of proliferation was not in the peripheral retina, and stable macrophages and glial cells located among neurons of the inner nuclear layer had the ability to divide. The antiGFAP antibody showed an increase in GFAP fibers in the rat retina as well as in the newt retina. Recoverin translocated into photoreceptor perikaryons and the outer plexiform layer in cultivated rat retina. Interestingly, some cells with probably de novo expression of recoverin were discovered in rat and newt retinas.
Marcucci, Florencia; Murcia-Belmonte, Veronica; Coca, Yaiza; Ferreiro-Galve, Susana; Wang, Qing; Kuwajima, Takaaki; Khalid, Sania; Ross, M. Elizabeth; Herrera, Eloisa; Mason, Carol
Summary The retina of lower vertebrates grows continuously by integrating new neurons generated from progenitors in the ciliary margin zone (CMZ). Whether the mammalian CMZ provides the neural retina with retinal cells is controversial. Live-imaging of embryonic retina expressing eGFP in the CMZ shows that cells migrate laterally from the CMZ to the neural retina where differentiated retinal ganglion cells (RGCs) reside. As Cyclin D2, a cell-cycle regulator, is enriched in ventral CMZ, we analyzed Cyclin D2−/− mice to test whether the CMZ is a source of retinal cells. Neurogenesis is diminished in Cyclin D2 mutants, leading to a reduction of RGCs in the ventral retina. In line with these findings, in the albino retina, the decreased production of ipsilateral RGCs is correlated with fewer Cyclin D2+ cells. Together, these results implicate the mammalian CMZ as a neurogenic site that produces RGCs and whose proper generation depends on Cyclin D2 activity. PMID:28009286
Signorovitch, James; Raviola, Elio; Pawlyk, Basil; Li, Tiansen; Weitz, Charles J.
SUMMARY Circadian clocks are widely distributed in mammalian tissues, but little is known about the physiological functions of clocks outside the suprachiasmatic nucleus of the brain. The retina has an intrinsic circadian clock, but its importance for vision is unknown. Here we show that mice lacking Bmal1, a gene required for clock function, had abnormal retinal transcriptional responses to light and defective inner retinal electrical responses to light, but normal photoreceptor responses to light and retinas that appeared structurally normal by light and electron microscopy. We generated mice with a retina-specific genetic deletion of Bmal1, and they had defects of retinal visual physiology essentially identical to those of mice lacking Bmal1 in all tissues and lacked a circadian rhythm of inner retinal electrical responses to light. Our findings indicate that the intrinsic circadian clock of the retina regulates retinal visual processing in vivo. PMID:17719549
... biology of the retina. In: Tasman W, Jaeger EA, eds. Duane's Foundations of Clinical Ophthalmology . 2013 ed. ... and retina with ultrasound. In: Tasman W, Jaeger EA, eds. Duane's Ophthalmology . 2013 ed. Philadelphia, PA: Lippincott ...
Venugopalan, Praseeda; Wang, Yan; Nguyen, Tu; Huang, Abigail; Muller, Kenneth J; Goldberg, Jeffrey L
Retinal ganglion cells (RGCs) degenerate in diseases like glaucoma and are not replaced in adult mammals. Here we investigate whether transplanted RGCs can integrate into the mature retina. We have transplanted GFP-labelled RGCs into uninjured rat retinas in vivo by intravitreal injection. Transplanted RGCs acquire the general morphology of endogenous RGCs, with axons orienting towards the optic nerve head of the host retina and dendrites growing into the inner plexiform layer. Preliminary data show in some cases GFP(+) axons extending within the host optic nerves and optic tract, reaching usual synaptic targets in the brain, including the lateral geniculate nucleus and superior colliculus. Electrophysiological recordings from transplanted RGCs demonstrate the cells' electrical excitability and light responses similar to host ON, ON-OFF and OFF RGCs, although less rapid and with greater adaptation. These data present a promising approach to develop cell replacement strategies in diseased retinas with degenerating RGCs.
Ho, T; Vessey, K A; Fletcher, E L
Extracellular adenosine 5'-triphosphate (eATP) acts as a neurotransmitter within the retina and brain, activating a range of ionotropic P2X and metabotropic P2Y receptors. In this study, the specific localization of the P2X4 receptor (P2X4-R) subunit was evaluated in the retina using fluorescence immunohistochemistry and pre-embedding immuno-electron microscopy. Punctate P2X4-R labeling was largely localized to the inner and outer plexiform layers of mouse, rat and cat retinae. In the mouse outer retina, double-labeling of P2X4-R with the horizontal cell marker, calbindin, revealed P2X4-R immunoreactivity (P2X4-R-IR) on horizontal cell somata and processes. In the inner retina, P2X4-R expression was found closely associated with rod and cone bipolar cell terminals, and the punctate labeling was observed on calretinin-positive amacrine cells. Using immuno-electron microscopy, P2X4-Rs were observed on processes post-synaptic to photoreceptor and bipolar cell terminals, likely representing horizontal, amacrine and ganglion cells, respectively. Furthermore, P2X4-R expression was also observed on Müller cells, astrocytes and microglia. These data suggest a role for P2X4-Rs in the lateral inhibitory pathways of the retina, modulating neuronal function of photoreceptors and bipolar cells. The expression on macro- and microglial cells implicates a role for P2X4-Rs in glial signaling, tissue homeostasis and immunosurveillance within the mammalian retina.
Ding, Huayu; Smith, Robert G; Poleg-Polsky, Alon; Diamond, Jeffrey S; Briggman, Kevin L
Directionally tuned signalling in starburst amacrine cell (SAC) dendrites lies at the heart of the circuit that detects the direction of moving stimuli in the mammalian retina. The relative contributions of intrinsic cellular properties and network connectivity to SAC direction selectivity remain unclear. Here we present a detailed connectomic reconstruction of SAC circuitry in mouse retina and describe two previously unknown features of synapse distributions along SAC dendrites: input and output synapses are segregated, with inputs restricted to proximal dendrites; and the distribution of inhibitory inputs is fundamentally different from that observed in rabbit retina. An anatomically constrained SAC network model suggests that SAC–SAC wiring differences between mouse and rabbit retina underlie distinct contributions of synaptic inhibition to velocity and contrast tuning and receptive field structure. In particular, the model indicates that mouse connectivity enables SACs to encode lower linear velocities that account for smaller eye diameter, thereby conserving angular velocity tuning. These predictions are confirmed with calcium imaging of mouse SAC dendrites responding to directional stimuli.
Ding, Huayu; Smith, Robert G.; Poleg-Polsky, Alon; Diamond, Jeffrey S.; Briggman, Kevin L.
Summary Directionally tuned signaling in starburst amacrine cell (SAC) dendrites lies at the heart of the direction selective (DS) circuit in the mammalian retina. The relative contributions of intrinsic cellular properties and network connectivity to SAC DS remain unclear. We present a detailed connectomic reconstruction of SAC circuitry in mouse retina and describe previously unknown features of synapse distributions along SAC dendrites: 1) input and output synapses are segregated, with inputs restricted to proximal dendrites; 2) the distribution of inhibitory inputs is fundamentally different from that observed in rabbit retina. An anatomically constrained SAC network model suggests that SAC-SAC wiring differences between mouse and rabbit retina underlie distinct contributions of synaptic inhibition to velocity and contrast tuning and receptive field structure. In particular, the model indicates that mouse connectivity enables SACs to encode lower linear velocities that account for smaller eye diameter, thereby conserving angular velocity tuning. These predictions are confirmed with calcium imaging of mouse SAC dendrites in response to directional stimuli. PMID:27350241
This study is directed toward the cytochemical localization of cholinergic markers in a mammalian (cat) retina and biochemical characterization of the interactions of cholinergic neurons with other neurotransmitters in the retina. Particular attention is paid to localization of acetylcholinesterase and the effects of anticholinesterase organophosphates on normal retinal function. Studies to date have shown the presence of newly synthesized acetylcholine in amacrine and displaced amacrine cells. Acetylcholinesterase was localized in both amacrine and ganglion cells. The presumed cholinotoxin, AF64A, causes severe destruction in the cat retina, involving both amacrine and ganglion cells. Although the evidence to date indicates that only amacrine cells are cholinergic, ganglion cells appear to play a major role in cholinergic or related pathways and may be particularly susceptible to organophosphate poisoning. The biochemical component of the study has centered on the development of a superfusion system in which to monitor the release of various amino acid transmitters in response to application of acetylcholine. Preliminary experiments suggest that cholinergic amacrine cells are presynaptic to glycinergic cells in the cat retina. After the normal pattern has been established, it should be possible to investigate the effects of changes in the level of acetylcholinesterase on these responses.
Sun, Lanfang; Kawano-Yamashita, Emi; Nagata, Takashi; Tsukamoto, Hisao; Furutani, Yuji; Koyanagi, Mitsumasa; Terakita, Akihisa
Mammals contain 1 melanopsin (Opn4) gene that is expressed in a subset of retinal ganglion cells to serve as a photopigment involved in non-image-forming vision such as photoentrainment of circadian rhythms. In contrast, most nonmammalian vertebrates possess multiple melanopsins that are distributed in various types of retinal cells; however, their functions remain unclear. We previously found that the lamprey has only 1 type of mammalian-like melanopsin gene, which is similar to that observed in mammals. Here we investigated the molecular properties and localization of melanopsin in the lamprey and other cyclostome hagfish retinas, which contribute to visual functions including image-forming vision and mainly to non-image-forming vision, respectively. We isolated 1 type of mammalian-like melanopsin cDNA from the eyes of each species. We showed that the recombinant lamprey melanopsin was a blue light-sensitive pigment and that both the lamprey and hagfish melanopsins caused light-dependent increases in calcium ion concentration in cultured cells in a manner that was similar to that observed for mammalian melanopsins. We observed that melanopsin was distributed in several types of retinal cells, including horizontal cells and ganglion cells, in the lamprey retina, despite the existence of only 1 melanopsin gene in the lamprey. In contrast, melanopsin was almost specifically distributed to retinal ganglion cells in the hagfish retina. Furthermore, we found that the melanopsin-expressing horizontal cells connected to the rhodopsin-containing short photoreceptor cells in the lamprey. Taken together, our findings suggest that in cyclostomes, the global distribution of melanopsin in retinal cells might not be related to the melanopsin gene number but to the extent of retinal contribution to visual function.
Sun, Lanfang; Kawano-Yamashita, Emi; Nagata, Takashi; Tsukamoto, Hisao; Furutani, Yuji; Koyanagi, Mitsumasa; Terakita, Akihisa
Mammals contain 1 melanopsin (Opn4) gene that is expressed in a subset of retinal ganglion cells to serve as a photopigment involved in non-image-forming vision such as photoentrainment of circadian rhythms. In contrast, most nonmammalian vertebrates possess multiple melanopsins that are distributed in various types of retinal cells; however, their functions remain unclear. We previously found that the lamprey has only 1 type of mammalian-like melanopsin gene, which is similar to that observed in mammals. Here we investigated the molecular properties and localization of melanopsin in the lamprey and other cyclostome hagfish retinas, which contribute to visual functions including image-forming vision and mainly to non-image-forming vision, respectively. We isolated 1 type of mammalian-like melanopsin cDNA from the eyes of each species. We showed that the recombinant lamprey melanopsin was a blue light-sensitive pigment and that both the lamprey and hagfish melanopsins caused light-dependent increases in calcium ion concentration in cultured cells in a manner that was similar to that observed for mammalian melanopsins. We observed that melanopsin was distributed in several types of retinal cells, including horizontal cells and ganglion cells, in the lamprey retina, despite the existence of only 1 melanopsin gene in the lamprey. In contrast, melanopsin was almost specifically distributed to retinal ganglion cells in the hagfish retina. Furthermore, we found that the melanopsin-expressing horizontal cells connected to the rhodopsin-containing short photoreceptor cells in the lamprey. Taken together, our findings suggest that in cyclostomes, the global distribution of melanopsin in retinal cells might not be related to the melanopsin gene number but to the extent of retinal contribution to visual function. PMID:25251771
Sánchez-Farías, Nuria; Candal, Eva
Neural stem cells give rise to transient progenitors termed neuroepithelial cells (NECs) and radial glial cells (RGCs). RGCs represent the major source of neurons, glia and adult stem cells in several regions of the central nervous system (CNS). RGCs are mostly transient in mammals, but they are widely maintained in the adult CNS of fishes, where they continue to be morphologically similar to RGCs in the mammalian brain and fulfill similar roles as progenitors and guide for migrating neurons. The retina of fishes offers an exceptional model to approach the study of adult neurogenesis because of the presence of constitutive proliferation from the ciliary marginal zone (CMZ), containing NECs, and from adult glial cells with radial morphology (the Müller glia). However, the cellular hierarchies and precise contribution of different types of progenitors to adult neurogenesis remain unsolved. We have analyzed the transition from NECs to RGCs and RGC differentiation in the retina of the cartilaginous fish Scyliorhinus canicula, which offers a particularly good spatial and temporal frame to investigate this process. We have characterized progenitor and adult RGCs by immunohistochemical detection of glial markers as glial fibrillary acidic protein (GFAP) and glutamine synthetase (GS). We have compared the emergence and localization of glial markers with that of proliferating cell nuclear antigen (PCNA, a proliferation maker) and Doublecortin (DCX, which increases at early stages of neuronal differentiation). During retinal development, GFAP-immunoreactive NECs located in the most peripheral CMZ (CMZp) codistribute with DCX-immunonegative cells. GFAP-immunoreactive RGCs and Müller cells are located in successive more central parts of the retina and codistribute with DCX- and DCX/GS-immunoreactive cells, respectively. The same types of progenitors are found in juveniles, suggesting that the contribution of the CMZ to adult neurogenesis implies a transition through the
Rodriguez, Allen R.; de Sevilla Müller, Luis Pérez; Brecha, Nicholas C.
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
Hufnagel, Robert B.; Le, Tien T.; Riesenberg, Ashley L.; Brown, Nadean L.
In the mammalian retina, neuronal differentiation begins in the dorso-central optic cup and sweeps peripherally and ventrally. While certain extrinsic factors have been implicated, little is known about the intrinsic factors that direct this process. In this study, we evaluate the expression and function of proneural bHLH transcription factors during the onset of mouse retinal neurogenesis. Dorso-central retinal progenitor cells that give rise to the first postmitotic neurons express Neurog2/Ngn2 and Atoh7/Math5. In the absence of Neurog2, the spread of neurogenesis stalls, along with Atoh7 expression and RGC differentiation. However, neurogenesis is eventually restored, and at birth Neurog2 mutant retinas are reduced in size, with only a slight increase in the retinal ganglion cell population. We find that the re-establishment of neurogenesis coincides with the onset of Ascl1 expression, and that Ascl1 can rescue the early arrest of neural development in the absence of Neurog2. Together, this study supports the hypothesis that the intrinsic factors Neurog2 and Ascl1 regulate the temporal progression of retinal neurogenesis by directing overlapping waves of neuron formation. PMID:20144606
Gamal, Eman M; Aly, Eman M; Mahmoud, Sherif S; Talaat, Mona S; Sallam, Abdel Sattar M
Ginkgo biloba extract has been therapeutically used for several decades to increase peripheral and cerebral blood flow as well as for the treatment of dementia. The extract contains multiple compounds such as flavonoids and terpenoids that are thought to contribute to its neuroprotective and vasotropic effects. In this study, we investigated the effect of prolonged administration of EGb 761, up to 10 weeks, on mammalian retina using Fourier transform infrared spectroscopy (FTIR). Two main groups were involved in this study: the normal group (n = 10); and EGb-administrated group (n = 50) that received-orally-a dose of 40 mg/kg/day EGb 761. The results demonstrated that EGb administration was associated with different beneficial effects on the retinal constituents especially the underlying amide I protein secondary structure components as well as the NH-OH region. It concluded that the optimum daily administration period of EGb (40 mg/kg) for ophthalmic applications that targeting the retina ranges from 5 to 8 weeks.
Pires, Susana S; Hughes, Steven; Turton, Michael; Melyan, Zare; Peirson, Stuart N; Zheng, Lei; Kosmaoglou, Maria; Bellingham, James; Cheetham, Michael E; Lucas, Robert J; Foster, Russell G; Hankins, Mark W; Halford, Stephanie
Melanopsin is the photopigment that confers photosensitivity to a subset of retinal ganglion cells (pRGCs) that regulate many non-image-forming tasks such as the detection of light for circadian entrainment. Recent studies have begun to subdivide the pRGCs on the basis of morphology and function, but the origin of these differences is not yet fully understood. Here we report the identification of two isoforms of melanopsin from the mouse Opn4 locus, a previously described long isoform (Opn4L) and a novel short isoform (Opn4S) that more closely resembles the sequence and structure of rat and human melanopsins. Both isoforms, Opn4L and Opn4S, are expressed in the ganglion cell layer of the retina, traffic to the plasma membrane and form a functional photopigment in vitro. Quantitative PCR revealed that Opn4S is 40 times more abundant than Opn4L. The two variants encode predicted proteins of 521 and 466 aa and only differ in the length of their C-terminal tails. Antibodies raised to isoform-specific epitopes identified two discrete populations of melanopsin-expressing RGCs, those that coexpress Opn4L and Opn4S and those that express Opn4L only. Recent evidence suggests that pRGCs show a range of anatomical subtypes, which may reflect the functional diversity reported for mouse Opn4-mediated light responses. The distinct isoforms of Opn4 described in this study provide a potential molecular basis for generating this diversity, and it seems likely that their differential expression plays a role in generating the variety of pRGC light responses found in the mammalian retina.
Farajian, Reza; Pan, Feng; Akopian, Abram; Völgyi, Béla; Bloomfield, Stewart A
Abstract A fundamental organizing feature of the visual system is the segregation of ON and OFF responses into parallel streams to signal light increment and decrement. However, we found that blockade of GABAergic inhibition unmasks robust ON responses in OFF α-ganglion cells (α-GCs). These ON responses had the same centre-mediated structure as the classic OFF responses of OFF α-GCs, but were abolished following disruption of the ON pathway with l-AP4. Experiments showed that both GABAA and GABAC receptors are involved in the masking inhibition of this ON response, located at presynaptic inhibitory synapses on bipolar cell axon terminals and possibly amacrine cell dendrites. Since the dendrites of OFF α-GCs are not positioned to receive excitatory inputs from ON bipolar cell axon terminals in sublamina-b of the inner plexiform layer (IPL), we investigated the possibility that gap junction-mediated electrical synapses made with neighbouring amacrine cells form the avenue for reception of ON signals. We found that the application of gap junction blockers eliminated the unmasked ON responses in OFF α-GCs, while the classic OFF responses remained. Furthermore, we found that amacrine cells coupled to OFF α-GCs display processes in both sublaminae of the IPL, thus forming a plausible substrate for the reception and delivery of ON signals to OFF α-GCs. Finally, using a multielectrode array, we found that masked ON and OFF signals are displayed by over one-third of ganglion cells in the rabbit and mouse retinas, suggesting that masked crossover excitation is a widespread phenomenon in the inner mammalian retina. PMID:21768265
Packer, Michael A.; Stasiv, Yuri; Benraiss, Abdellatif; Chmielnicki, Eva; Grinberg, Alexander; Westphal, Heiner; Goldman, Steven A.; Enikolopov, Grigori
Neural progenitor cells are widespread throughout the adult central nervous system but only give rise to neurons in specific loci. Negative regulators of neurogenesis have therefore been postulated, but none have yet been identified as subserving a significant role in the adult brain. Here we report that nitric oxide (NO) acts as an important negative regulator of cell proliferation in the adult mammalian brain. We used two independent approaches to examine the function of NO in adult neurogenesis. In a pharmacological approach, we suppressed NO production in the rat brain by intraventricular infusion of an NO synthase inhibitor. In a genetic approach, we generated a null mutant neuronal NO synthase knockout mouse line by targeting the exon encoding active center of the enzyme. In both models, the number of new cells generated in neurogenic areas of the adult brain, the olfactory subependyma and the dentate gyrus, was strongly augmented, which indicates that division of neural stem cells in the adult brain is controlled by NO and suggests a strategy for enhancing neurogenesis in the adult central nervous system.
Engelsberg, Karl; Ghosh, Fredrik
In this study we wanted to examine how an adult neuroretina from an animal with an eye similar to the human one survives in vitro. We also wanted to investigate how the culture process affects the adult retina when used in a transplantation paradigm. Full-thickness neuroretinal sheets from adult porcine eyes were dissected into pieces measuring 3 mm in diameter. These were kept in culture for 1-3 days. After this time, the explants were fixed or transplanted subretinally to adult pigs, which were killed after 72-74 days. Transplanted eyes, as well as tissue kept in culture only, were processed for hematoxylin and eosin staining and immunohistochemistry. Explants kept 1 day in vitro (DIV) displayed the normal morphology. In these specimens, single pyknotic cells were evident in the outer nuclear layer (ONL) and ganglion cell layer, but were more frequent in the inner nuclear layer (INL). After longer times in vitro, severe degenerative changes appeared. Transplanted explants kept 1 DIV prior to transplantation exhibited normal retinal lamination in two out of four specimens. Transducin and recoverin labeling revealed photoreceptors with inner segments in these grafts. Rod bipolar cells displayed a normal morphology. Vertically arranged Müller cells were also seen in the laminated grafts. Two of the three transplants kept 2 DIV displayed minimal lamination. Eyes with transplants kept 3 DIV prior to transplantation displayed degenerated grafts in all eyes. This study shows that adult porcine neuroretinal explants kept in culture for 1 day display a normal morphology in their major part. Additionally, 1-day explants can survive transplantation with retained morphology even after several months. This indicates the possibility of storing adult donor tissue between harvest and transplantation. The culture system may also be used in the future as a tool for manipulating retinal donor tissue prior to transplantation.
Gorsuch, Ryne A.; Hyde, David R.
This article examines our current knowledge underlying the mechanisms involved in neuronal regeneration in the adult zebrafish retina. Zebrafish, which has the capacity to regenerate a wide variety of tissues and organs (including the fins, kidney, heart, brain, and spinal cord), has become the premier model system to study retinal regeneration due to the robustness and speed of the response and the variety of genetic tools that can be applied to study this question. It is now well documented that retinal damage induces the resident Müller glia to dedifferentiate and reenter the cell cycle to produce neuronal progenitor cells that continue to proliferate, migrate to the damaged retinal layer and differentiate into the missing neuronal cell types. Increasing our understanding of how these cellular events are regulated and occur in response to neuronal damage may provide critical information that can be applied to stimulating a regeneration response in the mammalian retina. In this review, we will focus on the genes/proteins that regulate zebrafish retinal regeneration and will attempt to critically evaluate how these factors may interact to correctly orchestrate the definitive cellular events that occur during regeneration. PMID:23880528
Gorsuch, Ryne A; Hyde, David R
This article examines our current knowledge underlying the mechanisms involved in neuronal regeneration in the adult zebrafish retina. Zebrafish, which has the capacity to regenerate a wide variety of tissues and organs (including the fins, kidney, heart, brain, and spinal cord), has become the premier model system to study retinal regeneration due to the robustness and speed of the response and the variety of genetic tools that can be applied to study this question. It is now well documented that retinal damage induces the resident Müller glia to dedifferentiate and reenter the cell cycle to produce neuronal progenitor cells that continue to proliferate, migrate to the damaged retinal layer and differentiate into the missing neuronal cell types. Increasing our understanding of how these cellular events are regulated and occur in response to neuronal damage may provide critical information that can be applied to stimulating a regeneration response in the mammalian retina. In this review, we will focus on the genes/proteins that regulate zebrafish retinal regeneration and will attempt to critically evaluate how these factors may interact to correctly orchestrate the definitive cellular events that occur during regeneration.
Beach, Krista M.; Wang, Jianbo
In humans and other mammals, the neural retina does not spontaneously regenerate, and damage to the retina that kills retinal neurons results in permanent blindness. In contrast to embryonic stem cells, induced pluripotent stem cells, and embryonic/fetal retinal stem cells, Müller glia offer an intrinsic cellular source for regenerative strategies in the retina. Müller glia are radial glial cells within the retina that maintain retinal homeostasis, buffer ion flux associated with phototransduction, and form the blood/retinal barrier within the retina proper. In injured or degenerating retinas, Müller glia contribute to gliotic responses and scar formation but also show regenerative capabilities that vary across species. In the mammalian retina, regenerative responses achieved to date remain insufficient for potential clinical applications. Activation of JAK/STAT and MAPK signaling by CNTF, EGF, and FGFs can promote proliferation and modulate the glial/neurogenic switch. However, to achieve clinical relevance, additional intrinsic and extrinsic factors that restrict or promote regenerative responses of Müller glia in the mammalian retina must be identified. This review focuses on Müller glia and Müller glial-derived stem cells in the retina and phylogenetic differences among model vertebrate species and highlights some of the current progress towards understanding the cellular mechanisms regulating their regenerative response. PMID:28194183
Beach, Krista M; Wang, Jianbo; Otteson, Deborah C
In humans and other mammals, the neural retina does not spontaneously regenerate, and damage to the retina that kills retinal neurons results in permanent blindness. In contrast to embryonic stem cells, induced pluripotent stem cells, and embryonic/fetal retinal stem cells, Müller glia offer an intrinsic cellular source for regenerative strategies in the retina. Müller glia are radial glial cells within the retina that maintain retinal homeostasis, buffer ion flux associated with phototransduction, and form the blood/retinal barrier within the retina proper. In injured or degenerating retinas, Müller glia contribute to gliotic responses and scar formation but also show regenerative capabilities that vary across species. In the mammalian retina, regenerative responses achieved to date remain insufficient for potential clinical applications. Activation of JAK/STAT and MAPK signaling by CNTF, EGF, and FGFs can promote proliferation and modulate the glial/neurogenic switch. However, to achieve clinical relevance, additional intrinsic and extrinsic factors that restrict or promote regenerative responses of Müller glia in the mammalian retina must be identified. This review focuses on Müller glia and Müller glial-derived stem cells in the retina and phylogenetic differences among model vertebrate species and highlights some of the current progress towards understanding the cellular mechanisms regulating their regenerative response.
Kovács-Öller, Tamás; Debertin, Gábor; Balogh, Márton; Ganczer, Alma; Orbán, József; Nyitrai, Miklós; Balogh, Lajos; Kántor, Orsolya; Völgyi, Béla
Much knowledge about interconnection of human retinal neurons is inferred from results on animal models. Likewise, there is a lack of information on human retinal electrical synapses/gap junctions (GJ). Connexin36 (Cx36) forms GJs in both the inner and outer plexiform layers (IPL and OPL) in most species including humans. However, a comparison of Cx36 GJ distribution in retinas of humans and popular animal models has not been presented. To this end a multiple-species comparison was performed in retinas of 12 mammals including humans to survey the Cx36 distribution. Areas of retinal specializations were avoided (e.g., fovea, visual streak, area centralis), thus observed Cx36 distribution differences were not attributed to these species-specific architecture of central retinal areas. Cx36 was expressed in both synaptic layers in all examined retinas. Cx36 plaques displayed an inhomogenous IPL distribution favoring the ON sublamina, however, this feature was more pronounced in the human, swine and guinea pig while it was less obvious in the rabbit, squirrel monkey, and ferret retinas. In contrast to the relative conservative Cx36 distribution in the IPL, the labels in the OPL varied considerably among mammals. In general, OPL plaques were rare and rather small in rod dominant carnivores and rodents, whereas the human and the cone rich guinea pig retinas displayed robust Cx36 labels. This survey presented that the human retina displayed two characteristic features, a pronounced ON dominance of Cx36 plaques in the IPL and prevalent Cx36 plaque conglomerates in the OPL. While many species showed either of these features, only the guinea pig retina shared both. The observed similarities and subtle differences in Cx36 plaque distribution across mammals do not correspond to evolutionary distances but may reflect accomodation to lifestyles of examined species. PMID:28337128
Drew, Liam J.; Fusi, Stefano; Hen, René
In the adult mammalian brain, newly generated neurons are continuously incorporated into two networks: interneurons born in the subventricular zone migrate to the olfactory bulb, whereas the dentate gyrus (DG) of the hippocampus integrates locally born principal neurons. That the rest of the mammalian brain loses significant neurogenic capacity…
McMahon, Douglas G; Iuvone, P Michael; Tosini, Gianluca
The retinal circadian system represents a unique structure. It contains a complete circadian system and thus the retina represents an ideal model to study fundamental questions of how neural circadian systems are organized and what signaling pathways are used to maintain synchrony of the different structures in the system. In addition, several studies have shown that multiple sites within the retina are capable of generating circadian oscillations. The strength of circadian clock gene expression and the emphasis of rhythmic expression are divergent across vertebrate retinas, with photoreceptors as the primary locus of rhythm generation in amphibians, while in mammals clock activity is most robust in the inner nuclear layer. Melatonin and dopamine serve as signaling molecules to entrain circadian rhythms in the retina and also in other ocular structures. Recent studies have also suggested GABA as an important component of the system that regulates retinal circadian rhythms. These transmitter-driven influences on clock molecules apparently reinforce the autonomous transcription-translation cycling of clock genes. The molecular organization of the retinal clock is similar to what has been reported for the SCN although inter-neural communication among retinal neurons that form the circadian network is apparently weaker than those present in the SCN, and it is more sensitive to genetic disruption than the central brain clock. The melatonin-dopamine system is the signaling pathway that allows the retinal circadian clock to reconfigure retinal circuits to enhance light-adapted cone-mediated visual function during the day and dark-adapted rod-mediated visual signaling at night. Additionally, the retinal circadian clock also controls circadian rhythms in disk shedding and phagocytosis, and possibly intraocular pressure. Emerging experimental data also indicate that circadian clock is also implicated in the pathogenesis of eye disease and compelling experimental data
Liu, Qian; Guan, Liping; Huang, Bing; Li, Weihua; Su, Qiao; Yu, Minbin; Xu, Xiaoping; Luo, Ting; Lin, Shaochun; Sun, Xuerong; Chen, Mengfei; Chen, Xigu
Adult peripheral blood-derived cells are able to differentiate into a variety of cell types, including nerve cells, liver-like cells and epithelial cells. However, their differentiation into retina-like cells is controversial. In the present study, transdifferentiation potential of human adult peripheral blood mononuclear cells into retina-like cells and integration into the retina of mice were investigated. Freshly isolated adult peripheral blood mononuclear cells were divided into two groups: cells in group I were cultured in neural stem cell medium, and cells in group II were exposed to conditioned medium from rat retinal tissue culture. After 5 days, several distinct cell morphologies were observed, including standard mononuclear, neurons with one or two axons and elongated glial-like cells. Immunohistochemical analysis of neural stem cell, neuron and retina cell markers demonstrated that cells in both groups were nestin-, MAP2 (microtubule-associated protein)- and GFAP (glial fibrillary acidic protein)-positive. Flow cytometry results suggested a significant increase in nestin-, MAP2- and CD16-positive cells in group I and nestin-, GFAP-, MAP2-, vimentin- and rhodopsin-positive cells in group II. To determine survival, migration and integration in vivo, cell suspensions (containing group I or group II cells) were injected into the vitreous or the peritoneum. Tissue specimens were obtained and immunostained 4 weeks after transplantation. We found that cells delivered by intravitreal injection integrated into the retina. Labelled cells were not detected in the retina of mice receiving differentiated cells by intraperitoneal injection, but cells (groups I and II) were detected in the liver and spleen. Our findings revealed that human adult peripheral blood mononuclear cells could be induced to transdifferentiate into neural precursor cells and retinal progenitor cells in vitro, and the differentiated peripheral blood mononuclear cells can migrate and integrate
Chun, Myung-Hoon; Oh, Uhtaek; Kim, In-Beom
Calcium (Ca2+)-activated chloride (Cl−) channels (CaCCs) play a role in the modulation of action potentials and synaptic responses in the somatodendritic regions of central neurons. In the vertebrate retina, large Ca2+-activated Cl− currents (ICl(Ca)) regulate synaptic transmission at photoreceptor terminals; however, the molecular identity of CaCCs that mediate ICl(Ca) remains unclear. The transmembrane protein, TMEM16A, also called anoctamin 1 (ANO1), has been recently validated as a CaCC and is widely expressed in various secretory epithelia and nervous tissues. Despite the fact that tmem16a was first cloned in the retina, there is little information on its cellular localization and function in the mammalian retina. In this study, we found that ANO1 was abundantly expressed as puncta in 2 synaptic layers. More specifically, ANO1 immunoreactivity was observed in the presynaptic terminals of various retinal neurons, including photoreceptors. ICl(Ca) was first detected in dissociated rod bipolar cells expressing ANO1. ICl(Ca) was abolished by treatment with the Ca2+ channel blocker Co2+, the L-type Ca2+ channel blocker nifedipine, and the Cl− channel blockers 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) and niflumic acid (NFA). More specifically, a recently discovered ANO1-selective inhibitor, T16Ainh-A01, and a neutralizing antibody against ANO1 inhibited ICl(Ca) in rod bipolar cells. Under a current-clamping mode, the suppression of ICl(Ca) by using NPPB and T16Ainh-A01 caused a prolonged Ca2+ spike-like depolarization evoked by current injection in dissociated rod bipolar cells. These results suggest that ANO1 confers ICl(Ca) in retinal neurons and acts as an intrinsic regulator of the presynaptic membrane potential during synaptic transmission. PMID:23840801
Stella, Salvatore L.; Vila, Alejandro; Hung, Albert Y.; Rome, Michael E.; Huynh, Uyenchi; Sheng, Morgan; Kreienkamp, Hans-Juergen; Brecha, Nicholas C.
Photoreceptor terminals contain post-synaptic density (PSD) proteins e.g., PSD-95/PSD-93, but their role at photoreceptor synapses is not known. PSDs are generally restricted to post-synaptic boutons in central neurons and form scaffolding with multiple proteins that have structural and functional roles in neuronal signaling. The Shank family of proteins (Shank 1–3) functions as putative anchoring proteins for PSDs and is involved in the organization of cytoskeletal/signaling complexes in neurons. Specifically, Shank 1 is restricted to neurons and interacts with both receptors and signaling molecules at central neurons to regulate plasticity. However, it is not known whether Shank 1 is expressed at photoreceptor terminals. In this study we have investigated Shank 1A localization in the outer retina at photoreceptor terminals. We find that Shank 1A is expressed presynaptically in cone pedicles, but not rod spherules, and it is absent from mice in which the Shank 1 gene is deleted. Shank 1A co-localizes with PSD-95, peanut agglutinin, a marker of cone terminals, and glycogen phosphorylase, a cone specific marker. These findings provide convincing evidence for Shank 1A expression in both the inner and outer plexiform layers, and indicate a potential role for PSD-95/Shank 1 complexes at cone synapses in the outer retina. PMID:22984429
Prentice, Jason; Homann, Jan; Simmons, Kristy; Tkacik, Gasper; Balasubramanian, Vijay; Nelson, Philip
Multi-electrode array technology provides an efficient means of simultaneously recording from many neurons. However, as arrays become larger, a greater computational burden falls on the spike-sorting algorithm. We have developed a new method for sorting multi-electrode signals and applied it to retinal ganglion cells. Our method is explicitly designed to scale well with increasing array size. It can dissect temporally overlapping spikes and accommodate the amplitude variation seen in spike bursts. The broad outline of our method is to (1) identify spikes in the raw data, cluster a subset, generate template waveforms, then (2) fit the templates to all the data using an iterative Bayesian algorithm. Each of these two steps makes use of the 2D spatial arrangement of the ganglion cells and electrodes, and the locality of signals from each individual cell. We demonstrate the method on data recorded from guinea pig retina on a 30-electrode array.
Jin, Nan Ge
Rod photoreceptors are electrically coupled through gap junctions. Coupling is a key determinant of their light response properties, but whether rod electrical coupling is dynamically regulated remains elusive and controversial. Here, we have obtained direct measurements of the conductance between adjacent rods in mouse retina and present evidence that rod electrical coupling strength is dependent on the time of day, the lighting conditions, and the mouse strain. Specifically, we show in CBA/Ca mice that under circadian conditions, the rod junctional conductance has a median value of 98 pS during the subjective day and of 493 pS during the subjective night. In C57BL/6 mice, the median junctional conductance between dark-adapted rods is ∼140 pS, regardless of the time in the circadian cycle. Adaptation to bright light decreases the rod junctional conductance to ∼0 pS, regardless of the time of day or the mouse strain. Together, these results establish the high degree of plasticity of rod electrical coupling over the course of the day. Estimates of the rod coupling strength will provide a foundation for further investigations of rod interactions and the role of rod coupling in the ability of the visual system to anticipate, assimilate, and respond to the daily changes in ambient light intensity. SIGNIFICANCE STATEMENT Many cells in the CNS communicate via gap junctions, or electrical synapses, the regulation of which remains largely unknown. Here, we show that the strength of electrical coupling between rod photoreceptors of the retina is regulated by the time of day and the lighting conditions. This mechanism may help us understand some key aspects of day and night vision as well as some visual malfunctions. PMID:26740659
Bond, Allison M.; Ming, Guo-li; Song, Hongjun
Summary Adult somatic stem cells in various organs maintain homeostatic tissue regeneration and enhance plasticity. Since its initial discovery five decades ago, investigations of adult neurogenesis and neural stem cells have led to an established and expanding field that has significantly influenced many facets of neuroscience, developmental biology and regenerative medicine. Here we review recent progress and focus on questions related to adult mammalian neural stem cells that also apply to other somatic stem cells. We further discuss emerging topics that are guiding the field toward better understanding adult neural stem cells and ultimately applying these principles to improve human health. PMID:26431181
Kuhn, H Georg
The fact that continuous proliferation of stem cells and progenitors, as well as the production of new neurons, occurs in the adult mammalian central nervous system (CNS) raises several basic questions concerning the number of neurons required in a particular system. Can we observe continued growth of brain regions that sustain neurogenesis? Or does an elimination mechanism exist to maintain a constant number of cells? If so, are old neurons replaced, or are the new neurons competing for limited network access among each other? What signals support their survival and integration and what factors are responsible for their elimination? This review will address these and other questions regarding regulatory mechanisms that control cell-death and cell-survival mechanisms during neurogenesis in the intact adult mammalian brain.
Arisi, Gabriel M; Foresti, Maira L; Mukherjee, Sanjib; Shapiro, Lee A
Neurogenesis occurs in the adult mammalian brain in discrete regions related to olfactory sensory signaling and integration. The olfactory receptor cell population is in constant turn-over through local progenitor cells. Also, newborn neurons are added to the olfactory bulbs through a major migratory route from the subventricular zone, the rostral migratory stream. The olfactory bulbs project to different brain structures, including: piriform cortex, amygdala, entorhinal cortex, striatum and hippocampus. These structures play important roles in odor identification, feeding behavior, social interactions, reproductive behavior, behavioral reinforcement, emotional responses, learning and memory. In all of these regions neurogenesis has been described in normal and in manipulated mammalian brain. These data are reviewed in the context of a sensory-behavioral hypothesis on adult neurogenesis that olfactory input modulates neurogenesis in many different regions of the brain.
Li, Tianqing; Lewallen, Michelle; Chen, Shuyi; Yu, Wei; Zhang, Nian; Xie, Ting
Various stem cell types have been tested for their potential application in treating photoreceptor degenerative diseases, such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD). Only embryonic stem cells (ESCs) have so far been shown to generate functional photoreceptor cells restoring light response of photoreceptor-deficient mice, but there is still some concern of tumor formation. In this study, we have successfully cultured Nestin(+)Sox2(+)Pax6(+) multipotent retinal stem cells (RSCs) from the adult mouse retina, which are capable of producing functional photoreceptor cells that restore the light response of photoreceptor-deficient rd1 mutant mice following transplantation. After they have been expanded for over 35 passages in the presence of FGF and EGF, the cultured RSCs still maintain stable proliferation and differentiation potential. Under proper differentiation conditions, they can differentiate into all the major retinal cell types found in the adult retina. More importantly, they can efficiently differentiate into photoreceptor cells under optimized differentiation conditions. Following transplantation into the subretinal space of slowly degenerating rd7 mutant eyes, RSC-derived photoreceptor cells integrate into the retina, morphologically resembling endogenous photoreceptors and forming synapases with resident retinal neurons. When transplanted into eyes of photoreceptor-deficient rd1 mutant mice, a RP model, RSC-derived photoreceptors can partially restore light response, indicating that those RSC-derived photoreceptors are functional. Finally, there is no evidence for tumor formation in the photoreceptor-transplanted eyes. Therefore, this study has demonstrated that RSCs isolated from the adult retina have the potential of producing functional photoreceptor cells that can potentially restore lost vision caused by loss of photoreceptor cells in RP and AMD.
Singh, Ratnesh K; Mallela, Ramya K; Hayes, Abigail; Dunham, Nicholas R; Hedden, Morgan E; Enke, Raymond A; Fariss, Robert N; Sternberg, Hal; West, Michael D; Nasonkin, Igor O
Characterizing the role of epigenetic regulation in the mammalian retina is critical for understanding fundamental mechanisms of retinal development and disease. DNA methylation, an epigenetic modifier of genomic DNA, plays an important role in modulating networks of tissue and cell-specific gene expression. However, the impact of DNA methylation during retinal development and homeostasis of retinal neurons remains unclear. Here, we have created a tissue-specific DNA methyltransferase (Dnmt) triple mutant mouse in an effort to characterize the impact of DNA methylation in retinal development and homeostasis. An Rx-Cre transgene was used to drive targeted mutation of all three murine Dnmt genes in the mouse retina encoding major DNA methylation enzymes DNMT1, DNMT3A and DNMT3B. The triple mutant mice represent a hypomorph model since Dnmt1 catalytic activity was still present and excision of Dnmt3a and Dnmt3b had only about 90% efficiency. Disruption of all three Dnmts resulted in global genomic hypomethylation and dramatic reorganization of the photoreceptor and synaptic layers within retina. Transcriptome and proteomic analyses demonstrated enrichment of dysregulated phototransduction and synaptic genes. The 5 mC signal in triple mutant retina was confined to the central heterochromatin but reduced in the peripheral heterochromatin region of photoreceptor nuclei. In addition, we found a reduction of the 5 mC signal in ganglion cell nuclei. Collectively, this data suggests cooperation of all three Dnmts in the formation and homeostasis of photoreceptors and other retinal neurons within the mammalian retina, and highlight the relevance of epigenetic regulation to sensory retinal disorders and vision loss.
Sánchez-Farías, Nuria; Candal, Eva
Doublecortin (DCX) is a microtubule-associated protein that has been considered a marker for neuronal precursors and young migrating neurons during the development of the central nervous system and in adult neurogenic niches. The retina of fishes represents an accessible, continuously growing and highly structured (layered) part of the central nervous system and, therefore, offers an exceptional model to extend our knowledge on the possible role of DCX in promoting neurogenesis and migration to appropriate layers. We have analyzed the distribution of DCX in the embryonic and postembryonic retina of a small shark, the lesser spotted dogfish Scyliorhinus canicula, by means of immunohistochemistry. We investigated the relationship between DCX expression and the neurogenic state of DCX-labeled cells by exploring its co-localization with the proliferation marker PCNA (proliferating cell nuclear antigen) and the marker of neuronal differentiation HuC/D. Since radially migrating neurons use radial glial fibers as substrate, we explored the possible correlation between DCX expression and cell migration along radial glia by comparing its expression with that of the glial marker GFAP (glial fibrillary acidic protein). Additionally, we characterized DCX-expressing cells by double immunocytochemistry using antibodies against Calbindin (a marker for mature bipolar and horizontal cells in this species) and Pax6, which has been proposed as a regulator of cell proliferation, cell differentiation, and neuron diversification in the neural retina of sharks. Strong DCX immunoreactivity was observed in immature cells and cell processes, at a time when retinal cells were not yet organized into different laminae. DCX was also found in subsets of mature ganglion, amacrine, bipolar and horizontal cells long after they had exited the cell cycle, a pattern that was maintained in juveniles and adults. Our results on DCX expression in the retina are compatible with a role for DCX in cell
Eberle, Dominic; Santos-Ferreira, Tiago; Grahl, Sandra; Ader, Marius
Vision impairment and blindness due to the loss of the light-sensing cells of the retina, i.e. photoreceptors, represents the main reason for disability in industrialized countries. Replacement of degenerated photoreceptors by cell transplantation represents a possible treatment option in future clinical applications. Indeed, recent preclinical studies demonstrated that immature photoreceptors, isolated from the neonatal mouse retina at postnatal day 4, have the potential to integrate into the adult mouse retina following subretinal transplantation. Donor cells generated a mature photoreceptor morphology including inner and outer segments, a round cell body located at the outer nuclear layer, and synaptic terminals in close proximity to endogenous bipolar cells. Indeed, recent reports demonstrated that donor photoreceptors functionally integrate into the neural circuitry of host mice. For a future clinical application of such cell replacement approach, purified suspensions of the cells of choice have to be generated and placed at the correct position for proper integration into the eye. For the enrichment of photoreceptor precursors, sorting should be based on specific cell surface antigens to avoid genetic reporter modification of donor cells. Here we show magnetic-associated cell sorting (MACS) - enrichment of transplantable rod photoreceptor precursors isolated from the neonatal retina of photoreceptor-specific reporter mice based on the cell surface marker CD73. Incubation with anti-CD73 antibodies followed by micro-bead conjugated secondary antibodies allowed the enrichment of rod photoreceptor precursors by MACS to approximately 90%. In comparison to flow cytometry, MACS has the advantage that it can be easier applied to GMP standards and that high amounts of cells can be sorted in relative short time periods. Injection of enriched cell suspensions into the subretinal space of adult wild-type mice resulted in a 3-fold higher integration rate compared to
Ma, Dengke K; Marchetto, Maria Carolina; Guo, Junjie U; Ming, Guo-li; Gage, Fred H; Song, Hongjun
Epigenetic mechanisms regulate cell differentiation during embryonic development and also serve as important interfaces between genes and the environment in adulthood. Neurogenesis in adults, which generates functional neural cell types from adult neural stem cells, is dynamically regulated by both intrinsic state-specific cell differentiation cues and extrinsic neural niche signals. Epigenetic regulation by DNA and histone modifiers, non-coding RNAs and other self-sustained mechanisms can lead to relatively long-lasting biological effects and maintain functional neurogenesis throughout life in discrete regions of the mammalian brain. Here, we review recent evidence that epigenetic mechanisms carry out diverse roles in regulating specific aspects of adult neurogenesis and highlight the implications of such epigenetic regulation for neural plasticity and disorders. PMID:20975758
Drew, Liam J.; Fusi, Stefano; Hen, René
In the adult mammalian brain, newly generated neurons are continuously incorporated into two networks: interneurons born in the subventricular zone migrate to the olfactory bulb, whereas the dentate gyrus (DG) of the hippocampus integrates locally born principal neurons. That the rest of the mammalian brain loses significant neurogenic capacity after the perinatal period suggests that unique aspects of the structure and function of DG and olfactory bulb circuits allow them to benefit from the adult generation of neurons. In this review, we consider the distinctive features of the DG that may account for it being able to profit from this singular form of neural plasticity. Approaches to the problem of neurogenesis are grouped as “bottom-up,” where the phenotype of adult-born granule cells is contrasted to that of mature developmentally born granule cells, and “top-down,” where the impact of altering the amount of neurogenesis on behavior is examined. We end by considering the primary implications of these two approaches and future directions. PMID:24255101
Ferrer-Martín, Rosa M; Martín-Oliva, David; Sierra, Ana; Carrasco, Maria-Carmen; Martín-Estebané, María; Calvente, Ruth; Marín-Teva, José L; Navascués, Julio; Cuadros, Miguel A
Organotypic cultures of retinal explants allow the detailed analysis of microglial cells in a cellular microenvironment similar to that in the in situ retina, with the advantage of easy experimental manipulation. However, the in vitro culture causes changes in the retinal cytoarchitecture and induces a microglial response that may influence the results of these manipulations. The purpose of this study was to analyze the influence of the retinal age on changes in retinal cytoarchitecture, cell viability and death, and microglial phenotype and distribution throughout the in vitro culture of developing and adult retina explants. Explants from developing (3 and 10 postnatal days, P3 and P10) and adult (P60) mouse retinas were cultured for up to 10 days in vitro (div). Dead or dying cells were recognized by TUNEL staining, cell viability was determined by flow cytometry, and the numbers and distribution patterns of microglial cells were studied by flow cytometry and immunocytochemistry, respectively. The retinal cytoarchitecture was better preserved at prolonged culture times (10 div) in P10 retina explants than in P3 or adult explants. Particular patterns of cell viability and death were observed at each age: in general, explants from developing retinas showed higher cell viability and lower density of TUNEL-positive profiles versus adult retinas. The proportion of microglial cells relative to the whole population of retinal cells was higher in explants fixed immediately after their dissection (i.e., non-cultured) from adult retinas than in those from developing retinas. This proportion was always higher in non-cultured explants than in explants at 10 div, suggesting the death of some microglial cells during the culture. Activation of microglial cells, as revealed by their phenotypical appearance, was observed in both developing and adult retina explants from the beginning of the culture. Immunofluorescence with the anti-CD68 antibody showed that some activated
Hauser, Alexandra; Lepper, Marlen Franziska; Mayo, Rebecca
To date, the proteomic profiling of Müller cells, the dominant macroglia of the retina, has been hampered because of the absence of suitable enrichment methods. We established a novel protocol to isolate native, intact Müller cells from adult murine retinae at excellent purity which retain in situ morphology and are well suited for proteomic analyses. Two different strategies of sample preparation - an in StageTips (iST) and a subcellular fractionation approach including cell surface protein profiling were used for quantitative liquid chromatography-mass spectrometry (LC-MSMS) comparing Müller cell-enriched to depleted neuronal fractions. Pathway enrichment analyses on both data sets enabled us to identify Müller cell-specific functions which included focal adhesion kinase signaling, signal transduction mediated by calcium as second messenger, transmembrane neurotransmitter transport and antioxidant activity. Pathways associated with RNA processing, cellular respiration and phototransduction were enriched in the neuronal subpopulation. Proteomic results were validated for selected Müller cell genes by quantitative real time PCR, confirming the high expression levels of numerous members of the angiogenic and anti-inflammatory annexins and antioxidant enzymes (e.g. paraoxonase 2, peroxiredoxin 1, 4 and 6). Finally, the significant enrichment of antioxidant proteins in Müller cells was confirmed by measurements on vital retinal cells using the oxidative stress indicator CM-H2DCFDA. In contrast to photoreceptors or bipolar cells, Müller cells were most efficiently protected against H2O2-induced reactive oxygen species formation, which is in line with the protein repertoire identified in the proteomic profiling. Our novel approach to isolate intact glial cells from adult retina in combination with proteomic profiling enabled the identification of novel Müller glia specific proteins, which were validated as markers and for their functional impact in glial
Feliciano, David M; Bordey, Angélique; Bonfanti, Luca
Two decades after the discovery that neural stem cells (NSCs) populate some regions of the mammalian central nervous system (CNS), deep knowledge has been accumulated on their capacity to generate new neurons in the adult brain. This constitutive adult neurogenesis occurs throughout life primarily within remnants of the embryonic germinal layers known as "neurogenic sites." Nevertheless, some processes of neurogliogenesis also occur in the CNS parenchyma commonly considered as "nonneurogenic." This "noncanonical" cell genesis has been the object of many claims, some of which turned out to be not true. Indeed, it is often an "incomplete" process as to its final outcome, heterogeneous by several measures, including regional location, progenitor identity, and fate of the progeny. These aspects also strictly depend on the animal species, suggesting that persistent neurogenic processes have uniquely adapted to the brain anatomy of different mammals. Whereas some examples of noncanonical neurogenesis are strictly parenchymal, others also show stem cell niche-like features and a strong link with the ventricular cavities. This work will review results obtained in a research field that expanded from classic neurogenesis studies involving a variety of areas of the CNS outside of the subventricular zone (SVZ) and subgranular zone (SGZ). It will be highlighted how knowledge concerning noncanonical neurogenic areas is still incomplete owing to its regional and species-specific heterogeneity, and to objective difficulties still hampering its full identification and characterization.
Sun, Lei; Li, Ping; Carr, Aprell L.; Gorsuch, Ryne; Yarka, Clare; Li, Jingling; Bartlett, Michael; Pfister, Delaney; Hyde, David R.; Li, Lei
The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in vertebrate species. Previously, we identified a homolog of the Stil gene in zebrafish mutant (night blindness b, nbb), which showed neural defects in the retina (e.g. dopaminergic cell degeneration and/or lack of regeneration). In this research, we examined the roles of Stil in cell proliferation after degeneration in adult zebrafish retinas. We demonstrated that knockdown of Stil gene expression or inhibition of Sonic hedgehog (Shh) signaling transduction decreases the rate of cell proliferation. In contrast, activation of Shh signal transduction promotes cell proliferation. In nbb+/− retinas, inhibition of SUFU (a repressor in the Shh pathway) rescues the defects in cell proliferation due to down-regulation of Stil gene expression. The latter data suggest that Stil play a role in cell proliferation through the Shh signal transduction pathway. PMID:24469449
Sun, Lei; Li, Ping; Carr, Aprell L; Gorsuch, Ryne; Yarka, Clare; Li, Jingling; Bartlett, Michael; Pfister, Delaney; Hyde, David R; Li, Lei
The human oncogene SCL/TAL1 interrupting locus (Stil) is highly conserved in vertebrate species. Previously, we identified a homolog of the Stil gene in zebrafish mutant (night blindness b, nbb), which showed neural defects in the retina (e.g. dopaminergic cell degeneration and/or lack of regeneration). In this research, we examined the roles of Stil in cell proliferation after degeneration in adult zebrafish retinas. We demonstrated that knockdown of Stil gene expression or inhibition of Sonic hedgehog (Shh) signaling transduction decreases the rate of cell proliferation. In contrast, activation of Shh signal transduction promotes cell proliferation. In nbb(+/-) retinas, inhibition of SUFU (a repressor in the Shh pathway) rescues the defects in cell proliferation due to down-regulation of Stil gene expression. The latter data suggest that Stil play a role in cell proliferation through the Shh signal transduction pathway.
The present study investigated the temporal pattern and cellular localization of nestin in the adult mouse retina with pharmaceutically induced retinal degeneration using N-methyl-N-nitrosourea (MNU). After a single intraperitoneal injection of MNU in 8-week-old C57BL/6 mice, the animals were sacrificed at 1, 3, 5, 7, and 21 days (n = 6, in each stage). The eyes were examined by means of immunohistochemical tests using nestin, ionized calcium-binding adaptor molecule (Iba-1), CD11b, F4/80, and glial fibrillary acidic protein (GFAP). Western blot analysis and manual cell counting were performed for quantification. Nestin expression was increased after MNU administration. Nestin+/Iba-1+ cells were migrated into outer nuclear layer (ONL) and peaked at day 3 post injection (PI). Nestin+/CD11b+ cells were also mainly identified in ONL at day 3 PI and peaked at day 5. Nestin+/F4/80+ cells were shown in the subretinal space and peaked at day 3 PI. Nestin+/GFAP+ cells were distinctly increased at day 1 PI and peaked at day 5 PI. The up-regulation of nestin expression after MNU administration in adult mouse retinal microglia, and monocyte/macrophage suggests that when retinal degeneration progresses, these cells may revert to a more developmentally immature state. Müller cells also showed reactive gliosis and differentiational changes. PMID:28049248
Wong, R. O. L.; Chernjavsky, A.; Smith, S. J.; Shatz, C. J.
IN the adult mammalian retina, the principal direction of information flow is along a vertical pathway from photoreceptors to retinal interneurons to ganglion cells, the output neurons of the retina. We report here, however, that initially in development, at a time when the photoreceptors are not yet even present, there are already functionally defined networks within the retina. These networks are spontaneously active rather than visually driven, and they involve horizontal rather than vertical pathways. By means of optical recording using the calcium-sensitive dye Fura-2, we have found that sets of retinal ganglion cells and amacrine cells, a type of retinal interneuron, undergo synchronized oscillations in intracellular calcium concentration. These oscillations are highly correlated among subgroups of neighbouring cells, and spread in a wave-like fashion tangentially across the retina. Thus, in development of retinal circuitry, the initial patterning of neuronal function occurs in the horizontal domain before the adult pattern of vertical information transfer emerges.
Trost, A; Schroedl, F; Marschallinger, J; Rivera, F J; Bogner, B; Runge, C; Couillard-Despres, S; Aigner, L; Reitsamer, H A
Doublecortin (DCX) is predominantly expressed in neuronal precursor cells and young immature neurons of the developing and adult brain, where it is involved in neuronal differentiation, migration and plasticity. Moreover, its expression pattern reflects neurogenesis, and transgenic DCX promoter-driven reporter models have been previously used to investigate adult neurogenesis. In this study, we characterize dsRed2 reporter protein-expressing cells in the adult retina of the transgenic DCX promoter-dsRed2 rat model, with the aim to identify cells with putative neurogenic activity. Additionally, we confirmed the expression of the dsRed2 protein in DCX-expressing cells in the adult hippocampal dentate gyrus. Adult DCX-dsRed2 rat retinas were analyzed by immunohistochemistry for expression of DCX, NF200, Brn3a, Sox2, NeuN, calbindin, calretinin, PKC-a, Otx2, ChAT, PSA-NCAM and the glial markers GFAP and CRALBP, followed by confocal laser-scanning microscopy. In addition, brain sections of transgenic rats were analyzed for dsRed2 expression and co-localization with DCX, NeuN, GFAP and Sox2 in the cortex and dentate gyrus. Endogenous DCX expression in the adult retina was confined to horizontal cells, and these cells co-expressed the DCX promoter-driven dsRed2 reporter protein. In addition, we encountered dsRed2 expression in various other cell types in the retina: retinal ganglion cells (RGCs), a subpopulation of amacrine cells, a minority of bipolar cells and in perivascular cells. Since also RGCs expressed dsRed2, the DCX-dsRed2 rat model might offer a useful tool to study RGCs in vivo under various conditions. Müller glial cells, which have previously been identified as cells with stem cell features and with neurogenic potential, did express neither endogenous DCX nor the dsRed2 reporter. However, and surprisingly, we identified a perivascular glial cell type expressing the dsRed2 reporter, enmeshed with the glia/stem cell marker GFAP and colocalizing with the
Hickmott, Jack W; Chen, Chih-yu; Arenillas, David J; Korecki, Andrea J; Lam, Siu Ling; Molday, Laurie L; Bonaguro, Russell J; Zhou, Michelle; Chou, Alice Y; Mathelier, Anthony; Boye, Sanford L; Hauswirth, William W; Molday, Robert S; Wasserman, Wyeth W; Simpson, Elizabeth M
Current gene therapies predominantly use small, strong, and readily available ubiquitous promoters. However, as the field matures, the availability of small, cell-specific promoters would be greatly beneficial. Here we design seven small promoters from the human paired box 6 (PAX6) gene and test them in the adult mouse retina using recombinant adeno-associated virus. We chose the retina due to previous successes in gene therapy for blindness, and the PAX6 gene since it is: well studied; known to be driven by discrete regulatory regions; expressed in therapeutically interesting retinal cell types; and mutated in the vision-loss disorder aniridia, which is in need of improved therapy. At the PAX6 locus, 31 regulatory regions were bioinformatically predicted, and nine regulatory regions were constructed into seven MiniPromoters. Driving Emerald GFP, these MiniPromoters were packaged into recombinant adeno-associated virus, and injected intravitreally into postnatal day 14 mice. Four MiniPromoters drove consistent retinal expression in the adult mouse, driving expression in combinations of cell-types that endogenously express Pax6: ganglion, amacrine, horizontal, and Müller glia. Two PAX6-MiniPromoters drive expression in three of the four cell types that express PAX6 in the adult mouse retina. Combined, they capture all four cell types, making them potential tools for research, and PAX6-gene therapy for aniridia. PMID:27556059
Weber, Anke; Hochmann, Sarah; Cimalla, Peter; Gärtner, Maria; Kuscha, Veronika; Hans, Stefan; Geffarth, Michaela; Kaslin, Jan; Koch, Edmund; Brand, Michael
Light-induced lesions are a powerful tool to study the amazing ability of photoreceptors to regenerate in the adult zebrafish retina. However, the specificity of the lesion towards photoreceptors or regional differences within the retina are still incompletely understood. We therefore characterized the process of degeneration and regeneration in an established paradigm, using intense white light from a fluorescence lamp on swimming fish (diffuse light lesion). We also designed a new light lesion paradigm where light is focused through a microscope onto the retina of an immobilized fish (focused light lesion). Focused light lesion has the advantage of creating a locally restricted area of damage, with the additional benefit of an untreated control eye in the same animal. In both paradigms, cell death is observed as an immediate early response, and proliferation is initiated around 2 days post lesion (dpl), peaking at 3 dpl. We furthermore find that two photoreceptor subtypes (UV and blue sensitive cones) are more susceptible towards intense white light than red/green double cones and rods. We also observed specific differences within light lesioned areas with respect to the process of photoreceptor degeneration: UV cone debris is removed later than any other type of photoreceptor in light lesions. Unspecific damage to retinal neurons occurs at the center of a focused light lesion territory, but not in the diffuse light lesion areas. We simulated the fish eye optical properties using software simulation, and show that the optical properties may explain the light lesion patterns that we observe. Furthermore, as a new tool to study retinal degeneration and regeneration in individual fish in vivo, we use spectral domain optical coherence tomography. Collectively, the light lesion and imaging assays described here represent powerful tools for studying degeneration and regeneration processes in the adult zebrafish retina. PMID:24303018
Esquerdo, Mariona; Grau-Bové, Xavier; Garanto, Alejandro; Toulis, Vasileios; Garcia-Monclús, Sílvia; Millo, Erica; López-Iniesta, Ma José; Abad-Morales, Víctor; Ruiz-Trillo, Iñaki; Marfany, Gemma
Ubiquitination is a relevant cell regulatory mechanism to determine protein fate and function. Most data has focused on the role of ubiquitin as a tag molecule to target substrates to proteasome degradation, and on its impact in the control of cell cycle, protein homeostasis and cancer. Only recently, systematic assays have pointed to the relevance of the ubiquitin pathway in the development and differentiation of tissues and organs, and its implication in hereditary diseases. Moreover, although the activity and composition of ubiquitin ligases has been largely addressed, the role of the deubiquitinating enzymes (DUBs) in specific tissues, such as the retina, remains mainly unknown. In this work, we undertook a systematic analysis of the transcriptional levels of DUB genes in the adult mouse retina by RT-qPCR and analyzed the expression pattern by in situ hybridization and fluorescent immunohistochemistry, thus providing a unique spatial reference map of retinal DUB expression. We also performed a systematic phylogenetic analysis to understand the origin and the presence/absence of DUB genes in the genomes of diverse animal taxa that represent most of the known animal diversity. The expression landscape obtained supports the potential subfunctionalization of paralogs in those families that expanded in vertebrates. Overall, our results constitute a reference framework for further characterization of the DUB roles in the retina and suggest new candidates for inherited retinal disorders. PMID:26934049
Lieberwirth, Claudia; Wang, Zuoxin
Adult neurogenesis – the formation of new neurons in adulthood – has been shown to be modulated by a variety of endogenous (e.g., trophic factors, neurotransmitters, and hormones) as well as exogenous (e.g., physical activity and environmental complexity) factors. Research on exogenous regulators of adult neurogenesis has focused primarily on the non-social environment. More recently, however, evidence has emerged suggesting that the social environment can also affect adult neurogenesis. The present review details the effects of adult–adult (e.g., mating and chemosensory interactions) and adult–offspring (e.g., gestation, parenthood, and exposure to offspring) interactions on adult neurogenesis. In addition, the effects of a stressful social environment (e.g., lack of social support and dominant–subordinate interactions) on adult neurogenesis are reviewed. The underlying hormonal mechanisms and potential functional significance of adult-generated neurons in mediating social behaviors are also discussed. PMID:22586385
Salinas-Navarro, Manuel; Alarcón-Martínez, Luis; Valiente-Soriano, Francisco Javier; Ortín-Martínez, Arturo; Jiménez-López, Manuel; Avilés-Trigueros, Marcelino; Villegas-Pérez, María Paz; de la Villa, Pedro
Purpose To investigate the effects of laser photocoagulation (LP)-induced ocular hypertension (OHT) on the survival and retrograde axonal transport of retinal ganglion cells (RGC), as well as on the function of retinal layers. Methods Adult albino Swiss mice (35–45 g) received laser photocoagulation of limbal and episcleral veins in the left eye. Mice were sacrificed at 8, 17, 35, and 63 days. Intraocular pressure (IOP) in both eyes was measured with a Tono-Lab before LP and at various days after LP. Flash electroretinogram (ERG) scotopic threshold response (STR) and a- and b-wave amplitudes were recorded before LP and at various times after LP. RGCs were labeled with 10% hydroxystilbamidine methanesulfonate (OHSt) applied to both superior colliculi before sacrifice and in some mice, with dextran tetramethylrhodamine (DTMR) applied to the ocular stump of the intraorbitally transected optic nerve. Retinas were immunostained for RT97 or Brn3a. Retinas were prepared as whole-mounts and photographed under a fluorescence microscope. Labeled RGCs were counted using image analysis software, and an isodensity contour plot was generated for each retina. Results IOP increased to twice its basal values by 24 h and was maintained until day 5, after which IOP gradually declined to reach basal values by 1 wk. Similar IOP increases were observed in all groups. The mean total number of OHSt+ RGCs was 13,428±6,295 (n=12), 10,456±14,301 (n=13), 12,622±14,174 (n=21), and 10,451±13,949 (n=13) for groups I, II, III, and IV, respectively; these values represented 28%, 23%, 26%, and 22% of the values found in their contralateral fellow retinas. The mean total population of Brn3a+ RGCs was 24,343±5,739 (n=12) and 10,219±8,887 (n=9), respectively, for groups I and III; these values represented 49% and 20%, respectively, of the values found in their fellow eyes. OHT retinas showed an absence of OHSt+ and DTMR+ RGCs in both focal wedge-shaped and diffuse regions of the retina. By 1
Gonzalez-Cordero, Anai; West, Emma L; Pearson, Rachael A; Duran, Yanai; Carvalho, Livia S; Chu, Colin J; Naeem, Arifa; Blackford, Samuel J I; Georgiadis, Anastasios; Lakowski, Jorn; Hubank, Mike; Smith, Alexander J; Bainbridge, James W B; Sowden, Jane C; Ali, Robin R
Irreversible blindness caused by loss of photoreceptors may be amenable to cell therapy. We previously demonstrated retinal repair and restoration of vision through transplantation of photoreceptor precursors obtained from postnatal retinas into visually impaired adult mice. Considerable progress has been made in differentiating embryonic stem cells (ESCs) in vitro toward photoreceptor lineages. However, the capability of ESC-derived photoreceptors to integrate after transplantation has not been demonstrated unequivocally. Here, to isolate photoreceptor precursors fit for transplantation, we adapted a recently reported three-dimensional (3D) differentiation protocol that generates neuroretina from mouse ESCs. We show that rod precursors derived by this protocol and selected via a GFP reporter under the control of a Rhodopsin promoter integrate within degenerate retinas of adult mice and mature into outer segment-bearing photoreceptors. Notably, ESC-derived precursors at a developmental stage similar to postnatal days 4-8 integrate more efficiently compared with cells at other stages. This study shows conclusively that ESCs can provide a source of photoreceptors for retinal cell transplantation.
Machalińska, Anna; Zuba-Surma, Ewa K
The latest research reports revealed the presence of stem/progenitor cells located in different regions of matured eye. They are able to differentiate into retinal pigment epithelium cells as well as neural structure of retina. These cells were identified in neurosensory retina, pigment epithelium and within cilliary body and iris epithelium. Moreover, it has been proved that Muller glia possess the potential of differentiation into retinal cells. These findings indicate the presence of potential mechanisms enabling retinal cell repopulation and retinal tissue regeneration. In the present work, the recent reports documenting the presence of different stem cell populations in eye have been reviewed, particularly focusing on recently identified very small embryonic-like stem cells (VSEL-SCs). The potential clinical applications of the residing stem cells and limitations of such therapeutic strategies have been also discussed.
Lahne, Manuela; Li, Jingling; Marton, Rebecca M.
Loss of retinal neurons in adult zebrafish (Danio rerio) induces a robust regenerative response mediated by the reentry of the resident Müller glia into the cell cycle. Upon initiating Müller glia proliferation, their nuclei migrate along the apicobasal axis of the retina in phase with the cell cycle in a process termed interkinetic nuclear migration (INM). We examined the mechanisms governing this cellular process and explored its function in regenerating the adult zebrafish retina. Live-cell imaging revealed that the majority of Müller glia nuclei migrated to the outer nuclear layer (ONL) to divide. These Müller glia formed prominent actin filaments at the rear of nuclei that had migrated to the ONL. Inhibiting actin filament formation or Rho-associated coiled-coil kinase (Rock) activity, which is necessary for phosphorylation of myosin light chain and actin myosin-mediated contraction, disrupted INM with increased numbers of mitotic nuclei remaining in the basal inner nuclear layer, the region where Müller glia typically reside. Double knockdown of Rho-associated coiled-coil kinase 2a (Rock2a) and Rho-associated coiled-coil kinase 2b (Rock2b) similarly disrupted INM and reduced Müller glial cell cycle reentry. In contrast, Rock inhibition immediately before the onset of INM did not affect Müller glia proliferation, but subsequently reduced neuronal progenitor cell proliferation due to early cell cycle exit. Long-term, Rock inhibition increased the generation of mislocalized ganglion/amacrine cells at the expense of rod and cone photoreceptors. In summary, INM is driven by an actin-myosin-mediated process controlled by Rock2a and Rock2b activity, which is required for sufficient proliferation and regeneration of photoreceptors after light damage. SIGNIFICANCE STATEMENT The human retina does not replace lost or damaged neurons, ultimately causing vision impairment. In contrast, zebrafish are capable of regenerating lost neurons. Understanding the mechanisms
Vyas, Alka; Li, Zhaobo; Aspalter, Manuela; Feiner, Jeffrey; Hoke, Ahmet; Zhou, Chunhua; O'Daly, Andres; Abdullah, Madeel; Rohde, Charles; Brushart, Thomas M
The role of pathway-derived growth factors in the support of peripheral axon regeneration remains elusive. Few appropriate knock-out mice are available, and gene silencing techniques are rarely 100% effective. To overcome these difficulties, we have developed an in vitro organotypic co-culture system that accurately models peripheral nerve repair in the adult mammal. Spinal cord sections from P4 mice that express YFP in their neurons are used to innervate segments of P4 peripheral nerve. This reconstructed ventral root is then transected and joined to a nerve graft. Growth of axons across the nerve repair and into the graft can be imaged repeatedly with fluorescence microscopy to define regeneration speed, and parent neurons can be labeled in retrograde fashion to identify contributing neurons. Nerve graft harvested from adult mice remains viable in culture by both morphologic and functional criteria. Motoneurons are supported with GDNF for the first week in culture, after which they survive axotomy, and are thus functionally adult. This platform can be modified by using motoneurons from any genetically modified mouse that can be bred to express XFP, by harvesting nerve graft from any source, or by treating the culture systemically with antibodies, growth factors, or pathway inhibitors. The regeneration environment is controlled to a degree not possible in vivo, and the use of experimental animals is reduced substantially. The flexibility and control offered by this technique should thus make it a useful tool for the study of regeneration biology.
Sun, Gerald J.; Zhou, Yi; Stadel, Ryan P.; Moss, Jonathan; Yong, Jing Hui A.; Ito, Shiori; Kawasaki, Nicholas K.; Phan, Alexander T.; Oh, Justin H.; Modak, Nikhil; Reed, Randall R.; Toni, Nicolas; Song, Hongjun; Ming, Guo-li
In a classic model of mammalian brain formation, precursors of principal glutamatergic neurons migrate radially along radial glia fibers whereas GABAergic interneuron precursors migrate tangentially. These migration modes have significant implications for brain function. Here we used clonal lineage tracing of active radial glia-like neural stem cells in the adult mouse dentate gyrus and made the surprising discovery that proliferating neuronal precursors of glutamatergic granule neurons exhibit significant tangential migration along blood vessels, followed by limited radial migration. Genetic birthdating and morphological and molecular analyses pinpointed the neuroblast stage as the main developmental window when tangential migration occurs. We also developed a partial “whole-mount” dentate gyrus preparation and observed a dense plexus of capillaries, with which only neuroblasts, among the entire population of progenitors, are directly associated. Together, these results provide insight into neuronal migration in the adult mammalian nervous system. PMID:26170290
Lawrence, Jean M; Singhal, Shweta; Bhatia, Bhairavi; Keegan, David J; Reh, Thomas A; Luthert, Philip J; Khaw, Peng T; Limb, Gloria Astrid
Growing evidence suggests that glial cells may have a role as neural precursors in the adult central nervous system. Although it has been shown that Müller cells exhibit progenitor characteristics in the postnatal chick and rat retinae, their progenitor-like role in developed human retina is unknown. We first reported the Müller glial characteristics of the spontaneously immortalized human cell line MIO-M1, but recently we have derived similar cell lines from the neural retina of several adult eye donors. Since immortalization is one of the main properties of stem cells, we investigated whether these cells expressed stem cell markers. Cells were grown as adherent monolayers, responded to epidermal growth factor, and could be expanded indefinitely without growth factors under normal culture conditions. They could be frozen and thawed without losing their characteristics. In the presence of extracellular matrix and fibroblast growth factor-2 or retinoic acid, they acquired neural morphology, formed neurospheres, and expressed neural stem cell markers including betaIII tubulin, Sox2, Pax6, Chx10, and Notch 1. They also expressed markers of postmitotic retinal neurons, including peripherin, recoverin, calretinin, S-opsin, and Brn3. When grafted into the subretinal space of dystrophic Royal College of Surgeons rats or neonatal Lister hooded rats, immortalized cells migrated into the retina, where they expressed various markers of retinal neurons. These observations indicate that adult human neural retina harbors a population of cells that express both Müller glial and stem cell markers and suggest that these cells may have potential use for cell-based therapies to restore retinal function. Disclosure of potential conflicts of interest is found at the end of this article.
Bhatia, Bhairavi; Singhal, Shweta; Lawrence, Jean M; Khaw, Peng T; Limb, G Astrid
Much interest has been generated by the identification of neural stem cells in the human neural retina and ciliary body. However, it is not clear whether stem cells identified in these ocular compartments are of the same origin or whether they ontogenically derive from different cell populations. This study examined the in situ anatomical distribution of these cells within the neural retina and ciliary body, as well as their ability to proliferate in response to EGF. Human retinae and ciliary body were examined for co-expression of Nestin, cellular retinaldehyde binding (CRALBP) or Vimentin, and the stem cell markers SOX2, CHX10, NOTCH1 and SHH. Retinal explants were cultured with epidermal growth factor (EGF) to assess retinal cell proliferation. Intense Nestin and CRALBP staining was observed in the neural retinal margin, where cells formed bundles of spindle cells (resembling glial cells) that lacked lamination and co-stained for SOX2, CHX10 and SHH. This staining differentiated the neural retina from the ciliary epithelium, which expressed SOX2, CHX10 and NOTCH1 but not Nestin or CRALBP. Nestin and CRALBP expression decreased towards the posterior retina, where it anatomically identified a population of Müller glia. All Vimentin positive Müller glia co-stained for SOX2, but only few Vimentin positive cells expressed Nestin and SOX2. Cells of the retinal margin and the inner nuclear layer (INL), where the soma of Müller glia predominate, re-entered the cell cycle upon retinal explant culture with EGF. Lack of lamination and abundance of Müller glia expressing stem cell markers in the marginal region of the adult human retina resemble the ciliary marginal zone (CMZ) of fish and amphibians. The findings that cells in this CM-like zone, as well in the inner nuclear layer proliferate in response to EGF suggest that the adult human retina has regenerative potential. Identification of factors that may promote retinal regeneration in the adult human eye would
Brus, Maïna; Keller, Matthieu; Lévy, Frédéric
Production of new neurons continues throughout life in most invertebrates and vertebrates like crustaceans, fishes, reptiles, birds, and mammals including humans. Most studies have been carried out on rodent models and demonstrated that adult neurogenesis is located mainly in two structures, the dentate gyrus (DG) of the hippocampus and the sub-ventricular zone (SVZ). If adult neurogenesis is well preserved throughout evolution, yet there are however some features which differ between species. The present review proposes to target similarities and differences in the mechanism of mammalian adult neurogenesis by comparing selected species including humans. We will highlight the cellular composition and morphological organization of the SVZ in primates which differs from that of rodents and may be of functional relevance. We will particularly focus on the dynamic of neuronal maturation in rodents, primates, and humans but also in sheep which appears to be an interesting model due to its similarities with the primate brain. PMID:23935563
Chojnacka, Katarzyna; Zarzycka, Marta; Mruk, Dolores D
A healthy man typically produces between 50 × 10(6) and 200 × 10(6) spermatozoa per day by spermatogenesis; in the absence of Sertoli cells in the male gonad, this individual would be infertile. In the adult testis, Sertoli cells are sustentacular cells that support germ cell development by secreting proteins and other important biomolecules that are essential for germ cell survival and maturation, establishing the blood-testis barrier, and facilitating spermatozoa detachment at spermiation. In the fetal testis, on the other hand, pre-Sertoli cells form the testis cords, the future seminiferous tubules. However, the role of pre-Sertoli cells in this process is much less clear than the function of Sertoli cells in the adult testis. Within this framework, we provide an overview of the biology of the fetal, pubertal, and adult Sertoli cell, highlighting relevant cell biology studies that have expanded our understanding of mammalian spermatogenesis.
Weitz, Andrew C; Behrend, Matthew R; Lee, Nan Sook; Klein, Ronald L; Chiodo, Vince A; Hauswirth, William W; Humayun, Mark S; Weiland, James D; Chow, Robert H
Epiretinal implants for the blind are designed to stimulate surviving retinal neurons, thus bypassing the diseased photoreceptor layer. Single-unit or multielectrode recordings from isolated animal retina are commonly used to inform the design of these implants. However, such electrical recordings provide limited information about the spatial patterns of retinal activation. Calcium imaging overcomes this limitation, as imaging enables high spatial resolution mapping of retinal ganglion cell (RGC) activity as well as simultaneous recording from hundreds of RGCs. Prior experiments in amphibian retina have demonstrated proof of principle, yet experiments in mammalian retina have been hindered by the inability to load calcium indicators into mature mammalian RGCs. Here, we report a method for labeling the majority of ganglion cells in adult rat retina with genetically encoded calcium indicators, specifically GCaMP3 and GCaMP5G. Intravitreal injection of an adeno-associated viral vector targets ∼85% of ganglion cells with high specificity. Because of the large fluorescence signals provided by the GCaMP sensors, we can now for the first time visualize the response of the retina to electrical stimulation in real-time. Imaging transduced retinas mounted on multielectrode arrays reveals how stimulus pulse shape can dramatically affect the spatial extent of RGC activation, which has clear implications in prosthetic applications. Our method can be easily adapted to work with other fluorescent indicator proteins in both wild-type and transgenic mammals.
Ryu, Jae Ryun; Hong, Caroline Jeeyeon; Kim, Joo Yeon; Kim, Eun-Kyoung; Sun, Woong; Yu, Seong-Woon
The presence of neural stem cells (NSCs) and the production of new neurons in the adult brain have received great attention from scientists and the public because of implications to brain plasticity and their potential use for treating currently incurable brain diseases. Adult neurogenesis is controlled at multiple levels, including proliferation, differentiation, migration, and programmed cell death (PCD). Among these, PCD is the last and most prominent process for regulating the final number of mature neurons integrated into neural circuits. PCD can be classified into apoptosis, necrosis, and autophagic cell death and emerging evidence suggests that all three may be important modes of cell death in neural stem/progenitor cells. However, the molecular mechanisms that regulate PCD and thereby impact the intricate balance between self-renewal, proliferation, and differentiation during adult neurogenesis are not well understood. In this comprehensive review, we focus on the extent, mechanism, and biological significance of PCD for the control of adult neurogenesis in the mammalian brain. The role of intrinsic and extrinsic factors in the regulation of PCD at the molecular and systems levels is also discussed. Adult neurogenesis is a dynamic process, and the signals for differentiation, proliferation, and death of neural progenitor/stem cells are closely interrelated. A better understanding of how adult neurogenesis is influenced by PCD will help lead to important insights relevant to brain health and diseases.
Contartese, Daniela S.; Rolón, Federico; Sarotto, Anibal; Dorfman, Veronica B.; Loidl, Cesar F.; Martínez, Alfredo
Hypothermia has been proposed as a therapeutic intervention for some retinal conditions, including ischemic insults. Cold exposure elevates expression of cold-shock proteins (CSP), including RNA-binding motif protein 3 (RBM3) and cold inducible RNA-binding protein (CIRP), but their presence in mammalian retina is so far unknown. Here we show the effects of hypothermia on the expression of these CSPs in retina-derived cell lines and in the retina of newborn and adult rats. Two cell lines of retinal origin, R28 and mRPE, were exposed to 32°C for different time periods and CSP expression was measured by qRT-PCR and Western blotting. Neonatal and adult Sprague-Dawley rats were exposed to a cold environment (8°C) and expression of CSPs in their retinas was studied by Western blotting, multiple inmunofluorescence, and confocal microscopy. RBM3 expression was upregulated by cold in both R28 and mRPE cells in a time-dependent fashion. On the other hand, CIRP was upregulated in R28 cells but not in mRPE. In vivo, expression of CSPs was negligible in the retina of newborn and adult rats kept at room temperature (24°C). Exposure to a cold environment elicited a strong expression of both proteins, especially in retinal pigment epithelium cells, photoreceptors, bipolar, amacrine and horizontal cells, Müller cells, and ganglion cells. In conclusion, CSP expression rapidly rises in the mammalian retina following exposure to hypothermia in a cell type-specific pattern. This observation may be at the basis of the molecular mechanism by which hypothermia exerts its therapeutic effects in the retina. PMID:27556928
Larrayoz, Ignacio M; Rey-Funes, Manuel; Contartese, Daniela S; Rolón, Federico; Sarotto, Anibal; Dorfman, Veronica B; Loidl, Cesar F; Martínez, Alfredo
Hypothermia has been proposed as a therapeutic intervention for some retinal conditions, including ischemic insults. Cold exposure elevates expression of cold-shock proteins (CSP), including RNA-binding motif protein 3 (RBM3) and cold inducible RNA-binding protein (CIRP), but their presence in mammalian retina is so far unknown. Here we show the effects of hypothermia on the expression of these CSPs in retina-derived cell lines and in the retina of newborn and adult rats. Two cell lines of retinal origin, R28 and mRPE, were exposed to 32°C for different time periods and CSP expression was measured by qRT-PCR and Western blotting. Neonatal and adult Sprague-Dawley rats were exposed to a cold environment (8°C) and expression of CSPs in their retinas was studied by Western blotting, multiple inmunofluorescence, and confocal microscopy. RBM3 expression was upregulated by cold in both R28 and mRPE cells in a time-dependent fashion. On the other hand, CIRP was upregulated in R28 cells but not in mRPE. In vivo, expression of CSPs was negligible in the retina of newborn and adult rats kept at room temperature (24°C). Exposure to a cold environment elicited a strong expression of both proteins, especially in retinal pigment epithelium cells, photoreceptors, bipolar, amacrine and horizontal cells, Müller cells, and ganglion cells. In conclusion, CSP expression rapidly rises in the mammalian retina following exposure to hypothermia in a cell type-specific pattern. This observation may be at the basis of the molecular mechanism by which hypothermia exerts its therapeutic effects in the retina.
Valenzuela, D M; Economides, A N; Rojas, E; Lamb, T M; Nuñez, L; Jones, P; Lp, N Y; Espinosa, R; Brannan, C I; Gilbert, D J
The multiple roles of noggin during dorsal fate specification in Xenopus embryos, together with noggin's ability to directly induce neural tissue, inspired an effort to determine whether a similar molecule exists in mammals. Here we describe the identification of human and rat noggin and explore their expression patterns; we also localize the human NOGGIN gene to chromosome 17q22, and the mouse gene to a syntenic region of chromosome 11. Mammalian noggin is remarkably similar in its sequence to Xenopus noggin, and is similarly active in induction assays performed on Xenopus embryo tissues. In the adult mammal, noggin is most notably expressed in particular regions of the nervous system, such as the tufted cells of the olfactory bulb, the piriform cortex of the brain, and the Purkinje cells of the cerebellum, suggesting that one of the earliest acting neural inducers also has important roles in the adult nervous system.
Kainate receptors mediate glutamatergic signaling through both pre- and presynaptic receptors. Here, we studied the expression of the high affinity kainate receptor GluK5 in the mouse retina. Double-immunofluoresence labeling and electron microscopic analysis revealed a presynaptic localization of GluK5 in the outer plexiform layer. Unexpectedly, we found GluK5 almost exclusively localized to the presynaptic ribbon of photoreceptor terminals. Moreover, in GluK5-deficient mutant mice the structural integrity of synaptic ribbons was severely altered pointing to a novel function of GluK5 in organizing synaptic ribbons in the presynaptic terminals of rod photoreceptors. PMID:28235022
Porrello, Enzo R.; Mahmoud, Ahmed I.; Simpson, Emma; Johnson, Brett A.; Grinsfelder, David; Canseco, Diana; Mammen, Pradeep P.; Rothermel, Beverly A.; Olson, Eric N.; Sadek, Hesham A.
We recently identified a brief time period during postnatal development when the mammalian heart retains significant regenerative potential after amputation of the ventricular apex. However, one major unresolved question is whether the neonatal mouse heart can also regenerate in response to myocardial ischemia, the most common antecedent of heart failure in humans. Here, we induced ischemic myocardial infarction (MI) in 1-d-old mice and found that this results in extensive myocardial necrosis and systolic dysfunction. Remarkably, the neonatal heart mounted a robust regenerative response, through proliferation of preexisting cardiomyocytes, resulting in full functional recovery within 21 d. Moreover, we show that the miR-15 family of microRNAs modulates neonatal heart regeneration through inhibition of postnatal cardiomyocyte proliferation. Finally, we demonstrate that inhibition of the miR-15 family from an early postnatal age until adulthood increases myocyte proliferation in the adult heart and improves left ventricular systolic function after adult MI. We conclude that the neonatal mammalian heart can regenerate after myocardial infarction through proliferation of preexisting cardiomyocytes and that the miR-15 family contributes to postnatal loss of cardiac regenerative capacity. PMID:23248315
Sattler, Susanne; Rosenthal, Nadia
The immune system is a crucial player in tissue homeostasis and wound healing. A sophisticated cascade of events triggered upon injury ensures protection from infection and initiates and orchestrates healing. While the neonatal mammal can readily regenerate damaged tissues, adult regenerative capacity is limited to specific tissue types, and in organs such as the heart, adult wound healing results in fibrotic repair and loss of function. Growing evidence suggests that the immune system greatly influences the balance between regeneration and fibrotic repair. The neonate mammalian immune system has impaired pro-inflammatory function, is prone to T-helper type 2 responses and has an immature adaptive immune system skewed towards regulatory T cells. While these characteristics make infants susceptible to infection and prone to allergies, it may also provide an immunological environment permissive of regeneration. In this review we will give a comprehensive overview of the immune cells involved in healing and regeneration of the heart and explore differences between the adult and neonate immune system that may explain differences in regenerative ability. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.
Guinamard, Romain; Hof, Thomas; Sallé, Laurent
This chapter describes appropriate methods to investigate mammalian cardiac channels properties at the single channel level. Cell isolation is performed from new born or adult heart by enzymatic digestion on minced tissue or using the Langendorff apparatus. Isolation proceeding is suitable for rabbit, rat, and mouse hearts. In addition, isolation of human atrial cardiomyocytes is described. Such freshly isolated cells or cells maintained in primary culture are suitable for patch-clamp studies. Here we describe the single channel variants of the patch-clamp technique (cell-attached, inside-out, outside-out) used to investigate channel properties. Proceedings for the evaluation of biophysical properties such as conductance, ionic selectivity, regulations by extracellular and intracellular mechanisms are described. To illustrate the study, we provide an example by the characterization of a calcium-activated non-selective cation channel (TRPM4).
Proliferation of adult mammalian ventricular cardiomyocytes has been ruled out by some researchers, who have argued that these cells are terminally differentiated; however, this dogma has been rejected because other researchers have reported that these cells can present the processes necessary to proliferate, that is, DNA synthesis, mitosis and cytokinesis when the heart is damaged experimentally through pharmacological and surgical strategies or due to pathological conditions concerning the cardiovascular system. This review integrates some of the available works in the literature evaluating the DNA synthesis, mitosis and cytokinesis in these myocytes, when the myocardium is damaged, with the purpose of knowing if their proliferation can be considered as a feasible phenomenon. The review is concluded with a reflection about the perspectives of the knowledge generated in this area.
Eastlake, Karen; Heywood, Wendy E.; Tracey-White, Dhani; Aquino, Erika; Bliss, Emily; Vasta, Gerardo R.; Mills, Kevin; Khaw, Peng T.; Moosajee, Mariya; Limb, G. Astrid
Zebrafish spontaneously regenerate the retina after injury. Although the gene expression profile has been extensively studied in this species during regeneration, this does not reflect protein function. To further understand the regenerative process in the zebrafish, we compared the proteomic profile of the retina during injury and upon regeneration. Using two-dimensional difference gel electrophoresis (2D-DIGE) and label-free quantitative proteomics (quadrupole time of flight LC-MS/MS), we analysed the retina of adult longfin wildtype zebrafish at 0, 3 and 18 days after Ouabain injection. Gene ontology analysis indicates reduced metabolic processing, and increase in fibrin clot formation, with significant upregulation of fibrinogen gamma polypeptide, apolipoproteins A-Ib and A-II, galectin-1, and vitellogenin-6 during degeneration when compared to normal retina. In addition, cytoskeleton and membrane transport proteins were considerably altered during regeneration, with the highest fold upregulation observed for tubulin beta 2 A, histone H2B and brain type fatty acid binding protein. Key proteins identified in this study may play an important role in the regeneration of the zebrafish retina and investigations on the potential regulation of these proteins may lead to the design of protocols to promote endogenous regeneration of the mammalian retina following retinal degenerative disease. PMID:28300160
Zhang, Kuan; Chen, Chunhai; Yang, Zhiqi; He, Wenjing; Liao, Xiang; Ma, Qinlong; Deng, Ping; Lu, Jian; Li, Jingcheng; Wang, Meng; Li, Mingli; Zheng, Lianghong; Zhou, Zhuan; Sun, Wei; Wang, Liting; Jia, Hongbo; Yu, Zhengping; Zhou, Zhou; Chen, Xiaowei
Glial precursor transplantation provides a potential therapy for brain disorders. Before its clinical application, experimental evidence needs to indicate that engrafted glial cells are functionally incorporated into the existing circuits and become essential partners of neurons for executing fundamental brain functions. While previous experiments supporting for their functional integration have been obtained under in vitro conditions using slice preparations, in vivo evidence for such integration is still lacking. Here, we utilized in vivo two-photon Ca2+ imaging along with immunohistochemistry, fluorescent indicator labeling-based axon tracing and correlated light/electron microscopy to analyze the profiles and the functional status of glial precursor cell-derived astrocytes in adult mouse neocortex. We show that after being transplanted into somatosensory cortex, precursor-derived astrocytes are able to survive for more than a year and respond with Ca2+ signals to sensory stimulation. These sensory-evoked responses are mediated by functionally-expressed nicotinic receptors and newly-established synaptic contacts with the host cholinergic afferents. Our results provide in vivo evidence for a functional integration of transplanted astrocytes into adult mammalian neocortex, representing a proof-of-principle for sensory cortex remodeling through addition of essential neural elements. Moreover, we provide strong support for the use of glial precursor transplantation to understand glia-related neural development in vivo. PMID:27405333
Vergara, M Natalia; Smiley, Laura K; Del Rio-Tsonis, Katia; Tsonis, Panagiotis A
Adult newts are able to regenerate their retina and lens after injury or complete removal through transdifferentiation of the pigmented epithelial tissues of the eye. This process needs to be tightly controlled, and several different mechanisms are likely to be recruited for this function. The Na(+)/K(+) ATPase is a transmembrane protein that establishes electrochemical gradients through the transport of Na(+) and K(+) and has been implicated in the modulation of key cellular processes such as cell division, migration and adhesion. Even though it is expressed in all cells, its isoform composition varies with cell type and is tightly controlled during development and regeneration. In the present study we characterize the expression pattern of Na(+)/K(+) ATPase alpha1 in the adult newt eye and during the process of lens and retina regeneration. We show that this isoform is up-regulated in undifferentiated cells during transdifferentiation. Such change in composition could be one of the mechanisms that newt cells utilize to modulate this process.
Schwalbe, Ruth A; Corey, Melissa J; Cartwright, Tara A
The N-glycan pool of mammalian brain contains remarkably high levels of sialylated N-glycans. This study provides the first evidence that voltage-gated K+ channels Kv3.1, Kv3.3, and Kv3.4, possess distinct sialylated N-glycan structures throughout the central nervous system of the adult rat. Electrophoretic migration patterns of Kv3.1, Kv3.3, and Kv3.4 glycoproteins from spinal cord, hypothalamus, thalamus, cerebral cortex, hippocampus, and cerebellum membranes digested with glycosidases were used to identify the various glycoforms. Differences in the migration of Kv3 proteins were attributed to the desialylated N-glycans. Expression levels of the Kv3 proteins were highest in cerebellum, whereas those of Kv3.1 and Kv3.3 were much lower in the other 5 regions. The lowest level of Kv3.1 was expressed in the hypothalamus, whereas the lowest levels of Kv3.3 were expressed in both thalamus and hypothalamus. The other regions expressed intermediate levels of Kv3.3, with spinal cord expressing the highest. The expression level of Kv3.4 in the hippocampus was slightly lower than that in cerebellum, and was closely followed by the other 4 regions, with spinal cord expressing the lowest level. We suggest that novel Kv3 glycoforms may endow differences in channel function and expression among regions throughout the central nervous system.
Mayazur Rahman, S; Reichenbach, Andreas; Zink, Mareike; Mayr, Stefan G
Development of neuronal tissue, such as folding of the brain, and formation of the fovea centralis in the human retina are intimately connected with the mechanical properties of the underlying cells and the extracellular matrix. In particular for neuronal tissue as complex as the vertebrate retina, mechanical properties are still a matter of debate due to their relation to numerous diseases as well as surgery, where the tension of the retina can result in tissue detachment during cutting. However, measuring the elasticity of adult retina wholemounts is difficult and until now only the mechanical properties at the surface have been characterized with micrometer resolution. Many processes, however, such as pathological changes prone to cause tissue rupture and detachment, respectively, are reflected in variations of retina elasticity at smaller length scales at the protein level. In the present work we demonstrate that freely oscillating cantilevers composed of nanostructured TiO2 scaffolds can be employed to study the frequency-dependent mechanical response of adult mammalian retina explants at the nanoscale. Constituting highly versatile scaffolds with strong tissue attachment for long-term organotypic culture atop, these scaffolds perform damped vibrations as fingerprints of the mechanical tissue properties that are derived using finite element calculations. Since the tissue adheres to the nanostructures via constitutive proteins on the photoreceptor side of the retina, the latter are stretched and compressed during vibration of the underlying scaffold. Probing mechanical response of individual proteins within the tissue, the proposed mechanical spectroscopy approach opens the way for studying tissue mechanics, diseases and the effect of drugs at the protein level.
Krishna, Sanjay [Albuquerque, NM; Hayat, Majeed M [Albuquerque, NM; Tyo, J Scott [Tucson, AZ; Jang, Woo-Yong [Albuquerque, NM
Exemplary embodiments provide an infrared (IR) retinal system and method for making and using the IR retinal system. The IR retinal system can include adaptive sensor elements, whose properties including, e.g., spectral response, signal-to-noise ratio, polarization, or amplitude can be tailored at pixel level by changing the applied bias voltage across the detector. "Color" imagery can be obtained from the IR retinal system by using a single focal plane array. The IR sensor elements can be spectrally, spatially and temporally adaptive using quantum-confined transitions in nanoscale quantum dots. The IR sensor elements can be used as building blocks of an infrared retina, similar to cones of human retina, and can be designed to work in the long-wave infrared portion of the electromagnetic spectrum ranging from about 8 .mu.m to about 12 .mu.m as well as the mid-wave portion ranging from about 3 .mu.m to about 5 .mu.m.
Balenci, Laurent; van der Kooy, Derek
Understanding the mechanisms regulating retinal stem cell (RSC) activity is fundamental for future stem cell-based therapeutic purposes. By combining gain and loss of function approaches, we addressed whether Notch signaling may play a selective role in retinal stem versus retinal progenitor cells in both developing and adult eyes. Inhibition of either Notch or fibroblast growth factor signaling reduced proliferation of retinal stem and retinal progenitor cells, and inhibited RSC self-renewal. Conversely, exogenous Delta-like 3 and direct intrinsic Notch activation stimulated expansionary symmetric divisions in adult RSCs with the concomitant upregulation of Hes5. Knocking down Hes5 expression specifically decreased the numbers, but not the diameters, of adult RSC primary spheres, indicating that HES5 is the downstream effector of Notch receptor in controlling adult RSC proliferation. In addition, constitutive Notch activation induced retinal stem-like asymmetric self-renewal properties, with no expansion (no symmetrical division) in perinatal neural retina progenitor cells. These findings highlight central roles of Notch signaling activity in regulating the modes of division of retinal stem and retinal progenitor cells.
Melicharek, David; Shah, Arpit; DiStefano, Ginnene; Gangemi, Andrew J.; Orapallo, Andrew; Vrailas-Mortimer, Alysia D.; Marenda, Daniel R.
Atonal is a Drosophila proneural protein required for the proper formation of the R8 photoreceptor cell, the founding photoreceptor cell in the developing retina. Proper expression and refinement of the Atonal protein is essential for the proper formation of the Drosophila adult eye. In vertebrates, expression of transcription factors orthologous to Drosophila Atonal (MATH5/Atoh7, XATH5, and ATH5) and their progressive restriction are also involved in specifying the retinal ganglion cell, the founding neural cell type in the mammalian retina. Thus, identifying factors that are involved in regulating the expression of Atonal during development are important to fully understand how retinal neurogenesis is accomplished. We have performed a chemical mutagenesis screen for autosomal dominant enhancers of a loss-of-function atonal eye phenotype. We report here the identification of five genes required for proper Atonal expression, three of which are novel regulators of Atonal expression in the Drosophila retina. We characterize the role of the daughterless, kismet, and roughened eye genes on atonal transcriptional regulation in the developing retina and show that each gene regulates atonal transcription differently within the context of retinal development. Our results provide additional insights into the regulation of Atonal expression in the developing Drosophila retina. PMID:18832354
Dopamine is the most abundant catecholamine in the vertebrate retina. Despite the description of retinal dopaminergic cells three decades ago, many aspects of their function in the retina remain unclear. There is no consensus among the authors about the stimulus conditions for dopamine release (darkness, steady or flickering light) as well as about its action upon the various types of retinal cells. Many contradictory results exist concerning the dopamine effect on the gross electrical activity of the retina [reflected in electroretinogram (ERG)] and the receptors involved in its action. This review summarized current knowledge about the types of the dopaminergic neurons and receptors in the retina as well as the effects of dopamine receptor agonists and antagonists on the light responses of photoreceptors, horizontal and bipolar cells in both nonmammalian and mammalian retina. Special focus of interest concerns their effects upon the diffuse ERG as a useful tool for assessment of the overall function of the distal retina. An attempt is made to reveal some differences between the dopamine actions upon the activity of the ON versus OFF channel in the distal retina. The author has included her own results demonstrating such differences.
Rezzola, Sara; Belleri, Mirella; Gariano, Giuseppina; Ribatti, Domenico; Costagliola, Ciro; Semeraro, Francesco; Presta, Marco
Pathological angiogenesis of the retina is a key component of irreversible causes of blindness, as observed in proliferative diabetic retinopathy, age-related macular degeneration, and retinopathy of prematurity. Seminal studies in the early 1980 s about the angiogenic activity exerted by mammalian retinal tissue extracts on the chick embryo chorioallantoic membrane and the later discovery of vascular endothelial growth factor (VEGF) accumulation in eyes of patients with diabetic retinopathy paved the way for the development of anti-angiogenic VEGF blockers for the treatment of retinal neovascularization. Since then, numerous preclinical and clinical studies about diabetic retinopathy and other retinal disorders have opened new lines of angiogenesis inquiry, indicating that limitations to anti-VEGF therapies may exist. Moreover, the production of growth factors other than VEGF may affect the response to anti-VEGF approaches. Thus, experimental models of retinal angiogenesis remain crucial for investigating novel anti-angiogenic therapies and bringing them to patients. To this aim, in vitro and ex vivo angiogenesis assays may be suitable for a rapid screening of potential anti-angiogenic molecules before in vivo validation of the putative lead compounds. This review focuses on the different in vitro and ex vivo angiogenesis assays that have been developed over the years based on the isolation of endothelial cells from the retina of various animal species and ex vivo cultures of neonatal and adult retina explants. Also, recent observations have shown that eye neovascularization in zebrafish (Danio rerio) embryos, an in vivo animal platform experimentally analogous to in vitro/ex vivo models, may represent a novel target for the identification of angiogenesis inhibitors. When compared to in vivo assays, in vitro and ex vivo models of retina neovascularization, including zebrafish embryo, may represent cost-effective and rapid tools for the screening of novel anti
Li, Chunyi; Yang, Fuhe; Sheppard, Allan
Mammalian organ regeneration is the "Holy Grail" of modern regenerative biology and medicine. The most dramatic organ replacement is known as epimorphic regeneration. To date our knowledge of epimorphic regeneration has come from studies of amphibians. Notably, these animals have the ability to reprogram phenotypically committed cells at the amputation plane toward an embryonic-like cell phenotype (dedifferentiation). The capability of mammals to initiate analogous regeneration, and whether similar mechanisms would be involved if it were to occur, remain unclear. Deer antlers are the only mammalian appendages capable of full renewal, and therefore offer a unique opportunity to explore how nature has solved the problem of mammalian epimorphic regeneration. Following casting of old hard antlers, new antlers regenerate from permanent bony protuberances, known as pedicles. Studies through morphological and histological examinations, tissue deletion and transplantation, and cellular and molecular techniques have demonstrated that antler renewal is markedly different from that of amphibian limb regeneration (dedifferentiation-based), being a stem cell-based epimorphic process. Antler stem cells reside in the pedicle periosteum. We envisage that epimorphic regeneration of mammalian appendages, other than antler, could be made possible by recreating comparable milieu to that which supports the elaboration of that structure from the pedicle periosteum.
Garza-Lombó, Carla; Gonsebatt, María E.
The kinase mammalian target of rapamycin (mTOR) integrates signals triggered by energy, stress, oxygen levels, and growth factors. It regulates ribosome biogenesis, mRNA translation, nutrient metabolism, and autophagy. mTOR participates in various functions of the brain, such as synaptic plasticity, adult neurogenesis, memory, and learning. mTOR is present during early neural development and participates in axon and dendrite development, neuron differentiation, and gliogenesis, among other processes. Furthermore, mTOR has been shown to modulate lifespan in multiple organisms. This protein is an important energy sensor that is present throughout our lifetime its role must be precisely described in order to develop therapeutic strategies and prevent diseases of the central nervous system. The aim of this review is to present our current understanding of the functions of mTOR in neural development, the adult brain and aging. PMID:27378854
de Curtis, M; Paré, D; Llinás, R R
The viability and general electrophysiological properties of the limbic system in the adult mammalian brain isolated and maintained in vitro by arterial perfusion are described. The isolated brain preparation combines the advantages of intact synaptic connectivity and accessibility of different areas of the encephalic mass with those of the in vitro approach, i.e., stability and control of the ionic environment. Extracellular field potential as well as intracellular recordings were performed at different levels in the limbic system of isolated adult guinea pig brains. The results demonstrate that in the piriform, entorhinal, and hippocampal cortices, the intrinsic electrical properties of individual cells as well as the spontaneous and evoked electrical activity in the neuronal ensembles they comprise, were virtually identical to those observed in vivo. The properties of the limbic system loop were determined.
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Centanin, Lázaro; Ander, Janina-J; Hoeckendorf, Burkhard; Lust, Katharina; Kellner, Tanja; Kraemer, Isabel; Urbany, Cedric; Hasel, Eva; Harris, William A; Simons, Benjamin D; Wittbrodt, Joachim
The potency of post-embryonic stem cells can only be addressed in the living organism, by labeling single cells after embryonic development and following their descendants. Recently, transplantation experiments involving permanently labeled cells revealed multipotent neural stem cells (NSCs) of embryonic origin in the medaka retina. To analyze whether NSC potency is affected by developmental progression, as reported for the mammalian brain, we developed an inducible toolkit for clonal labeling and non-invasive fate tracking. We used this toolkit to address post-embryonic stem cells in different tissues and to functionally differentiate transient progenitor cells from permanent, bona fide stem cells in the retina. Using temporally controlled clonal induction, we showed that post-embryonic retinal NSCs are exclusively multipotent and give rise to the complete spectrum of cell types in the neural retina. Intriguingly, and in contrast to any other vertebrate stem cell system described so far, long-term analysis of clones indicates a preferential mode of asymmetric cell division. Moreover, following the behavior of clones before and after external stimuli, such as injuries, shows that NSCs in the retina maintained the preference for asymmetric cell division during regenerative responses. We present a comprehensive analysis of individual post-embryonic NSCs in their physiological environment and establish the teleost retina as an ideal model for studying adult stem cell biology at single cell resolution.
Czekaj, Magdalena; Haas, Jochen; Gebhardt, Marlen; Müller-Reichert, Thomas; Humphries, Peter; Farrar, Jane; Bartsch, Udo; Ader, Marius
Cell transplantation to treat retinal degenerative diseases represents an option for the replacement of lost photoreceptor cells. In vitro expandable cells isolated from the developing mammalian retina have been suggested as a potential source for the generation of high numbers of donor photoreceptors. In this study we used standardized culture conditions based on the presence of the mitogens FGF-2 and EGF to generate high numbers of cells in vitro from the developing mouse retina. These presumptive 'retinal stem cells' ('RSCs') can be propagated as monolayer cultures over multiple passages, express markers of undifferentiated neural cells, and generate neuronal and glial cell types upon withdrawal of mitogens in vitro or following transplantation into the adult mouse retina. The proportion of neuronal differentiation can be significantly increased by stepwise removal of mitogens and inhibition of the notch signaling pathway. However, 'RSCs', by contrast to their primary counterparts in vivo, i.e. retinal progenitor cells, loose the expression of retina-specific progenitor markers like Rax and Chx10 after passaging and fail to differentiate into photoreceptors both in vitro or after intraretinal transplantation. Notably, 'RSCs' can be induced to differentiate into myelinating oligodendrocytes, a cell type not generated by primary retinal progenitor cells. Based on these findings we conclude that 'RSCs' expanded in high concentrations of FGF-2 and EGF loose their retinal identity and acquire features of in vitro expandable neural stem-like cells making them an inappropriate cell source for strategies aimed at replacing photoreceptor cells in the degenerated retina.
Pinsky, Ehud; Donchin, Opher; Segev, Ronen
Direction selective cells have been found in the retina, the first level of the visual system, in mammals and recently also in the archer fish. These cells are involved in a variety of fast neural computation processes, from the control of eye movements to the detection of prey by the archer fish. The standard model for this mechanism in mammalian retina is well understood and is based on the asymmetry of inhibitory and excitatory inputs to the retinal ganglion cells. However, it remains unclear whether the mechanism that underlies direction selectivity is similar across animal classes. This study reports a pharmacological investigation designed to elucidate the mechanism that underlies motion detection in the archer fish retina. Direction selectivity in the retina was characterized under the influence of specific channel blockers that are known to be present in the different types of neurons of the retina. The results show that the direction-selective mechanism in the archer fish retina is modified only when the inhibitory channels of GABA and Glycine are manipulated. This suggests that the mechanism of direction selectivity in the archer fish retina is fundamentally different from the mechanism of direction selectivity in the mammalian retina.
Schmeer, Christian W; Wohl, Stefanie G; Isenmann, Stefan
Visual impairment severely affects the quality of life of patients and their families and is also associated with a deep economic impact. The most common pathologies responsible for visual impairment and legally defined blindness in developed countries include age-related macular degeneration, glaucoma and diabetic retinopathy. These conditions share common pathophysiological features: dysfunction and loss of retinal neurons. To date, two main approaches are being taken to develop putative therapeutic strategies: neuroprotection and cell replacement. Cell replacement is a novel therapeutic approach to restore visual capabilities to the degenerated adult neural retina and represents an emerging field of regenerative neurotherapy. The discovery of a population of proliferative cells in the mammalian retina has raised the possibility of harnessing endogenous retinal stem cells to elicit retinal repair. Furthermore, the development of suitable protocols for the reprogramming of differentiated somatic cells to a pluripotent state further increases the therapeutic potential of stem-cell-based technologies for the treatment of major retinal diseases. Stem-cell transplantation in animal models has been most effectively used for the replacement of photoreceptors, although this therapeutic approach is also being used for inner retinal pathologies. In this review, we discuss recent advances in the development of cell-replacement approaches for the treatment of currently incurable degenerative retinal diseases.
Yeo, Jia Hao; McAllan, Bronwyn M; Fraser, Stuart T
Erythroblastic islands are multicellular clusters in which a central macrophage supports the development and maturation of red blood cell (erythroid) progenitors. These clusters play crucial roles in the pathogenesis observed in animal models of hematological disorders. The precise structure and function of erythroblastic islands is poorly understood. Here, we have combined scanning electron microscopy and immuno-gold labeling of surface proteins to develop a better understanding of the ultrastructure of these multicellular clusters. The erythroid-specific surface antigen Ter-119 and the transferrin receptor CD71 exhibited distinct patterns of protein sorting during erythroid cell maturation as detected by immuno-gold labeling. During electron microscopy analysis we observed two distinct classes of erythroblastic islands. The islands varied in size and morphology, and the number and type of erythroid cells interacting with the central macrophage. Assessment of femoral marrow isolated from a cavid rodent species (guinea pig, Cavis porcellus) and a marsupial carnivore species (fat-tailed dunnarts, Sminthopsis crassicaudata) showed that while the morphology of the central macrophage varied, two different types of erythroblastic islands were consistently identifiable. Our findings suggest that these two classes of erythroblastic islands are conserved in mammalian evolution and may play distinct roles in red blood cell production.
McCutcheon, Victoria; Park, Eugene; Liu, Elaine; Sobhebidari, Pooya; Tavakkoli, Jahan; Wen, Xiao-Yan; Baker, Andrew J
Traumatic brain injury (TBI) is a leading cause of death and morbidity in industrialized countries with considerable associated health care costs. The cost and time associated with pre-clinical development of TBI therapeutics is lengthy and expensive with a poor track record of successful translation to the clinic. The zebrafish is an emerging model organism in research with unique technical and genomic strengths in the study of disease and development. Its high degree of genetic homology and cell signaling pathways relative to mammalian species and amenability to high and medium throughput assays has potential to accelerate the rate of therapeutic drug identification. Accordingly, we developed a novel closed-head model of TBI in adult zebrafish using a targeted, pulsed, high-intensity focused ultrasound (pHIFU) to induce mechanical injury of the brain. Western blot results indicated altered microtubule and neurofilament expression as well as increased expression of cleaved caspase-3 and beta APP (β-APP; p < 0.05). We used automated behavioral tracking software to evaluate locomotor deficits 24 and 48 h post-injury. Significant behavioral impairment included decreased swim distance and velocity (p < 0.05), as well as heightened anxiety and altered group social dynamics. Responses to injury were pHIFU dose-dependent and modifiable with MK-801, MDL-28170, or temperature modulation. Together, results indicate that the zebrafish exhibits responses to injury and intervention similar to mammalian TBI pathophysiology and suggest the potential for use to rapidly evaluate therapeutic compounds with high efficiency.
Yannas, I. V.; Lee, E.; Orgill, D. P.; Skrabut, E. M.; Murphy, G. F.
Regeneration of the dermis does not occur spontaneously in the adult mammal. The epidermis is regenerated spontaneously provided there is a dermal substrate over which it can migrate. Certain highly porous, crosslinked collagen--glycosaminoglycan copolymers have induced partial morphogenesis of skin when seeded with dermal and epidermal cells and then grafted on standard, full-thickness skin wounds in the adult guinea pig. A mature epidermis and a nearly physiological dermis, which lacked hair follicles but was demonstrably different from scar, were regenerated over areas as large as 16 cm2. These chemical analogs of extracellular matrices were morphogenetically active provided that the average pore diameter ranged between 20 and 125 μ m, the resistance to degradation by collagenase exceeded a critical limit, and the density of autologous dermal and epidermal cells inoculated therein was >5 × 104 cells per cm2 of wound area. Unseeded copolymers with physical structures that were within these limits delayed the onset of wound contraction by about 10 days but did not eventually prevent it. Seeded copolymers not only delayed contraction but eventually arrested and reversed it while new skin was being regenerated. The data identify a model extracellular matrix that acts as if it were an insoluble growth factor with narrowly specified physicochemical structure, functioning as a transient basal lamina during morphogenesis of skin.
Yannas, I V; Lee, E; Orgill, D P; Skrabut, E M; Murphy, G F
Regeneration of the dermis does not occur spontaneously in the adult mammal. The epidermis is regenerated spontaneously provided there is a dermal substrate over which it can migrate. Certain highly porous, crosslinked collagen-glycosaminoglycan copolymers have induced partial morphogenesis of skin when seeded with dermal and epidermal cells and then grafted on standard, full-thickness skin wounds in the adult guinea pig. A mature epidermis and a nearly physiological dermis, which lacked hair follicles but was demonstrably different from scar, were regenerated over areas as large as 16 cm2. These chemical analogs of extracellular matrices were morphogenetically active provided that the average pore diameter ranged between 20 and 125 microns, the resistance to degradation by collagenase exceeded a critical limit, and the density of autologous dermal and epidermal cells inoculated therein was greater than 5 x 10(4) cells per cm2 of wound area. Unseeded copolymers with physical structures that were within these limits delayed the onset of wound contraction by about 10 days but did not eventually prevent it. Seeded copolymers not only delayed contraction but eventually arrested and reversed it while new skin was being regenerated. The data identify a model extracellular matrix that acts as if it were an insoluble growth factor with narrowly specified physiochemical structure, functioning as a transient basal lamina during morphogenesis of skin. Images PMID:2915988
Lubin, Farah D
Brain-derived neurotrophic factor (bdnf) is one of numerous gene products necessary for long-term memory formation and dysregulation of bdnf has been implicated in the pathogenesis of cognitive and mental disorders. Recent work indicates that epigenetic-regulatory mechanisms including the markings of histone proteins and associated DNA remain labile throughout the life-span and represent an attractive molecular process contributing to gene regulation in the brain. In this review, important information will be discussed on epigenetics as a set of newly identified dynamic transcriptional mechanisms serving to regulate gene expression changes in the adult brain with particular emphasis on bdnf transcriptional readout in learning and memory formation. This review will also highlight evidence for the role of epigenetics in aberrant bdnf gene regulation in the pathogenesis of cognitive dysfunction associated with seizure disorders, Rett syndrome, Schizophrenia, and Alzheimer's disease. Such research offers novel concepts for understanding epigenetic transcriptional mechanisms subserving adult cognition and mental health, and furthermore promises novel avenues for therapeutic approach in the clinic.
Hollyfield, J.G.; Frederick, J.M.; Rayborn, M.E.
Human retinal tissue from a newborn was examined autoradiographically for the presence of high-affinity uptake and localization of the following putative neurotransmitters: dopamine, glycine, GABA, aspartate, and glutamate. In addition, the dopamine content of this newborn retina was measured by high pressure liquid chromatography. Our study reveals that specific uptake mechanisms for /sup 3/H-glycine, /sup 3/H-dopamine, and /sup 3/H-GABA are present at birth. However, the number and distribution of cells labeled with each of these /sup 3/H-transmitters are not identical to those observed in adult human retinas. Furthermore, the amount of endogenous dopamine in the newborn retina is approximately 1/20 the adult level. Photoreceptor-specific uptake of /sup 3/H-glutamate and /sup 3/H-aspartate are not observed. These findings indicate that, while some neurotransmitter-specific properties are present at birth, significant maturation of neurotransmitter systems occurs postnatally.
Liu, Sha; Wang, Mei-Xia; Mao, Cheng-Jie; Cheng, Xiao-Yu; Wang, Chen-Tao; Huang, Jian; Zhong, Zhao-Min; Hu, Wei-Dong; Wang, Fen; Hu, Li-Fang; Wang, Han; Liu, Chun-Feng
It has been demonstrated that acid sensing ionic channels (ASICs) are present in the central and peripheral nervous system of mammals, including the retina. However, it remains unclear whether the zebrafish retina also expresses ASICs. In the present study, the expression and distribution of zasic1 were examined in the retina of zebrafish. Both zasic1 mRNA and protein expressions were detected in the adult zebrafish retina. A wide distribution of ASIC1 in zebrafish retina was confirmed using whole mount in situ hybridization and immunohistochemistry study. Acidosis-induced currents in the isolated retinal ganglion cells (RGCs) were also recorded using whole cell patch clamping. Moreover, blockade of ASICs channel significantly reduced the locomotion of larval zebrafish in response to light exposure. In sum, our data demonstrate the presence of ASIC1 and its possible functional relevance in the retina of zebrafish.
Parte, Seema; Telang, Jyoti; Daithankar, Vinita; Salvi, Vinita; Zaveri, Kusum; Hinduja, Indira
The present study was undertaken to detect, characterize, and study differentiation potential of stem cells in adult rabbit, sheep, monkey, and menopausal human ovarian surface epithelium (OSE). Two distinct populations of putative stem cells (PSCs) of variable size were detected in scraped OSE, one being smaller and other similar in size to the surrounding red blood cells in the scraped OSE. The smaller 1–3 μm very small embryonic-like PSCs were pluripotent in nature with nuclear Oct-4 and cell surface SSEA-4, whereas the bigger 4–7 μm cells with cytoplasmic localization of Oct-4 and minimal expression of SSEA-4 were possibly the tissue committed progenitor stem cells. Pluripotent gene transcripts of Oct-4, Oct-4A, Nanog, Sox-2, TERT, and Stat-3 in human and sheep OSE were detected by reverse transcriptase–polymerase chain reaction. The PSCs underwent spontaneous differentiation into oocyte-like structures, parthenote-like structures, embryoid body-like structures, cells with neuronal-like phenotype, and embryonic stem cell-like colonies, whereas the epithelial cells transformed into mesenchymal phenotype by epithelial–mesenchymal transition in 3 weeks of OSE culture. Germ cell markers like c-Kit, DAZL, GDF-9, VASA, and ZP4 were immuno-localized in oocyte-like structures. In conclusion, as opposed to the existing view of OSE being a bipotent source of oocytes and granulosa cells, mammalian ovaries harbor distinct very small embryonic-like PSCs and tissue committed progenitor stem cells population that have the potential to develop into oocyte-like structures in vitro, whereas mesenchymal fibroblasts appear to form supporting granulosa-like somatic cells. Research at the single-cell level, including complete gene expression profiling, is required to further confirm whether postnatal oogenesis is a conserved phenomenon in adult mammals. PMID:21291304
Parte, Seema; Bhartiya, Deepa; Telang, Jyoti; Daithankar, Vinita; Salvi, Vinita; Zaveri, Kusum; Hinduja, Indira
The present study was undertaken to detect, characterize, and study differentiation potential of stem cells in adult rabbit, sheep, monkey, and menopausal human ovarian surface epithelium (OSE). Two distinct populations of putative stem cells (PSCs) of variable size were detected in scraped OSE, one being smaller and other similar in size to the surrounding red blood cells in the scraped OSE. The smaller 1-3 μm very small embryonic-like PSCs were pluripotent in nature with nuclear Oct-4 and cell surface SSEA-4, whereas the bigger 4-7 μm cells with cytoplasmic localization of Oct-4 and minimal expression of SSEA-4 were possibly the tissue committed progenitor stem cells. Pluripotent gene transcripts of Oct-4, Oct-4A, Nanog, Sox-2, TERT, and Stat-3 in human and sheep OSE were detected by reverse transcriptase-polymerase chain reaction. The PSCs underwent spontaneous differentiation into oocyte-like structures, parthenote-like structures, embryoid body-like structures, cells with neuronal-like phenotype, and embryonic stem cell-like colonies, whereas the epithelial cells transformed into mesenchymal phenotype by epithelial-mesenchymal transition in 3 weeks of OSE culture. Germ cell markers like c-Kit, DAZL, GDF-9, VASA, and ZP4 were immuno-localized in oocyte-like structures. In conclusion, as opposed to the existing view of OSE being a bipotent source of oocytes and granulosa cells, mammalian ovaries harbor distinct very small embryonic-like PSCs and tissue committed progenitor stem cells population that have the potential to develop into oocyte-like structures in vitro, whereas mesenchymal fibroblasts appear to form supporting granulosa-like somatic cells. Research at the single-cell level, including complete gene expression profiling, is required to further confirm whether postnatal oogenesis is a conserved phenomenon in adult mammals.
Griesinger, Claudius B; Richards, Chistopher D; Ashmore, Jonathan F
Neural transmission of complex sounds demands fast and sustained rates of synaptic release from the primary cochlear receptors, the inner hair cells (IHCs). The cells therefore require efficient membrane recycling. Using two-photon imaging of the membrane marker FM1-43 in the intact sensory epithelium within the cochlear bone of the adult guinea pig, we show that IHCs possess fast calcium-dependent membrane uptake at their apical pole. FM1-43 did not permeate through the stereocilial mechanotransducer channel because uptake kinetics were neither changed by the blockers dihydrostreptomycin and d-tubocurarine nor by treatment of the apical membrane with BAPTA, known to disrupt mechanotransduction. Moreover, the fluid phase marker Lucifer Yellow produced a similar labeling pattern to FM1-43, consistent with FM1-43 uptake via endocytosis. We estimate the membrane retrieval rate at approximately 0.5% of the surface area of the cell per second. Labeled membrane was rapidly transported to the base of IHCs by kinesin-dependent trafficking and accumulated in structures that resembled synaptic release sites. Using confocal imaging of FM1-43 in excised strips of the organ of Corti, we show that the time constants of fluorescence decay at the basolateral pole of IHCs and apical endocytosis were increased after depolarization of IHCs with 40 mm potassium, a stimulus that triggers calcium influx and increases synaptic release. Blocking calcium channels with either cadmium or nimodipine during depolarization abolished the rate increase of apical endocytosis. We suggest that IHCs use fast calcium-dependent apical endocytosis for activity-associated replenishment of synaptic membrane.
... for RGC reprogramming is understanding the cues that direct their maturation and integration with other cells. The ... the retina. The report appears in Translational Vision Science and Technology. Learn more about the NEI AGI ...
Divakaran, Vijay G.; Evans, Sarah; Topkara, Veli K.; Diwan, Abhinav; Burchfield, Jana; Gao, Feng; Dong, Jianwen; Tzeng, Huei-Ping; Sivasubramanian, Natarajan; Barger, Philip M.; Mann, Douglas L.
Background Tumor necrosis factor (TNF) superfamily ligands that provoke a dilated cardiac phenotype signal through a common scaffolding protein termed TNF receptor associated factor 2 (TRAF2); however, virtually nothing is known with regard to TRAF2 signaling in the adult mammalian heart. Methods and Results We generated multiple founder lines of mice with cardiac restricted overexpression of TRAF2 and characterized the phenotype of mice with higher expression levels of TRAF2 (MHC-TRAF2HC). MHC-TRAF2HC transgenic mice developed a time-dependent increase in cardiac hypertrophy, LV dilation and adverse LV remodeling, and a significant decrease in LV +dP/dt and −dP/dt when compared to littermate (LM) controls (p < 0.05 compared to LM). During the early phases of LV remodeling there was a significant increase in total matrix metalloproteinase (MMP) activity that corresponded with a decrease in total myocardial fibrillar collagen content. As the MHC-TRAF2HC mice aged, there was a significant decrease in total MMP activity accompanied by an increase in total fibrillar collagen content and an increase in myocardial tissue inhibitor of metalloproteinase-1 levels. There was a significant increase in NF-κB activation at 4 – 12 weeks and JNK activation at 4 weeks in the MHCs TRAF2HC mice. Transciptional profiling revealed that > 95% of the hypertrophic/dilated cardiomyopathy-related genes that were significantly upregulated genes in the MHC-TRAF2HC hearts contained κB elements in their promoters. Conclusions These results show for the first time that targeted overexpression of TRAF2 is sufficient to mediate adverse cardiac remodeling in the heart. PMID:23493088
Jaeger, Catherine; Sandu, Cristina; Malan, André; Mellac, Katell; Hicks, David; Felder-Schmittbuhl, Marie-Paule
Rhythmic physiology is central to retinal function and survival and adapts vision to daily light intensity changes. Mammalian retina rhythmically releases melatonin when cultured under constant conditions, and the occurrence of clock gene [e.g., Period (Per)] expression has been shown for most cellular layers. However, contribution of the distinct layers to genesis of circadian rhythms within the retina is still debated. To characterize their endogenous oscillatory capacity and their communication at the whole-tissue level, we used a vibratome-based method to isolate individual or paired retina cellular layers from the mPer2(Luc) mouse and Per1-luciferase (Per1-Luc) rat, and real-time recorded bioluminescence. We report that each layer of the mouse retina harbors a self-sustained oscillator whose period is significantly longer (∼ 26 hours) than in whole-retina explants (∼ 22.9 hours), indicating that the period is correlated with the degree of coupling. Accordingly, the maximal period (∼ 29 hours) is reached upon complete enzymatic dissociation of the retina. By using pharmacological approaches, we demonstrate that connection between retina oscillators involves gap junctions but only minor contribution from the main retina neurochemicals. Taken together with results from Per1-Luc rats, these data show that mammalian retina consists of a network of layer-specific oscillators whose period is determined by their connectivity.
Santos-Carvalho, Ana; Ambrósio, António Francisco; Cavadas, Cláudia
The retina is a highly complex structure where several types of cells communicate through countless different molecules to codify visual information. Each type of cells plays unique roles in the retina, presenting a singular expression of neurotransmitters. Some neurotransmitter systems in the retina are well understood, while others need to be better explored to unravel the intricate signaling system involved. Neuropeptide Y (NPY), a 36 amino acid peptide, is one of the most common peptide neurotransmitter in the CNS and a highly conserved peptide among species. We review the localization of NPY and NPY receptors (mainly NPY Y1, Y2, Y4 and Y5) in retinal cells. Common features of the expression of NPY and NPY receptors in mammalian and non-mammalian species indicate universal roles of this system in the retina. In the present review, we highlight the putative roles of NPY receptor activation in the retina, discussing, in particular, their involvement in retinal development, neurotransmitter release modulation, neuroprotection, microglia and Muller cells function, retinal pigmented epithelium changes, retinal endothelial physiology and proliferation of retinal progenitor cells. Further studies are needed to confirm that targeting the NPY system might be a potential therapeutic strategy for retinal degenerative diseases.
Inhibitory interneurons participate in all neuronal circuits in the mammalian brain, including the circadian clock system, and are indispensable for their effective function. Although the clock neurons have different molecular and electrical properties, their main function is the generation of circadian oscillations. Here we review the circadian plasticity of GABAergic interneurons in several areas of the mammalian brain, suprachiasmatic nucleus, neocortex, hippocampus, olfactory bulb, cerebellum, striatum, and in the retina. PMID:28367335
Henning, Yoshiyuki; Szafranski, Karol
The thyroid hormones (TH) triiodothyronine (T3) and its prohormone thyroxine (T4) are crucial for retinal development and function, and increasing evidence points at TH dysregulation as a cause for retinal degenerative diseases. Thus, precise regulation of retinal TH supply is required for proper retinal function, but knowledge on these mechanisms is still fragmentary. Several transmembrane transporters have been described as key regulators of TH availability in target tissues of which the monocarboxylate transporter 8 (MCT8), a high affinity transporter for T4 and T3, plays an essential role in the central nervous system. Moreover, in the embryonic chicken retina, MCT8 is highly expressed, but the postnatal availability of MCT8 in the mammalian retina was not reported to date. In the present study, spatiotemporal retinal MCT8 availability was examined in mice of different age. For this purpose, we quantified expression levels of Mct8 via Real-Time Reverse-Transcriptase PCR in mouse eyecups (C57BL/6) of juvenile and adult age groups. Additionally, age-dependent MCT8 protein levels were quantified via Western blotting and localized via immunofluorescence confocal microscopy. While no difference in Mct8 expression levels could be detected between age groups, MCT8 protein levels in juvenile animals were about two times higher than in adult animals based on Western blot analyses. Immunohistochemical analyses showed that MCT8 immunoreactivity in the eyecup was restricted to the retina and the retinal pigment epithelium. In juvenile mice, MCT8 was broadly observed along the apical membrane of the retinal pigment epithelium, tightly surrounding photoreceptor outer segments. Distinct immunopositive staining was also detected in the inner nuclear layer and the ganglion cell layer. However, in adult specimens, immunoreactivity visibly declined in all layers, which was in line with Western blot analyses. Since MCT8 was abundantly present in juvenile and about twofold lower in
Henning, Yoshiyuki; Szafranski, Karol
The thyroid hormones (TH) triiodothyronine (T3) and its prohormone thyroxine (T4) are crucial for retinal development and function, and increasing evidence points at TH dysregulation as a cause for retinal degenerative diseases. Thus, precise regulation of retinal TH supply is required for proper retinal function, but knowledge on these mechanisms is still fragmentary. Several transmembrane transporters have been described as key regulators of TH availability in target tissues of which the monocarboxylate transporter 8 (MCT8), a high affinity transporter for T4 and T3, plays an essential role in the central nervous system. Moreover, in the embryonic chicken retina, MCT8 is highly expressed, but the postnatal availability of MCT8 in the mammalian retina was not reported to date. In the present study, spatiotemporal retinal MCT8 availability was examined in mice of different age. For this purpose, we quantified expression levels of Mct8 via Real-Time Reverse-Transcriptase PCR in mouse eyecups (C57BL/6) of juvenile and adult age groups. Additionally, age-dependent MCT8 protein levels were quantified via Western blotting and localized via immunofluorescence confocal microscopy. While no difference in Mct8 expression levels could be detected between age groups, MCT8 protein levels in juvenile animals were about two times higher than in adult animals based on Western blot analyses. Immunohistochemical analyses showed that MCT8 immunoreactivity in the eyecup was restricted to the retina and the retinal pigment epithelium. In juvenile mice, MCT8 was broadly observed along the apical membrane of the retinal pigment epithelium, tightly surrounding photoreceptor outer segments. Distinct immunopositive staining was also detected in the inner nuclear layer and the ganglion cell layer. However, in adult specimens, immunoreactivity visibly declined in all layers, which was in line with Western blot analyses. Since MCT8 was abundantly present in juvenile and about twofold lower in
This review examines the biological background to the development of ideas on rapid eye movement sleep (REM sleep), so-called paradoxical sleep (PS), and its relation to dreaming. Aspects of the phenomenon which are discussed include physiological changes and their anatomical location, the effects of total and selective sleep deprivation in the human and animal, and REM sleep behavior disorder, the latter with its clinical manifestations in the human. Although dreaming also occurs in other sleep phases (non-REM or NREM sleep), in the human, there is a contingent relation between REM sleep and dreaming. Thus, REM is taken as a marker for dreaming and as REM is distributed ubiquitously throughout the mammalian class, it is suggested that other mammals also dream. It is suggested that the overall function of REM sleep/dreaming is more important than the content of the individual dream; its function is to place the dreamer protagonist/observer on the topographical world. This has importance for the developing infant who needs to develop a sense of self and separateness from the world which it requires to navigate and from which it is separated for long periods in sleep. Dreaming may also serve to maintain a sense of ‘I’ness or “self” in the adult, in whom a fragility of this faculty is revealed in neurological disorders.
Arbogast, Patrick; Flamant, Frédéric; Godement, Pierre; Glösmann, Martin
Thyroid hormone is a crucial regulator of gene expression in the developing and adult retina. Here we sought to map sites of thyroid hormone signaling at the cellular level using the transgenic FINDT3 reporter mouse model in which neurons express β-galactosidase (β-gal) under the control of a hybrid Gal4-TRα receptor when triiodothyronine (T3) and cofactors of thyroid receptor signaling are present. In the adult retina, nearly all neurons of the ganglion cell layer (GCL, ganglion cells and displaced amacrine cells) showed strong β-gal labeling. In the inner nuclear layer (INL), a minority of glycineric and GABAergic amacrine cells showed β-gal labeling, whereas the majority of amacrine cells were unlabeled. At the level of amacrine types, β-gal labeling was found in a large proportion of the glycinergic AII amacrines, but only in a small proportion of the cholinergic/GABAergic ‘starburst’ amacrines. At postnatal day 10, there also was a high density of strongly β-gal-labeled neurons in the GCL, but only few amacrine cells were labeled in the INL. There was no labeling of bipolar cells, horizontal cells and Müller glia cells at both stages. Most surprisingly, the photoreceptor somata in the outer nuclear layer also showed no β-gal label, although thyroid hormone is known to control cone opsin expression. This is the first record of thyroid hormone signaling in the inner retina of an adult mammal. We hypothesize that T3 levels in photoreceptors are below the detection threshold of the reporter system. The topographical distribution of β-gal-positive cells in the GCL follows the overall neuron distribution in that layer, with more T3-signaling cells in the ventral than the dorsal half-retina. PMID:27942035
Constable, Simon; Pirmohamed, Munir
The retina is relatively protected from systemic drug administration because of the blood-retinal barrier, a highly selective mechanism adapted to providing a regulated homeostatic environment for this highly specialised tissue. However, a number of drugs have been associated with retinal toxicity. Vigabatrin, as an adjunctive therapy for the management of partial epilepsy, is associated with visual field defects in approximately 40% of patients. Hydroxychloroquine, used in the treatment of rheumatoid arthritis and systemic lupus erythematosus, is also associated with a retinopathy. In view of this, ophthalmological screening and monitoring is recommended during prescription of both of these drugs. In these cases, the retina is the site for an adverse drug reaction and the dose of therapy may be important in determining the likelihood of retinal toxicity. However, in the case of cytomegalovirus retinitis, the retina is the intended site for pharmacological action. The treatment of this condition with the antiviral agents ganciclovir, valganciclovir, foscarnet and cidofovir, can also be associated with significant systemic toxicity.
Walsh, C.; Polley, E.H.
The ganglion cells of the cat's retina form several classes distinguishable in terms of soma size, axon diameter, dendritic morphology, physiological properties, and central connections. Labeling with (/sup 3/H)thymidine shows that the ganglion cells which survive in the adult are produced as several temporally shifted, overlapping waves: medium-sized cells are produced before large cells, whereas the smallest ganglion cells are produced throughout the period of ganglion cell generation. Large cells and medium-sized cells show the same distinctive pattern of production, forming rough spirals around the area centralis. The oldest cells tend to lie superior and nasal to the area centralis, whereas cells in the inferior nasal retina and inferior temporal retina are, in general, progressively younger. Within each retinal quadrant, cells nearer the area centralis tend to be older than cells in the periphery, but there is substantial overlap. The retinal raphe divides the superior temporal quadrant into two zones with different patterns of cell addition. Superior temporal retina near the vertical meridian adds cells only slightly later than superior nasal retina, whereas superior temporal retina near the horizontal meridian adds cells very late, contemporaneously with inferior temporal retina. The broader wave of production of smaller ganglion cells seems to follow this same spiral pattern at its beginning and end. The presence of the area centralis as a nodal point about which ganglion cell production in the retinal quadrants pivots suggests that the area centralis is already an important retinal landmark even at the earliest stages of retinal development.
Zaghloul, Kareem A.; Boahen, Kwabena
Prosthetic devices may someday be used to treat lesions of the central nervous system. Similar to neural circuits, these prosthetic devices should adapt their properties over time, independent of external control. Here we describe an artificial retina, constructed in silicon using single-transistor synaptic primitives, with two forms of locally controlled adaptation: luminance adaptation and contrast gain control. Both forms of adaptation rely on local modulation of synaptic strength, thus meeting the criteria of internal control. Our device is the first to reproduce the responses of the four major ganglion cell types that drive visual cortex, producing 3600 spiking outputs in total. We demonstrate how the responses of our device's ganglion cells compare to those measured from the mammalian retina. Replicating the retina's synaptic organization in our chip made it possible to perform these computations using a hundred times less energy than a microprocessor—and to match the mammalian retina in size and weight. With this level of efficiency and autonomy, it is now possible to develop fully implantable intraocular prostheses.
Lee, Sammy C.S.; Meyer, Arndt; Schubert, Timm; Hüser, Laura; Dedek, Karin; Haverkamp, Silke
Amacrine cells comprise ~30 morphological types in the mammalian retina. The synaptic connectivity and function of a few GABAergic wide-field amacrine cells have recently been studied, however, with the exception of the rod pathway-specific AII amacrine cell the connectivity of glycinergic small-field amacrine cells has not been investigated in the mouse retina. Here, we studied the morphology and connectivity pattern of the small-field A8 amacrine cell. A8 cells in mouse retina are bistratified with lobular processes in the ON sublamina and arboreal dendrites in the OFF sublamina of the inner plexiform layer. The distinct bistratified morphology was first visible at postnatal day 8, reaching the adult shape at P13, around eye opening. The connectivity of A8 cells to bipolar cells and ganglion cells was studied by double and triple immunolabeling experiments using various cell markers combined with synaptic markers. Our data suggest that A8 amacrine cells receive glutamatergic input from both OFF and ON cone bipolar cells. Furthermore, A8 cells are coupled to ON cone bipolar cells by gap junctions, and provide inhibitory input via glycine receptor (GlyR) subunit α1 to OFF cone bipolar cells and to ON A-type ganglion cells. Measurements of spontaneous glycinergic postsynaptic currents and GlyR immunolabeling revealed that A8 cells express GlyRs containing the α2 subunit. Taken together, the bistratified A8 cell makes very similar synaptic contacts with cone bipolar cells as the rod pathway-specific AII amacrine cell. However, unlike AII cells, A8 amacrine cells provide glycinergic input to ON A-type ganglion cells. PMID:25630271
Feller, Kathryn D; Cohen, Jonathan H; Cronin, Thomas W
Stomatopod eye development is unusual among crustaceans. Just prior to metamorphosis, an adult retina and associated neuro-processing structures emerge adjacent to the existing material in the larval compound eye. Depending on the species, the duration of this double-retina eye can range from a few hours to several days. Although this developmental process occurs in all stomatopod species observed to date, the retinal physiology and extent to which each retina contributes to the animal's visual sensitivity during this transition phase is unknown. We investigated the visual physiology of stomatopod double retinas using microspectrophotometry and electroretinogram recordings from different developmental stages of the Western Atlantic species Squilla empusa. Though microspectrophotometry data were inconclusive, we found robust ERG responses in both larval and adult retinas at all sampled time points indicating that the adult retina responds to light from the very onset of its emergence. We also found evidence of an increase in the response dynamics with ontogeny as well as an increase in sensitivity of retinal tissue during the double-retina phase relative to single retinas. These data provide an initial investigation into the ontogeny of vision during stomatopod double-retina eye development.
Wang, Jin-shan; Nymark, Soile; Frederiksen, Rikard; Estevez, Maureen E.; Shen, Susan Q.; Corbo, Joseph C.; Cornwall, M. Carter
Efficient regeneration of visual pigment following its destruction by light is critical for the function of mammalian photoreceptors. Here, we show that misexpression of a subset of cone genes in the rd7 mouse hybrid rods enables them to access the normally cone-specific retina visual cycle. The rapid supply of chromophore by the retina visual cycle dramatically accelerated the mouse rod dark adaptation. At the same time, the competition between rods and cones for retina-derived chromophore slowed cone dark adaptation, indicating that the cone specificity of the retina visual cycle is key for rapid cone dark adaptation. Our findings demonstrate that mammalian photoreceptor dark adaptation is dominated by the supply of chromophore. Misexpression of cone genes in rods may represent a novel approach to treating visual disorders associated with mutations of visual cycle proteins or with reduced retinal pigment epithelium function due to aging. PMID:25143602
Sugitani, Kayo; Koriyama, Yoshiki; Ogai, Kazuhiro; Wakasugi, Keisuke; Kato, Satoru
Neuroglobin (Ngb) is a new member of the family of heme proteins and is specifically expressed in neurons of the central and peripheral nervous systems in all vertebrates. In particular, the retina has a 100-fold higher concentration of Ngb than do other nervous tissues. The role of Ngb in the retina is yet to be clarified. Therefore, to understand the functional role of Ngb in the retina after optic nerve injury (ONI), we used two types of retina, from zebrafish and mice, which have permissible and non-permissible capacity for nerve regeneration after ONI, respectively. After ONI, the Ngb protein in zebrafish was upregulated in the amacrine cells within 3 days, whereas in the mouse retina, Ngb was downregulated in the retinal ganglion cells (RGCs) within 3 days. Zebrafish Ngb (z-Ngb) significantly enhanced neurite outgrowth in retinal explant culture. According to these results, we designed an overexpression experiment with the mouse Ngb (m-Ngb) gene in RGC-5 cells (retinal precursor cells). The excess of m-Ngb actually rescued RGC-5 cells under hypoxic conditions and significantly enhanced neurite outgrowth in cell culture. These data suggest that mammalian Ngb has positive neuroprotective and neuritogenic effects that induce nerve regeneration after ONI.
Cornide-Petronio, María Eugenia; Anadón, Ramón; Barreiro-Iglesias, Antón; Rodicio, María Celina
The dual development of the retina of lampreys is exceptional among vertebrates and offers an interesting EvoDevo (evolutionary developmental biology) model for understanding the origin and evolution of the vertebrate retina. Only a single type of photoreceptor, ganglion cell and bipolar cell are present in the early-differentiated central retina of lamprey prolarvae. A lateral retina appears later in medium-sized larvae (about 3 years after hatching in the sea lamprey), growing and remaining largely neuroblastic until metamorphosis. In this lateral retina, only ganglion cells and optic fibers differentiate in larvae, whereas differentiation of amacrine, horizontal, photoreceptor and bipolar cells mainly takes place during metamorphosis, which gives rise to the adult retina. Serotonin (5-hydroxytryptamine, 5-HT) is a neurotransmitter found in the retina of vertebrates whose synthesis is mediated by the rate-limiting enzyme tryptophan hydroxylase (TPH). TPH is also the first enzyme in the biosynthetic pathways of melatonin in photoreceptor cells. The serotonin 1A receptor (5-HT1A) is a major determinant of the activity of both serotonergic cells and their targets due to its pre- and post-synaptic location. Here, we report the developmental pattern of expression of tph and 5-ht1a transcripts in the sea lamprey retina by means of in situ hybridization. In larvae, strong tph mRNA signal was observed in photoreceptors and putative ganglion cells of the central retina, and in some neuroblasts of the lateral retina. In adults, strong tph expression was observed in bipolar, amacrine and ganglion cells and in photoreceptors. In the prolarval (central) retina, all the differentiated retinal cells expressed 5-ht1a transcripts, which were not observed in undifferentiated cells. In larvae, photoreceptors, bipolar cells and ganglion cells in the central retina, and neuroblasts in the lateral retina, showed 5-ht1a expression. In the adult retina, expression of 5-ht1a transcript
Weinl, Christine; Wasylyk, Christine; Garcia Garrido, Marina; Sothilingam, Vithiyanjali; Beck, Susanne C; Riehle, Heidemarie; Stritt, Christine; Roux, Michel J; Seeliger, Mathias W; Wasylyk, Bohdan; Nordheim, Alfred
Serum Response Factor (SRF) fulfills essential roles in post-natal retinal angiogenesis and adult neovascularization. These functions have been attributed to the recruitment by SRF of the cofactors Myocardin-Related Transcription Factors MRTF-A and -B, but not the Ternary Complex Factors (TCFs) Elk1 and Elk4. The role of the third TCF, Elk3, remained unknown. We generated a new Elk3 knockout mouse line and showed that Elk3 had specific, non-redundant functions in the retinal vasculature. In Elk3(-/-) mice, post-natal retinal angiogenesis was transiently delayed until P8, after which it proceeded normally. Interestingly, tortuous arteries developed in Elk3(-/-) mice from the age of four weeks, and persisted into late adulthood. Tortuous vessels have been observed in human pathologies, e.g. in ROP and FEVR. These human disorders were linked to altered activities of vascular endothelial growth factor (VEGF) in the affected eyes. However, in Elk3(-/-) mice, we did not observe any changes in VEGF or several other potential confounding factors, including mural cell coverage and blood pressure. Instead, concurrent with the post-natal transient delay of radial outgrowth and the formation of adult tortuous arteries, Elk3-dependent effects on the expression of Angiopoietin/Tie-signalling components were observed. Moreover, in vitro microvessel sprouting and microtube formation from P10 and adult aortic ring explants were reduced. Collectively, these results indicate that Elk3 has distinct roles in maintaining retinal artery integrity. The Elk3 knockout mouse is presented as a new animal model to study retinal artery tortuousity in mice and human patients.
Gatta, Claudia; Castaldo, Luciana; Cellerino, Alessandro; de Girolamo, Paolo; Lucini, Carla; D'Angelo, Livia
BDNF plays an important role in the development and maintenance of visual circuitries in the retina and brain visual centers. In adulthood, BDNF signaling is involved in neural protection and regeneration of retina. In this survey, we investigated the expression of BDNF in the retina of adult Nothobranchius furzeri, a teleost fish employed for age research. After describing the retina of N. furzeri and confirming that the structure is organized in layers as in all vertebrates, we have studied the localization of BDNF mRNA and protein throughout the retinal layers. BDNF mRNA is detectable in all layers, whereas the protein is lacking in the photoreceptors. The occurrence of BDNF provides new insights on its role in the retina, particularly in view of age-related disease of retina.
Amrein, Irmgard; Isler, Karin; Lipp, Hans-Peter
Adult hippocampal neurogenesis is a prominent event in rodents. In species with longer life expectancies, newly born cells in the adult dentate gyrus of the hippocampal formation are less abundant or can be completely absent. Several lines of evidence indicate that the regulatory mechanisms of adult neurogenesis differ between short- and long-lived mammals. After a critical appraisal of the factors and problems associated with comparing different species, we provide a quantitative comparison derived from seven laboratory strains of mice (BALB, C57BL/6, CD1, outbred) and rats (F344, Sprague-Dawley, Wistar), six other rodent species of which four are wild-derived (wood mouse, vole, spiny mouse and guinea pig), three non-human primate species (marmoset and two macaque species) and one carnivore (red fox). Normalizing the number of proliferating cells to total granule cell number, we observe an overall exponential decline in proliferation that is chronologically equal between species and orders and independent of early developmental processes and life span. Long- and short-lived mammals differ with regard to major life history stages; at the time points of weaning, age at first reproduction and average life expectancy, long-lived primates and foxes have significantly fewer proliferating cells than rodents. Although the database for neuronal differentiation is limited, we find indications that the extent of neuronal differentiation is subject to species-specific selective adaptations. We conclude that absolute age is the critical factor regulating cell genesis in the adult hippocampus of mammals. Ontogenetic and ecological factors primarily influence the regulation of neuronal differentiation rather than the rate of cell proliferation.
Stone, Jonathan; van Driel, Diana; Valter, Krisztina; Rees, Sandra; Provis, Jan
Adult mammalian photoreceptors are elongated cells, and their mitochondria are sequestered to the ends of the cell, to the inner segments and (in some species) to axon terminals in the outer plexiform layer (OPL). We hypothesised that mitochondria migrate to these locations towards sources of oxygen, from the choroid and (in some species) from the deep capillaries of the retinal circulation. Six mammalian species were surveyed, using electron and light microscopy, including immunohistochemistry for the mitochondrial enzyme cytochrome oxidase (CO). In all 6 species, mitochondria were absent from photoreceptor somas and were numerous in inner segments. Mitochondria were prominent in axon terminals in 3 species (mouse, rat, human) with a retinal circulation and were absent from those terminals in 3 species (wallaby, rat, guinea pig) with avascular retinas. Further, in a human developmental series, it was evident that mitochondria migrate within rods and cones, towards and eventually past the outer limiting membrane (OLM), into the inner segment. In Müller and RPE cells also, mitochondria concentrated at the external surface of the cells. Neurones located in the inner layers of avascular retinas have mitochondria, but their expression of CO is low. Mitochondrial locations in photoreceptors, Müller and RPE cells are economically explained as the result of migration within the cell towards sources of oxygen. In photoreceptors, this migration results in a separation of mitochondria from the nuclear genome; this separation may be a factor in the vulnerability of photoreceptors to mutations, toxins and environmental stresses, which other retinal neurones survive.
Zhang, Pingbo; Dufresne, Craig; Turner, Randi; Ferri, Sara; Venkatraman, Vidya; Karani, Rabia; Lutty, Gerard A; Van Eyk, Jennifer E; Semba, Richard D
The retina is a delicate tissue that detects light, converts photochemical energy into neural signals, and transmits the signals to the visual cortex of the brain. A detailed protein inventory of the proteome of the normal human eye may provide a foundation for new investigations into both the physiology of the retina and the pathophysiology of retinal diseases. To provide an inventory, proteins were extracted from five retinas of normal eyes and fractionated using SDS-PAGE. After in-gel digestion, peptides were analyzed in duplicate using LC-MS/MS on an Orbitrap Elite mass spectrometer. A total of 3436 nonredundant proteins were identified in the human retina, including 20 unambiguous protein isoforms, of which eight have not previously been demonstrated to exist at the protein level. The proteins identified in the retina included most of the enzymes involved in the visual cycle and retinoid metabolism. One hundred and fifty-eight proteins that have been associated with age-related macular degeneration were identified in the retina. The MS proteome database of the human retina may serve as a valuable resource for future investigations of retinal biology and disease. All MS data have been deposited in the ProteomeXchange with identifier PXD001242 (http://proteomecentral.proteomexchange.org/dataset/PXD001242).
Wan, Qun-Fang; Heidelberger, Ruth
Bipolar cells play a vital role in the transfer of visual information across the vertebrate retina. The synaptic output of these neurons is regulated by factors that are extrinsic and intrinsic. Relatively little is known about the intrinsic factors that regulate neurotransmitter exocytosis. Much of what we know about intrinsic presynaptic mechanisms that regulate glutamate release has come from the study of the unusually large and accessible synaptic terminal of the goldfish rod-dominant bipolar cell, the Mb1 bipolar cell. However, over the past several years, examination of presynaptic mechanisms governing neurotransmitter release has been extended to the mammalian rod bipolar cell. In this review, we discuss the recent advances in our understanding of synaptic vesicle dynamics and neurotransmitter release in rodent rod bipolar cells and consider how these properties help shape the synaptic output of the mammalian retina. PMID:21272392
Rath, Martin F; Muñoz, Estela; Ganguly, Surajit; Morin, Fabrice; Shi, Qiong; Klein, David C; Møller, Morten
Otx2 is a vertebrate homeobox gene, which has been found to be essential for the development of rostral brain regions and appears to play a role in the development of retinal photoreceptor cells and pinealocytes. In this study, the temporal expression pattern of Otx2 was revealed in the rat brain, with special emphasis on the pineal gland throughout late embryonic and postnatal stages. Widespread high expression of Otx2 in the embryonic brain becomes progressively restricted in the adult to the pineal gland. Crx (cone-rod homeobox), a downstream target gene of Otx2, showed a pineal expression pattern similar to that of Otx2, although there was a distinct lag in time of onset. Otx2 protein was identified in pineal extracts and found to be localized in pinealocytes. Total pineal Otx2 mRNA did not show day-night variation, nor was it influenced by removal of the sympathetic input, indicating that the level of Otx2 mRNA appears to be independent of the photoneural input to the gland. Our results are consistent with the view that pineal expression of Otx2 is required for development and we hypothesize that it plays a role in the adult in controlling the expression of the cluster of genes associated with phototransduction and melatonin synthesis.
Fallon, James; Reid, Steve; Kinyamu, Richard; Opole, Isaac; Opole, Rebecca; Baratta, Janie; Korc, Murray; Endo, Tiffany L.; Duong, Alexander; Nguyen, Gemi; Karkehabadhi, Masoud; Twardzik, Daniel; Loughlin, Sandra
The development of an in vivo procedure for the induction of massive proliferation, directed migration, and neurodifferentiation (PMD) in the damaged adult central nervous system would hold promise for the treatment of human neurodegenerative disorders such as Parkinson's disease. We investigated the in vivo induction of PMD in the forebrain of the adult rat by using a combination of 6-hydroxydopamine lesion of the substantia nigra dopaminergic neurons and infusions of transforming growth factor α (TGFα) into forebrain structures. Only in animals with both lesion and infusion of TGFα was there a rapid proliferation of forebrain stem cells followed by a timed migration of a ridge of neuronal and glial progenitors directed toward the region of the TGFα infusion site. Subsequently, increasing numbers of differentiated neurons were observed in the striatum. In behavioral experiments, there was a significant reduction of apomorphine-induced rotations in animals receiving the TGFα infusions. These results show that the brain contains stem cells capable of PMD in response to an exogenously administered growth factor. This finding has significant implications with respect to the development of treatments for both acute neural trauma and neurodegenerative diseases. PMID:11121069
Hirano, Arlene A.; Brandstätter, Johann Helmut; Morgans, Catherine W.; Brecha, Nicholas C.
Horizontal cells mediate inhibitory feedforward and feedback lateral interactions in the outer retina at photoreceptor terminals and bipolar cell dendrites; however, the mechanisms that underlie synaptic transmission from mammalian horizontal cells are poorly understood. The localization of a vesicular γ-aminobutyric acid (GABA) transporter (VGAT) to horizontal cell processes in primate and rodent retinae suggested that mammalian horizontal cells release transmitter in a vesicular manner. Toward determining whether the molecular machinery for vesicular transmitter release is present in horizontal cells, we investigated the expression of SNAP25 (synaptosomal-associated protein of 25 kDa), a key SNARE protein, by immunocytochemistry with cell type-specific markers in the retinae of mouse, rat, rabbit, and monkey. Different commercial antibodies to SNAP25 were tested on vertical sections of retina. We report the robust expression of SNAP25 in both plexiform layers. Double labeling with SNAP25 and calbindin antibodies demonstrated that horizontal cell processes and their endings in photoreceptor triad synapses were strongly labeled for both proteins in mouse, rat, rabbit, and monkey retinae. Double labeling with parvalbumin antibodies in monkey retina verified SNAP25 immunoreactivity in all horizontal cells. Pre-embedding immunoelectron microscopy in rabbit retina confirmed expression of SNAP25 in lateral elements within photoreceptor triad synapses. The SNAP25 immunoreactivity in the plexiform layers and outer nuclear layer fell into at least three patterns depending on the antibody, suggesting a differential distribution of SNAP25 isoforms. The presence of SNAP25a and SNAP25b isoforms in mouse retina was established by reverse transcriptase-polymerase chain reaction. SNAP25 expression in mammalian horizontal cells along with other SNARE proteins is consistent with vesicular exocytosis. PMID:21280047
Jelcick, Austin S.; Yuan, Yang; Leehy, Barrett D.; Cox, Lakeisha C.; Silveira, Alexandra C.; Qiu, Fang; Schenk, Sarah; Sachs, Andrew J.; Morrison, Margaux A.; Nystuen, Arne M.; DeAngelis, Margaret M.; Haider, Neena B.
Variation in genetic background can significantly influence the phenotypic outcome of both disease and non-disease associated traits. Additionally, differences in temporal and strain specific gene expression can also contribute to phenotypes in the mammalian retina. This is the first report of microarray based cross-strain analysis of gene expression in the retina investigating genetic background effects. Microarray analyses were performed on retinas from the following mouse strains: C57BL6/J, AKR/J, CAST/EiJ, and NOD.NON-H2-nb1 at embryonic day 18.5 (E18.5) and postnatal day 30.5 (P30.5). Over 3000 differentially expressed genes were identified between strains and developmental stages. Differential gene expression was confirmed by qRT-PCR, Western blot, and immunohistochemistry. Three major gene networks were identified that function to regulate retinal or photoreceptor development, visual perception, cellular transport, and signal transduction. Many of the genes in these networks are implicated in retinal diseases such as bradyopsia, night-blindness, and cone-rod dystrophy. Our analysis revealed strain specific variations in cone photoreceptor cell patterning and retinal function. This study highlights the substantial impact of genetic background on both development and function of the retina and the level of gene expression differences tolerated for normal retinal function. These strain specific genetic variations may also be present in other tissues. In addition, this study will provide valuable insight for the development of more accurate models for human retinal diseases. PMID:21779340
Edgerton, V. R.; Roy, R. R.; Hodgson, J. A.; Prober, R. J.; de Guzman, C. P.; de Leon, R.
The neural circuitry of the lumbar spinal cord can generate alternating extension and flexion of the hindlimbs. The hindlimbs of adult cats with complete transection of the spinal cord at a low thoracic level (T12-T13) can perform full weight-supporting locomotion on a treadmill belt moving at a range of speeds. Some limitations in the locomotor capacity can be associated with a deficit in the recruitment level of the fast extensors during the stance phase and the flexors during the swing phase of a step cycle. The level of locomotor performance, however, can be enhanced by daily training on a treadmill while emphasizing full weight-support stepping and by providing appropriately timed sensory stimulation, loading, and/or pharmacologic stimulation of the hindlimb neuromuscular apparatus. Furthermore, there appears to be an interactive effect of these interventions. For example, the maximum treadmill speed that a spinal adult cat can attain and maintain is significantly improved with daily full weight-supporting treadmill training, but progressive recruitment of fast extensors becomes apparent only when the hindlimbs are loaded by gently pulling down on the tail during the stepping. Stimulation of the sural nerve at the initiation of the flexion phase of the step cycle can likewise markedly improve the locomotor capability. Administration of clonidine, in particular in combination with an elevated load, resulted in the most distinct and consistent alternating bursts of electromyographic activity during spinal stepping. These data indicate that the spinal cord has the ability to execute alternating activation of the extensor and flexor musculature of the hindlimbs (stepping) and that this ability can be improved by several interventions such as training, sensory stimulation, and use of some pharmacologic agents. Thus, it appears that the spinal cord, without supraspinal input, is highly plastic and has the potential to "learn," that is, to acquire and improve its
Stanke, Jennifer; Moose, Holly E; El-Hodiri, Heithem M; Fischer, Andy J
Little is known about the expression of Pax2 in mature retina or optic nerve. Here we probed for the expression of Pax2 in late stages of embryonic development and in mature chick retina. We find two distinct Pax2 isoforms expressed by cells within the retina and optic nerve. Surprisingly, Müller glia in central regions of the retina express Pax2, and levels of expression are decreased with increasing distance from the nerve head. In Müller glia, the expression levels of Pax2 are increased by acute retinal damage or treatment with growth factors. At the optic nerve, Pax2 is expressed by peripapillary glia, at the junction of the neural retina and optic nerve head and by glia within the optic nerve. In addition, we assayed for Pax2 expression in glial cells in mammalian retinas. In mammalian retinas, unlike the case in chick retina, the Müller glia do not express Pax2. Pax2-expressing cells are found in the optic nerve and astrocytes within the mouse retina. By comparison, Pax2-positive cells are not found within the guinea pig retina; Pax2-expressing glia are confined to the optic nerve. In dog and monkey (Macaca fascicularis), Pax2 is expressed by astrocytes that are scattered across inner retinal layers and by numerous glia within the optic nerve. Interestingly, Pax2-positive glial cells are found at the peripheral edge of the dog retina, but only in older animals. We conclude that the expression of Pax2 in the vertebrate eye is restricted to retinal astrocytes, peripapillary glia, and glia within the optic nerve.
Wu, Ji; Ding, Xinbao; Wang, Jian
Stem cells have great value in clinical application because of their ability to self-renew and their potential to differentiate into many different cell types. Mammalian gonads, including testes for males and ovaries for females, are composed of germline and somatic cells. In male mammals, spermatogonial stem cells maintain spermatogenesis which occurs continuously in adult testis. Likewise, a growing body of evidence demonstrated that female germline stem cells could be found in mammalian ovaries. Meanwhile, prior studies have shown that somatic stem cells exist in both testes and ovaries. In this chapter, we focus on mammalian gonad stem cells and discuss their characteristics as well as differentiation potentials.
Mamczur, Piotr; Mazurek, Jakub
To shed some light on gluconeogenesis in mammalian retina, we have focused on fructose-1,6-bisphosphatase (FBPase), a regulatory enzyme of the process. The abundance of the enzyme within the layers of the rat retina suggests that, in mammals in contrast to amphibia, gluconeogenesis is not restricted to one specific cell of the retina. We propose that FBPase, in addition to its gluconeogenic role, participates in the protection of the retina against reactive oxygen species. Additionally, the nuclear localization of FBPase and of its binding partner, aldolase, in the retinal cells expressing the proliferation marker Ki-67 indicates that these two gluconeogenic enzymes are involved in non-enzymatic nuclear processes. Electronic supplementary material The online version of this article (doi:10.1007/s00441-010-1008-2) contains supplementary material, which is available to authorized users. PMID:20614135
Kaltschmidt, Barbara; Kaltschmidt, Christian
The transcription factor nuclear factor kappaB (NF-κB) is a well-known regulator of inflammation, stress, and immune responses as well as cell survival. In the nervous system, NF-κB is one of the crucial components in the molecular switch that converts short- to long-term memory—a process that requires de novo gene expression. Here, the researches published on NF-κB and downstream target genes in mammals will be reviewed, which are necessary for structural plasticity and long-term memory, both under normal and pathological conditions in the brain. Genetic evidence has revealed that NF-κB regulates neuroprotection, neuronal transmission, and long-term memory. In addition, after genetic ablation of all NF-κB subunits, a severe defect in hippocampal adult neurogenesis was observed during aging. Proliferation of neural precursors is increased; however, axon outgrowth, synaptogenesis, and tissue homeostasis of the dentate gyrus are hampered. In this process, the NF-κB target gene PKAcat and other downstream target genes such as Igf2 are critically involved. Therefore, NF-κB activity seems to be crucial in regulating structural plasticity and replenishment of granule cells within the hippocampus throughout the life. In addition to the function of NF-κB in neurons, we will discuss on a neuroinflammatory role of the transcription factor in glia. Finally, a model for NF-κB homeostasis on the molecular level is presented, in order to explain seemingly the contradictory, the friend or foe, role of NF-κB in the nervous system. PMID:26635522
Liberles, Stephen D
Mammalian pheromones control a myriad of innate social behaviors and acutely regulate hormone levels. Responses to pheromones are highly robust, reproducible, and stereotyped and likely involve developmentally predetermined neural circuits. Here, I review several facets of pheromone transduction in mammals, including (a) chemosensory receptors and signaling components of the main olfactory epithelium and vomeronasal organ involved in pheromone detection; (b) pheromone-activated neural circuits subject to sex-specific and state-dependent modulation; and (c) the striking chemical diversity of mammalian pheromones, which range from small, volatile molecules and sulfated steroids to large families of proteins. Finally, I review (d) molecular mechanisms underlying various behavioral and endocrine responses, including modulation of puberty and estrous; control of reproduction, aggression, suckling, and parental behaviors; individual recognition; and distinguishing of own species from predators, competitors, and prey. Deconstruction of pheromone transduction mechanisms provides a critical foundation for understanding how odor response pathways generate instinctive behaviors.
Liberles, Stephen D.
Mammalian pheromones control a myriad of innate social behaviors and acutely regulate hormone levels. Responses to pheromones are highly robust, reproducible, and stereotyped and likely involve developmentally predetermined neural circuits. Here, I review several facets of pheromone transduction in mammals, including (a) chemosensory receptors and signaling components of the main olfactory epithelium and vomeronasal organ involved in pheromone detection; (b) pheromone-activated neural circuits subject to sex-specific and state-dependent modulation; and (c) the striking chemical diversity of mammalian pheromones, which range from small, volatile molecules and sulfated steroids to large families of proteins. Finally, I review (d ) molecular mechanisms underlying various behavioral and endocrine responses, including modulation of puberty and estrous; control of reproduction, aggression, suckling, and parental behaviors; individual recognition; and distinguishing of own species from predators, competitors, and prey. Deconstruction of pheromone transduction mechanisms provides a critical foundation for understanding how odor response pathways generate instinctive behaviors. PMID:23988175
Eckmiller, Rolf; Neumann, Dirk; Baruth, Oliver
Current research towards retina implants for partial restoration of vision in blind humans with retinal degenerative dysfunctions focuses on implant and stimulation experiments and technologies. In contrast, our approach takes the availability of an epiretinal multi-electrode neural interface for granted and studies the conditions for successful joint information processing of both retinal prosthesis and brain. Our proposed learning retina encoder (RE) includes information processing modules to simulate the complex mapping operation of parts of the 5-layered neural retina and to provide an iterative, perception-based dialog between RE and human subject. Alternative information processing technologies in the learning RE are being described, which allow an individual optimization of the RE mapping operation by means of iterative tuning with learning algorithms in a dialog between implant wearing subject and RE. The primate visual system is modeled by a retina module (RM) composed of spatio-temporal (ST) filters and a central visual system module (VM). RM performs a mapping 1 of an optical pattern P1 in the physical domain onto a retinal output vector R1(t) in a neural domain, whereas VM performs a mapping 2 of R1(t) in a neural domain onto a visual percept P2 in the perceptual domain. Retinal ganglion cell properties represent non-invertible ST filters in RE, which generate ambiguous output signals. VM generates visual percepts only if the corresponding R1(t) is properly encoded, contains sufficient information, and can be disambiguated. Based on the learning RE and the proposed visual system model, a novel retina encoder (RE*) is proposed, which considers both ambiguity removal and miniature eye movements during fixation. Our simulation results suggest that VM requires miniature eye movements under control of the visual system to retrieve unambiguous patterns P2 corresponding to P1. For retina implant applications, RE* can be tuned to generate optimal ganglion cell
Curcio, Christine A.; Sloan, Kenneth R.; Packer, Orin; Hendrickson, Anita E.; Kalina, Robert E.
The distribution of photoreceptors is known for only one complete human retina and for the cardinal meridians only in the macaque monkey retina. Cones can be mapped in computer-reconstructed whole mounts of human and monkey retina. A 2.9-fold range in maximum cone density in the foveas of young adult human eyes may contribute to individual differences in acuity. Cone distribution is radially asymmetrical about the fovea in both species, as previously described for the distribution of retinal ganglion cells and for lines of visual isosensitivity. Cone density was greater in the nasal than in the temporal peripheral retina, and this nasotemporal asymmetry was more pronounced in monkey than in human retina.
Hamilton, L K; Truong, M K V; Bednarczyk, M R; Aumont, A; Fernandes, K J L
A stem cell's microenvironment, or "niche," is a critical regulator of its behaviour. In the adult mammalian spinal cord, central canal ependymal cells possess latent neural stem cell properties, but the ependymal cell niche has not yet been described. Here, we identify important similarities and differences between the central canal ependymal zone and the forebrain subventricular zone (SVZ), a well-characterized niche of neural stem cells. First, direct immunohistochemical comparison of the spinal cord ependymal zone and the forebrain SVZ revealed distinct patterns of neural precursor marker expression. In particular, ependymal cells in the spinal cord were found to be bordered by a previously uncharacterized sub-ependymal layer, which is relatively less elaborate than that of the SVZ and comprised of small numbers of astrocytes, oligodendrocyte progenitors and neurons. Cell proliferation surrounding the central canal occurs in close association with blood vessels, but unlike in the SVZ, involves mainly ependymal rather than sub-ependymal cells. These proliferating ependymal cells typically self-renew rather than produce transit-amplifying progenitors, as they generate doublets of progeny that remain within the ependymal layer and show no evidence of a lineage relationship to sub-ependymal cells. Interestingly, the dorsal pole of the central canal was found to possess a sub-population of tanycyte-like cells that express markers of both ependymal cells and neural precursors, and their presence correlates with higher numbers of dorsally proliferating ependymal cells. Together, these data identify key features of the spinal cord ependymal cell niche, and suggest that dorsal ependymal cells possess the potential for stem cell activity. This work provides a foundation for future studies aimed at understanding ependymal cell regulation under normal and pathological conditions.
Wilhelm, M; Zhu, B; Gábriel, R; Straznicky, C
Serotonin-synthesizing neurons in the retinas of goldfish, axolotl, turtle, chick, rabbit and cat were identified using double labelling with anti-serotonin and anti-phenylalanine hydroxylase antibodies. The latter antibody recognizes tryptophan 5-hydroxylase, one of the synthesizing enzymes for serotonin. Neurons labelled by both markers were considered to be serotonin-synthesizing neurons, while those only with serotonin-immunoreactivity were assumed to be serotonin-accumulating neurons. In the goldfish and chick retinas, all serotonin-immunoreactive amacrine cells (S1) were positive for phenylalanine hydroxylase. In the axolotl and turtle retinas, all the S1 amacrine cells, and only 52.8% and 40.5% of S2 amacrine cells were double-labelled. Although serotonin-immunoreactive bipolar cells were observed in the turtle and chick retinas, the synthesizing enzyme for serotonin could not be detected in these cells. In the rabbit and cat retinas, tryptophan hydroxylase could not be revealed in any cell type with immunocytochemistry. In control experiments SLI neurons in the raphe nuclei of the brain stem always exhibited PH-LI in all species examined, including mammals, indicating that our anti-PH antibody is able to recognize tryptophan hydroxylase across vertebrate classes. These results indicate that the majority of serotonin-immunoreactive amacrine cells are able to synthesize serotonin and are the source of endogenous serotonin in the non-mammalian retina, while some serotonin-immunoreactive amacrine and bipolar cells possibly only accumulate serotonin. We also suggest that serotonin may not be a primary neurotransmitter in the serotonin-accumulating bipolar and amacrine cells of the non-mammalian retina.
Parker, Alice C.; Azar, Adi N.
Connectivity in the human retina is complex. Over one hundred million photoreceptors transduce light into electrical signals. These electrical signals are sent to the ganglion cells through amacrine and bipolar cells. Lateral connections involving horizontal and amacrine cells span throughout the outer plexiform layer and inner plexiform layer respectively. Horizontal cells are important for photoreceptor regulation by depolarizing them after an illumination occurs. Horizontal cells themselves form an electrical network that communicates by gap junctions, and these cells exhibit plasticity (change in behavior and structure) with respect to glycine receptors. The bipolar and amacrine cells transfer electrical signals from photoreceptors to the ganglion cells. Furthermore, amacrine cells are responsible for further processing the retinal image. Finally, the ganglion cells receive electrical signals from the bipolar and amacrine cells and will spike at a faster rate if there is a change in the overall intensity for a group of photoreceptors, sending a signal to the brain. Dramatic progress is being made with respect to retinal prostheses, raising hope for an entire synthetic retina in the future. We propose a bio-inspired 3D hierarchical pyramidal architecture for a synthetic retina that mimics the overall structure of the human retina. We chose to use a 3D architecture to facilitate connectivity among retinal cells, maintaining a hierarchical structure similar to that of the biological retina. The first layer of the architecture contains electronic circuits that model photoreceptors and horizontal cells. The second layer contains amacrine and bipolar electronic cells, and the third layer contains ganglion cells. Layer I has the highest number of cells, and layer III has the lowest number of cells, resulting in a pyramidal architecture. In our proposed architecture we intend to use photodetectors to transduce light into electrical signals. We propose to employ
Greferath, Ursula; Vessey, Kirstan A; Jobling, Andrew I; Mills, Samuel A; Bui, Bang V; He, Zheng; Nag, Nupur; Ohtsu, Hiroshi; Fletcher, Erica L
The role of histamine in the retina is not well understood, despite it regulating a number of functions within the brain, including sleep, feeding, energy balance, and anxiety. In this study we characterized the structure and function of the retina in mice that lacked expression of the rate limiting enzyme in the formation of histamine, histidine decarboxylase (Hdc-/- mouse). Using laser capture microdissection, Hdc mRNA expression was assessed in the inner and outer nuclear layers of adult C57Bl6J wildtype (WT) and Hdc(-/-)-retinae. In adult WT and Hdc(-/-)-mice, retinal fundi were imaged, retinal structure was assessed using immunocytochemistry and function was probed by electroretinography. Blood flow velocity was assessed by quantifying temporal changes in the dynamic fluorescein angiography in arterioles and venules. In WT retinae, Hdc gene expression was detected in the outer nuclear layer, but not the inner nuclear layer, while the lack of Hdc expression was confirmed in the Hdc-/- retina. Preliminary examination of the fundus and retinal structure of the widely used Hdc-/- mouse strain revealed discrete lesions across the retina that corresponded to areas of photoreceptor abnormality reminiscent of the rd8 (Crb1) mutation. This was confirmed after genotyping and the strain designated Hdcrd8/rd8. In order to determine the effect of the lack of Hdc-alone on the retina, Hdc-/- mice free of the Crb1 mutation were bred. Retinal fundi appeared normal in these animals and there was no difference in retinal structure, macrogliosis, nor any change in microglial characteristics in Hdc-/- compared to wildtype retinae. In addition, retinal function and retinal blood flow dynamics showed no alterations in the Hdc-/- retina. Overall, these results suggest that histamine plays little role in modulating retinal structure and function.
The mostly cause of blindness in the developed countries is a degeneration of the retina. For restoring this loss of vision one possible approach is the substitution of the lost functions by means of an electronic implant. This approach is based on MEMS technologies. It has been shown that electrical stimulation of retinal ganglion cells yield visual sensations1. Therefore, an artificial retina for blind humans based on this concept seems to be feasible. Besides electrical stimulation of retinal ganglion cells also the direct electrical stimulation of the optic nerve2 and the visual cortex3 have been under investigation. This paper wants to give an overview about the activities on the retinal ganglion cell stimulation.
Somers, Robert L.; Klein, David C.
Rhodopsin kinase, an enzyme involved in photochemical transduction in the retina, has been found in the mammalian pineal gland in amounts equal to those in the retina; other tissues had 7 percent of this amount, or less. This finding suggests that, in mammals, rhodopsin kinase functions in the pineal gland and other tissues to phosphorylate rhodopsin-like integral membrane receptors and is thereby involved in signal transduction.
Workman, Alan D; Charvet, Christine J; Clancy, Barbara; Darlington, Richard B; Finlay, Barbara L
A general model of neural development is derived to fit 18 mammalian species, including humans, macaques, several rodent species, and six metatherian (marsupial) mammals. The goal of this work is to describe heterochronic changes in brain evolution within its basic developmental allometry, and provide an empirical basis to recognize equivalent maturational states across animals. The empirical data generating the model comprises 271 developmental events, including measures of initial neurogenesis, axon extension, establishment, and refinement of connectivity, as well as later events such as myelin formation, growth of brain volume, and early behavioral milestones, to the third year of human postnatal life. The progress of neural events across species is sufficiently predictable that a single model can be used to predict the timing of all events in all species, with a correlation of modeled values to empirical data of 0.9929. Each species' rate of progress through the event scale, described by a regression equation predicting duration of development in days, is highly correlated with adult brain size. Neural heterochrony can be seen in selective delay of retinogenesis in the cat, associated with greater numbers of rods in its retina, and delay of corticogenesis in all species but rodents and the rabbit, associated with relatively larger cortices in species with delay. Unexpectedly, precocial mammals (those unusually mature at birth) delay the onset of first neurogenesis but then progress rapidly through remaining developmental events.
Workman, Alan D.; Charvet, Christine J.; Clancy, Barbara; Darlington, Richard B.
A general model of neural development is derived to fit 18 mammalian species, including humans, macaques, several rodent species, and six metatherian (marsupial) mammals. The goal of this work is to describe heterochronic changes in brain evolution within its basic developmental allometry, and provide an empirical basis to recognize equivalent maturational states across animals. The empirical data generating the model comprises 271 developmental events, including measures of initial neurogenesis, axon extension, establishment, and refinement of connectivity, as well as later events such as myelin formation, growth of brain volume, and early behavioral milestones, to the third year of human postnatal life. The progress of neural events across species is sufficiently predictable that a single model can be used to predict the timing of all events in all species, with a correlation of modeled values to empirical data of 0.9929. Each species' rate of progress through the event scale, described by a regression equation predicting duration of development in days, is highly correlated with adult brain size. Neural heterochrony can be seen in selective delay of retinogenesis in the cat, associated with greater numbers of rods in its retina, and delay of corticogenesis in all species but rodents and the rabbit, associated with relatively larger cortices in species with delay. Unexpectedly, precocial mammals (those unusually mature at birth) delay the onset of first neurogenesis but then progress rapidly through remaining developmental events. PMID:23616543
Gramage, E; Li, J; Hitchcock, P
The functional role of midkine during development, following injury and in disease has been studied in a variety of tissues. In this review, we summarize what is known about midkine in the vertebrate retina, focusing largely on recent studies utilizing the zebrafish (Danio rerio) as an animal model. Zebrafish are a valuable animal model for studying the retina, due to its very rapid development and amazing ability for functional neuronal regeneration following neuronal cell death. The zebrafish genome harbours two midkine paralogues, midkine-a (mdka) and midkine-b (mdkb), which, during development, are expressed in nested patterns among different cell types. mdka is expressed in the retinal progenitors and mdkb is expressed in newly post-mitotic cells. Interestingly, studies of loss-and gain-of-function in zebrafish larvae indicate that midkine-a regulates cell cycle kinetics. Moreover, both mdka and mdkb are expressed in different cell types in the normal adult zebrafish retina, but after light-induced death of photoreceptors, both are up-regulated and expressed in proliferating Müller glia and photoreceptor progenitors, suggesting an important and (perhaps) coincident role for these cytokines during stem cell-based neuronal regeneration. Based on its known role in other tissues and the expression and function of the midkine paralogues in the zebrafish retina, we propose that midkine has an important functional role both during development and regeneration in the retina. Further studies are needed to understand this role and the mechanisms that underlie it. Linked Articles This article is part of a themed section on Midkine. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171.issue-4 PMID:24460673
Martínez-Cañada, Pablo; Morillas, Christian; Pino, Begoña; Ros, Eduardo; Pelayo, Francisco
Computational simulations of the retina have led to valuable insights about the biophysics of its neuronal activity and processing principles. A great number of retina models have been proposed to reproduce the behavioral diversity of the different visual processing pathways. While many of these models share common computational stages, previous efforts have been more focused on fitting specific retina functions rather than generalizing them beyond a particular model. Here, we define a set of computational retinal microcircuits that can be used as basic building blocks for the modeling of different retina mechanisms. To validate the hypothesis that similar processing structures may be repeatedly found in different retina functions, we implemented a series of retina models simply by combining these computational retinal microcircuits. Accuracy of the retina models for capturing neural behavior was assessed by fitting published electrophysiological recordings that characterize some of the best-known phenomena observed in the retina: adaptation to the mean light intensity and temporal contrast, and differential motion sensitivity. The retinal microcircuits are part of a new software platform for efficient computational retina modeling from single-cell to large-scale levels. It includes an interface with spiking neural networks that allows simulation of the spiking response of ganglion cells and integration with models of higher visual areas.
Heimann, H.; Bornfeld, N.; Vij, O.; Coupland, S.; Bechrakis, N.; Kellner, U.; Foerster, M.
BACKGROUND—Vasoproliferative tumours of the retina (VPTR) are benign tumours of unknown origin, occurring mostly in otherwise healthy patients. VPTR may be associated with other chorioretinal diseases, such as uveitis. The tumours, which histologically represent reactive gliovascular proliferations, are characterised by a pink to yellow appearance on funduscopy and are accompanied by exudative and haemorrhagic changes of the retina. METHODS—22 cases of VPTR in 21 patients were examined with a follow up period between 1 month and 6 years. Ophthalmological changes associated with VPTR were intraretinal and subretinal exudations (n=18), exudative detachments of the surrounding sensory retina (n=13), intraretinal and subretinal haemorrhages (n=10), exudative changes within the macula (n=10), hyperpigmentation of the retinal pigment epithelium at the border of the exudative retinal changes (n=9), and vitreous haemorrhages (n=4). Tumour biopsy was performed in two cases. Treatment consisted of plaque radiotherapy (n=14), plaque radiotherapy and cryotherapy (two), cryotherapy only (two), observation (three), and enucleation in one case of a blind and painful eye. RESULTS—Regression of the tumour and the associated exudative changes could be observed in all treated cases. Visual acuity at last follow up improved two lines or more in two cases, remained within two lines of the initial visual acuity in 15 cases, and worsened in the remaining five. Histopathological examination of the biopsy specimens and the tumour of the enucleated eye showed massive capillary proliferation with perivascular spindle-shaped glial cells of retinal origin. CONCLUSION—The correct diagnosis of VPTR is of importance as these lesions may lead to visual loss. Further, VPTR must be differentiated from angiomas associated with von Hippel-Lindau disease as well as from ocular and systemic malignancies. Regression of tumour thickness and associated retinal changes can be achieved with
Xia, Xiaohuan; Ahmad, Iqbal
Müller glia (MG) are the primary support cells in the vertebrate retina, regulating homeostasis in one of the most metabolically active tissues. In lower vertebrates such as fish, they respond to injury by proliferating and reprogramming to regenerate retinal neurons. In mammals, MG may also react to injury by proliferating, but they fail to initiate regeneration. The barriers to regeneration could be intrinsic to mammalian MG or the function of the niche that cannot support the MG reprogramming required for lineage conversion or both. Understanding these mechanisms in light of those being discovered in fish may lead to the formulation of strategies to unlock the neurogenic potential of MG and restore regeneration in the mammalian retina. PMID:27572710
The effort to isolate and maintain alive in vitro an intact mammalian eye is rewarded by the full control provided over the arterial input and exclusion of systemic regulatory or compensatory mechanisms. Electrical recording of typical light-evoked field potentials from retina and optic nerve can be complemented by single-cell recording. Thus, light-induced electrical activity reflects the function of the retinal pigment epithelium, of the layers of the retina and of the ganglion cells or their axons. Retinal function in vitro is documented by electrophysiological and morphological methods revealing subtle features of retinal information processing as well as optic nerve signals that approach-at threshold stimulus intensity-the human psychophysical threshold. Such sensitivity of third-order retinal neurons is described for the first time. This well controlled in vitro preparation has been used successfully for biophysical, metabolic and pharmacological studies. Examples are provided that demonstrate the marked sensibility of the rod system to changes in glucose supply. Moreover, histochemical identification of glycogen stores revealed labeling of the second- and third-order neurons subserving the rod system, in addition to labeling of Müller (glial) cells in the cat retina. The glycogen content of the cat retina is augmented by prolonged anesthesia, largely depleted by ischemia after enucleation and enhanced by insulin. Pharmacological experiments using agonists and antagonists of putative retinal neurotransmitters are summarized and outlined using the muscarinic cholinergic agonist QNB as an example. Actions and uptake of the neuromodulator adenosine are presented in detail, including inhibitory effects on physiologically characterized ganglion cells. Neuronal effects of adenosine are distinguished from those resulting from vasodilatation and from glycogenolysis induced by the neuromodulator. To open the blood-retina barrier, a hyperosmotic challenge can be
Su, Y.Y.; Fry, K.R.; Lam, D.M.; Watt, C.B.
Enkephalin-like immunoreactive amacrine cells were visualized using the highly sensitive avidin-biotin method. The somas of these cells were situated in the inner nuclear and ganglion cell layers. Enkephalin-stained processes were observed in layers 1, 3, and 5 of the inner plexiform layer. The biosynthesis of sulfur-containing compounds in the goldfish retina was studied by means of a pulse-chase incubation with /sup 35/S-methionine. A /sup 35/S-labeled compound, which comigrated with authentic Met5-enkephalin on high-performance liquid chromatography (HPLC), was synthesized and was bound competitively by antibodies to enkephalin and by opiate receptors. This compound was tentatively identified as Met5-enkephalin. The newly synthesized /sup 35/S-Met5-enkephalin was released upon depolarization of the retina with a high K+ concentration. This K+-stimulated release was greatly suppressed by 5 mM Co/sup 2 +/, suggesting that the release was Ca/sup 2 +/ dependent. Using a double-label technique, enkephalin immunoreactivity and gamma-aminobutyric acid (GABA) uptake were colocalized to some amacrine cells, whereas others labeled only for enkephalin or GABA. The possible significance of enkephalin-GABA interactions is also discussed.
Masland, Richard H.
Examines research related to the retina's coding of visual input with emphasis on the organization of two kinds of ganglion cell receptive fields. Reviews current techniques for examining the shapes and arrangement in the retina of entire populations of nerve cells. (ML)
Hutchins, J.B.; Hollyfield, J.G.
Evidence for a population of acetylcholine (ACh) receptors in the human retina is presented. The authors have used the irreversible ligand TH-propylbenzilylcholine mustard (TH-PrBCM) to label muscarinic receptors. TH- or SVI-alpha-bungarotoxin (alpha-BTx) was used to label putative nicotinic receptors. Muscarinic receptors are apparently present in the inner plexiform layer of the retina. Autoradiographic grain densities are reduced in the presence of saturating concentrations of atropine, quinuclidinyl benzilate or scopolamine; this indicates that TH-PrBCM binding is specific for a population of muscarinic receptors in the human retina. Binding sites for radiolabeled alpha-BTx are found predominantly in the inner plexiform layer of the retina. Grain densities are reduced in the presence of d-tubocurarine, indicating that alpha-BTx may bind to a pharmacologically relevant nicotinic ACh receptor. This study provides evidence for cholinergic neurotransmission in the human retina.
Zhang, Xian-Shi; Gao, Shao-Bing; Li, Chao-Yi; Li, Yong-Jie
The mammalian retina seems far smarter than scientists have believed so far. Inspired by the visual processing mechanisms in the retina, from the layer of photoreceptors to the layer of retinal ganglion cells (RGCs), we propose a computational model for haze removal from a single input image, which is an important issue in the field of image enhancement. In particular, the bipolar cells serve to roughly remove the low-frequency of haze, and the amacrine cells modulate the output of cone bipolar cells to compensate the loss of details by increasing the image contrast. Then the RGCs with disinhibitory receptive field surround refine the local haze removal as well as the image detail enhancement. Results on a variety of real-world and synthetic hazy images show that the proposed model yields results comparative to or even better than the state-of-the-art methods, having the advantage of simultaneous dehazing and enhancing of single hazy image with simple and straightforward implementation. PMID:26733857
Sia, Paul Ikgan; Luiten, André N; Stace, Thomas M; Wood, John Pm; Casson, Robert J
The emerging field of quantum biology has led to a greater understanding of biological processes at the microscopic level. There is recent evidence to suggest that non-trivial quantum features such as entanglement, tunnelling and coherence have evolved in living systems. These quantum features are particularly evident in supersensitive light-harvesting systems such as in photosynthesis and photoreceptors. A biomimetic strategy utilizing biological quantum phenomena might allow new advances in the field of quantum engineering, particularly in quantum information systems. In addition, a better understanding of quantum biological features may lead to novel medical diagnostic and therapeutic developments. In the present review, we discuss the role of quantum physics in biological systems with an emphasis on the retina.
La Vail, M.M.; Rapaport, D.H.; Rakic, P. )
Time of cell origin in the retina of the rhesus monkey (Macaca mulatta) was studied by plotting the number of heavily radiolabeled nuclei in autoradiograms prepared from 2- to 6-month-old animals, each of which was exposed to a pulse of 3H-thymidine (3H-TdR) on a single embryonic (E) or postnatal (P) day. Cell birth in the monkey retina begins just after E27, and approximately 96% of cells are generated by E120. The remaining cells are produced during the last (approximately 45) prenatal days and into the first several weeks after birth. Cell genesis begins near the fovea, and proceeds towards the periphery. Cell division largely ceases in the foveal and perifoveal regions by E56. Despite extensive overlap, a class-specific sequence of cell birth was observed. Ganglion and horizontal cells, which are born first, have largely congruent periods of cell genesis with the peak between E38 and E43, and termination around E70. The first labeled cones were apparent by E33, and their highest density was achieved between E43 and E56, tapering to low values at E70, although some cones are generated in the far periphery as late as E110. Amacrine cells are next in the cell birth sequence and begin genesis at E43, reach a peak production between E56 and E85, and cease by E110. Bipolar cell birth begins at the same time as amacrines, but appears to be separate from them temporally since their production reaches a peak between E56 and E102, and persists beyond the day of birth. Mueller cells and rod photoreceptors, which begin to be generated at E45, achieve a peak, and decrease in density at the same time as bipolar cells, but continue genesis at low density on the day of birth. Thus, bipolar, Mueller, and rod cells have a similar time of origin.
Coffe, Víctor; Carbajal, Raymundo C; Salceda, Rocío
It has been reported that glycogen levels in retina vary with retinal vascularization. However, the electrical activity of isolated retina depends on glucose supply, suggesting that it does not contain energetic reserves. We determined glycogen levels and pyruvate and lactate production under various conditions in isolated retina. Ex vivo retinas from light- and dark-adapted rats showed values of 44 +/- 0.3 and 19.5 +/- 0.4 nmol glucosyl residues/mg protein, respectively. The glycogen content of retinas from light-adapted animals was reduced by 50% when they were transferred to darkness. Glycogen levels were low in retinas incubated in glucose-free media and increased in the presence of glucose. The highest glycogen values were found in media containing 20 mm of glucose. A rapid increase in lactate production was observed in the presence of glucose. Surprisingly, glycogen levels were the lowest and lactate production was also very low in the presence of 30 mm glucose. Our results suggest that glycogen can be used as an immediate accessible energy reserve in retina. We speculate on the possibility that gluconeogenesis may play a protective role by removal of lactic acid.
Khavinson, V Kh; Proniaeva, V E; Lin'kova, N S; Trofimova, S V; Umnpv, R S
Peptide's bioregulators promotes restoration of the physiological activity of the retina in retinitis pigmentosa in older adults and in animal models. The molecular mechanism of physiological activity of peptides is connected with its ability to regulate synthesis of protein markers of differentiation of neurons and retinal pigment epithelium epigenetically.
Gao, Feng-Juan; Zhang, Sheng-Hai; Li, Ting-Ting; Wu, Ji-Hong; Wu, Qiang
Mesencephalic astrocyte-derived neurotrophic factor (MANF), otherwise named Arginine-Rich, Mutated in Early-stage Tumors (ARMET), is a secretory endoplasmic reticulum stress (ERS) protein that is widely expressed in mammalian tissues. To date, little is known about the distribution and expression of MANF in the retina and optic nerve (ON). Therefore, we studied the expression and distribution of MANF in the ON and retina by real-time PCR, immunofluorescence staining and western blotting. Results from rat and mouse were highly consistent in the retina. MANF was detected in both tissues in rat, wherein it was principally localized to the ganglion cell layer (GCL), followed by the inner nuclear layer (INL). The MANF protein levels in the rat retina were 3.33-fold higher than in the rat ON. Additionally, MANF was robustly expressed by retinal ganglion cells (RGCs) in the human retina. In human ON, MANF was partially co-localized with glial fibrillary acidic protein (GFAP), suggesting that it was not restricted to astrocytes. In vitro studies confirmed that MANF could be robustly expressed in RGCs and was found principally within the cytoplasm. Hypoxia can stimulate up-regulation by of MANF expression over time, suggesting that MANF may play a vital role in the functional regulation of RGCs both in health and disease. We believe that the present study improves our understanding of the distribution and expression of MANF in the retina and ON and could help in further analysis of its interact and correlate with the relevant ophthalmic diseases. PMID:28154531
Gonzalez-Nunez, Veronica; Nocco, Valentina; Budd, Aidan
There is a clear need to develop novel tools to help improve our understanding of stem cell biology, and potentially also the utility of stem cells in regenerative medicine. We report the cloning, functional, and bioinformatic characterization of a novel stem cell marker in the zebrafish retina, drCol 15a1b. The expression pattern of drCol 15a1b is restricted to stem cell niches located in the central nervous system, whereas other collagen XVs are associated with muscle and endothelial tissues. Knocking down drCol 15a1b expression causes smaller eyes, ear defects, and brain edema. Microscopic analysis reveals enhanced proliferation in the morphant eye, with many mitotic nuclei located in the central retina, together with a delayed differentiation of the mature retinal cell types. Besides, several markers known to be expressed in the ciliary marginal zone display broader expression areas in morpholino-injected embryos, suggesting an anomalous diffusion of signaling effectors from the sonic hedgehog and notch pathways. These results indicate that drCol 15a1b is a novel stem cell marker in the central nervous system that has a key role in homing stem cells into specialized niches in the adult organism. Moreover, mutations in the hCol 18a1 gene are responsible for the Knobloch syndrome, which affects brain and retinal structures, suggesting that drCol 15a1b may function similarly to mammalian Col 18a1. Thus, our results shed new light on the signaling pathways that underlie the maintenance of stem cells in the adult organism while helping us to understand the role of extracellular matrix proteins in modulating the signals that determine stem cell differentiation, cell cycle exit and apoptosis.
Berta, Agnes I; Kiss, Anna L; Lukáts, Akos; Szabó, Arnold; Szél, Agoston
The distribution of caveolin isoforms was previously evaluated in the retinas of different species, but has not yet been described in the primate retina. In this study, the distribution of caveolins was assessed via immunochemistry using isoform-specific antibodies in the retina of the black-and-white ruffed lemur. Here, we report the presence of a variety of caveolin isoforms in many layers of the lemur retina. As normal human retinas were not available for research and the retinas of primates are fairly similar to those of humans, the lemur retina can be utilized as a model for caveolin distribution in normal humans.
Du, Lucia Y.; Chang, Lily Y-L.; Ardiles, Alvaro O.; Tapia-Rojas, Cheril; Araya, Joaquin; Inestrosa, Nibaldo C.
New studies show that the retina also undergoes pathological changes during the development of Alzheimer’s disease (AD). While transgenic mouse models used in these previous studies have offered insight into this phenomenon, they do not model human sporadic AD, which is the most common form. Recently, the Octodon degus has been established as a sporadic model of AD. Degus display age-related cognitive impairment associated with Aβ aggregates and phosphorylated tau in the brain. Our aim for this study was to examine the expression of AD-related proteins in young, adult and old degus retina using enzyme-linked or fluorescence immunohistochemistry and to quantify the expression using slot blot and western blot assays. Aβ4G8 and Aβ6E10 detected Aβ peptides in some of the young animals but the expression was higher in the adults. Aβ peptides were observed in the inner and outer segment of the photoreceptors, the nerve fiber layer (NFL) and ganglion cell layer (GCL). Expression was higher in the central retinal region than in the retinal periphery. Using an anti-oligomer antibody we detected Aβ oligomer expression in the young, adult and old retina. Immunohistochemical labeling showed small discrete labeling of oligomers in the GCL that did not resemble plaques. Congo red staining did not result in green birefringence in any of the animals analyzed except for one old (84 months) animal. We also investigated expression of tau and phosphorylated tau. Expression was seen at all ages studied and in adults it was more consistently observed in the NFL-GCL. Hyperphosphorylated tau detected with AT8 antibody was significantly higher in the adult retina and it was localized to the GCL. We confirm for the first time that Aβ peptides and phosphorylated tau are expressed in the retina of degus. This is consistent with the proposal that AD biomarkers are present in the eye. PMID:26267479
Theurl, Milan; Song, Delu; Clark, Esther; Sterling, Jacob; Grieco, Steve; Altamura, Sandro; Galy, Bruno; Hentze, Matthias; Muckenthaler, Martina U; Dunaief, Joshua L
Because ferroportin (Fpn) is the only known mammalian cellular iron exporter, understanding its localization and regulation within the retina would shed light on the direction of retinal iron flux. The hormone hepcidin may regulate retinal Fpn, as it triggers Fpn degradation in the gut. Immunofluorescence was used to label Fpn in retinas of mice with 4 different genotypes (wild type; Fpn C326S, a hepcidin-resistant Fpn; hepcidin knockout; and ceruloplasmin/hephaestin double knockout). No significant difference in Fpn levels was observed in these retinas. Fpn localized to the abluminal side of the outer plexiform vascular endothelial cells, Müller glia cells, and the basolateral side of the retinal pigment epithelium. Adeno-associated virus (AAV)-hepcidin was injected into the eyes of hepcidin knockout mice, while AAV-lacZ was injected into the contralateral eyes as a control. AAV-hepcidin injected eyes had increased ferritin immunolabeling in retinal vascular endothelial cells. Fpn C326S mice had systemic iron overload compared to wild type and had the fastest retinal iron accumulation of any hereditary model studied to date. The results suggest that physiologic hepcidin levels are insufficient to alter Fpn levels within the retinal pigment epithelium and Müller cells, but may limit iron transport into the retina from vascular endothelial cells.
Renna, Jordan M; Chellappa, Deepa K; Ross, Christopher L; Stabio, Maureen E; Berson, David M
Melanopsin ganglion cells express the photopigment melanopsin and are the first functional photoreceptors to develop in the mammalian retina. They have been shown to play a variety of important roles in visual development and behavior in the early postnatal period (Johnson et al., 2010; Kirkby and Feller, 2013; Rao et al., 2013; Renna et al., 2011). Here, we probed the maturation of the dendritic arbors of melanopsin ganglion cells during this developmental period in mice. We found that some melanopsin ganglion cells (mainly the M1-subtype) transiently extend their dendrites not only into the inner plexiform layer (where they receive synaptic inputs from bipolar and amacrine cells) but also into the outer plexiform layer, where in mature retina, rod and cone photoreceptors are thought to contact only bipolar and horizontal cells. Thus, some immature melanopsin ganglion cells are biplexiform. This feature is much less common although still present in the mature retina. It reaches peak incidence 8-12 days after birth, before the eyes open and bipolar cells are sufficiently mature to link rods and cones to ganglion cells. At this age, some outer dendrites of melanopsin ganglion cells lie in close apposition to the axon terminals of cone photoreceptors and express a postsynaptic marker of glutamatergic transmission, postsynaptic density-95 protein (PSD-95). These findings raise the possibility of direct, monosynaptic connections between cones and melanopsin ganglion cells in the early postnatal retina. We provide a detailed description of the developmental profile of these processes and consider their possible functional and evolutionary significance.
Deshmukh, Nikhil Rajiv
Elucidating the general principles of computation in neural circuits is a difficult problem requiring both a tractable model circuit as well as sophisticated measurement tools. This thesis advances our understanding of complex computation in the salamander retina and its underlying circuitry and furthers the development of advanced tools to enable detailed study of neural circuits. The retina provides an ideal model system for neural circuits in general because it is capable of producing complex representations of the visual scene, and both its inputs and outputs are accessible to the experimenter. Chapter 2 describes the biophysical mechanisms that give rise to the omitted stimulus response in retinal ganglion cells described in Schwartz et al., (2007) and Schwartz and Berry, (2008). The extra response to omitted flashes is generated at the input to bipolar cells, and is separable from the characteristic latency shift of the OSR apparent in ganglion cells, which must occur downstream in the circuit. Chapter 3 characterizes the nonlinearities at the first synapse of the ON pathway in response to high contrast flashes and develops a phenomenological model that captures the effect of synaptic activation and intracellular signaling dynamics on flash responses. This work is the first attempt to model the dynamics of the poorly characterized mGluR6 transduction cascade unique to ON bipolar cells, and explains the second lobe of the biphasic flash response. Complementary to the study of neural circuits, recent advances in wafer-scale photolithography have made possible new devices to measure the electrical and mechanical properties of neurons. Chapter 4 reports a novel piezoelectric sensor that facilitates the simultaneous measurement of electrical and mechanical signals in neural tissue. This technology could reveal the relationship between the electrical activity of neurons and their local mechanical environment, which is critical to the study of mechanoreceptors
Williams, David R.
A quarter century ago, we were limited to a macroscopic view of the retina inside the living eye. Since then, new imaging technologies, including confocal scanning laser ophthalmoscopy, optical coherence tomography, and adaptive optics fundus imaging, transformed the eye into a microscope in which individual cells can now be resolved noninvasively. These technologies have enabled a wide range of studies of the retina that were previously impossible. PMID:21596053
Tu, Hung-Ya; Hsu, Chih-Chun; Chen, Yu-Jiun
SHORT ABSTRACT This protocol demonstrates how to perform whole-cell patch clamp recording on retinal neurons from a flat-mount preparation. LONG ABSTRACT The mammalian retina is a layered tissue composed of multiple neuronal types. To understand how visual signals are processed within its intricate synaptic network, electrophysiological recordings are frequently used to study connections among individual neurons. We have optimized a flat-mount preparation for patch clamp recording of genetically marked neurons in both GCL (ganglion cell layer) and INL (inner nuclear layer) of mouse retinas. Recording INL neurons in flat-mounts is favored over slices because both vertical and lateral connections are preserved in the former configuration, allowing retinal circuits with large lateral components to be studied. We have used this procedure to compare responses of mirror-partnered neurons in retinas such as the cholinergic starburst amacrine cells (SACs). PMID:27768050
Yang, Y S; Hanke, J H; Carayannopoulos, L; Craft, C M; Capra, J D; Tucker, P W
We have cloned the ubiquitous form of an octamer-binding, 60-kDa protein (NonO) that appears to be the mammalian equivalent of the Drosophila visual and courtship song behavior protein, no-on-transient A/dissonance (nonAdiss). A region unprecedently rich in aromatic amino acids containing two ribonuclear protein binding motifs is highly conserved between the two proteins. A ubiquitous form of NonO is present in all adult tissues, whereas lymphocytes and retina express unique forms of NonO mRNA. The ubiquitous form contains a potential helix-turn-helix motif followed by a highly charged region but differs from prototypic octamer-binding factors by lacking the POU DNA-binding domain. In addition to its conventional octamer duplex-binding, NonO binds single-stranded DNA and RNA at a site independent of the duplex site.
Sierra, Ana; Navascués, Julio; Cuadros, Miguel A; Calvente, Ruth; Martín-Oliva, David; Ferrer-Martín, Rosa M; Martín-Estebané, María; Carrasco, María-Carmen; Marín-Teva, José L
Inducible nitric oxide synthase (iNOS), which produce large amounts of nitric oxide (NO), is induced in macrophages and microglia in response to inflammatory mediators such as LPS and cytokines. Although iNOS is mainly expressed by microglia that become activated in different pathological and experimental situations, it was recently reported that undifferentiated amoeboid microglia can also express iNOS during normal development. The aim of this study was to investigate the pattern of iNOS expression in microglial cells during normal development and after their activation with LPS by using the quail retina as model. iNOS expression was analyzed by iNOS immunolabeling, western-blot, and RT-PCR. NO production was determined by using DAR-4M AM, a reliable fluorescent indicator of subcellular NO production by iNOS. Embryonic, postnatal, and adult in situ quail retinas were used to analyze the pattern of iNOS expression in microglial cells during normal development. iNOS expression and NO production in LPS-treated microglial cells were investigated by an in vitro approach based on organotypic cultures of E8 retinas, in which microglial cell behavior is similar to that of the in situ retina, as previously demonstrated in our laboratory. We show here that amoeboid microglia in the quail retina express iNOS during normal development. This expression is stronger in microglial cells migrating tangentially in the vitreal part of the retina and is downregulated, albeit maintained, when microglia differentiate and become ramified. LPS treatment of retina explants also induces changes in the morphology of amoeboid microglia compatible with their activation, increasing their lysosomal compartment and upregulating iNOS expression with a concomitant production of NO. Taken together, our findings demonstrate that immature microglial cells express iNOS during normal development, suggesting a certain degree of activation. Furthermore, LPS treatment induces overactivation of amoeboid
Miloudi, Khalil; Dejda, Agnieszka; Binet, François; Lapalme, Eric; Cerani, Agustin; Sapieha, Przemyslaw
The rodent retina is perhaps the most accessible mammalian system in which to investigate neurovascular interplay within the central nervous system (CNS). It is increasingly being recognized that several neurodegenerative diseases such as Alzheimer's, multiple sclerosis, and amyotrophic lateral sclerosis present elements of vascular compromise. In addition, the most prominent causes of blindness in pediatric and working age populations (retinopathy of prematurity and diabetic retinopathy, respectively) are characterized by vascular degeneration and failure of physiological vascular regrowth. The aim of this technical paper is to provide a detailed protocol to study CNS vascular regeneration in the retina. The method can be employed to elucidate molecular mechanisms that lead to failure of vascular growth after ischemic injury. In addition, potential therapeutic modalities to accelerate and restore healthy vascular plexuses can be explored. Findings obtained using the described approach may provide therapeutic avenues for ischemic retinopathies such as that of diabetes or prematurity and possibly benefit other vascular disorders of the CNS.
Zlateski, Aleksandar; Lee, Kisuk; Richardson, Mark; Turaga, Srinivas C.; Purcaro, Michael; Balkam, Matthew; Robinson, Amy; Behabadi, Bardia F.; Campos, Michael; Denk, Winfried; Seung, H. Sebastian
How does the mammalian retina detect motion? This classic problem in visual neuroscience has remained unsolved for 50 years. In search of clues, we reconstructed Off-type starburst amacrine cells (SACs) and bipolar cells (BCs) in serial electron microscopic images with help from EyeWire, an online community of “citizen neuroscientists.” Based on quantitative analyses of contact area and branch depth in the retina, we found evidence that one BC type prefers to wire with a SAC dendrite near the SAC soma, while another BC type prefers to wire far from the soma. The near type is known to lag the far type in time of visual response. A mathematical model shows how such “space-time wiring specificity” could endow SAC dendrites with receptive fields that are oriented in space-time and therefore respond selectively to stimuli that move in the outward direction from the soma. PMID:24805243
Retina implants are currently being developed by several interdisciplinary research consortia worldwide for blind humans with various retinal degenerative diseases. It is the aim of our retina implant project to develop a novel type of visual prosthesis to regain a moderate amount of vision such as perception of location and shape of large objects in the first stage and to approach reading quality in a subsequent stage. In our planned retina implant, a retina encoder (RE) outside the eye has to replace the information processing of the retina. A retina stimulator (RS), implanted adjacently to the retinal ganglion cell layer, has to contact a sufficient number of retinal ganglion cells/fibers for electrical elicitation of spikes. A wireless signal and energy transmission system has to provide the communication between the RE and RS. This paper outlines the retina implant project of our consortium of 14 expert groups and describes first results of the learning RE. The RE approximates the typical receptive field (RF) properties of primate retinal ganglion cells by means of individually tunable spatiotemporal RF filters. The RE as a cluster of RF filters maps visual patterns onto spike trains for a number of contacted ganglion cells. A concept is presented to train the individual RF filters in an unsupervised learning process, which employs neural networks in a dialog with the individual human subject. The desired aim of this dialog is an optimization of the visual perception by matching the various RF filter properties with those 'expected' by the central visual system for each contacted ganglion cell.
Zanello, Susana B.; Wotring, V.; Theriot, C.; Ploutz-Snyder, R.; Zhang, Y.; Wu, H.
Exposure to cosmic radiation implies a risk of tissue degeneration. Radiation retinopathy is a complication of radiotherapy and exhibits common features with other retinopathies and neuropathies. Exposure to a low radiation dose elicits protective cellular events (radioadaptive response), reducing the stress of a subsequent higher dose. To assess the risk of radiation-induced retinal changes and the extent to which a small priming dose reduces this risk, we used a mouse model exposed to a source of Cs-137-gamma radiation. Gene expression profiling of retinas from non-irradiated control C57BL/6J mice (C) were compared to retinas from mice treated with a low 50 mGy dose (LD), a high 6 Gy dose (HD), and a combined treatment of 50 mGy (priming) and 6 Gy (challenge) doses (LHD). Whole retina RNA was isolated and expression analysis for selected genes performed by RTqPCR. Relevant target genes associated with cell death/survival, oxidative stress, cellular stress response and inflammation pathways, were analyzed. Cellular stress response genes were upregulated at 4 hr after the challenge dose in LHD retinas (Sirt1: 1.5 fold, Hsf1: 1.7 fold, Hspa1a: 2.5 fold; Hif1a: 1.8 fold, Bag1: 1.7). A similar trend was observed in LD animals. Most antioxidant enzymes (Hmox1, Sod2, Prdx1, Cygb, Cat1) and inflammatory mediators (NF B, Ptgs2 and Tgfb1) were upregulated in LHD and LD retinas. Expression of the pro-survival gene Bcl2 was upregulated in LD (6-fold) and LHD (4-fold) retinas. In conclusion, cytoprotective gene networks activation in the retina suggests a radioadaptive response to a priming irradiation dose, with mitigation of the deleterious effects of a subsequent high dose exposure. The enhancement of these cytoprotective mechanisms has potential value as a countermeasure to ocular alterations caused by radiation alone or in combination with other factors in spaceflight environments.
Singhal, Shweta; Lawrence, Jean M; Bhatia, Bhairavi; Ellis, James S; Kwan, Anthony S; Macneil, Angus; Luthert, Philip J; Fawcett, James W; Perez, Maria-Thereza; Khaw, Peng T; Limb, G Astrid
At present, there are severe limitations to the successful migration and integration of stem cells transplanted into the degenerated retina to restore visual function. This study investigated the potential role of chondroitin sulfate proteoglycans (CSPGs) and microglia in the migration of human Müller glia with neural stem cell characteristics following subretinal injection into the Lister hooded (LH) and Royal College of Surgeons (RCS) rat retinae. Neonate LH rat retina showed minimal baseline microglial accumulation (CD68-positive cells) that increased significantly 2 weeks after transplantation (p < .001), particularly in the ganglion cell layer (GCL) and inner plexiform layer. In contrast, nontransplanted 5-week-old RCS rat retina showed considerable baseline microglial accumulation in the outer nuclear layer (ONL) and photoreceptor outer segment debris zone (DZ) that further increased (p < .05) throughout the retina 2 weeks after transplantation. Marked deposition of the N-terminal fragment of CSPGs, as well as neurocan and versican, was observed in the DZ of 5-week-old RCS rat retinae, which contrasted with the limited expression of these proteins in the GCL of the adult and neonate LH rat retinae. Staining for CSPGs and CD68 revealed colocalization of these two molecules in cells infiltrating the ONL and DZ of the degenerating RCS rat retina. Enhanced immune suppression with oral prednisolone and intraperitoneal injections of indomethacin caused a reduction in the number of microglia but did not facilitate Müller stem cell migration. However, injection of cells with chondroitinase ABC combined with enhanced immune suppression caused a dramatic increase in the migration of Müller stem cells into all the retinal cell layers. These observations suggest that both microglia and CSPGs constitute a barrier for stem cell migration following transplantation into experimental models of retinal degeneration and that control of matrix deposition and the innate
Morrow, Eric M; Chen, C-M Amy; Cepko, Constance L
Background Retinal bipolar cells comprise a diverse group of neurons. Cone bipolar cells and rod bipolar cells are so named for their connections with cone and rod photoreceptors, respectively. Morphological criteria have been established that distinguish nine types of cone bipolar cells and one type of rod bipolar cell in mouse and rat. While anatomical and physiological aspects of bipolar types have been actively studied, little is known about the sequence of events that leads to bipolar cell type specification and the potential relationship this process may have with synapse formation in the outer plexiform layer. In this study, we have examined the birth order of rod and cone bipolar cells in the developing mouse and rat in vivo. Results Using retroviral lineage analysis with the histochemical marker alkaline phosphatase, the percentage of cone and rod bipolar cells born on postnatal day 0 (P0), P4, and P6 were determined, based upon the well characterized morphology of these cells in the adult rat retina. In this in vivo experiment, we have demonstrated that cone bipolar genesis clearly precedes rod bipolar genesis. In addition, in the postnatal mouse retina, using a combination of tritiated-thymidine birthdating and immunohistochemistry to distinguish bipolar types, we have similarly found that cone bipolar genesis precedes rod bipolar genesis. The tritiated-thymidine birthdating studies also included quantification of the birth of all postnatally generated retinal cell types in the mouse. Conclusion Using two independent in vivo methodologies in rat and mouse retina, we have demonstrated that there are distinct waves of genesis of the two major bipolar cell types, with cone bipolar genesis preceding rod bipolar genesis. These waves of bipolar genesis correspond to the order of genesis of the presynaptic photoreceptor cell types. PMID:18215319
Sibille, Estelle; Berdeaux, Olivier; Martine, Lucy; Bron, Alain M.; Creuzot-Garcher, Catherine P.; He, Zhiguo; Thuret, Gilles; Bretillon, Lionel; Masson, Elodie A. Y.
Gangliosides make a wide family of glycosphingolipids, highly heterogeneous in both the ceramide moiety and the oligosaccharide chain. While ubiquitously expressed in mammalian tissues, they are particularly abundant in the brain and the peripheral nervous system. Gangliosides are known to play a crucial role in the development, maintenance and functional integrity of the nervous system. However, the expression and roles of gangliosides in the retina, although often considered as a window on the brain, has been far less studied. We performed an in-depth analysis of gangliosides of the human retina, especially using powerful LC/MS methods. We compared the pattern of ganglioside classes and ceramide molecular species of this tissue with other ocular structures and with brain and plasma in elderly human individuals. About a hundred of ganglioside molecular species among 15 distinct classes were detected illustrating the huge structural diversity of these compounds. The retina exhibited a very diverse ganglioside profile and shared several common features with the brain (prominence of tetraosylgangliosides, abundance of d20:1 long chain base and 18:0 fatty acid…). However, the retina stood out with the specific expression of GD3, GT3 and AcGT3, which further presented a peculiar molecular species distribution. The unique ganglioside pattern we observed in the human retina suggests that these ganglioside species play a specific role in the structure and function of this tissue. This lipidomic study, by highlighting retina specific ganglioside species, opens up novel research directions for a better understanding of the biological role of gangliosides in the retina. PMID:27997589
Zieger, Marina; Ahnelt, Peter K.; Uhrin, Pavel
We aimed to investigate fractalkine (CX3CL1) protein expression in wild type (wt) retina and its alterations during retinal degeneration in mouse model (rd10) of retinitis pigmentosa. Forms of retinal protein CX3CL1, total protein and mRNA levels of CX3CL1 were analyzed at postnatal days (P) 5, 10, 14, 22, 30, 45, and 60 by Western blotting and real-time PCR. Cellular sources of CX3CL1 were investigated by in situ hybridization histochemistry (ISH) and using transgenic (CX3CL1cherry) mice. The immunoblots revealed that in both, wt and rd10 retinas, a membrane integrated ∼100 kDa CX3CL1 form and a cleaved ∼85 kDa CX3CL1 form were present at P5. At P10, accumulation of another presumably intra-neuronal ∼95 kDa form and a decrease in the ∼85-kDa form were observed. From P14, a ∼95 kDa form became principal in wt retina, while in rd10 retinas a soluble ∼85 kDa form increased at P45 and P60. In comparison, retinas of rd10 mice had significantly lower levels of total CX3CL1 protein (from P10 onwards) and lower CX3CL1 mRNA levels (from P14), even before the onset of primary rod degeneration. ISH and mCherry reporter fluorescence showed neurons in the inner retina layers as principal sites of CX3CL1 synthesis both in wt and rd10 retinas. In conclusion, our results demonstrate that CX3CL1 has a distinctive course of expression and functional regulation in rd10 retina starting at P10. The biological activity of CX3CL1 is regulated by conversion of a membrane integrated to a soluble form during neurogenesis and in response to pathologic changes in the adult retinal milieu. Viable mature neurons in the inner retina likely exhibit a dynamic intracellular storage depot of CX3CL1. PMID:25191897
Lange, Christina; Caprara, Christian; Tanimoto, Naoyuki; Beck, Susanne; Huber, Gesine; Samardzija, Marijana; Seeliger, Mathias; Grimm, Christian
Loss of vision and blindness in human patients is often caused by the degeneration of neuronal cells in the retina. In mouse models, photoreceptors can be protected from death by hypoxic preconditioning. Preconditioning in low oxygen stabilizes and activates hypoxia inducible transcription factors (HIFs), which play a major role in the hypoxic response of tissues including the retina. We show that a tissue-specific knockdown of von Hippel-Lindau protein (VHL) activated HIF transcription factors in normoxic conditions in the retina. Sustained activation of HIF1 and HIF2 was accompanied by persisting embryonic vasculatures in the posterior eye and the iris. Embryonic vessels persisted into adulthood and led to a severely abnormal mature vessel system with vessels penetrating the photoreceptor layer in adult mice. The sustained hypoxia-like response also activated the leukemia inhibitory factor (LIF)-controlled endogenous molecular cell survival pathway. However, this was not sufficient to protect the retina against massive cell death in all retinal layers of adult mice. Caspases 1, 3 and 8 were upregulated during the degeneration as were several VHL target genes connected to the extracellular matrix. Misregulation of these genes may influence retinal structure and may therefore facilitate growth of vessels into the photoreceptor layer. Thus, an early and sustained activation of a hypoxia-like response in retinal cells leads to abnormal vasculature and severe retinal degeneration in the adult mouse retina.
Duong, Timothy Q.; Muir, Eric R.
This paper reviews recent developments in high-resolution magnetic resonance imaging (MRI) and its application to image anatomy, physiology, and function in the retina of animals. It describes technical issues and solutions in performing retinal MRI, anatomical MRI, blood oxygenation level-dependent functional MRI (fMRI), and blood-flow MRI both of normal retinas and of retinal degeneration. MRI offers unique advantages over existing retinal imaging techniques, including the ability to image multiple layers without depth limitation and to provide multiple clinically relevant data in a single setting. Retinal MRI has the potential to complement existing retinal imaging techniques. PMID:19763752
Spencer, Clarellen; Pajovic, Sanja; Devlin, Hollie; Dinh, Quynh-Dao; Corson, Timothy W; Gallie, Brenda L
Although RB1 function is disrupted in the majority of human cancers, an undefined cell of developing human retina is uniquely sensitive to cancer induction when the RB1 tumor suppressor gene is lost. Murine retinoblastoma is initiated only when two of the RB family of genes, RB1 and p107 or p130, are inactivated. Although whole embryonic retina shows RB family gene expression by several techniques, when E14 developing retina was depleted of the earliest differentiating cells, ganglion cells, the remaining proliferating murine embryonic retinal progenitor cells clearly did not express RB1 or p130, while the longer splice form of p107 was expressed. Each retinal cell type expressed some member of the RB family at some stage of differentiation. Rod photoreceptors stained for the RB1 protein product, pRB, and p107 in only a brief window of postnatal murine development, with no detectable staining for any of the RB family proteins in adult human and mouse rod photoreceptors. Adult mouse and human Muller glia, ganglion and rare horizontal cells, and adult human, but not adult mouse, cone photoreceptors stained for pRB. The RB gene family is dynamically and variably expressed through retinal development in specific retinal cells.
Lagman, David; Callado-Pérez, Amalia; Franzén, Ilkin E.
Gene duplications provide raw materials that can be selected for functional adaptations by evolutionary mechanisms. We describe here the results of 350 million years of evolution of three functionally related gene families: the alpha, beta and gamma subunits of transducins, the G protein involved in vision. Early vertebrate tetraploidisations resulted in separate transducin heterotrimers: gnat1/gnb1/gngt1 for rods, and gnat2/gnb3/gngt2 for cones. The teleost-specific tetraploidisation generated additional duplicates for gnb1, gnb3 and gngt2. We report here that the duplicates have undergone several types of subfunctionalisation or neofunctionalisation in the zebrafish. We have found that gnb1a and gnb1b are co-expressed at different levels in rods; gnb3a and gnb3b have undergone compartmentalisation restricting gnb3b to the dorsal and medial retina, however, gnb3a expression was detected only at very low levels in both larvae and adult retina; gngt2b expression is restricted to the dorsal and medial retina, whereas gngt2a is expressed ventrally. This dorsoventral distinction could be an adaptation to protect the lower part of the retina from intense light damage. The ontogenetic analysis shows earlier onset of expression in the pineal complex than in the retina, in accordance with its earlier maturation. Additionally, gnb1a but not gnb1b is expressed in the pineal complex, and gnb3b and gngt2b are transiently expressed in the pineal during ontogeny, thus showing partial temporal subfunctionalisation. These retina-pineal distinctions presumably reflect their distinct functional roles in vision and circadian rhythmicity. In summary, this study describes several functional differences between transducin gene duplicates resulting from the teleost-specific tetraploidisation. PMID:25806532
Lagman, David; Callado-Pérez, Amalia; Franzén, Ilkin E; Larhammar, Dan; Abalo, Xesús M
Gene duplications provide raw materials that can be selected for functional adaptations by evolutionary mechanisms. We describe here the results of 350 million years of evolution of three functionally related gene families: the alpha, beta and gamma subunits of transducins, the G protein involved in vision. Early vertebrate tetraploidisations resulted in separate transducin heterotrimers: gnat1/gnb1/gngt1 for rods, and gnat2/gnb3/gngt2 for cones. The teleost-specific tetraploidisation generated additional duplicates for gnb1, gnb3 and gngt2. We report here that the duplicates have undergone several types of subfunctionalisation or neofunctionalisation in the zebrafish. We have found that gnb1a and gnb1b are co-expressed at different levels in rods; gnb3a and gnb3b have undergone compartmentalisation restricting gnb3b to the dorsal and medial retina, however, gnb3a expression was detected only at very low levels in both larvae and adult retina; gngt2b expression is restricted to the dorsal and medial retina, whereas gngt2a is expressed ventrally. This dorsoventral distinction could be an adaptation to protect the lower part of the retina from intense light damage. The ontogenetic analysis shows earlier onset of expression in the pineal complex than in the retina, in accordance with its earlier maturation. Additionally, gnb1a but not gnb1b is expressed in the pineal complex, and gnb3b and gngt2b are transiently expressed in the pineal during ontogeny, thus showing partial temporal subfunctionalisation. These retina-pineal distinctions presumably reflect their distinct functional roles in vision and circadian rhythmicity. In summary, this study describes several functional differences between transducin gene duplicates resulting from the teleost-specific tetraploidisation.
Tokuda, Kazuhiro; Kuramitsu, Yasuhiro; Baron, Byron; Kitagawa, Takao; Tokuda, Nobuko; Kobayashi, Masaaki; Kimura, Kazuhiro; Sonoda, Koh-Hei; Nakamura, Kazuyuki
Understanding how energy metabolism and related proteins influence neural progenitor cells in adult tissues is critical for developing new strategies in clinical tissue regeneration therapy. We have recently reported that a subtoxic concentration of glutamate-induced neural progenitor cells in the mature ex vivo rat retina. We herein explore changes in the metabolic pathways during the process. We firstly observed an increase in lactate and lactate dehydrogenase concentration in the glutamate-treated retina. We then investigated the levels of glycolytic enzymes and confirmed significant upregulation of pyruvate kinase M type (PKM), especially PKM2, enolase, phosphoglycerate mutase 1 (PGAM1), and inosine-5'-monophosphate dehydrogenase (IMPDH1) in the glutamate-treated retina compared to the untreated retina. An analysis of the subcellular localization of PKM2 revealed nuclear translocation in the treated retina, which has been reported to regulate cell cycle proliferation and glycolytic enzymes. Our findings indicate that the mature rat retina undergoes an increase in aerobic glycolysis. PKM2, both in the cytoplasm and in the nucleus, may thus play an important role during neural progenitor cell induction, as it does in other proliferating cells.
Ventura, D F; Zana, Y; de Souza, J M; DeVoe, R D
We have examined the functional architecture of the turtle Pseudemys scripta elegans retina with respect to colour processing, extending spectral stimulation into the ultraviolet, which has not been studied previously in the inner retina. We addressed two questions. (i) Is it possible to deduce the ultraviolet cone spectral sensitivity function through horizontal cell responses? (ii) Is there evidence for tetrachromatic neural mechanisms, i.e. UV/S response opponency? Using a constant response methodology we have isolated the ultraviolet cone input into the S/LM horizontal cell type and described it in fine detail. Monophasic (luminosity), biphasic L/M (red-green) and triphasic S/LM (yellow-blue) horizontal cells responded strongly to ultraviolet light. The blue-adapted spectral sensitivity function of a S/LM cell peaked in the ultraviolet and could be fitted to a porphyropsin cone template with a peak at 372 nm. In the inner retina eight different combinations of spectral opponency were found in the centre of the receptive field of ganglion cells. Among amacrine cells the only types found were UVSM-L+ and its reverse. One amacrine and four ganglion cells were also opponent in the receptive field surround. UV/S opponency, seen in three different types of ganglion cell, provides a neural basis for discrimination of ultraviolet colours. In conclusion, the results strongly suggest that there is an ultraviolet channel and a neural basis for tetrachromacy in the turtle retina.
Fulton, Anne B; Hansen, Ronald M; Moskowitz, Anne; Akula, James D
The continuing worldwide epidemic of retinopathy of prematurity (ROP), a leading cause of childhood visual impairment, strongly motivates further research into mechanisms of the disease. Although the hallmark of ROP is abnormal retinal vasculature, a growing body of evidence supports a critical role for the neural retina in the ROP disease process. The age of onset of ROP coincides with the rapid developmental increase in rod photoreceptor outer segment length and rhodopsin content of the retina with escalation of energy demands. Using a combination of non-invasive electroretinographic (ERG), psychophysical, and image analysis procedures, the neural retina and its vasculature have been studied in prematurely born human subjects, both with and without ROP, and in rats that model the key vascular and neural parameters found in human ROP subjects. These data are compared to comprehensive numeric summaries of the neural and vascular features in normally developing human and rat retina. In rats, biochemical, anatomical, and molecular biological investigations are paired with the non-invasive assessments. ROP, even if mild, primarily and persistently alters the structure and function of photoreceptors. Post-receptor neurons and retinal vasculature, which are intimately related, are also affected by ROP; conspicuous neurovascular abnormalities disappear, but subtle structural anomalies and functional deficits may persist years after clinical ROP resolves. The data from human subjects and rat models identify photoreceptor and post-receptor targets for interventions that promise improved outcomes for children at risk for ROP.
Shanmugam, P Mahesh; Ramanjulu, Rajesh
Vascular tumors of the retina and choroid can be seen occasionally. In the following article, the key clinical and diagnostic features of the major retinal and choroidal vascular tumors, their systemic associations, and the literature pertaining to the most currently available treatment strategies are reviewed.
Shanmugam, P Mahesh; Ramanjulu, Rajesh
Vascular tumors of the retina and choroid can be seen occasionally. In the following article, the key clinical and diagnostic features of the major retinal and choroidal vascular tumors, their systemic associations, and the literature pertaining to the most currently available treatment strategies are reviewed. PMID:25827544
GUO, CHENYING; STELLA, SALVATORE L.; HIRANO, ARLENE A.; BRECHA, NICHOLAS C.
Plasmalemmal and vesicular γ-aminobutyric acid (GABA) transporters influence neurotransmission by regulating high-affinity GABA uptake and GABA release into the synaptic cleft and extracellular space. Postnatal expression of the plasmalemmal GABA transporter-1 (GAT-1), GAT-3, and the vesicular GABA/glycine transporter (VGAT) were evaluated in the developing mouse retina by using immunohistochemistry with affinity-purified antibodies. Weak transporter immunoreactivity was observed in the inner retina at postnatal day 0 (P0). GAT-1 immunostaining at P0 and at older ages was in amacrine and displaced amacrine cells in the inner nuclear layer (INL) and ganglion cell layer (GCL), respectively, and in their processes in the inner plexiform layer (IPL). At P10, weak GAT-1 immunostaining was in Müller cell processes. GAT-3 immunostaining at P0 and older ages was in amacrine cells and their processes, as well as in Müller cells and their processes that extended radially across the retina. At P10, Müller cell somata were observed in the middle of the INL. VGAT immunostaining was present at P0 and older ages in amacrine cells in the INL as well as processes in the IPL. At P5, weak VGAT immunostaining was also observed in horizontal cell somata and processes. By P15, the GAT and VGAT immunostaining patterns appear similar to the adult immunostaining patterns; they reached adult levels by about P20. These findings demonstrate that GABA uptake and release are initially established in the inner retina during the first postnatal week and that these systems subsequently mature in the outer retina during the second postnatal week. PMID:18975268
Brandli, Alice; Gerhart, Jacquelyn; Sutera, Christopher K.; Purushothuman, Sivaraman; George-Weinstein, Mindy; Stone, Jonathan; Bravo-Nuevo, Arturo
Purpose To identify Myo/Nog cells in the adult retina and test their role in protecting retinal photoreceptors from light damage. Methods Light damage was induced by exposing albino rats raised in dim cyclic light to 1000 lux light for 24 hours. In one group of rats, Myo/Nog cells were purified from rat brain tissue by magnetic cell sorting following binding of the G8 monoclonal antibody (mAb). These cells were injected into the vitreous humour of the eye within 2 hours following bright light exposure. Retinal function was assessed using full-field, flash electroretinogram (ERG) before and after treatment. The numbers of Myo/Nog cells, apoptotic photoreceptors, and the expression of glial fibrillary acidic protein (GFAP) in Muller cells were assessed by immunohistochemistry. Results Myo/Nog cells were present in the undamaged retina in low numbers. Light induced damage increased their numbers, particularly in the choroid, ganglion cell layer and outer plexiform layer. Intravitreal injection of G8-positive (G8+) cells harvested from brain mitigated all the effects of light damage examined, i.e. loss of retinal function (ERG), death of photoreceptors and the stress-induced expression of GFAP in Muller cells. Some of the transplanted G8+ cells were integrated into the retina from the vitreous. Conclusions Myo/Nog cells are a subpopulation of cells that are present in the adult retina. They increase in number in response to light induced stress. Intravitreal injection of Myo/Nog cells was protective to the retina, in part, by reducing retinal stress as measured by the Muller cell response. These results suggest that Myo/Nog cells, or the factors they produce, are neuroprotective and may be therapeutic in neurodegenerative retinal diseases. PMID:28099524
Díaz, Nicolás M; Morera, Luis P; Guido, Mario E
Melanopsin (Opn4), a member of the G-protein-coupled receptor family, is a vitamin A-based opsin in the vertebrate retina that has been shown to be involved in the synchronization of circadian rhythms, pupillary light reflexes, melatonin suppression and other light-regulated tasks. In nonmammalian vertebrates there are two Opn4 genes, Opn4m and Opn4x, the mammalian and Xenopus orthologs respectively. Opn4x is only expressed in nonmammalian vertebrates including reptiles, fish and birds, while Opn4m is found in a subset of retinal ganglion cells (RGCs), the intrinsically photosensitive (ip) RGCs of the inner retina of both mammals and nonmammalian vertebrates. All opsins described utilize retinaldehyde as chromophore, photoisomerized from 11-cis- to all-trans-retinal upon light exposure. Visual retinal photoreceptor cones and rods, responsible for day and night vision respectively, recycle retinoids through a process called the visual cycle that involves the retinal pigment epithelium or glial Müller cells. Although Opn4 has been characterized as a bistable photopigment, little is known about the mechanism/s involved in its chromophore regeneration. In this review, we will attempt to shed light on the visual cycle taking place in the inner retina and discuss the state of the art in the nonvisual photochemistry of vertebrates.
Masihzadeh, Omid; Ammar, David A.; Kahook, Malik Y.; Lei, Tim C.
Purpose. To image the cellular and noncellular structures of the retina in an intact mouse eye without the application of exogenous fluorescent labels using noninvasive, nondestructive techniques. Methods. Freshly enucleated mouse eyes were imaged using two nonlinear optical techniques: coherent anti-Stokes Raman scattering (CARS) and two-photon autofluorescence (TPAF). Cross sectional transverse sections and sequential flat (en face) sagittal sections were collected from a region of sclera approximately midway between the limbus and optic nerve. Imaging proceeded from the surface of the sclera to a depth of ∼60 μm. Results. The fluorescent signal from collagen fibers within the sclera was evident in the TPAF channel; the scleral collagen fibers showed no organization and appeared randomly packed. The sclera contained regions lacking TPAF and CARS fluorescence of ∼3 to 15 μm in diameter that could represent small vessels or scleral fibroblasts. Intense punctate CARS signals from the retinal pigment epithelial layer were of a size and shape of retinyl storage esters. Rod outer segments could be identified by the CARS signal from their lipid-rich plasma membranes. Conclusions. CARS microscopy can be used to image the outer regions of the mammalian retina without the use of a fluorescent dye or exogenously expressed recombinant protein. With technical advancements, CARS/TPAF may represent a new avenue for noninvasively imaging the retina and might complement modalities currently used in clinical practice. PMID:23580484
Mariani, A P
The retinas of most vertebrates contain two or more morphologically distinct types of horizontal cell, and usually one of these types lacks an axon. Among mammals, in which two types are observed, primates are exceptional in that both types of horizontal cell have axons. It then seemed of interest to study the horizontal cells of tree shrews (Tupaia glis), insectivores thought to be closely related to primates. Golgi impregnations of whole, flat-preparations revealed two types of horizontal cell. Uniaxonal cells have a compact dendritic organization with clusters of terminals, and a single thin axon with short collaterals and a few terminals, located along its length. Multiaxonal cells have a relatively large dendritic tree, and arising from the tips of about four to eight dendrites of an individual cell are thin axonlike processes which terminate as profusely branched telodendritic arborizations. This identification of the multiaxonal horizontal cells in Tupaia retina is the first time any vertebrate horizontal cell has been found to possess more than a single axon. A comparison of horizontal cells in tree shrew, monkey, cat, and squirrel retinas shows a remarkable morphological diversity within this class of mammalian retinal neuron.
Intrinsically photosensitive retinal ganglion cells (ipRGCs) have been well characterized in mammalian systems, both morphologically and electrophysiologically. They show slow, sustained responses to bright light in the absence of photoreceptor-based input, mediated by the photopigment melanopsin. Only one mammalian melanopsin gene is expressed in a small fraction of the retinal ganglion cell population, but there are two genes for melanopsin among nonmammalian vertebrates that are widely expressed in a variety of retinal and extraretinal cell types, along with other photosensitive pigments. The current study provides an electrophysiological study of ipRGCs in the larval tiger salamander (Ambystoma tigrinum), a nonmammalian vertebrate with a well-characterized retina. The results show that the ipRGC population is equivalent to the ON ganglion cell population in the tiger salamander retina. This sheds light on the evolutionary trajectory and functional significance of intrinsic photosensitivity through the vertebrate lineage and also affects our understanding of ON cell activity and development. We have characterized the nature of the intrinsic responses of the ON cell population, compared intrinsic and synaptically based receptive fields, and quantified the spectrum of the intrinsic activity. A wider action spectrum of intrinsic photosensitivity was obtained than would be expected for a single opsin photopigment, suggesting the expression of multiple photopigments in the salamander ipRGC. J. Comp. Neurol., 2012. © 2011 Wiley Periodials, Inc.
Saint-Geniez, Magali; Jiang, Aihua; Abend, Stephanie; Liu, Laura; Sweigard, Harry; Connor, Kip M; Arany, Zoltan
Neovascular diseases of the eye are the most common causes of blindness worldwide. The mechanisms underlying pathological neovascularization in the retina remain incompletely understood. PGC-1α is a transcriptional coactivator that plays a central role in the regulation of cellular metabolism. In skeletal muscle, PGC-1α induces VEGFA expression and powerfully promotes angiogenesis, suggesting a similar role in other tissues. This study investigates the role of PGC-1α during normal and pathological vascularization in the retina. We show that PGC-1α induces the expression of VEGFA in numerous retinal cells, and that PGC-1α expression is strongly induced during postnatal retinal development, coincident with VEGFA expression and angiogenesis. PGC-1α(-/-) mice have a significant reduction of early retinal vascular outgrowth, and reduced density of capillaries and number of main arteries and veins as adults. In the oxygen-induced retinopathy model of retinopathy of prematurity, PGC-1α expression is dramatically induced in the inner nuclear layer of the retina, suggesting that PGC-1α drives pathological neovascularization. In support of this, PGC-1α(-/-) mice subjected to oxygen-induced retinopathy had decreased expression of VEGFA and were protected against pathological neovascularization. These results demonstrate that PGC-1α regulates VEGFA in the retina and is required for normal vessel development and for pathological neovascularization. The data highlight PGC-1α as a novel target in the treatment of neovascular diseases of the eye.
Chowdhury, Rebecca; Trimarchi, Jeffrey M.
The entire repertoire of intrinsic factors that control the cell fate determination process of specific retinal neurons has yet to be fully identified. Single cell transcriptome profiling experiments of retinal progenitor cells revealed considerable gene expression heterogeneity between individual cells, especially among different classes of transcription factors. In this study, we show that two of those factors, Onecut1 and Onecut2, are expressed during mouse retinal development. Using mice that are deficient for each of these transcription factors, we further demonstrate a significant loss (∼70–80%) of horizontal cells in the absence of either of these proteins, while the other retinal cells appear at normal numbers. Microarray profiling experiments performed on knockout retinas revealed defects in horizontal cell genes as early as E14.5. Additional profiling assays showed an upregulation of several stress response genes in the adult Onecut2 knockout, suggesting that the integrity of the retina is compromised in the absence of normal numbers of horizontal cells. Interestingly, melanopsin, the gene coding for the photopigment found in photosensitive ganglion cells, was observed to be upregulated in Onecut1 deficient retinas, pointing to a possible regulatory role for Onecut1. Taken together, our data show that similar to Onecut1, Onecut2 is also necessary for the formation of normal numbers of horizontal cells in the developing retina. PMID:25313862
de Souza, Clairton F; Acosta, Monica L; Polkinghorne, Philip J; McGhee, Charles N J; Kalloniatis, Michael
We localised amino acids in the mid-peripheral aged human retina and a retina that had undergone radiation treatment 10 years earlier. The distribution pattern of glutamate, γ-amino butyric acid (GABA), glycine, glutamine and taurine, reflected patterns established in the primate retina. The retina that had undergone radiation exposure displayed both anatomical and neurochemical remodelling. The proximal retina comprised around 40 to 45 per cent of the total retina and neuronal kinesis and aberrant neuronal projections were also present. Amino acid neurochemistry was strikingly different with Müller cells displaying GABA loading, glycinergic neurons displaced and displaying a very high level of glycine labelling. We conclude that radiation exposure triggered these changes in the human retina and likely reflects general remodelling of structure and function following ischaemic damage to endothelial cells.
Liu, Xiaoqin; Zhang, Zhijing; Ribelayga, Christophe P.
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
Merriman, Dana K; Sajdak, Benjamin S; Li, Wei; Jones, Bryan W
With a photoreceptor mosaic containing ∼85% cones, the ground squirrel is one of the richest known mammalian sources of these important retinal cells. It also has a visual ecology much like the human's. While the ground squirrel retina is understandably prominent in the cone biochemistry, physiology, and circuitry literature, far less is known about the remodeling potential of its retinal pigment epithelium, neurons, macroglia, or microglia. This review aims to summarize the data from ground squirrel retina to this point in time, and to relate them to data from other brain areas where appropriate. We begin with a survey of the ground squirrel visual system, making comparisons with traditional rodent models and with human. Because this animal's status as a hibernator often goes unnoticed in the vision literature, we then present a brief primer on hibernation biology. Next we review what is known about ground squirrel retinal remodeling concurrent with deep torpor and with rapid recovery upon re-warming. Notable here is rapidly-reversible, temperature-dependent structural plasticity of cone ribbon synapses, as well as pre- and post-synaptic plasticity throughout diverse brain regions. It is not yet clear if retinal cell types other than cones engage in torpor-associated synaptic remodeling. We end with the small but intriguing literature on the ground squirrel retina's remodeling responses to insult by retinal detachment. Notable for widespread loss of (cone) photoreceptors, there is surprisingly little remodeling of the RPE or Müller cells. Microglial activation appears minimal, and remodeling of surviving second- and third-order neurons seems absent, but both require further study. In contrast, traumatic brain injury in the ground squirrel elicits typical macroglial and microglial responses. Overall, the data to date strongly suggest a heretofore unrecognized, natural checkpoint between retinal deafferentiation and RPE and Müller cell remodeling events. As we
Solursh, M; Langille, R M; Wood, J; Sampath, T K
Osteogenic Protein-1 (OP-1/BMP-7) is a bone morphogenetic protein in the transforming growth factor-beta superfamily and has been shown to be expressed temporally and spatially during epithelial-mesenchymal interactions mediating tissue morphogenesis in early embryogenesis. In order to identify the primary role(s) for OP-1 in development, we carried out whole rat embryo cultures, over a 72-h period from primitive streak stages to early limb bud stages, in rat sera containing either OP-1 blocking antibodies (10 micrograms/ml) or nonreactive IgG. Rat embryos cultured with control antibodies developed normally, while those cultured with anti-OP-1 antibodies consistently exhibited over-all reduced size and absence of eyes. Histological sections revealed a greater reduction in neural retina development in the embryos treated with anti-OP-1 blocking antibodies. In situ hybridization and immunolocalization analyses indicate that OP-1 is expressed in the neuroepithelium of the optic vesicle at E11.5, is limited to the presumptive neural retina and developing lens placode, and is subsequently expressed in the neural retina, lens and developing cornea at E12.5-E13.5. Our results indicate that OP-1 mediates the inductive signals involved in mammalian eye development.
Bernardos, Rebecca L; Barthel, Linda K; Meyers, Jason R; Raymond, Pamela A
Neuronal progenitors in the mammalian brain derive from radial glia or specialized astrocytes. In developing neural retina, radial glia-like Müller cells are generated late in neurogenesis and are not considered to be neuronal progenitors, but they do proliferate after injury and can express neuronal markers, suggesting a latent neurogenic capacity. To examine the neurogenic capacity of retinal glial cells, we used lineage tracing in transgenic zebrafish with a glial-specific promoter (gfap, for glial fibrillary acid protein) driving green fluorescent protein in differentiated Müller glia. We found that all Müller glia in the zebrafish retina express low levels of the multipotent progenitor marker Pax6 (paired box gene 6), and they proliferate at a low frequency in the intact, uninjured retina. Müller glia-derived progenitors express Crx (cone rod homeobox) and are late retinal progenitors that generate the rod photoreceptor lineage in the postembryonic retina. These Müller glia-derived progenitors also remain competent to produce earlier neuronal lineages, in that they respond to loss of cone photoreceptors by specifically regenerating the missing neurons. We conclude that zebrafish Müller glia function as multipotent retinal stem cells that generate retinal neurons by homeostatic and regenerative developmental mechanisms.
Sony, Parul; Venkatesh, Pradeep; Gadaginamath, Shailesh; Garg, Sat Pal
A 16-year-old boy presented with diminished visual acuity of 6/60 following blunt trauma to his right eye with a cricket ball. Fundus examination showed commotio retinae. Optical coherence tomography (OCT) demonstrated increased reflectivity with small optically clear spaces in the area corresponding to the photoreceptor outer segment. At 2-month follow up the visual acuity improved to 6/6. A small area of retinal opacification persisted nasally, and OCT of the corresponding area continued to show increased reflectivity in the area of photoreceptor outer segment. Increased reflectivity on OCT in eyes with commotio retinae probably denotes photoreceptor outer segment disruption and seems to be reversible to a variable extent.
LLNL has assisted in the development of the first long-term retinal prosthesis - called an artificial retina - that can function for years inside the harsh biological environment of the eye. This work has been done in collaboration with four national laboratories (Argonne, Los Alamos, Oak Ridge and Sandia), four universities (the California Institute of Technology, the Doheny Eye Institute at USC, North Carolina State University and the University of California, Santa Cruz), an industrial partner (Second SightÂ® Medical Products Inc. of Sylmar, Calif.) and the U.S. Department of Energy. With this device, application-specific integrated circuits transform digital images from a camera into electric signals in the eye that the brain uses to create a visual image. In clinical trials, patients with vision loss were able to successfully identify objects, increase mobility and detect movement using the artificial retina.
LLNL has assisted in the development of the first long-term retinal prosthesis - called an artificial retina - that can function for years inside the harsh biological environment of the eye. This work has been done in collaboration with four national laboratories (Argonne, Los Alamos, Oak Ridge and Sandia), four universities (the California Institute of Technology, the Doheny Eye Institute at USC, North Carolina State University and the University of California, Santa Cruz), an industrial partner (Second Sight® Medical Products Inc. of Sylmar, Calif.) and the U.S. Department of Energy. With this device, application-specific integrated circuits transform digital images from a camera into electric signals in the eye that the brain uses to create a visual image. In clinical trials, patients with vision loss were able to successfully identify objects, increase mobility and detect movement using the artificial retina.
Engelhardt, Maren; Bogdahn, Ulrich; Aigner, Ludwig
The adult mammalian retina is devoid of any detectable neurogenesis. However, different cell types have been suggested to potentially act as neural progenitors in the adult mammalian retina in vitro, such as ciliary body (CB), Muller glia, and retinal pigment epithelium (RPE) cells. In rodents and humans, strong evidence for neural stem or progenitor properties exists only for CB-derived cells, but not for other retinal cell types. Here, we provide a comparative analysis of adult rat CB- and RPE-derived cells suggesting that the two cell types share certain neural progenitor properties in vitro. CB and RPE cells expressed neural progenitor markers such as Nestin, Flk-1, Hes1, and Musashi. They proliferated under adherent and neurosphere conditions and showed limited self-renewal. Moreover, they differentiated into neuronal and glial cells based on the expression of differentiation markers such as the young neuronal marker beta-III tubulin and the glial and progenitor markers GFAP and NG2. Expression of beta-III tubulin was found in cells with neuronal and non-neuronal morphology. A subpopulation of RPE- and CB-derived progenitor cells expressed the neurogenesis-specific protein doublecortin (DCX). Interestingly, DCX expression defined a beta-III tubulin-positive CB and RPE fraction with a distinct neuronal morphology. In summary, the data suggest that RPE cells share with CB cells the potential to de-differentiate into a cell type with neural progenitor-like identity. In addition, DCX expression might define the neuronal-differentiating RPE- and CB-derived progenitor population.
Brown, R Lane; Xiong, Wei-Hong; Peters, James H; Tekmen-Clark, Merve; Strycharska-Orczyk, Iwona; Reed, Brian T; Morgans, Catherine W; Duvoisin, Robert M
Transient receptor potential (TRP) channels constitute a large family of cation permeable ion channels that serve crucial functions in sensory systems by transducing environmental changes into cellular voltage and calcium signals. Within the retina, two closely related members of the melastatin TRP family, TRPM1 and TRPM3, are highly expressed. TRPM1 has been shown to be required for the depolarizing response to light of ON-bipolar cells, but the role of TRPM3 in the retina is unknown. Immunohistochemical staining of mouse retina with an antibody directed against the C-terminus of TRPM3 labeled the inner plexiform layer (IPL) and a subset of cells in the ganglion cell layer. Within the IPL, TRPM3 immunofluorescence was markedly stronger in the OFF sublamina than in the ON sublamina. Electroretinogram recordings showed that the scotopic and photopic a- and b-waves of TRPM3(-/-) mice are normal indicating that TRPM3 does not play a major role in visual processing in the outer retina. TRPM3 activity was measured by calcium imaging and patch-clamp recording of immunopurified retinal ganglion cells. Application of the TRPM3 agonist, pregnenolone sulfate (PS), stimulated increases in intracellular calcium in ~40% of cells from wild type and TRPM1(‑/‑) mice, and the PS-stimulated increases in calcium were blocked by co-application of mefenamic acid, a TRPM3 antagonist. No PS-stimulated changes in fluorescence were observed in ganglion cells from TRPM3(-/-) mice. Similarly, PS-stimulated currents that could be blocked by mefenamic acid were recorded from wild type retinal ganglion cells but were absent in ganglion cells from TRPM3-/- mice.
MALCOLM, J. E.
A mathematical model for study of blood flow has been derived from the avian egg, utilizing the theories of crystallography and photosynthesis. The model is employed to explain the form of the eye and the function of the cells of the human retina, with special reference to colour vision and the pathology of migraine. ImagesFig. 1Fig. 4Fig. 5Fig. 7Fig. 8Fig. 9Fig. 10Fig. 11 PMID:4736600
Strohbehn, K.; Jenkins, R. E.; Sun, X.; Andreou, A. G.
There are a host of position sensors, such as quadcells and CCD's, which are candidates for detecting optical position errors and providing error signals for a mirror positioning loop. We are developing a novel, very high bandwidth, biologically inspired position sensor for optical position tracking systems. We present recent test results and design issues for the use of biologically inspired silicon retinas for spaceborne optical position tracking systems.
The visual system is beautifully crafted to transmit information of the external world to visual processing and cognitive centers in the brain. For visual information to be relayed to the brain, a series of axon pathfinding events must take place to ensure that the axons of retinal ganglion cells, the only neuronal cell type in the retina that sends axons out of the retina, find their way out of the eye to connect with targets in the brain. In the past few decades, the power of molecular and genetic tools, including the generation of genetically manipulated mouse lines, have multiplied our knowledge about the molecular mechanisms involved in the sculpting of the visual system. Here, we review major advances in our understanding of the mechanisms controlling the differentiation of RGCs, guidance of their axons from the retina to the primary visual centers, and the refinement processes essential for the establishment of topographic maps and eye-specific axon segregation. Human disorders, such as albinism and achiasmia, that impair RGC axon growth and guidance and, thus, the establishment of a fully functioning visual system will also be discussed. PMID:25504540
Lorber, Barbara; Hsiao, Wen-Kai; Martin, Keith R.
Purpose of review Biological three-dimensional printing has received a lot of media attention over recent years with advances made in printing cellular structures, including skin and heart tissue for transplantation. Although limitations exist in creating functioning organs with this method, the hope has been raised that creating a functional retina to cure blindness is within reach. The present review provides an update on the advances made toward this goal. Recent findings It has recently been shown that two types of retinal cells, retinal ganglion cells and glial cells, can be successfully printed using a piezoelectric inkjet printer. Importantly, the cells remained viable and did not change certain phenotypic features as a result of the printing process. In addition, recent advances in the creation of complex and viable three-dimensional cellular structures have been made. Summary Some first promising steps toward the creation of a functional retina have been taken. It now needs to be investigated whether recent findings can be extended to other cells of the retina, including those derived from human tissue, and if a complex and viable retinal structure can be created through three-dimensional printing. PMID:27045545
Hansen, Ronald M; Moskowitz, Anne; Akula, James D; Fulton, Anne B
Retinopathy of prematurity (ROP) is a neurovascular disease that affects prematurely born infants and is known to have significant long term effects on vision. We conducted the studies described herein not only to learn more about vision but also about the pathogenesis of ROP. The coincidence of ROP onset and rapid developmental elongation of the rod photoreceptor outer segments motivated us to consider the role of the rods in this disease. We used noninvasive electroretinographic (ERG), psychophysical, and retinal imaging procedures to study the function and structure of the neurosensory retina. Rod photoreceptor and post-receptor responses are significantly altered years after the preterm days during which ROP is an active disease. The alterations include persistent rod dysfunction, and evidence of compensatory remodeling of the post-receptor retina is found in ERG responses to full-field stimuli and in psychophysical thresholds that probe small retinal regions. In the central retina, both Mild and Severe ROP delay maturation of parafoveal scotopic thresholds and are associated with attenuation of cone mediated multifocal ERG responses, significant thickening of post-receptor retinal laminae, and dysmorphic cone photoreceptors. These results have implications for vision and control of eye growth and refractive development and suggest future research directions. These results also lead to a proposal for noninvasive management using light that may add to the currently invasive therapeutic armamentarium against ROP.
Puller, Christian; Ondreka, Katharina; Haverkamp, Silke
Parallel processing of an image projected onto the retina starts at the first synapse, the cone pedicle, and each cone feeds its light signal into a minimum of eight different bipolar cell types. Hence, the morphological classification of bipolar cells is a prerequisite for analyzing retinal circuitry. Here we applied common bipolar cell markers to the cone-dominated ground squirrel retina, studied the labeling by confocal microscopy and electron microscopy, and compared the resulting bipolar cell types with those of the mouse (rod dominated) and primate retina. Eight different cone bipolar cell types (three OFF and five ON) and one rod bipolar cell were distinguished. The major criteria for classifying the cells were their immunocytochemical identity, their dendritic branching pattern, and the shape and stratification level of their axons in the inner plexiform layer (IPL). Immunostaining with antibodies against Gγ13, a marker for ON bipolar cells, made it possible to separate OFF and ON bipolars. Recoverin-positive OFF bipolar cells partly overlapped with ON bipolar axon terminals at the ON/OFF border of the IPL. Antibodies against HCN4 labeled the S-cone selective (bb) bipolar cell. The calcium-binding protein CaB5 was expressed in two OFF and two ON cone bipolar cell types, and CD15 labeled a widefield ON cone bipolar cell comparable to the DB6 in primate.
Xu, Lili; Ruan, Guoxiang; Dai, Heng; Liu, Andrew C.; Penn, John
Purpose To test whether Müller glia of the mammalian retina have circadian rhythms. Methods We used Müller glia cultures isolated from mouse lines or from humans and bioluminescent reporters of circadian clock genes to monitor molecular circadian rhythms. The clock gene dependence of the Müller cell rhythms was tested using clock gene knockout mouse lines or with siRNA for specific clock genes. Results We demonstrated that retinal Müller glia express canonical circadian clock genes, are capable of sustained circadian oscillations in isolation from other cell types, and exhibit unique features of their molecular circadian clock compared to the retina as a whole. Mouse and human Müller cells demonstrated circadian clock function; however, they exhibited species-specific differences in the gene dependence of their clocks. Conclusions Müller cells are the first mammalian retinal cell type in which sustained circadian rhythms have been demonstrated in isolation from other retinal cells. PMID:27081298
Sánchez-Chávez, Gustavo; Hernández-Berrones, Jethro; Luna-Ulloa, Luis Bernardo; Coffe, Víctor; Salceda, Rocío
Glucose is the main fuel for energy metabolism in retina. The regulatory mechanisms that maintain glucose homeostasis in retina could include hormonal action. Retinopathy is one of the chemical manifestations of long-standing diabetes mellitus. In order to better understand the effect of hyperglycemia in retina, we studied glycogen content as well as glycogen synthase and phosphorylase activities in both normal and streptozotocin-induced diabetic rat retina and compared them with other tissues. Glycogen levels in normal rat retina are low (46 +/- 4.0 nmol glucosyl residues/mg protein). However, high specific activity of glycogen synthase was found in retina, indicating a substantial capacity for glycogen synthesis. In diabetic rats, glycogen synthase activity increased between 50% and 100% in retina, brain cortex and liver of diabetic rats, but only retina exhibited an increase in glycogen content. Although, total and phosphorylated glycogen synthase levels were similar in normal and diabetic retina, activation of glycogen synthase by glucose-6-P was remarkable increased. Glycogen phosphorylase activity decreased 50% in the liver of diabetic animals; it was not modified in the other tissues examined. We conclude that the increase in glycogen levels in diabetic retina was due to alterations in glycogen synthase regulation.
Nakayama, Yoshiaki; Miyake, Ayumi; Nakagawa, Yu; Mido, Tomotaka; Yoshikawa, Maya; Konishi, Morichika; Itoh, Nobuyuki
Fgf signaling plays crucial roles in morphogenesis. Fgf19 is required for zebrafish forebrain development. Here, we examined the roles of Fgf19 in the formation of the lens and retina in zebrafish. Knockdown of Fgf19 caused a size reduction of the lens and the retina, failure of closure of the choroids fissure, and a progressive expansion of the retinal tissue to the midline of the forebrain. Fgf19 expressed in the nasal retina and lens was involved in cell survival but not cell proliferation during embryonic lens and retina development. Fgf19 was essential for the differentiation of lens fiber cells in the lens but not for the neuronal differentiation and lamination in the retina. Loss of nasal fate in the retina caused by the knockdown of Fgf19, expansion of nasal fate in the retina caused by the overexpression of Fgf19 and eye transplantation indicated that Fgf19 in the retina was crucial for the nasal-temporal patterning of the retina that is critical for the guidance of retinal ganglion cell axons. Knockdown of Fgf19 also caused incorrect axon pathfinding. The present findings indicate that Fgf19 positively regulates the patterning and growth of the retina, and the differentiation and growth of the lens in zebrafish.
Pahlberg, Johan; Sampath, Alapakkam P
In sensory biology, a major outstanding question is how sensory receptor cells minimize noise while maximizing signal to set the detection threshold. This optimization could be problematic because the origin of both the signals and the limiting noise in most sensory systems is believed to lie in stimulus transduction. Signal processing in receptor cells can improve the signal-to-noise ratio. However, neural circuits can further optimize the detection threshold by pooling signals from sensory receptor cells and processing them using a combination of linear and nonlinear filtering mechanisms. In the visual system, noise limiting light detection has been assumed to arise from stimulus transduction in rod photoreceptors. In this context, the evolutionary optimization of the signal-to-noise ratio in the retina has proven critical in allowing visual sensitivity to approach the limits set by the quantal nature of light. Here, we discuss how noise in the mammalian retina is mitigated to allow for highly sensitive night vision.
Rojas, Blanca; Ramírez, Ana I.; de Hoz, Rosa; Salazar, Juan J.; Triviño, Alberto; Ramírez, José M.
Proliferation of microglial cells has been considered a sign of glial activation and a hallmark of ongoing neurodegenerative diseases. Microglia activation is analyzed in animal models of different eye diseases. Numerous retinal samples are required for each of these studies to obtain relevant data of statistical significance. Because manual quantification of microglial cells is time consuming, the aim of this study was develop an algorithm for automatic identification of retinal microglia. Two groups of adult male Swiss mice were used: age-matched controls (naïve, n = 6) and mice subjected to unilateral laser-induced ocular hypertension (lasered; n = 9). In the latter group, both hypertensive eyes and contralateral untreated retinas were analyzed. Retinal whole mounts were immunostained with anti Iba-1 for detecting microglial cell populations. A new algorithm was developed in MATLAB for microglial quantification; it enabled the quantification of microglial cells in the inner and outer plexiform layers and evaluates the area of the retina occupied by Iba-1+ microglia in the nerve fiber-ganglion cell layer. The automatic method was applied to a set of 6,000 images. To validate the algorithm, mouse retinas were evaluated both manually and computationally; the program correctly assessed the number of cells (Pearson correlation R = 0.94 and R = 0.98 for the inner and outer plexiform layers respectively). Statistically significant differences in glial cell number were found between naïve, lasered eyes and contralateral eyes (P<0.05, naïve versus contralateral eyes; P<0.001, naïve versus lasered eyes and contralateral versus lasered eyes). The algorithm developed is a reliable and fast tool that can evaluate the number of microglial cells in naïve mouse retinas and in retinas exhibiting proliferation. The implementation of this new automatic method can enable faster quantification of microglial cells in retinal pathologies. PMID:26580208
Lahouaoui, Hasna; Coutanson, Christine; Cooper, Howard M.; Bennis, Mohamed
Purpose Diabetic retinopathy is one of the most common consequences of diabetes that affects millions of working-age adults worldwide and leads to progressive degeneration of the retina, visual loss, and blindness. Diabetes is associated with circadian disruption of the central and peripheral circadian clocks, but the mechanisms responsible for such alterations are unknown. Using a streptozotocin (STZ)-induced model of diabetes, we investigated whether diabetes alters 1) the circadian regulation of clock genes in the retina and in the central clocks, 2) the light response of clock genes in the retina, and/or 3) light-driven retinal dopamine (DA), a major output marker of the retinal clock. Methods To quantify circadian expression of clock and clock-controlled genes, retinas and suprachiasmatic nucleus (SCN) from the same animals were collected every 4 h in circadian conditions, 12 weeks post-diabetes. Induction of Per1, Per2, and c-fos mRNAs was quantified in the retina after the administration of a pulse of monochromatic light (480 nm, 1.17×1014 photons/cm2/s, 15 min) at circadian time 16. Gene expression was assessed with real-time reverse transcription PCR (RT–PCR). Pooled retinas from the control and STZ-diabetic mice were collected 2 h after light ON and light OFF (Zeitgeber time (ZT)2 and ZT14), and DA and its metabolite were analyzed with high-performance liquid chromatography (HPLC). Results We found variable effects of diabetes on the expression of clock genes in the retina and only slight differences in phase and/or amplitude in the SCN. c-fos and Per1 induction by a 480 nm light pulse was abolished in diabetic animals at 12 weeks post-induction of diabetes in comparison with the control mice, suggesting a deficit in light-induced neuronal activation of the retinal clock. Finally, we quantified a 56% reduction in the total number of tyrosine hydroxylase (TH) immunopositive cells, associated with a decrease in DA levels during the subjective day (ZT2
Ronca, April E.
Life on Earth, and thus the reproductive and ontogenetic processes of all extant species and their ancestors, evolved under the constant influence of the Earth's l g gravitational field. These considerations raise important questions about the ability of mammals to reproduce and develop in space. In this chapter, I review the current state of our knowledge of spaceflight effects on developing mammals. Recent studies are revealing the first insights into how the space environment affects critical phases of mammalian reproduction and development, viz., those events surrounding fertilization, embryogenesis, pregnancy, birth, postnatal maturation and parental care. This review emphasizes fetal and early postnatal life, the developmental epochs for which the greatest amounts of mammalian spaceflight data have been amassed. The maternal-offspring system, the coordinated aggregate of mother and young comprising mammalian development, is of primary importance during these early, formative developmental phases. The existing research supports the view that biologically meaningful interactions between mothers and offspring are changed in the weightlessness of space. These changes may, in turn, cloud interpretations of spaceflight effects on developing offspring. Whereas studies of mid-pregnant rats in space have been extraordinarily successful, studies of young rat litters launched at 9 days of postnatal age or earlier, have been encumbered with problems related to the design of in-flight caging and compromised maternal-offspring interactions. Possibilities for mammalian birth in space, an event that has not yet transpired, are considered. In the aggregate, the results indicate a strong need for new studies of mammalian reproduction and development in space. Habitat development and systematic ground-based testing are important prerequisites to future research with young postnatal rodents in space. Together, the findings support the view that the environment within which young
Ronca, April E
Life on Earth, and thus the reproductive and ontogenetic processes of all extant species and their ancestors, evolved under the constant influence of the Earth's l g gravitational field. These considerations raise important questions about the ability of mammals to reproduce and develop in space. In this chapter, I review the current state of our knowledge of spaceflight effects on developing mammals. Recent studies are revealing the first insights into how the space environment affects critical phases of mammalian reproduction and development, viz., those events surrounding fertilization, embryogenesis, pregnancy, birth, postnatal maturation and parental care. This review emphasizes fetal and early postnatal life, the developmental epochs for which the greatest amounts of mammalian spaceflight data have been amassed. The maternal-offspring system, the coordinated aggregate of mother and young comprising mammalian development, is of primary importance during these early, formative developmental phases. The existing research supports the view that biologically meaningful interactions between mothers and offspring are changed in the weightlessness of space. These changes may, in turn, cloud interpretations of spaceflight effects on developing offspring. Whereas studies of mid-pregnant rats in space have been extraordinarily successful, studies of young rat litters launched at 9 days of postnatal age or earlier, have been encumbered with problems related to the design of in-flight caging and compromised maternal-offspring interactions. Possibilities for mammalian birth in space, an event that has not yet transpired, are considered. In the aggregate, the results indicate a strong need for new studies of mammalian reproduction and development in space. Habitat development and systematic ground-based testing are important prerequisites to future research with young postnatal rodents in space. Together, the findings support the view that the environment within which young
Hara, Akira; Taguchi, Ayako; Aoki, Hitomi; Hatano, Yuichiro; Niwa, Masayuki; Yamada, Yasuhiro; Kunisada, Takahiro
Transplanted embryonic stem (ES) cells can be integrated into the retinas of adult mice as well-differentiated neuroretinal cells. However, the transplanted ES cells also have a tumorigenic activity as they have the ability for multipotent differentiation to various types of tissues. In the present study, human ES (hES) cells were transplanted into adult nude mouse retinas by intravitreal injections 20 h after intravitreal N-methyl-D-aspartate (NMDA) administration. After the transplantation of hES cells, the folate antagonist, methotrexate (MTX) was administrated in order to control the differentiation of the transplanted hES cells. Neuronal differentiation and teratogenic potential of hES cells were examined immunohistochemically 5 weeks after transplantation. The proliferative activity of transplanted cells was determined by both the mitotic index and the Ki-67 proliferative index. Disappearance of Oct-4-positive hES cells showing undifferentiated morphology was observed after intraperitoneal MTX treatment daily, for 15 days. Decreased mitotic and Ki-67 proliferative indices, and increased neuronal differentiation were detected in the surviving hES cells after the MTX treatment. These results suggest two important effects of intraperitoneal MTX treatment for hES cells transplanted into nude mouse retina: (1) MTX treatment following transplantation induces neuronal differentiation, and (2) MTX decreases proliferative activity and tumorigenic potential.
Vuong, Helen E.; de Sevilla Müller, Luis Pérez; Hardi, Claudia N.; McMahon, Douglas G.; Brecha, Nicholas C.
Transgenic mouse lines are essential tools for understanding the connectivity, physiology and function of neuronal circuits, including those in the retina. This report compares transgene expression in the retina of a tyrosine hydroxylase (TH)-red fluorescent protein (RFP) line with three catecholamine-related Cre recombinase lines [TH-bacterial artificial chromosome (BAC)-, TH-, and dopamine transporter (DAT)-Cre] that were crossed with a ROSA26-tdTomato reporter line. Retinas were evaluated and immunostained with commonly used antibodies including those directed to TH, GABA and glycine to characterize the RFP or tdTomato fluorescent-labeled amacrine cells, and an antibody directed to RNA-binding protein with multiple splicing to identify ganglion cells. In TH-RFP retinas, types 1 and 2 dopamine (DA) amacrine cells were identified by their characteristic cellular morphology and type 1 DA cells by their expression of TH immunoreactivity. In the TH-BAC-, TH-, and DAT-tdTomato retinas, less than 1%, ~6%, and 0%, respectively, of the fluorescent cells were the expected type 1 DA amacrine cells. Instead, in the TH-BAC-tdTomato retinas, fluorescently labeled AII amacrine cells were predominant, with some medium somal diameter ganglion cells. In TH-tdTomato retinas, fluorescence was in multiple neurochemical amacrine cell types, including four types of polyaxonal amacrine cells. In DAT-tdTomato retinas, fluorescence was in GABA immunoreactive amacrine cells, including two types of bistratified and two types of monostratified amacrine cells. Although each of the Cre lines were generated with the intent to specifically label DA cells, our findings show a cellular diversity in Cre expression in the adult retina and indicate the importance of careful characterization of transgene labeling patterns. These mouse lines with their distinctive cellular labeling patterns will be useful tools for future studies of retinal function and visual processing. PMID:26335381
Ungureanu, Constantin; Corniencu, Felicia
Biometric is automated method of recognizing a person based on physiological or behavior characteristics. Among the features measured are retina scan, voice, and fingerprint. A retina-based biometric involves the analysis of the blood vessels situated at the back of the eye. In this paper we present a method, which uses the fractal analysis to characterize the retina images. The Fractal Dimension (FD) of retina vessels was measured for a number of 20 images and have been obtained different values of FD for each image. This algorithm provides a good accuracy is cheap and easy to implement.
Hill, Andrew J; Zwart, Isabel; Tam, Henry H; Chan, Jane; Navarrete, Cristina; Jen, Ling-Sun; Navarrete, Roberto
This study investigated the ability of mesenchymal stem cells (MSCs) derived from full-term human umbilical cord blood to survive, integrate and differentiate after intravitreal grafting to the degenerating neonatal rat retina following intracranial optic tract lesion. MSCs survived for 1 week in the absence of immunosuppression. When host animals were treated with cyclosporin A and dexamethasone to suppress inflammatory and immune responses, donor cells survived for at least 3 weeks, and were able to spread and cover the entire vitreal surface of the host retina. However, MSCs did not significantly integrate into or migrate through the retina. They also maintained their human antigenicity, and no indication of neural differentiation was observed in retinas where retinal ganglion cells either underwent severe degeneration or were lost. These results have provided the first in vivo evidence that MSCs derived from human umbilical cord blood can survive for a significant period of time when the host rat response is suppressed even for a short period. These results, together with the observation of a lack of neuronal differentiation and integration of MSCs after intravitreal grafting, has raised an important question as to the potential use of MSCs for neural repair through the replacement of lost neurons in the mammalian retina and central nervous system.
Hu, Sherry Shu-Jung; Arnold, Andy; Hutchens, Jacqueline M.; Radicke, Josh; Cravatt, Benjamin F.; Wager-Miller, Jim; Mackie, Ken; Straiker, Alex
Cannabinoid receptors and their ligands constitute an endogenous signaling system that is found throughout the body, including the eye. This system can be activated by Δ9-tetrahydrocannabinol, a major drug of abuse. Cannabinoids offer considerable therapeutic potential in modulating ocular immune and inflammatory responses and in regulating intraocular pressure. The location of cannabinoid receptors 1 (CB1) in the retina is known, but recently a constellation of proteins has been identified that produce and break down endocannabinoids (eCBs) and modulate CB1 function. Localization of these proteins is critical to defining specific cannabinoid signaling circuitry in the retina. Here we show the localization of diacylglycerol lipase α and β (DGLα/β), implicated in the production of the eCB 2-arachidonoyl glycerol (2-AG); monoacylglycerol lipase (MGL) and α/β-hydrolase domain 6 (ABHD6), both implicated in the breakdown of 2-AG; cannabinoid receptor interacting protein 1a (CRIP1a), a protein that may modulate CB1 function; Fatty acid amide hydrolase (FAAH) and N-acylethanolamine-hydrolyzing acid amidase (NAAA) which have been shown to break down the eCB anandamide and related acyl amides. In our most prominent finding, DGLα is present in post-synaptic Type 1 OFF cone bipolar cells juxtaposed to CB1-containing cone photoreceptor terminals. Interestingly, CRIP1a is reliably presynaptic to DGLα, consistent with a possible role in cannabinoid signaling, NAAA is restricted to retinal pigment epithelium (RPE), while DGLβ is limited to retinal blood vessels. These results taken together with previous anatomic and functional studies define specific cannabinoid circuitry likely to modulate eCB signaling at the first synapse of the retina as well as in the inner plexiform layer (IPL). PMID:20653038
This award supported the investigation on electromagnetic and thermal effects associated with the artificial retina, designed in collaboration with national laboratories, universities, and private companies. Our work over the two years of support under this award has focused mainly on 1) Design of new telemetry coils for optimal power and data transfer between the implant and the external device while achieving a significant size reduction with respect to currently used coils; 2) feasibility study of the virtual electrode configuration 3) study the effect of pulse shape and duration on the stimulation efficacy.
Clark, Clifton D.
This dissertation presents recent progress in several areas related to modeling laser damage to the retina. In Chapter 3, we consider the consequences of using the Arrhenius damage model to predict the damage thresholds of multiple pulse, or repetitive pulse, exposures. We have identified a few fundamental trends associated with the multiple pulse damage predictions made by the Arrhenius model. These trends differ from what would be expected by non-thermal mechanisms, and could prove useful in differentiating thermal and non-thermal damage. Chapter 4 presents a new rate equation damage model hypothesized to describe photochemical damage. The model adds a temperature dependent term to the simple rate equation implied by the principle of reciprocity that is characteristic of photochemical damage thresholds. A recent damage threshold study, conducted in-vitro, has revealed a very sharp transition between thermal and photochemical damage threshold trends. For the wavelength used in the experiment (413 nm), thermal damage thresholds were observed at exposure levels that were twice the expected photochemical damage threshold, based on the traditional understanding of photochemical damage. Our model accounts for this observed trend by introducing a temperature dependent quenching, or repair, rate to the photochemical damage rate. For long exposures that give a very small temperature rise, the model reduces to the principle of reciprocity. Near the transition region between thermal and photochemical damage, the model allows the damage threshold to be set by thermal mechanisms, even at exposure above the reciprocity exposure. In Chapter 5, we describe a retina damage model that includes thermal lensing in the eye by coupling beam propagation and heat transfer models together. Thermal lensing has recently been suggested as a contributing factor to the large increase in measured retinal damage thresholds in the near infrared. The transmission of the vitreous decreases
Yen, Hao-Ren; Su, Guo-Dung J.
In this paper, we propose a multi-channel imaging system which combines the principles of an insect's compound eye and optical cluster eye. The system consists of two curved structure lens arrays with different pitches. Both of them have the same curvature and the radiuses of the lenses in the arrays are optimized to focus rays on the retina. The optical axes of different channels are tilted to each other in order to reduce the optical system volume and transmit a wide field of view. Each channel of an array of multiple optical system transfers only a part of the field of view. Each partial image passes through each channel and stitches together on the retina to reconstruct a complete image. In order to simulate the image stitching, we also build an eye model. The thickness from the panel to the last surface of lens group is less than 25mm. The panel size is designed to be 4 inch which is the scale of eyeglass. The system can provide a large field of view about 150 degrees which is much wider than the commercial products. By using the 3D printer, we can make a model of lens array to achieve our design.
Hojman, Anne S; Otzen, Louise W D; Schrøder-Hansen, Lise Maj; Wegener, Karen M
Incidental findings in the rat eye are not uncommon in acute and long-term toxicological studies. These findings can be associated with a number of causes unrelated to treatment with the test article, including congenital malformation, trauma, infection, metabolic disease, genetic predisposition, and age-related changes. The occurrence of pigment deposition in the retina of Wistar Hannover (Crl:WI (Han)) rats in a 4-week toxicity study is reported in this communication. The microscopic examination of the eyes in the 4-week toxicity study revealed focal yellow-brown pigment deposits in the retina, mainly located in the ganglion cell layer. The retinal pigment deposits were randomly distributed in the control and treated groups and were considered incidental. The deposits were clearly positive for ferric iron in the Perls' stain but not for lipofuscin by the Schmorl's and Long Ziehl-Neelsen methods. The iron-containing pigment is likely to represent hemosiderin accumulation after retinal micro-hemorrhage or could be indicative of the normal intraretinal iron transport and turnover.
Ozawa, Yoko; Sasaki, Mariko; Takahashi, Noriko; Kamoshita, Mamoru; Miyake, Seiji; Tsubota, Kazuo
Although a large variety of pharmaceutical therapies for treating disease have been developed in recent years, there has been little progress in disease prevention. In particular, the protection of neural tissue is essential, because it is hardly regenerated. The use of nutraceuticals for maintaining the health has been supported by several clinical studies, including cross-sectional and interventional studies for age-related macular disease. However, mechanistic evidence for their effects at the molecular level has been very limited. In this review, we focus on lutein, which is a xanthophyll type of carotenoid. Lutein is not synthesized in mammals, and must be obtained from the diet. It is delivered to the retina, and in humans, it is concentrated in the macula. Here, we describe the neuroprotective effects of lutein and their underlying molecular mechanisms in animal models of vision-threatening diseases, such as innate retinal inflammation, diabetic retinopathy, and light-induced retinal degeneration. In lutein-treated mouse ocular disease models, oxidative stress in the retina is reduced, and its downstream pathological signals are inhibited. Furthermore, degradation of the functional proteins, rhodopsin (a visual substance) and synaptophysin (a synaptic vesicle protein also influenced in other neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease), the depletion of brain-derived neurotrophic factor (BDNF), and DNA damage are prevented by lutein, which preserves visual function. We discuss the possibility of using lutein, an antioxidant, as a neuroprotective treatment for humans.
Ozawa, Yoko; Sasaki, Mariko; Takahashi, Noriko; Kamoshita, Mamoru; Miyake, Seiji; Tsubota, Kazuo
Although a large variety of pharmaceutical therapies for treating disease have been developed in recent years, there has been little progress in disease prevention. In particular, the protection of neural tissue is essential, because it is hardly regenerated. The use of nutraceuticals for maintaining the health has been supported by several clinical studies, including cross-sectional and interventional studies for age-related macular disease. However, mechanistic evidence for their effects at the molecular level has been very limited. In this review, we focus on lutein, which is a xanthophyll type of carotenoid. Lutein is not synthesized in mammals, and must be obtained from the diet. It is delivered to the retina, and in humans, it is concentrated in the macula. Here, we describe the neuroprotective effects of lutein and their underlying molecular mechanisms in animal models of vision-threatening diseases, such as innate retinal inflammation, diabetic retinopathy, and light-induced retinal degeneration. In lutein-treated mouse ocular disease models, oxidative stress in the retina is reduced, and its downstream pathological signals are inhibited. Furthermore, degradation of the functional proteins, rhodopsin (a visual substance) and synaptophysin (a synaptic vesicle protein also influenced in other neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease), the depletion of brain-derived neurotrophic factor (BDNF), and DNA damage are prevented by lutein, which preserves visual function. We discuss the possibility of using lutein, an antioxidant, as a neuroprotective treatment for humans. PMID:22211688
Li, Lu; Eter, Nicole; Heiduschka, Peter
The microglia are the immune cells of the central nervous system and, also the retina. They fulfil several tasks of surveillance in the healthy retina. In case of an injury or disease, microglia become activated and tries to repair the damage. However, in a lot of cases it does not work, and microglia deteriorate the situation by releasing toxic and pro-inflammatory compounds. Moreover, they further promote degenerative processes by attacking and phagocytosing damaged neurones and photoreceptors that otherwise would possibly have the chance to survive. Such deleterious action of the microglia has been observed in degeneration of retinal ganglion cells and photoreceptors, and it takes place in hereditary diseases, infections as well as in case of traumatic or light injuries. Therefore, a number of attempts has been undertaken so far to inhibit the microglia, with varying success. The task remains to study behaviour of the microglia and their interaction with other retinal cell populations in more detail with respect to released factors and expressed receptors including the time points of the corresponding events. The goal has to be to find a better balance between helpful and detrimental actions of the microglia.
Hurley, James B.; Lindsay, Kenneth J.; Du, Jianhai
The vertebrate retina has specific functions and structures that give it a unique set of constraints on the way in which it can produce and use metabolic energy. The retina’s response to illumination influences its energy requirements, and the retina’s laminated structure influences the extent to which neurons and glia can access metabolic fuels. There are fundamental differences between energy metabolism in retina and that in brain. The retina relies on aerobic glycolysis much more than the brain does, and morphological differences between retina and brain limit the types of metabolic relationships that are possible between neurons and glia. This Mini-Review summarizes the unique metabolic features of the retina with a focus on the role of lactate shuttling. PMID:25801286
Du, Jianhai; Linton, Jonathan D; Hurley, James B
Vertebrate retinas have several characteristics that make them particularly interesting from a metabolic perspective. The retinas have a highly laminated structure, high energy demands, and they share several metabolic features with tumors, such as a strong Warburg effect and abundant pyruvate kinase M2 isoform expression. The energy demands of retinas are both qualitatively and quantitatively different in light and darkness and metabolic dysfunction could cause retinal degeneration. Stable isotope-based metabolic analysis with mass spectrometry is a powerful tool to trace the dynamic metabolic reactions and reveal novel metabolic pathways within cells and between cells in retina. Here, we describe methods to quantify retinal metabolism in intact retinas and discuss applications of these methods to the understanding of neuron-glia interaction, light and dark adaptation, and retinal degenerative diseases.
Abd-El-Barr, Muhammad M.; Pennesi, Mark E.; Saszik, Shannon M.; Barrow, Andrew J.; Lem, Janis; Bramblett, Debra E.; Paul, David L.; Frishman, Laura J.; Wu, Samuel M.
A monumental task of the mammalian retina is to encode an enormous range (>109-fold) of light intensities experienced by the animal in natural environments. Retinal neurons carry out this task by dividing labor into many parallel rod and cone synaptic pathways. Here we study the operational plan of various rod- and cone-mediated pathways by analyzing electroretinograms (ERGs), primarily b-wave responses, in dark-adapted wildtype, connexin36 knockout, depolarizing rod–bipolar cell (DBCR) knockout, and rod transducin alpha-subunit knockout mice [WT, Cx36(−/−), Bhlhb4(−/−), and Trα(−/−)]. To provide additional insight into the cellular origins of various components of the ERG, we compared dark-adapted ERG responses with response dynamic ranges of individual retinal cells recorded with patch electrodes from dark-adapted mouse retinas published from other studies. Our results suggest that the connexin36-mediated rod–cone coupling is weak when light stimulation is weak and becomes stronger as light stimulation increases in strength and that rod signals may be transmitted to some DBCCs via direct chemical synapses. Moreover, our analysis indicates that DBCR responses contribute about 80% of the overall DBC response to scotopic light and that rod and cone signals contribute almost equally to the overall DBC responses when stimuli are strong enough to saturate the rod bipolar cell response. Furthermore, our study demonstrates that analysis of ERG b-wave of dark-adapted, pathway-specific mutants can be used as an in vivo tool for dissecting rod and cone synaptic pathways and for studying the functions of pathway-specific gene products in the retina. PMID:19587322
Marth, Jamey D; Grewal, Prabhjit K
Glycosylation produces a diverse and abundant repertoire of glycans, which are collectively known as the glycome. Glycans are one of the four fundamental macromolecular components of all cells, and are highly regulated in the immune system. Their diversity reflects their multiple biological functions that encompass ligands for proteinaceous receptors known as lectins. Since the discovery that selectins and their glycan ligands are important for the regulation of leukocyte trafficking, it has been shown that additional features of the vertebrate immune system are also controlled by endogenous cellular glycosylation. This Review focuses on the emerging immunological roles of the mammalian glycome.
Grimes, William N.; Li, Wei; Chávez, Andrés E.; Diamond, Jeffrey S.
In the mammalian retina, A17 amacrine cells provide reciprocal inhibitory feedback to rod bipolar cells, thereby shaping the time course of visual signaling in vivo. Previous results indicate that A17 feedback can be triggered by Ca2+ influx through Ca2+ permeable AMPARs and can occur independently of voltage-gated Ca2+ (Cav) channels, whose presence and functional role in A17 dendrites have not been explored. Here, we combine electrophysiology, calcium imaging and immunohistochemistry to show that L-type Cav channels in rat A17 amacrine cells are located at the sites of reciprocal synaptic feedback, but their contribution to GABA release is diminished by large-conductance Ca2+-activated potassium (BK) channels, which suppress postsynaptic depolarization in A17s and limit Cav channel activation. We also show that BK channels, by limiting GABA release from A17s, regulate the flow of excitatory synaptic transmission through the rod pathway. PMID:19363492
Rohde, Kristian; Klein, David C; Møller, Morten; Rath, Martin F
Retina and anterior neural fold homeobox (Rax) gene encodes a transcription factor essential for vertebrate eye development. Recent microarray studies indicate that Rax is expressed in the adult rat pineal gland and retina. The present study reveals that Rax expression levels in the rat change significantly during retinal development with a peak occurring at embryonic day 18, whereas Rax expression in the pineal is relatively delayed and not detectable until embryonic day 20. In both tissues, Rax is expressed throughout postnatal development into adulthood. In the mature rat pineal gland, the abundance of Rax transcripts increases 2-fold during the light period with a peak occurring at dusk. These findings are consistent with the evidence that Rax is of functional importance in eye development and suggest a role of Rax in the developing pineal gland. In addition, it would appear possible that Rax contributes to phenotype maintenance in the mature retina and pineal gland and may facilitate 24-h changes in the pineal transcriptome.
Castanheira, Paula; Torquetti, Leonardo Torquetti; Magalhãs, Débora Rodrigues Soares; Nehemy, Marcio B; Goes, Alfredo M
To evaluate DAPI (4',6-diamidino-2-phenylindole) as a nuclear tracer of stem cell migration and incorporation it was observed the pattern of retinal integration and differentiation of mesenchymal stem cells (MSCs) injected into the vitreous cavity of rat eyes with retinal injury. For this purpose adult rat retinas were submitted to laser damage followed by transplantation of DAPI-labeled BM-MSCs grafts and double-labeled DAPI and quantum dot-labeled BM-MSCs. To assess a possible DAPI diffusion as well as the integration and differentiation of DAPI-labeled BM-MSCs in laser-injured retina, host retinas were evaluated 8 weeks after injury/transplantation. It was demonstrated that, 8 weeks after the transplant, most of the retinal cells in all neural retinal presented nuclear DAPI labeling, specifically in the outer nuclear layer (ONL), inner nuclear layer (INL), and ganglion cell layer (GCL). Meanwhile, at this point, most of the double-labeled BM-MSCs (DAPI and quantum dot) remained in the vitreous cavity and no retinal cells presented the quantum dot marker. Based on these evidences we concluded that DAPI diffused to adjacent retinal cells while the nanocrystals remained labeling only the transplanted BM-MSCs. Therefore, DAPI is not a useful marker for stem cells in vivo tracing experiments because the DAPI released from dying cells in moment of the transplant are taken up by host cells in the tissue.
Tokuda, Kazuhiro; Kuramitsu, Yasuhiro; Byron, Baron; Kitagawa, Takao; Tokuda, Nobuko; Kobayashi, Daiki; Nagayama, Megumi; Araki, Norie; Sonoda, Koh-Hei; Nakamura, Kazuyuki
Glutamate has been shown to induce neural progenitor cells in the adult vertebrate retina. However, protein dynamics during progenitor cell induction by glutamate are not fully understood. To identify specific proteins involved in the process, we employed two-dimensional electrophoresis-based proteomics on glutamate untreated and treated retinal ex vivo sections. Rat retinal tissues were incubated with 1 mM glutamate for 1 h, followed by incubation in glutamate-free media for a total of 24 h. Consistent with prior reports, it was found that mitotic cells appeared in the outer nuclear layer without any histological damage. Immunohistological evaluations and immunoblotting confirmed the emergence of neuronal progenitor cells in the mature retina treated with glutamate. Proteomic analysis revealed the up-regulation of dihydropyrimidinase-related protein 3 (DRP-3), DRP-2 and stress-induced-phosphoprotein 1 (STIP1) during neural progenitor cell induction by glutamate. Moreover, mRNA expression of DRP-3, especially, its long isoform, robustly increased in the treated retina compared to that in the untreated retina. These results may indicate that glutamate induces neural progenitor cells in the mature rat retina by up-regulating the proteins which mediate cell mitosis and neurite growth. - Highlights: • Glutamate induced neuronal progenitor cells in the mature rat retina. • Proteomic analysis revealed the up-regulation of DRP-3, DRP-2 and STIP1. • mRNA expression of DRP-3, especially, its long isoform, robustly increased.
Gonzalez-Santos, Juana Maria; Cao, Huibi; Wang, Anan; Koehler, David R.; Martin, Bernard; Navab, Roya; Hu, Jim
Our progress in understanding mammalian gene function has lagged behind that of gene identification. New methods for mammalian gene functional analysis are needed to accelerate the process. In yeast, the powerful genetic shuffle system allows deletion of any chromosomal gene by homologous recombination and episomal expression of a mutant allele in the same cell. Here, we report a method for mammalian cells, which employs a helper-dependent adenoviral (HD-Ad) vector to synthesize small hairpin (sh) RNAs to knock-down the expression of an endogenous gene by targeting untranslated regions (UTRs). The vector simultaneously expresses an exogenous version of the same gene (wild-type or mutant allele) lacking the UTRs for functional analysis. We demonstrated the utility of the method by using PRPF3, which encodes the human RNA splicing factor Hprp3p. Recently, missense mutations in PRPF3 were found to cause autosomal-dominant Retinitis Pigmentosa, a form of genetic eye diseases affecting the retina. We knocked-down endogenous PRPF3 in multiple cell lines and rescued the phenotype (cell death) with exogenous PRPF3 cDNA, thereby creating a genetic complementation method. Because Ad vectors can efficiently transduce a wide variety of cell types, and many tissues in vivo, this method could have a wide application for gene function studies. PMID:15944448
The ON-OFF direction selective ganglion cells (DSGCs) in the mammalian retina code image motion by responding much more strongly to movement in one direction. They do so by receiving inhibitory inputs selectively from a particular sector of processes of the overlapping starburst amacrine cells, a type of retinal interneuron. The mechanisms of establishment and regulation of this selective connection are unknown. Here, we report that in the rat retina, the morphology, physiology of the ON-OFF DSGCs and the circuitry for coding motion directions develop normally with pharmacological blockade of GABAergic, cholinergic activity and/or action potentials for over two weeks from birth. With recent results demonstrating light independent formation of the retinal DS circuitry, our results strongly suggest the formation of the circuitry, i.e., the connections between the second and third order neurons in the visual system, can be genetically programmed, although emergence of direction selectivity in the visual cortex appears to require visual experience. PMID:21573161
Hausselt, Susanne E.; Breuninger, Tobias; Castell, Xavier; Denk, Winfried; Margolis, David J.; Detwiler, Peter B.
Dendritic signals play an essential role in processing visual information in the retina. To study them in neurites too small for electrical recording, we developed an instrument that combines a multi-photon (MP) microscope with a through-the-objective high-resolution visual stimulator. An upright microscope was designed that uses the objective lens for both MP imaging and delivery of visual stimuli to functionally intact retinal explants or eyecup preparations. The stimulator consists of a miniature liquid-crystal-on-silicon display coupled into the optical path of an infrared-excitation laser-scanning microscope. A pair of custom-made dichroic filters allows light from the excitation laser and three spectral bands (‘colors’) from the stimulator to reach the retina, leaving two intermediate bands for fluorescence imaging. Special optics allow displacement of the stimulator focus relative to the imaging focus. Spatially resolved changes in calcium-indicator fluorescence in response to visual stimuli were recorded in dendrites of different types of mammalian retinal neurons. PMID:19023590
Liao, Meng-Lin; Peng, Wei-Hau; Kan, Daphne; Chien, Chung-Liang
α-Internexin is a member of the neuronal intermediate filament (nIF) protein family, which also includes peripherin and neurofilament (NF) triplet proteins. Previous studies found that expression of α-internexin precedes that of the NF triplet proteins in mammals and suggested that α-internexin plays a key role in the neuronal cytoskeleton network during development. In this study, we aimed to analyze the expression patterns and function of internexin neuronal intermediate filament protein-alpha a (inaa), the encoding gene of which is a homolog of the mammalian α-internexin, during retinal development in zebrafish. Via in vitro and in vivo studies, we demonstrated that zebrafish inaa is an α-internexin homolog that shares characteristics with nIFs. An immunohistochemical analysis of zebrafish revealed that inaa was distributed dynamically in the developing retina. It was widely localized in retinal neuroepithelial cells at 1 day postfertilization (dpf), and was mainly found in the ganglion cell layer (GCL) and inner part of the inner nuclear layer (INL) from 3-9 dpf; after 14 dpf, it was restricted to the outer nuclear layer (ONL). Moreover, we demonstrated for the first time that inaa acted distinctively from the cytoskeletal scaffold of zebrafish cone photoreceptors during development. In conclusion, we demonstrated the morphological features of a novel nIF, inaa, and illustrated its developmental expression pattern in the zebrafish retina. J. Comp. Neurol. 524:3810-3826, 2016. © 2016 Wiley Periodicals, Inc.
Yu, D Y; Cringle, S J
Maintenance of an adequate oxygen supply to the retina is critical for retinal function. In species with vascularised retinas, such as man, oxygen is delivered to the retina via a combination of the choroidal vascular bed, which lies immediately behind the retina, and the retinal vasculature, which lies within the inner retina. The high-oxygen demands of the retina, and the relatively sparse nature of the retinal vasculature, are thought to contribute to the particular vulnerability of the retina to vascular disease. A large proportion of retinal blindness is associated with diseases having a vascular component, and disrupted oxygen supply to the retina is likely to be a critical factor. Much attention has therefore been directed at determining the intraretinal oxygen environment in healthy and diseased eyes. Measurements of oxygen levels within the retina have largely been restricted to animal studies in which oxygen sensitive microelectrodes can be used to obtain high-resolution measurements of oxygen tension as a function of retinal depth. Such measurements can immediately identify which retinal layers are supplied with oxygen from the different vascular elements. Additionally, in the outer retinal layers, which do not have any intrinsic oxygen sources, the oxygen distribution can be analysed mathematically to quantify the oxygen consumption rate of specific retinal layers. This has revealed a remarkable heterogeneity of oxygen requirements of different components of the outer retina, with the inner segments of the photoreceptors being the dominant oxygen consumers. Since the presence of the retinal vasculature precludes such a simple quantitative analysis of local oxygen consumption within the inner retina, our understanding of the oxygen needs of the inner retinal components is much less complete. Although several lines of evidence suggest that in the more commonly studied species such as cat, pig, and rat, the oxygen demands of the inner retina as a whole is
Mollema, Nissa J; Yuan, Yang; Jelcick, Austin S; Sachs, Andrew J; von Alpen, Désirée; Schorderet, Daniel; Escher, Pascal; Haider, Neena B
The majority of diseases in the retina are caused by genetic mutations affecting the development and function of photoreceptor cells. The transcriptional networks directing these processes are regulated by genes such as nuclear hormone receptors. The nuclear hormone receptor gene Rev-erb alpha/Nr1d1 has been widely studied for its role in the circadian cycle and cell metabolism, however its role in the retina is unknown. In order to understand the role of Rev-erb alpha/Nr1d1 in the retina, we evaluated the effects of loss of Nr1d1 to the developing retina and its co-regulation with the photoreceptor-specific nuclear receptor gene Nr2e3 in the developing and mature retina. Knock-down of Nr1d1 expression in the developing retina results in pan-retinal spotting and reduced retinal function by electroretinogram. Our studies show that NR1D1 protein is co-expressed with NR2E3 in the outer neuroblastic layer of the developing mouse retina. In the adult retina, NR1D1 is expressed in the ganglion cell layer and is co-expressed with NR2E3 in the outer nuclear layer, within rods and cones. Several genes co-targeted by NR2E3 and NR1D1 were identified that include: Nr2c1, Recoverin, Rgr, Rarres2, Pde8a, and Nupr1. We examined the cyclic expression of Nr1d1 and Nr2e3 over a twenty-four hour period and observed that both nuclear receptors cycle in a similar manner. Taken together, these studies reveal a novel role for Nr1d1, in conjunction with its cofactor Nr2e3, in regulating transcriptional networks critical for photoreceptor development and function.
Gage, M J
Understanding the adaptive significance of sperm form and function has been a challenge to biologists because sperm are highly specialized cells operating at a microscopic level in a complex environment. A fruitful course of investigation has been to use the comparative approach. This comparative study attempts to address some fundamental questions of the evolution of mammalian sperm morphometry. Data on sperm morphometry for 445 mammalian species were collated from published sources. I use contemporary phylogenetic analysis to control for the inherent non-independence of species and explore relationships between the morphometric dimensions of the three essential spermatozoal components: head, mid-piece and flagellum. Energy for flagellar action is metabolized by the mitochondrial-dense mid-piece and these combine to propel the sperm head, carrying the male haplotype, to the ovum. I therefore search for evolutionary associations between sperm morphometry and body mass, karyotype and the duration of oestrus. In contrast to previous findings, there is no inverse correlation between body weight and sperm length. Sperm mid-piece and flagellum lengths are positively associated with both head length and area, and the slopes of these relationships are discussed. Flagellum length is positively associated with mid-piece length but, in contrast to previous research and after phylogenetic control, I find no relationship between flagellum length and the volume of the mitochondrial sheath. Sperm head dimensions are not related to either genome mass or chromosome number, and there are no relationships between sperm morphometry and the duration of oestrus. PMID:9474794
Casini, G; Trasarti, L; Andolfi, L; Bagnoli, P
Tachykinin (TK) peptides, which include substance P, neurokinin A, two neurokinin A-related peptides and neurokinin B, are widely present in the nervous system, including the retina, where they act as neurotransmitters/modulators as well as growth factors. In the present study, we investigated the maturation of TK-immunoreactive (IR) cells in the rabbit retina with the aim of further contributing to the knowledge of the development of transmitter-identified retinal cell populations. In the adult retina, the pattern of TK immunostaining is consistent with the presence of TK peptides in amacrine, displaced amacrine, interplexiform and ganglion cells. In the newborn retina, intensely immunostained TK-IR somata are located in the ganglion cell layer (GCL) and in the inner nuclear layer (INL) adjacent to the inner plexiform layer (IPL). They are characterized by an oval-shaped cell body originating a single process without ramifications. TK-IR processes are occasionally observed in the IPL and in the outer plexiform layer (OPL). Long TK-IR fiber bundles are observed in the ganglion cell axon layer. TK-IR profiles resembling small somata are rarely observed in the INL adjacent to the OPL. At postnatal day (PND) 2, some TK-IR cells display more complex morphologic features, including processes with secondary ramifications. Long TK-IR processes in the IPL are often seen to terminate with growth cones. Between PND 6 and PND 11 (eye opening), there is a dramatic increase in the number of immunolabeled processes with growth cones both in the IPL and in the OPL and the mature lamination of TK-IR fibers in laminae 1, 3 and 5 of the IPL is established. TK-IR cells attain mature morphological characteristics and the rare, putative TK-IR somata in the distal INL are no longer observed. After eye opening, growth cones are not present and the pattern typical of the adult is reached. These observations indicate that the development of TK-IR cells can be divided into an early phase
Safdari, Reza; Mokhtaran, Mehrshad; Tahmasebian, Shahram
Introduction: Electronic medical records as one of major parts of electronic health records is an important application of Medical Informatics. EMR includes different types of data, Graphical items being one of these data types. To this end, a standard structure for storing and recovering and finally exchanging this data type is required. In order to standardize information items in this research, UMLS standard is used. In this research, graphical information from fondues designing in retina surgery forms is used for the task of implementation. Implementation: Three-layer software architecture is used for implementation of this system, which includes user interface, data base access and business logic. XML database is used for storing and exchanging of data. User interface is designed by the means of Adobe Flash. Also in the user interface for eye examinations, appropriate icons compatible with current pathologies in retina examinations are considered and UMLS codes are used for standardizations purposes. Results: As this project is independently implemented in Adobe Flash, it can be run in most of electronic patient records software. For evaluation purposes of this research, an EMR system for eye clinics is used. Tree structure is used for data entry and finally a text report based on the entered data will be generated. By storing graphical items in this software editing and searching in medical concepts and also comparing features will be available. Conclusion: One of the data items that we encounter in various medical records is graphical data. In order to cover the patient’s complete electronic medical records, the Electronic Implementation of this information is important. For this purpose, graphical items in retina surgery forms were used and finally a software application for drawing retina picture was developed. Also, XML files were used for the purpose of storing valuable medical data from the pictures, and also UMLS were applied for the standardization
Reese, Benjamin E.
Our understanding of the development of the retina and visual pathways has seen enormous advances during the past twenty-five years. New imaging technologies, coupled with advances in molecular biology, have permitted a fuller appreciation of the histotypical events associated with proliferation, fate determination, migration, differentiation, pathway navigation, target innervation, synaptogenesis and cell death, and in many instances, in understanding the genetic, molecular, cellular and activity-dependent mechanisms underlying those developmental changes. The present review considers those advances associated with the lineal relationships between retinal nerve cells, the production of retinal nerve cell diversity, the migration, patterning and differentiation of different types of retinal nerve cells, the determinants of the decussation pattern at the optic chiasm, the formation of the retinotopic map, and the establishment of ocular domains within the thalamus. PMID:20647017
Battu, Rajani; Dabir, Supriya; Khanna, Anjani; Kumar, Anupama Kiran; Roy, Abhijit Sinha
Adaptive optics is a relatively new tool that is available to ophthalmologists for study of cellular level details. In addition to the axial resolution provided by the spectral-domain optical coherence tomography, adaptive optics provides an excellent lateral resolution, enabling visualization of the photoreceptors, blood vessels and details of the optic nerve head. We attempt a mini review of the current role of adaptive optics in retinal imaging. PubMed search was performed with key words Adaptive optics OR Retina OR Retinal imaging. Conference abstracts were searched from the Association for Research in Vision and Ophthalmology (ARVO) and American Academy of Ophthalmology (AAO) meetings. In total, 261 relevant publications and 389 conference abstracts were identified. PMID:24492503
Battu, Rajani; Dabir, Supriya; Khanna, Anjani; Kumar, Anupama Kiran; Roy, Abhijit Sinha
Adaptive optics is a relatively new tool that is available to ophthalmologists for study of cellular level details. In addition to the axial resolution provided by the spectral-domain optical coherence tomography, adaptive optics provides an excellent lateral resolution, enabling visualization of the photoreceptors, blood vessels and details of the optic nerve head. We attempt a mini review of the current role of adaptive optics in retinal imaging. PubMed search was performed with key words Adaptive optics OR Retina OR Retinal imaging. Conference abstracts were searched from the Association for Research in Vision and Ophthalmology (ARVO) and American Academy of Ophthalmology (AAO) meetings. In total, 261 relevant publications and 389 conference abstracts were identified.
Tao, Zui; Zhao, Chen; Jian, Qian; Gillies, Mark; Xu, Haiwei; Yin, Zheng Qin
Retinal regeneration and repair are severely impeded in higher mammalian animals. Although Müller cells can be activated and show some characteristics of progenitor cells when injured or under pathological conditions, they quickly form gliosis scars. Unfortunately, the basic mechanisms that impede retinal regeneration remain unknown. We studied retinas from Royal College of Surgeon (RCS) rats and found that let-7 family molecules, let-7e and let-7i, were significantly overexpressed in Müller cells of degenerative retinas. It demonstrated that down-regulation of the RNA binding protein Lin28B was one of the key factors leading to the overexpression of let-7e and let-7i. Lin28B ectopic expression in the Müller cells suppressed overexpression of let-7e and let-7i, stimulated and mobilized Müller glia de-differentiation, proliferation, promoted neuronal commitment, and inhibited glial fate acquisition of de-differentiated Müller cells. ERG recordings revealed that the amplitudes of a-wave and b-wave were improved significantly after Lin28B was delivered into the subretinal space of RCS rats. In summary, down-regulation of Lin28B as well as up-regulation of let-7e and let-7i may be the main factors that impede Müller cell de-differentiation and proliferation in the retina of RCS rats. PMID:27384999
Luft, Veronika; Reinhard, Jacqueline; Shibuya, Masabumi; Fischer, Klaus D; Faissner, Andreas
The seven main cell types in the mammalian retina arise from multipotent retinal progenitor cells, a process that is tightly regulated by intrinsic and extrinsic signals. However, the molecular mechanisms that control proliferation, differentiation and cell-fate decisions of retinal progenitor cells are not fully understood yet. Here, we report that the guanine nucleotide exchange factor Vav3, a regulator of Rho-GTPases, is involved in retinal development. We demonstrate that Vav3 is expressed in the mouse retina during the embryonic period. In order to study the role of Vav3 in the developing retina, we generate Vav3-deficient mice. The loss of Vav3 results in an accelerated differentiation of retinal ganglion cells and cone photoreceptors during early and late embryonic development. We provide evidence that more retinal progenitor cells express the late progenitor marker Sox9 in Vav3-deficient mice than in wild-types. This premature differentiation is compensated during the postnatal period and late-born cell types such as bipolar cells and Müller glia display normal numbers. Taken together, our data imply that Vav3 is a regulator of retinal progenitor cell differentiation, thus highlighting a novel role for guanine nucleotide exchange factors in retinogenesis.
Thoreson, Wallace B; Mangel, Stuart C
Lateral interactions in the outer retina, particularly negative feedback from horizontal cells to cones and direct feed-forward input from horizontal cells to bipolar cells, play a number of important roles in early visual processing, such as generating center-surround receptive fields that enhance spatial discrimination. These circuits may also contribute to post-receptoral light adaptation and the generation of color opponency. In this review, we examine the contributions of horizontal cell feedback and feed-forward pathways to early visual processing. We begin by reviewing the properties of bipolar cell receptive fields, especially with respect to modulation of the bipolar receptive field surround by the ambient light level and to the contribution of horizontal cells to the surround. We then review evidence for and against three proposed mechanisms for negative feedback from horizontal cells to cones: 1) GABA release by horizontal cells, 2) ephaptic modulation of the cone pedicle membrane potential generated by currents flowing through hemigap junctions in horizontal cell dendrites, and 3) modulation of cone calcium currents (I(Ca)) by changes in synaptic cleft proton levels. We also consider evidence for the presence of direct horizontal cell feed-forward input to bipolar cells and discuss a possible role for GABA at this synapse. We summarize proposed functions of horizontal cell feedback and feed-forward pathways. Finally, we examine the mechanisms and functions of two other forms of lateral interaction in the outer retina: negative feedback from horizontal cells to rods and positive feedback from horizontal cells to cones.
Thoreson, Wallace B.; Mangel, Stuart C.
Lateral interactions in the outer retina, particularly negative feedback from horizontal cells to cones and direct feed-forward input from horizontal cells to bipolar cells, play a number of important roles in early visual processing, such as generating center-surround receptive fields that enhance spatial discrimination. These circuits may also contribute to post-receptoral light adaptation and the generation of color opponency. In this review, we examine the contributions of horizontal cell feedback and feed-forward pathways to early visual processing. We begin by reviewing the properties of bipolar cell receptive fields, especially with respect to modulation of the bipolar receptive field surround by the ambient light level and to the contribution of horizontal cells to the surround. We then review evidence for and against three proposed mechanisms for negative feedback from horizontal cells to cones: 1) GABA release by horizontal cells, 2) ephaptic modulation of the cone pedicle membrane potential generated by currents flowing through hemigap junctions in horizontal cell dendrites, and 3) modulation of cone calcium currents (ICa) by changes in synaptic cleft proton levels. We also consider evidence for the presence of direct horizontal cell feed-forward input to bipolar cells and discuss a possible role for GABA at this synapse. We summarize proposed functions of horizontal cell feedback and feed-forward pathways. Finally, we examine the mechanisms and functions of two other forms of lateral interaction in the outer retina: negative feedback from horizontal cells to rods and positive feedback from horizontal cells to cones. PMID:22580106
Osakada, Fumitaka; Hirami, Yasuhiko; Takahashi, Masayo
Embryonic stem (ES) cells, which are derived from the inner cell mass of mammalian blastocyst stage embryos, have the ability to differentiate into any cell type in the body and to grow indefinitely while maintaining pluripotency. During development, cells undergo progressive and irreversible differentiation into specialized adult cell types. Remarkably, in spite of this restriction in potential, adult somatic cells can be reprogrammed and returned to the naive state of pluripotency found in the early embryo simply by forcing expression of a defined set of transcription factors. These induced pluripotent stem (iPS) cells are molecularly and functionally equivalent to ES cells and provide powerful in vitro models for development, disease, and drug screening, as well as material for cell replacement therapy. Since functional impairment results from cell loss in most central nervous system (CNS) diseases, recovery of lost cells is an important treatment strategy. Although adult neurogenesis occurs in restricted regions, the CNS has poor potential for regeneration to compensate for cell loss. Thus, cell transplantation into damaged or diseased CNS tissues is a promising approach to treating various neurodegenerative disorders. Transplantation of photoreceptors or retinal pigment epithelium cells derived from human ES cells can restore some visual function. Patient-specific iPS cells may lead to customized cell therapy. However, regeneration of retinal function will require a detailed understanding of eye development, visual system circuitry, and retinal degeneration pathology. Here, we review the current progress in retinal regeneration, focusing on the therapeutic potential of pluripotent stem cells.
Bose, Sayantan; Schönenbrücher, Holger; Richt, Jürgen A; Casey, Thomas A; Rasmussen, Mark A; Kehrli, Marcus E; Petrich, Jacob W
Recently, we have proposed that the fluorescence spectra of sheep retina can be well correlated with the presence or absence of scrapie. Scrapie is the most widespread TSE (transmissible spongiform encephalopathy) affecting sheep and goats worldwide. Mice eyes have been previously reported as a model system to study age-related accumulation of lipofuscin, which has been investigated by monitoring the increasing fluorescence with age covering its entire life span. The current work aims at developing mice retina as a convenient model system to diagnose scrapie and other fatal TSE diseases in animals such as sheep and cows. The objective of the research reported here was to determine whether the spectral features are conserved between two different species namely mice and sheep, and whether an appropriate small animal model system could be identified for diagnosis of scrapie based on the fluorescence intensity in retina. The results were consistent with the previous reports on fluorescence studies of healthy and scrapie-infected retina of sheep. The fluorescence from the retinas of scrapie-infected sheep was significantly more intense and showed more heterogeneity than that from the retinas of uninfected mice. Although the structural characteristics of fluorescence spectra of scrapie-infected sheep and mice eyes are slightly different, more importantly, murine retinas reflect the enhancement of fluorescence intensity upon infecting the mice with scrapie, which is consistent with the observations in sheep eyes.
The inability of proteins to potently penetrate mammalian cells limits their usefulness as tools and therapeutics. When fused to superpositively charged GFP, proteins rapidly (within minutes) entered five different types of mammalian cells with potency up to ∼100-fold greater than that of corresponding fusions with known protein transduction domains (PTDs) including Tat, oligoarginine, and penetratin. Ubiquitin-fused supercharged GFP when incubated with human cells was partially deubiquitinated, suggesting that proteins delivered with supercharged GFP can access the cytosol. Likewise, supercharged GFP delivered functional, nonendosomal recombinase enzyme with greater efficiencies than PTDs in vitro and also delivered functional recombinase enzyme to the retinae of mice when injected in vivo. PMID:20545362
Borst, Alexander; Helmstaedter, Moritz
Motion-sensitive neurons have long been studied in both the mammalian retina and the insect optic lobe, yet striking similarities have become obvious only recently. Detailed studies at the circuit level revealed that, in both systems, (i) motion information is extracted from primary visual information in parallel ON and OFF pathways; (ii) in each pathway, the process of elementary motion detection involves the correlation of signals with different temporal dynamics; and (iii) primary motion information from both pathways converges at the next synapse, resulting in four groups of ON-OFF neurons, selective for the four cardinal directions. Given that the last common ancestor of insects and mammals lived about 550 million years ago, this general strategy seems to be a robust solution for how to compute the direction of visual motion with neural hardware.
HIRANO, ARLENE A.; BRANDSTÄTTER, JOHANN H.; BRECHA, NICHOLAS C.
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
Frankenberg, Stephen R; de Barros, Flavia R O; Rossant, Janet; Renfree, Marilyn B
The blastocyst is a mammalian invention that carries the embryo from cleavage to gastrulation. For such a simple structure, it exhibits remarkable diversity in its mode of formation, morphology, longevity, and intimacy with the uterine endometrium. This review explores this diversity in the light of the evolution of viviparity, comparing the three main groups of mammals: monotremes, marsupials, and eutherians. The principal drivers in blastocyst evolution were loss of yolk coupled with evolution of the placenta. An important outcome of blastocyst development is differentiation of two extraembryonic lineages (trophoblast and hypoblast) that contribute to the placenta. While in many species trophoblast segregation is often coupled with blastocyst formation, in marsupials and at least some Afrotherians, these events do not coincide. Thus, many questions regarding the conservation of molecular mechanisms controlling these events are of great interest but currently unresolved. For further resources related to this article, please visit the WIREs website.
Wan, Yinan; Almeida, Alexandra D.; Rulands, Steffen; Chalour, Naima; Muresan, Leila; Wu, Yunmin; Simons, Benjamin D.; He, Jie; Harris, William A.
Clonal analysis is helping us understand the dynamics of cell replacement in homeostatic adult tissues (Simons and Clevers, 2011). Such an analysis, however, has not yet been achieved for continuously growing adult tissues, but is essential if we wish to understand the architecture of adult organs. The retinas of lower vertebrates grow throughout life from retinal stem cells (RSCs) and retinal progenitor cells (RPCs) at the rim of the retina, called the ciliary marginal zone (CMZ). Here, we show that RSCs reside in a niche at the extreme periphery of the CMZ and divide asymmetrically along a radial (peripheral to central) axis, leaving one daughter in the peripheral RSC niche and the other more central where it becomes an RPC. We also show that RPCs of the CMZ have clonal sizes and compositions that are statistically similar to progenitor cells of the embryonic retina and fit the same stochastic model of proliferation. These results link embryonic and postembryonic cell behaviour, and help to explain the constancy of tissue architecture that has been generated over a lifetime. PMID:26893352
Wan, Yinan; Almeida, Alexandra D; Rulands, Steffen; Chalour, Naima; Muresan, Leila; Wu, Yunmin; Simons, Benjamin D; He, Jie; Harris, William A
Clonal analysis is helping us understand the dynamics of cell replacement in homeostatic adult tissues (Simons and Clevers, 2011). Such an analysis, however, has not yet been achieved for continuously growing adult tissues, but is essential if we wish to understand the architecture of adult organs. The retinas of lower vertebrates grow throughout life from retinal stem cells (RSCs) and retinal progenitor cells (RPCs) at the rim of the retina, called the ciliary marginal zone (CMZ). Here, we show that RSCs reside in a niche at the extreme periphery of the CMZ and divide asymmetrically along a radial (peripheral to central) axis, leaving one daughter in the peripheral RSC niche and the other more central where it becomes an RPC. We also show that RPCs of the CMZ have clonal sizes and compositions that are statistically similar to progenitor cells of the embryonic retina and fit the same stochastic model of proliferation. These results link embryonic and postembryonic cell behaviour, and help to explain the constancy of tissue architecture that has been generated over a lifetime.
of and posterior to the ORCC are prominent features in these retinal degenerations. This dissertation represents the first report of quantitative OCT results in hereditary retinal degenerations in experimental animals and humans. Considering that OCT is being used increasingly to investigate, diagnose, and monitor human retinal diseases, insight gained in these studies may direct further studies in mammalian and human retinas and advance this technology to its full potential in ophthalmology.
Nery, Joseph R.; Urich, Mark; Puddifoot, Clare A.; Johnson, Nicholas D.; Lucero, Jacinta; Huang, Yun; Dwork, Andrew J.; Schultz, Matthew D.; Yu, Miao; Tonti-Filippini, Julian; Heyn, Holger; Hu, Shijun; Wu, Joseph C.; Rao, Anjana; Esteller, Manel; He, Chuan; Haghighi, Fatemeh G.; Sejnowski, Terrence J.; Behrens, M. Margarita; Ecker, Joseph R.
DNA methylation is implicated in mammalian brain development and plasticity underlying learning and memory. We report the genome-wide composition, patterning, cell specificity, and dynamics of DNA methylation at single-base resolution in human and mouse frontal cortex throughout their lifespan. Widespread methylome reconfiguration occurs during fetal to young adult development, coincident with synaptogenesis. During this period, highly conserved non-CG methylation (mCH) accumulates in neurons, but not glia, to become the dominant form of methylation in the human neuronal genome. Moreover, we found an mCH signature that identifies genes escaping X-chromosome inactivation. Last, whole-genome single-base resolution 5-hydroxymethylcytosine (hmC) maps revealed that hmC marks fetal brain cell genomes at putative regulatory regions that are CG-demethylated and activated in the adult brain and that CG demethylation at these hmC-poised loci depends on Tet2 activity. PMID:23828890
Many simple experiments can be performed in the classroom to explore the physics of vision. Students can learn of the two types of receptive cells (rods and cones), their distribution on the retina and the existence of the blind spot.
Tian, N; Slaughter, M M
Amacrine and ganglion cells in the amphibian retina contain GABAB, as well as GABAA, receptors. Baclofen, a GABAB agonist, hyperpolarizes the dark membrane potential of these third order neurons and makes their light responses more transient. GABAB receptors in the retina have a similar agonist profile to GABAB receptors described at other sites in the brain. Namely, preferential activation by the R-enantiomer of baclofen, and agonist sensitivity in the order 3-aminopropylphosphinic acid > baclofen > 3-aminopropylphosphonic acid. The GABAB receptor was not activated by 4-aminobutylphosphonic acid. Several antagonists, such as phaclofen, saclofen, and 2-hydroxysaclofen, were ineffective in the amphibian retina. However, CGP35348 blocked the action of applied baclofen and produced effects on the light response that were opposite to those of baclofen. Applied agonists and antagonists support the hypothesis that GABAB receptors serve to regulate the balance of sustained and transient signals to the inner retina.
Bobu, Corina; Lahmam, Mohamed; Vuillez, Patrick; Ouarour, Ali; Hicks, David
To characterize rod and cone distribution and composition in two diurnal mouse-like rodents, retinas from adult Arvicanthis ansorgei and Lemniscomys barbarus were processed for immunohistochemistry using multiple rod- and cone-specific antibodies. Antibodies tested included rhodopsin, cone opsins, pan-arrestin and cone arrestin, recoverin, and cGMP dependent ion channel. In both species, retinas were composed of approximately 33% cones, and most antibodies gave similar staining patterns. Data show these two diurnal rodents possess large numbers of cones, organised in a strict anatomical array. This suggests that diurnal rodents in general possess elevated cone numbers and could constitute valuable models for investigating cone pathophysiology.
Tooker, Ryan E; Vigh, Jozsef
Nitric oxide (NO) synthesis in the retina is triggered by light stimulation. NO has been shown to modulate visual signal processing at multiple sites in the vertebrate retina, via activation of the most sensitive target of NO signaling, soluble guanylate cyclase. NO can also alter protein structure and function and exert biological effects directly by binding to free thiol groups of cysteine residues in a chemical reaction called S-nitrosylation. However, in the central nervous system, including the retina, this reaction has not been considered to be significant under physiological conditions. Here we provide immunohistochemical evidence for extensive S-nitrosylation that takes place in the goldfish and mouse retinas under physiologically relevant light intensities, in an intensity-dependent manner, with a strikingly similar pattern in both species. Pre-treatment with NEM, which occludes S-nitrosylation, or with TRIM, an inhibitor of neuronal NO synthase, eliminated the light-evoked increase in S-nitrosylated protein immunofluorescence (SNI) in the retinas of both species. Similarly, light did not increase SNI, above basal levels, in retinas of transgenic mice lacking neuronal NO synthase. Qualitative analysis of the light-adapted mouse retina with mass spectrometry revealed more than 300 proteins that were S-nitrosylated upon illumination, many of which are known to participate directly in retinal signal processing. Our data strongly suggest that in the retina, light-evoked NO production leads to extensive S-nitrosylation and that this process is a significant post-translational modification affecting a wide range of proteins under physiological conditions. PMID:25823749
Tooker, Ryan E; Vigh, Jozsef
Nitric oxide (NO) synthesis in the retina is triggered by light stimulation. NO has been shown to modulate visual signal processing at multiple sites in the vertebrate retina, via activation of the most sensitive target of NO signaling, soluble guanylate cyclase. NO can also alter protein structure and function and exert biological effects directly by binding to free thiol groups of cysteine residues in a chemical reaction called S-nitrosylation. However, in the central nervous system, including the retina, this reaction has not been considered to be significant under physiological conditions. Here we provide immunohistochemical evidence for extensive S-nitrosylation that takes place in the goldfish and mouse retinas under physiologically relevant light intensities, in an intensity-dependent manner, with a strikingly similar pattern in both species. Pretreatment with N-ethylmaleimide (NEM), which occludes S-nitrosylation, or with 1-(2-trifluromethylphenyl)imidazole (TRIM), an inhibitor of neuronal NO synthase, eliminated the light-evoked increase in S-nitrosylated protein immunofluorescence (SNI) in the retinas of both species. Similarly, light did not increase SNI, above basal levels, in retinas of transgenic mice lacking neuronal NO synthase. Qualitative analysis of the light-adapted mouse retina with mass spectrometry revealed more than 300 proteins that were S-nitrosylated upon illumination, many of which are known to participate directly in retinal signal processing. Our data strongly suggest that in the retina light-evoked NO production leads to extensive S-nitrosylation and that this process is a significant posttranslational modification affecting a wide range of proteins under physiological conditions.
Firth, Sally I.; Varela, Carolina; De La Villa, Pedro; Marshak, David W.
High levels of endogenous cholecystokinin (CCK) are present in the rat retina (Eskay & Beinfeld, 1982), but the cellular localization and physiological actions of CCK in the rat retina are uncertain. The goals of this study were to characterize the cells containing CCK, identify cell types that interact with CCK cells, and investigate the effects of CCK on rod bipolar cells. Rat retinas were labeled with antibody to gastrin-CCK (gCCK) using standard immunofluorescence techniques. Patch-clamp methods were used to record from dissociated rod bipolar cells from rats and mice. Gastrin-CCK immunoreactive (-IR) axons were evenly distributed throughout the retina in stratum 5 of the inner plexiform layer of the rat retina. However, the gCCK-IR somata were only detected in the ganglion cell layer in the peripheral retina. The gCCK-IR cells contained glutamate decarboxylase, and some of them also contained immunoreactive substance P. Labeled axons contacted PKC-IR rod bipolar cells, and recoverin-IR ON-cone bipolar cells. CCK-octapeptide inhibits GABAC but not GABAA mediated currents in dissociated rod bipolar cells. PMID:12511085
Chen, Xiaoyan; Lane, Stephen
We have used Monte Carlo simulation of autofluorescence in the retina to determine that noninvasive detection of nutritional iron deficiency is possible. Nutritional iron deficiency (which leads to iron deficiency anemia) affects more than 2 billion people worldwide, and there is an urgent need for a simple, noninvasive diagnostic test. Zinc protoporphyrin (ZPP) is a fluorescent compound that accumulates in red blood cells and is used as a biomarker for nutritional iron deficiency. We developed a computational model of the eye, using parameters that were identified either by literature search, or by direct experimental measurement to test the possibility of detecting ZPP non-invasively in retina. By incorporating fluorescence into Steven Jacques' original code for multi-layered tissue, we performed Monte Carlo simulation of fluorescence in the retina and determined that if the beam is not focused on a blood vessel in a neural retina layer or if part of light is hitting the vessel, ZPP fluorescence will be 10-200 times higher than background lipofuscin fluorescence coming from the retinal pigment epithelium (RPE) layer directly below. In addition we found that if the light can be focused entirely onto a blood vessel in the neural retina layer, the fluorescence signal comes only from ZPP. The fluorescence from layers below in this second situation does not contribute to the signal. Therefore, the possibility that a device could potentially be built and detect ZPP fluorescence in retina looks very promising.
Kalsbeek, Andries; Yi, Chun-Xia; Cailotto, Cathy; la Fleur, Susanne E; Fliers, Eric; Buijs, Ruud M
In mammals many behaviours (e.g. sleep-wake, feeding) as well as physiological (e.g. body temperature, blood pressure) and endocrine (e.g. plasma corticosterone concentration) events display a 24 h rhythmicity. These 24 h rhythms are induced by a timing system that is composed of central and peripheral clocks. The highly co-ordinated output of the hypothalamic biological clock not only controls the daily rhythm in sleep-wake (or feeding-fasting) behaviour, but also exerts a direct control over many aspects of hormone release and energy metabolism. First, we present the anatomical connections used by the mammalian biological clock to enforce its endogenous rhythmicity on the rest of the body, especially the neuro-endocrine and energy homoeostatic systems. Subsequently, we review a number of physiological experiments investigating the functional significance of this neuro-anatomical substrate. Together, this overview of experimental data reveals a highly specialized organization of connections between the hypothalamic pacemaker and neuro-endocrine system as well as the pre-sympathetic and pre-parasympathetic branches of the autonomic nervous system.
Neubauer, Katharina; Zieger, Barbara
Septins are GTP-binding and membrane-interacting proteins with a highly conserved domain structure involved in various cellular processes, including cytoskeleton organization, cytokinesis, and membrane dynamics. To date, 13 different septin genes have been identified in mammals (SEPT1 to SEPT12 and SEPT14), which can be classified into four distinct subgroups based on the sequence homology of their domain structure (SEPT2, SEPT3, SEPT6, and SEPT7 subgroup). The family members of these subgroups have a strong affinity for other septins and form apolar tri-, hexa-, or octameric complexes consisting of multiple septin polypeptides. The first characterized core complex is the hetero-trimer SEPT2-6-7. Within these complexes single septins can be exchanged in a subgroup-specific manner. Hexamers contain SEPT2 and SEPT6 subgroup members and SEPT7 in two copies each whereas the octamers additionally comprise two SEPT9 subgroup septins. The various isoforms seem to determine the function and regulation of the septin complex. Septins self-assemble into higher-order structures, including filaments and rings in orders, which are typical for different cell types. Misregulation of septins leads to human diseases such as neurodegenerative and bleeding disorders. In non-dividing cells such as neuronal tissue and platelets septins have been associated with exocytosis. However, many mechanistic details and roles attributed to septins are poorly understood. We describe here some important mammalian septin interactions with a special focus on the clinically relevant septin interactions. PMID:28224124
A new carotenoid has been isolated from the chicken retina for which the name galloxanthin is proposed. This substance has the properties of a hydroxy carotenoid or xanthophyll. It has not yet been crystallized. On a chromatogram of calcium carbonate it is adsorbed just below astaxanthin and above lutein. The absorption spectrum of galloxanthin lies in a region where natural carotenoids have not ordinarily been found. Its main, central absorption band falls at about 400 mmicro. The position of its spectrum suggests a conjugated system of eight double bonds. This relatively short polyene structure must be reconciled with very strong adsorption affinities. With antimony trichloride, galloxanthin yields a deep blue product, possessing a main absorption band at 785 to 795 mmicro, and a secondary maximum at about 710 mmicro which may not be due to galloxanthin itself. Galloxanthin appears to be one of the carotenoid filter pigments associated with cone vision in the chicken. It may act as an auxiliary to the other filter pigments in differentiating colors; or its primary function may be to exclude violet and near ultraviolet radiations for which the eye has a large chromatic aberration.
Bernard, Thierry M.
A digital programmable artificial retina (PAR) is a functional extension of a CMOS imager, in which every pixel is fitted with a local ADC and a tiny digital programmable processor. From an architectural viewpoint, a PAR is an SIMD array processor with local optical input. A PAR is aimed at processing images on-site until they can be output from the array under concentrated form. The overall goal is to get compact, fast and inexpensive vision systems, in particular for robotics applications. A 256 by 256 PAR with up to a few tens bits of local memory per pixel is now within reach at reasonable cost. However, whereas the local memory size benefits quadratically from the feature size decrease, wiring density improvement can only be linear, at best. So control should become more complex with the danger of a growing proportion of the digital pixel area being devoted to instruction or address decoding. We propose efficient scalable solutions to this problem at the architectural, circuit and topological levels, which attempt to minimize both silicon area and power consumption.
Ikuta, Koichi; Tamura, Toshiyuki; Tanaka, Ken-ichi; Kyuma, Kazuo
The AR chip is a versatile CMOS image sensor, functions are not only normal image acquisition but also on-chip image processing. Such features can accelerate algorithms of image processing and the controls of proper image. We have developed the low-cost and compact vehicle detection system using he AR chips. The system is composed of a processing module and an AR camera module. The AR Camera module has dual artificial retina chips to cover the wide dynamic range of the outdoor brightness environment. The ND filter is coated on the lens of one of the chips, each AR chip covers different range of the brightness. The control algorithm of image acquisition is designed to select an adequate chip based on the image quality. The images of the selected chip are processed by on-chip functions for pre-processing and they are transferred to the processing module. Finally the processing module judges the existence of vehicles and detects several kinds of attributive information of the detected vehicle such as moving direction. In our paper, we describe details of the system and the algorithm and we show several result data through field experiments under the real road environment.
Cheung, Carol Yim-Lui; Ikram, M Kamran; Chen, Christopher; Wong, Tien Yin
With increase in life expectancy, the number of persons suffering from common age-related brain diseases, including neurodegenerative (e.g., dementia) and cerebrovascular (e.g., stroke) disease is expected to rise substantially. As current neuro-imaging modalities such as magnetic resonance imaging may not be able to detect subtle subclinical changes (resolution <100-500 μm) in dementia and stroke, there is an urgent need for other complementary techniques to probe the pathophysiology of these diseases. The retina - due to its anatomical, embryological and physiological similarities with the brain - offers a unique and accessible "window" to study correlates and consequences of subclinical pathology in the brain. Retinal components such as the microvasculature and retinal ganglion cell axons can now be visualized non-invasively using different retinal imaging techniques e.g., ocular fundus photography and optical coherence tomography. Advances in retinal imaging may provide new and potentially important insights into cerebrovascular neurodegenerative processes in addition to what is currently possible with neuro-imaging. In this review, we present an overview of the current literature on the application of retinal imaging in the study of dementia and stroke. We discuss clinical implications of these studies, novel state-of-the-art retinal imaging techniques and future directions aimed at evaluating whether retinal imaging can be an additional investigation tool in the study of dementia and stroke.
Prentice, Jason; Simmons, Kristina; Tkacik, Gasper; Homann, Jan; Yee, Heather; Palmer, Stephanie; Nelson, Phillip; Balasubramanian, Vijay
Correlations in the responses of sensory neurons seem to waste neural resources, but can carry cues about structured stimuli and help the brain correct for response errors. To assess how the retina negotiates this tradeoff, we measured simultaneous responses from many retinal ganglion cells presented with natural and artificial stimuli that varied in correlation structure. Responding to spatio-temporally structured stimuli such as natural movies, pairs of ganglion cells were more correlated than in response to white noise checkerboards, but were much less correlated than predicted by a non-adapting functional model of retinal response. Meanwhile, responding to stimuli with purely spatial correlations, pairs of ganglion cells showed increased correlations consistent with a static, non-adapting receptive field and nonlinearity. We found that in response to spatio- temporally correlated stimuli, ganglion cells had faster temporal kernels and tended to have stronger surrounds. These properties of individual cells, along with gain changes that opposed changes in effective contrast at the ganglion cell input, largely explained the pattern of correlations across stimuli.
Nadeau, J H; Grant, P L; Mankala, S; Reiner, A H; Richardson, J E; Eppig, J T
The Mammalian Comparative Database provides genetic maps of mammalian species. Comparative maps are valuable aids for predicting linkages, developing animal models and studying genome organization and evolution.
Watson, Adam J.; Henson, Kyle; Dorsey, Susan G.
Brain-derived neurotrophic factor (BDNF) is a neurotrophin hypothesized to play an important role in mammalian sleep expression and regulation. In order to investigate the role of the truncated receptor for BDNF, TrkB.T1, in mammalian sleep, we examined sleep architecture and sleep regulation in adult mice constitutively lacking this receptor. We find that TrkB.T1 knockout mice have increased REM sleep time, reduced REM sleep latency, and reduced sleep continuity. These results demonstrate a novel role for the TrkB.T1 receptor in sleep expression and provide new insights into the relationship between BDNF, psychiatric illness, and sleep. PMID:25502751
Blankenship, Aaron G; Hamby, Aaron M; Firl, Alana; Vyas, Shri; Maxeiner, Stephan; Willecke, Klaus; Feller, Marla B
Gap junction coupling synchronizes activity among neurons in adult neural circuits, but its role in coordinating activity during development is less known. The developing retina exhibits retinal waves--spontaneous depolarizations that propagate among retinal interneurons and drive retinal ganglion cells (RGCs) to fire correlated bursts of action potentials. During development, two connexin isoforms, connexin 36 (Cx36) and Cx45, are expressed in bipolar cells and RGCs, and therefore provide a potential substrate for coordinating network activity. To determine whether gap junctions contribute to retinal waves, we compared spontaneous activity patterns using calcium imaging, whole-cell recording, and multielectrode array recording in control, single-knock-out (ko) mice lacking Cx45 and double-knock-out (dko) mice lacking both isoforms. Wave frequency, propagation speed, and bias in propagation direction were similar in control, Cx36ko, Cx45ko, and Cx36/45dko retinas. However, the spontaneous firing rate of individual retinal ganglion cells was elevated in Cx45ko retinas, similar to Cx36ko retinas (Hansen et al., 2005; Torborg and Feller, 2005), a phenotype that was more pronounced in Cx36/45dko retinas. As a result, spatial correlations, as assayed by nearest-neighbor correlation and functional connectivity maps, were significantly altered. In addition, Cx36/45dko mice had reduced eye-specific segregation of retinogeniculate afferents. Together, these findings suggest that although Cx36 and Cx45 do not play a role in gross spatial and temporal propagation properties of retinal waves, they strongly modulate the firing pattern of individual RGCs, ensuring strongly correlated firing between nearby RGCs and normal patterning of retinogeniculate projections.
Sweigard, J. Harry; Cashman, Siobhan M.
Purpose. Gene therapy for a number of retinal diseases necessitates efficient transduction of photoreceptor cells. Whereas adenovirus (Ad) serotype 5 (Ad5) does not transduce photoreceptors efficiently, previous studies have demonstrated improved photoreceptor transduction by Ad5 pseudotyped with Ad35 (Ad5/F35) or Ad37 (Ad5/F37) fiber or by the deletion of the RGD domain in the Ad5 penton base (Ad5ΔRGD). However, each of these constructs contained a different transgene cassette, preventing the evaluation of the relative performance of these vectors, an important consideration before the use of these vectors in the clinic. The aim of this study was to evaluate these vectors in the retina and to attempt photoreceptor-specific transgene expression. Methods. Three Ad5-based vectors containing the same expression cassette were generated and injected into the subretinal space of adult mice. Eyes were analyzed for green fluorescence protein expression in flat-mounts, cross-sections, quantitative RT-PCR, and a modified stereological technique. A 257-bp fragment derived from the mouse opsin promoter was analyzed in the context of photoreceptor-specific transgene expression. Results. Each virus tested efficiently transduced the retinal pigment epithelium. The authors found no evidence that Ad5/F35 or Ad5/F37 transduced photoreceptors. Instead, they found that Ad5/F37 transduced Müller cells. Robust photoreceptor transduction by Ad5ΔRGD was detected. Photoreceptor-specific transgene expression from the 257-bp mouse opsin promoter in the context of Ad5ΔRGD vectors was found. Conclusions. Adenovirus vectors may be designed with tropism to distinct cell populations. Robust photoreceptor-specific transgene expression can be achieved in the context of Ad5ΔRGD vectors. PMID:19892875
Hevner, Robert F
The dentate gyrus (DG), a part of the hippocampal formation, has important functions in learning, memory, and adult neurogenesis. Compared with homologous areas in sauropsids (birds and reptiles), the mammalian DG is larger and exhibits qualitatively different phenotypes: 1) folded (C- or V-shaped) granule neuron layer, concave toward the hilus and delimited by a hippocampal fissure; 2) nonperiventricular adult neurogenesis; and 3) prolonged ontogeny, involving extensive abventricular (basal) migration and proliferation of neural stem and progenitor cells (NSPCs). Although gaps remain, available data indicate that these DG traits are present in all orders of mammals, including monotremes and marsupials. The exception is Cetacea (whales, dolphins, and porpoises), in which DG size, convolution, and adult neurogenesis have undergone evolutionary regression. Parsimony suggests that increased growth and convolution of the DG arose in stem mammals concurrently with nonperiventricular adult hippocampal neurogenesis and basal migration of NSPCs during development. These traits could all result from an evolutionary change that enhanced radial migration of NSPCs out of the periventricular zones, possibly by epithelial-mesenchymal transition, to colonize and maintain nonperiventricular proliferative niches. In turn, increased NSPC migration and clonal expansion might be a consequence of growth in the cortical hem (medial patterning center), which produces morphogens such as Wnt3a, generates Cajal-Retzius neurons, and is regulated by Lhx2. Finally, correlations between DG convolution and neocortical gyrification (or capacity for gyrification) suggest that enhanced abventricular migration and proliferation of NSPCs played a transformative role in growth and folding of neocortex as well as archicortex.
Forbes-Osborne, Marie A.; Wilson, Stephen G.; Morris, Ann C.
The zinc-finger transcription factor Insulinoma-associated 1 (Insm1, previously IA-1) is expressed in the developing nervous and neuroendocrine systems, and is required for cell type specific differentiation. Expression of Insm1 is largely absent in the adult, although it is present in neurogenic regions of the adult brain and zebrafish retina. While expression of Insm1 has also been observed in the embryonic retina of numerous vertebrate species, its function during retinal development has remained unexplored. Here, we demonstrate that in the developing zebrafish retina, insm1a is required for photoreceptor differentiation. Insm1a-deficient embryos were microphthalmic and displayed defects in rod and cone photoreceptor differentiation. Rod photoreceptor cells were more sensitive to loss of insm1a expression than were cone photoreceptor cells. Additionally, we provide evidence that insm1a regulates cell cycle progression of retinoblasts, and functions upstream of the bHLH transcription factors ath5/atoh7 and neurod, and the photoreceptor specification genes crx and nr2e3. Finally, we show that insm1a is negatively regulated by Notch-Delta signaling. Taken together, our data demonstrate that Insm1 influences neuronal subtype differentiation during retinal development. PMID:23747542
Tokuda, Kazuhiro; Kuramitsu, Yasuhiro; Byron, Baron; Kitagawa, Takao; Tokuda, Nobuko; Kobayashi, Daiki; Nagayama, Megumi; Araki, Norie; Sonoda, Koh-Hei; Nakamura, Kazuyuki
Glutamate has been shown to induce neural progenitor cells in the adult vertebrate retina. However, protein dynamics during progenitor cell induction by glutamate are not fully understood. To identify specific proteins involved in the process, we employed two-dimensional electrophoresis-based proteomics on glutamate untreated and treated retinal ex vivo sections. Rat retinal tissues were incubated with 1 mM glutamate for 1 h, followed by incubation in glutamate-free media for a total of 24 h. Consistent with prior reports, it was found that mitotic cells appeared in the outer nuclear layer without any histological damage. Immunohistological evaluations and immunoblotting confirmed the emergence of neuronal progenitor cells in the mature retina treated with glutamate. Proteomic analysis revealed the up-regulation of dihydropyrimidinase-related protein 3 (DRP-3), DRP-2 and stress-induced-phosphoprotein 1 (STIP1) during neural progenitor cell induction by glutamate. Moreover, mRNA expression of DRP-3, especially, its long isoform, robustly increased in the treated retina compared to that in the untreated retina. These results may indicate that glutamate induces neural progenitor cells in the mature rat retina by up-regulating the proteins which mediate cell mitosis and neurite growth.
Simpson, Jeremy C; Mateos, Alvaro; Pepperkok, Rainer
A recent use of quantitative proteomics to determine the constituents of the endoplasmic reticulum and Golgi complex is discussed in the light of other available methodologies for cataloging the proteins associated with the mammalian secretory pathway. PMID:17472737
The Gordon Research Conference (GRC) on Mammalian DNA Repair was held at Harbortown Resort, Ventura Beach, CA. Emphasis was placed on current unpublished research and discussion of the future target areas in this field.
Fischer, Andy J.; Skorupa, Dana; Schonberg, David L.; Walton, Nathaniel A.
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
Chen, William C. W.; Wang, Zhouguang; Missinato, Maria Azzurra; Park, Dae Woo; Long, Daniel Ward; Liu, Heng-Jui; Zeng, Xuemei; Yates, Nathan A.; Kim, Kang; Wang, Yadong
Heart attack is a global health problem that leads to significant morbidity, mortality, and health care burden. Adult human hearts have very limited regenerative capability after injury. However, evolutionarily primitive species generally have higher regenerative capacity than mammals. The extracellular matrix (ECM) may contribute to this difference. Mammalian cardiac ECM may not be optimally inductive for cardiac regeneration because of the fibrotic, instead of regenerative, responses in injured adult mammalian hearts. Given the high regenerative capacity of adult zebrafish hearts, we hypothesize that decellularized zebrafish cardiac ECM (zECM) made from normal or healing hearts can induce mammalian heart regeneration. Using zebrafish and mice as representative species of lower vertebrates and mammals, we show that a single administration of zECM, particularly the healing variety, enables cardiac functional recovery and regeneration of adult mouse heart tissues after acute myocardial infarction. zECM-treated groups exhibit proliferation of the remaining cardiomyocytes and multiple cardiac precursor cell populations and reactivation of ErbB2 expression in cardiomyocytes. Furthermore, zECM exhibits pro-proliferative and chemotactic effects on human cardiac precursor cell populations in vitro. These contribute to the structural preservation and correlate with significantly higher cardiac contractile function, notably less left ventricular dilatation, and substantially more elastic myocardium in zECM-treated hearts than control animals treated with saline or decellularized adult mouse cardiac ECM. Inhibition of ErbB2 activity abrogates beneficial effects of zECM administration, indicating the possible involvement of ErbB2 signaling in zECM-mediated regeneration. This study departs from conventional focuses on mammalian ECM and introduces a new approach for cardiac tissue regeneration. PMID:28138518
Park, Ko Uoon; Randazzo, Grace; Jones, Kenneth L.; Brzezinski, Joseph A.
Purpose How retinal bipolar cell interneurons are specified and assigned to specialized subtypes is only partially understood. In part, this is due to a lack of early pan- and subtype-specific bipolar cell markers. To discover these factors, we identified genes that were upregulated in Blimp1 (Prdm1) mutant retinas, which exhibit precocious bipolar cell development. Methods Postnatal day (P)2 retinas from Blimp1 conditional knock-out (CKO) mice and controls were processed for RNA sequencing. Genes that increased at least 45% and were statistically different between conditions were considered candidate bipolar-specific factors. Candidates were further evaluated by RT-PCR, in situ hybridization, and immunohistochemistry. Knock-in Tmem215-LacZ mice were used to better trace retinal expression. Results A comparison between Blimp1 CKO and control RNA-seq datasets revealed approximately 40 significantly upregulated genes. We characterized the expression of three genes that have no known function in the retina, Gsg1 (germ cell associated gene), Trnp1 (TMF-regulated nuclear protein), and Tmem215 (a predicted transmembrane protein). Germ cell associated gene appeared restricted to a small subset of cone bipolars while Trnp1 was seen in all ON type bipolar cells. Using Tmem215-LacZ heterozygous knock-in mice, we observed that β-galactosidase expression started early in bipolar cell development. In adults, Tmem215 was expressed by a subset of ON and OFF cone bipolar cells. Conclusions We have identified Gsg1, Tmem215, and Trnp1 as novel bipolar subtype-specific genes. The spatial and temporal pattern of their expression is consistent with a role in controlling bipolar subtype fate choice, differentiation, or physiology. PMID:28199486
Huang, Jie; Liu, Ying; Oltean, Alina; Beebe, David C
Previous studies of mouse embryos concluded that after the optic vesicle evaginates from the ventral forebrain and contacts the surface ectoderm, signals from the ectoderm specify the distal region of the optic vesicle to become retina and signals from the optic vesicle induce the lens. Germline deletion of Bmp4 resulted in failure of lens formation. We performed conditional deletion of Bmp4 from the optic vesicle to test the function of Bmp4 in murine eye development. The optic vesicle evaginated normally and contacted the surface ectoderm. Lens induction did not occur. The optic cup failed to form and the expression of retina-specific genes decreased markedly in the distal optic vesicle. Instead, cells in the prospective retina expressed genes characteristic of the retinal pigmented epithelium. We conclude that Bmp4 is required for retina specification in mice. In the absence of Bmp4, formation of the retinal pigmented epithelium is the default differentiation pathway of the optic vesicle. Differences in the signaling pathways required for specification of the retina and retinal pigmented epithelium in chicken and mouse embryos suggest major changes in signaling during the evolution of the vertebrate eye.
Grigoryan, E. N.; Anton, H. J.; Mitashov, V. I.
Data on forelimb and eye lens regenerationin in urodeles under spaceflight conditions (SFC) have been obtained in our previous studies. Today, evidence is available that SFC stimulate regeneration in experimental animals rather than inhibit it. The results of control on-ground experiments with simulated microgravity suggest that the stimulatory effect of SFC is due largely to weightlessness. An original experimental model is proposed, which is convenient for comprehensively analyzing neural regeneration under SFC. The initial results described here concern regeneration of neural retina in Pleurodeles waltl newts exposed to microgravity simulated in radial clinostat. After clinorotation for seven days (until postoperation day 16), a positive effect of altered gravity on structural restoration of detached neural retina was confirmed by a number of criteria. Specifically, an increased number of Müllerian glial cells, an increased relative volume of the plexiform layers, reduced cell death, advanced redifferentiation of retinal pigment epithelium, and extended areas of neural retina reattachment were detected in experimental newts. Moreover, cell proliferation in the inner nuclear layer of neural retina increased as compared with control. Thus, low gravity appears to intensify natural cytological and molecular mechanisms of neural retina regeneration in lower vertebrates.
Smith, Peter; Wilhelm, Dagmar; Rodgers, Raymond J
Pre-natal and early post-natal ovarian development has become a field of increasing importance over recent years. The full effects of perturbations of ovarian development on adult fertility, through environmental changes or genetic anomalies, are only now being truly appreciated. Mitigation of these perturbations requires an understanding of the processes involved in the development of the ovary. Herein, we review some recent findings from mice, sheep, and cattle on the key events involved in ovarian development. We discuss the key process of germ cell migration, ovigerous cord formation, meiosis, and follicle formation and activation. We also review the key contributions of mesonephric cells to ovarian development and propose roles for these cells. Finally, we discuss polycystic ovary syndrome, premature ovarian failure, and pre-natal undernutrition; three key areas in which perturbations to ovarian development appear to have major effects on post-natal fertility.
Jin, Nan Ge; Chuang, Alice Z; Masson, Philippe J; Ribelayga, Christophe P
Key points Rod photoreceptors play a key role in vision in dim light; in the mammalian retina, although rods are anatomically connected or coupled by gap junctions, a type of electrical synapse, the functional importance and regulation of rod coupling has remained elusive. We have developed a new technique in the mouse: perforated patch-clamp recording of rod inner segments in isolated intact retinae maintained by superfusion. We find that rod electrical coupling is controlled by a circadian clock and dopamine, and is weak during the day and stronger at night. The results also indicate that the signal-to-noise ratio for a dim light response is increased at night because of coupling. Our observations will provide a framework for understanding the daily variations in human vision as well as the basis of specific retinal malfunctions. Abstract Rod single-photon responses are critical for vision in dim light. Electrical coupling via gap junction channels shapes the light response properties of vertebrate photoreceptors, but the regulation of rod coupling and its impact on the single-photon response have remained unclear. To directly address these questions, we developed a perforated patch-clamp recording technique and recorded from single rod inner segments in isolated intact neural mouse retinae, maintained by superfusion. Experiments were conducted at different times of the day or under constant environmental conditions, at different times across the circadian cycle. We show that rod electrical coupling is regulated by a circadian clock and dopamine, so that coupling is weak during the day and strong at night. Altogether, patch-clamp recordings of single-photon responses in mouse rods, tracer coupling, receptive field measurements and pharmacological manipulations of gap junction and dopamine receptor activity provide compelling evidence that rod coupling is modulated in a circadian manner. These data are consistent with computer modelling. At night, single
Langouet-Astrie, Christophe J.; Meinsen, Annamarie L.; Grunwald, Emily R.; Turner, Stephen D.; Enke, Raymond A.
RNA sequencing transcriptome analysis using massively parallel next generation sequencing technology provides the capability to understand global changes in gene expression throughout a range of tissue samples. Development of the vertebrate retina requires complex temporal orchestration of transcriptional activation and repression. The chicken embryo (Gallus gallus) is a classic model system for studying developmental biology and retinogenesis. Existing retinal transcriptome projects have been critical to the vision research community for studying aspects of murine and human retinogenesis, however, there are currently no publicly available data sets describing the developing chicken retinal transcriptome. Here we used Illumina RNA sequencing (RNA-seq) analysis to characterize the mRNA transcriptome of the developing chicken retina in an effort to identify genes critical for retinal development in this important model organism. These data will be valuable to the vision research community for characterizing global changes in gene expression between ocular tissues and critical developmental time points during retinogenesis in the chicken retina. PMID:27996968
Grossniklaus, Hans E; Shehata, Bahig; Sorensen, Poul; Bergstrom, Chris; Hubbard, G Baker
An 11-year-old boy underwent enucleation of his left eye for an intraocular tumor. Examination showed a small, round blue cell tumor arising in the peripheral retina near the ciliary body. Immunohistochemical stain results were positive for neuron-specific enolase, synaptophysin, cluster of differentiation 99 (CD99), Friend leukemia integration 1, and CD56. Ultrastructural findings included occasional intracytoplasmic dense core granules. Polymerase chain reaction of the tumor showed a Ewing sarcoma/Friend leukemia integration gene fusion product. The tumor was classified as a primitive neuroectodermal tumor/Ewing sarcoma of the retina and should be distinguished from retinoblastoma. To our knowledge, this is the first case of primary primitive neuroectodermal tumor of the retina.
Shah, Bhavin; Püschel, Andreas W
Small GTPases are central regulators of many cellular processes. The highly conserved Rap GTPases perform essential functions in the mammalian nervous system during development and in mature neurons. During neocortical development, Rap1 is required to regulate cadherin- and integrin-mediated adhesion. In the adult nervous system Rap1 and Rap2 regulate the maturation and plasticity of dendritic spine and synapses. Although genetic studies have revealed important roles of Rap GTPases in neurons, their regulation by guanine nucleotide exchange factors (GEFs) that activate them and GTPase activating proteins (GAPs) that inactivate them by stimulating their intrinsic GTPase activity is just beginning to be explored in vivo. Here we review how GEFs and GAPs regulate Rap GTPases in the nervous system with a focus on their in vivo function.
Ratliff, Charles P.
The retina streams visual information to the brain through parallel channels with highly stereotyped patterns of organization and connection. Much progress has been made toward identifying the types of neurons present, and their connectivity. A key problem is inferring the function of a neural system based on its known anatomy and physiology, and identifying the advantages conferred by its particular design. Often, characterizing its architecture reveals some strange features of its organization, and the utility of these features is not always explained easily. Here evidence is presented that several intriguing 'design features' of the retina can be explained by careful application of a single hypothesis: that the retina is organized to maximize the information transmitted about natural visual stimuli, subject to a set of biophysical constraints. Specifically, the input neurons to the retina (photoreceptors) and the output neurons (ganglion cells) exhibit the following interesting features: (1) In trichromats, cone photoreceptors with peak sensitivity to long (L), medium (M) and short (S) wavelengths of light are asymmetrically distributed, so that the ratio of L/M (red/green) cones is highly variable, and S (blue) cones are relatively scarce. (2) Ganglion cell receptive fields are organized so that 3-4 cells of the same type represent each point in a visual image. (3) The retina devotes more resources to ganglion cells selective for negative contrasts (OFF cells) than those selective for positive contrasts (ON cells). (4) The shape of ganglion cell center/surround receptive fields depends on their spatial scale, so that the ratio of surround size to center size decreases with the visual angle subtended by the receptive field. In each case, statistical properties of natural visual stimuli could be coupled with realistic biophysical constraints to account for the features described. The analyses here constitute progress toward long-standing questions concerning the
Ruiz-Cañada, C; Koh, Y H; Budnik, V; Tejedor, F J
Subcellular localization of ion channels is crucial for the transmission of electrical signals in the nervous system. Here we show that Discs-Large (DLG), a member of the MAGUK (membrane-associated guanylate kinases) family in Drosophila, co-localizes with Shaker potassium channels (Sh Kch) in most synaptic areas of the adult brain and in the outer membrane of photoreceptors. However, DLG is absent from axonal tracts in which Sh channels are concentrated. Truncation of the C-terminal of Sh (including the PDZ binding site) disturbs its pattern of distribution in both CNS and retina, while truncation of the guanylate kinase/C-terminal domain of DLG induces ectopic localization of these channels to neuronal somata in the CNS, but does not alter the distribution of channels in photoreceptors. Immunocytochemical, membrane fractionation and detergent solubilization analysis indicate that the C-terminal of Sh Kch is required for proper trafficking to its final destination. Thus, several major conclusions emerge from this study. First, DLG plays a major role in the localization of Sh channels in the CNS and retina. Second, localization of DLG in photoreceptors but not in the CNS seems to depend on its interaction with Sh. Third, the guanylate kinase/C-terminal domain of DLG is involved in the trafficking of Shaker channels but not of DLG in the CNS. Fourth, different mechanisms for the localization of Sh Kch operate in different cell types.
Ogura, Shuntaro; Kurata, Kaori; Hattori, Yuki; Takase, Hiroshi; Ishiguro-Oonuma, Toshina; Hwang, Yoonha; Ahn, Soyeon; Park, Inwon; Ikeda, Wataru; Kusuhara, Sentaro; Fukushima, Yoko; Nara, Hiromi; Sakai, Hideto; Fujiwara, Takashi; Matsushita, Jun; Ema, Masatsugu; Hirashima, Masanori; Minami, Takashi; Shibuya, Masabumi; Takakura, Nobuyuki; Kim, Pilhan; Miyata, Takaki; Ogura, Yuichiro; Uemura, Akiyoshi
In the central nervous system, endothelial cells (ECs) and pericytes (PCs) of blood vessel walls cooperatively form a physical and chemical barrier to maintain neural homeostasis. However, in diabetic retinopathy (DR), the loss of PCs from vessel walls is assumed to cause breakdown of the blood-retina barrier (BRB) and subsequent vision-threatening vascular dysfunctions. Nonetheless, the lack of adequate DR animal models has precluded disease understanding and drug discovery. Here, by using an anti-PDGFRβ antibody, we show that transient inhibition of the PC recruitment to developing retinal vessels sustained EC-PC dissociations and BRB breakdown in adult mouse retinas, reproducing characteristic features of DR such as hyperpermeability, hypoperfusion, and neoangiogenesis. Notably, PC depletion directly induced inflammatory responses in ECs and perivascular infiltration of macrophages, whereby macrophage-derived VEGF and placental growth factor (PlGF) activated VEGFR1 in macrophages and VEGFR2 in ECs. Moreover, angiopoietin-2 (Angpt2) upregulation and Tie1 downregulation activated FOXO1 in PC-free ECs locally at the leaky aneurysms. This cycle of vessel damage was shut down by simultaneously blocking VEGF, PlGF, and Angpt2, thus restoring the BRB integrity. Together, our model provides new opportunities for identifying the sequential events triggered by PC deficiency, not only in DR, but also in various neurological disorders.
Tian, Ning; Xu, Hong-ping; Wang, Ping
Retinal light responsiveness measured via electroretinography undergoes developmental modulation and is thought to be critically regulated by both visual experience and dopamine. The primary goal of this study is to determine whether the dopamine D2 receptor regulates the visual experience-dependent functional development of the retina. Accordingly, we recorded electroretinograms from wild type mice and mice with a genetic deletion of the gene that encodes the dopamine D2 receptor raised under normal cyclic light conditions and constant darkness. Our results demonstrate that mutation of the dopamine D2 receptors preferentially increases the amplitude of the inner retinal light responses evoked by high intensity light measured as oscillatory potentials in adult mice. During postnatal development, all three major components of electroretinograms, the a-wave, b-wave and oscillatory potentials, increase with age. Comparatively, mutation of the dopamine D2 receptors preferentially reduces the age-dependent increase of b-waves evoked by low intensity light. Light deprivation from birth reduces the amplitude of b-waves and completely diminishes the increased amplitude of oscillatory potentials. Taken together, these results demonstrate that the dopamine D2 receptor plays an important role in the activity-dependent functional development of the mouse retina. PMID:25393815
Ogura, Shuntaro; Kurata, Kaori; Hattori, Yuki; Takase, Hiroshi; Ishiguro-Oonuma, Toshina; Hwang, Yoonha; Ahn, Soyeon; Ikeda, Wataru; Kusuhara, Sentaro; Fukushima, Yoko; Nara, Hiromi; Sakai, Hideto; Fujiwara, Takashi; Matsushita, Jun; Ema, Masatsugu; Hirashima, Masanori; Shibuya, Masabumi; Takakura, Nobuyuki; Kim, Pilhan; Miyata, Takaki; Ogura, Yuichiro
In the central nervous system, endothelial cells (ECs) and pericytes (PCs) of blood vessel walls cooperatively form a physical and chemical barrier to maintain neural homeostasis. However, in diabetic retinopathy (DR), the loss of PCs from vessel walls is assumed to cause breakdown of the blood-retina barrier (BRB) and subsequent vision-threatening vascular dysfunctions. Nonetheless, the lack of adequate DR animal models has precluded disease understanding and drug discovery. Here, by using an anti-PDGFRβ antibody, we show that transient inhibition of the PC recruitment to developing retinal vessels sustained EC-PC dissociations and BRB breakdown in adult mouse retinas, reproducing characteristic features of DR such as hyperpermeability, hypoperfusion, and neoangiogenesis. Notably, PC depletion directly induced inflammatory responses in ECs and perivascular infiltration of macrophages, whereby macrophage-derived VEGF and placental growth factor (PlGF) activated VEGFR1 in macrophages and VEGFR2 in ECs. Moreover, angiopoietin-2 (Angpt2) upregulation and Tie1 downregulation activated FOXO1 in PC-free ECs locally at the leaky aneurysms. This cycle of vessel damage was shut down by simultaneously blocking VEGF, PlGF, and Angpt2, thus restoring the BRB integrity. Together, our model provides new opportunities for identifying the sequential events triggered by PC deficiency, not only in DR, but also in various neurological disorders. PMID:28194443
Scarpatetti, A; Forni, S; Niemeyer, G
Malattia leventinese was studied clinically and by electroretinography in 27 members of an affected family and in two additional cases. Typical changes in the central fundus consist of agglomerations of drusen, varying in size, number and localization. These distinct ophthalmoscopic changes occur in young adults, but characteristically remain asymptomatic for one to five decades. Then, metamorphopsia and diffuse color vision deficiency introduce a rapid decay in visual acuity. These changes in function are observed in parallel with large fields of confluent drusen, atrophy of the pigment epithelium and of the retina and with patchy pigmentations in the central fundus. The electroretinogram (ERG) of the subjects with drusen revealed "low normal" amplitudes of the b-waves of both, the cone- and rod system. These ERG-data suggest discrete but widespread functional abnormalities in the outer and middle layers of the retina well before the onset of the clinical symptomatology. In contrast, subnormal ERGs (b-wave amplitudes below 2 standard deviations of normal controls) were recorded in the advance cases, and reflected the severity of functional and structural changes.
Pool, B.L.; Brendler, S.Y.; Liegibel, U.M.; Schmezer, P. ); Tompa, A. )
This report focuses on the use of freshly isolated primary mammalian cells from different tissues and organs of the rat for the rapid and efficient analysis of toxic and genotoxic chemicals. The cells are either treated in vitro or they are isolated from treated animals. Viability by trypan blue exclusion and DNA damage as single-strand breaks are monitored in either case. Therefore, it is possible to compare in vitro and in vivo results directly. N-nitrosamines with unique organ-specific modes in carcinogenesis were studied in vitro using hepatocytes derived from three species (rat, hamster, and pig) and in rat lung and kidney cells. The sensitive detection of all carcinogenic nitrosamines was achieved, although a pattern of cell-specific activation was not observable. The new modification of the in vivo approach allowed the sensitive detection of NDMA genotoxicity in hepatic and in extrahepatic tissues. It is important to point out that the method is an efficient tool for toxicokinetic studies with genotoxic carcinogens in vivo.
Cao, Fengmei; Cao, Nan; Bai, Tingzhu; Song, Shengyu
For a new kind of retina-like senor camera, the image acquisition, coordinates transformation and interpolation need to be realized. Both of the coordinates transformation and interpolation are computed in polar coordinate due to the sensor's particular pixels distribution. The image interpolation is based on sub-pixel interpolation and its relative weights are got in polar coordinates. The hardware platform is composed of retina-like senor camera, image grabber and PC. Combined the MIL and OpenCV library, the software program is composed in VC++ on VS 2010. Experience results show that the system can realizes the real-time image acquisition, coordinate transformation and interpolation.
Nomura, Tadashi; Takahashi, Masanori; Osumi, Noriko
Over the last century, mammalian embryos have been used extensively as a common animal model to investigate fundamental questions in the field of developmental biology. More recently, the establishment of transgenic and gene-targeting systems in laboratory mice has enabled researchers to unveil the genetic mechanisms under lying complex developmental processes (Mak, 2007). However, our understanding of cell—cell interactions and their molecular basis in the early stages of mammalian embryogenesis is still very fragmentary. One of the major problems is the difficulty of precise manipulation and limited accessibility to mammalian embryos via uterus wall. Unfortunately, existing tissue and organotypic culture systems per se do not fully recapitulate three-dimensional, dynamic processes of organogenesis observed in vivo. Although transgenic animal technology and virus-mediated gene delivery are useful to manipulate gene expression, these techniques take much time and financial costs, which limit their use.
Pang, Ji-Jie; Gao, Fan; Lem, Janis; Bramblett, Debra E.; Paul, David L.; Wu, Samuel M.
Bipolar cells are the central neurons of the retina that transmit visual signals from rod and cone photoreceptors to third-order neurons in the inner retina and the brain. A dogma set forth by early anatomical studies is that bipolar cells in mammalian retinas receive segregated rod/cone synaptic inputs (either from rods or from cones), and here, we present evidence that challenges this traditional view. By analyzing light-evoked cation currents from morphologically identified depolarizing bipolar cells (DBCs) in the wild-type and three pathway-specific knockout mice (rod transducin knockout [Trα−/−], connexin36 knockout [Cx36−/−], and transcription factor beta4 knockout [Bhlhb4−/−]), we show that a subpopulation of rod DBCs (DBCR2s) receives substantial input directly from cones and a subpopulation of cone DBCs (DBCC1s) receives substantial input directly from rods. These results provide evidence of the existence of functional rod-DBCC and cone-DBCR synaptic pathways in the mouse retina as well as the previously proposed rod hyperpolarizing bipolar-cells pathway. This is grounds for revising the mammalian rod/cone bipolar cell dogma. PMID:20018684
Vinogradova, Iu V; Tronov, V A; Liakhova, K N; Poplinskaia, V A; Ostrovskiĭ, M A
The eye retina consists of terminally differentiated cells that have lost their ability to proliferate. The death of these cells leads tothe loss of sight. The mice retina is characterized by relatively high resistance to radiation, which is provided by its ability to repair damage caused by environmental factors. The aim of our work was to assess the damaging effect of ionizing radiation and methylnitrosourea (MNU) on the DNA structure in the mouse retina, the functional activity of the retina, and its ability to recover in vivo. The results confirm the ability of the mature retina to structural and functional recovery. Adapting influence of low dose chemical agent increases retina resistance to cytotoxic dose of genotoxicants and prevents degeneration of photoreceptor layer of the retina. The results show the possibility of neurohormesis effect in the mice retina after exposure to ionizing radiation and chemicals.
Wassmer, Sarah J; Carvalho, Livia S; György, Bence; Vandenberghe, Luk H; Maguire, Casey A
Widespread gene transfer to the retina is challenging as it requires vector systems to overcome physical and biochemical barriers to enter and diffuse throughout retinal tissue. We investigated whether exosome-associated adeno-associated virus, (exo-AAV) enabled broad retinal targeting following intravitreal (IVT) injection, as exosomes have been shown to traverse biological barriers and mediate widespread distribution upon systemic injection. We packaged an AAV genome encoding green fluorescent protein (GFP) into conventional AAV2 and exo-AAV2 vectors. Vectors were IVT injected into the eyes of adult mice. GFP expression was noninvasively monitored by fundus imaging and retinal expression was analyzed 4 weeks post-injection by qRT-PCR and histology. Exo-AAV2 outperformed conventional AAV2 in GFP expression based on fundus image analysis and qRT-PCR. Exo-AAV2 demonstrated deeper penetration in the retina, efficiently reaching the inner nuclear and outer plexiform, and to a lesser extent the outer nuclear layer. Cell targets were ganglion cells, bipolar cells, Müller cells, and photoreceptors. Exo-AAV2 serves as a robust gene delivery tool for murine retina, and the simplicity of production and isolation should make it widely applicable to basic research of the eye.
Wassmer, Sarah J.; Carvalho, Livia S.; György, Bence; Vandenberghe, Luk H.; Maguire, Casey A.
Widespread gene transfer to the retina is challenging as it requires vector systems to overcome physical and biochemical barriers to enter and diffuse throughout retinal tissue. We investigated whether exosome-associated adeno-associated virus, (exo-AAV) enabled broad retinal targeting following intravitreal (IVT) injection, as exosomes have been shown to traverse biological barriers and mediate widespread distribution upon systemic injection. We packaged an AAV genome encoding green fluorescent protein (GFP) into conventional AAV2 and exo-AAV2 vectors. Vectors were IVT injected into the eyes of adult mice. GFP expression was noninvasively monitored by fundus imaging and retinal expression was analyzed 4 weeks post-injection by qRT-PCR and histology. Exo-AAV2 outperformed conventional AAV2 in GFP expression based on fundus image analysis and qRT-PCR. Exo-AAV2 demonstrated deeper penetration in the retina, efficiently reaching the inner nuclear and outer plexiform, and to a lesser extent the outer nuclear layer. Cell targets were ganglion cells, bipolar cells, Müller cells, and photoreceptors. Exo-AAV2 serves as a robust gene delivery tool for murine retina, and the simplicity of production and isolation should make it widely applicable to basic research of the eye. PMID:28361998
Nagy, Kornél; Brahmbhatt, Viral Vishnuprasad; Berdeaux, Olivier; Bretillon, Lionel; Destaillats, Frédéric; Acar, Niyazi
The objective of this work was to detect and identify phosphatidylserine plasmalogen species in human ocular neurons represented by the retina and the optic nerve. Plasmalogens (vinyl-ether bearing phospholipids) are commonly found in the forms of phosphatidylcholine and phosphatidylethanolamine in numerous mammalian cell types, including the retina. Although their biological functions are unclear, the alteration of cellular plasmalogen content has been associated with several human disorders such as rhizomelic chondrodysplasia punctata Type 2 and primary open-angle glaucoma. By using liquid chromatography coupled to high-resolution and tandem mass spectrometry, we have identified for the first time several species of phosphatidylserine plasmalogens, including atypical forms having moieties with odd numbers of carbons and unsaturation in sn-2 position. Structural elucidation of the potential phosphatidylserine ether linked species was pursued by performing MS3 experiments, and three fragments are proposed as marker ions to deduce which fatty acid is linked as ether or ester on the glycerol backbone. Interpretation of the fragmentation patterns based on this scheme enabled the assignment of structures to the m/z values, thereby identifying the phosphatidylserine plasmalogens. PMID:22266369
Lee, Sherwin C.; Ishida, Andrew T.
Antisera directed against hyperpolarization-activated mixed-cation (“Ih”) and K+ (“Kir”) channels bind to some somata in the ganglion cell layer of rat and rabbit retina. Additionally, the termination of hyperpolarizing current injections can trigger spikes in some cat retinal ganglion cells, suggesting a rebound depolarization due to activation of Ih. However, patch-clamp studies have reported that rat ganglion cells lack inward rectification, or present an inwardly rectifying K+ current. We therefore tested whether hyperpolarization activates Ih in dissociated, adult rat retinal ganglion cell somata. We report here that while we found no inward rectification in some cells, and a Kir-like current in a few cells, hyperpolarization activated Ih in roughly 75% of the cells we recorded from in voltage clamp. We show that this current is blocked by Cs+ or ZD7288 and only slightly reduced by Ba2+, that the current amplitude and reversal potential are sensitive to extracellular Na+ and K+, and that we found no evidence of Kir in cells presenting Ih. In current clamp, injecting hyperpolarizing current induced a slowly relaxing membrane hyperpolarization that rebounded to a few action potentials when the hyperpolarizing current was stopped; both the membrane potential relaxation and rebound spikes were blocked by ZD7288. These results provide the first measurement of Ih in mammalian retinal ganglion cells, and indicate that the ion channels of rat retinal ganglion cells may vary in ways not expected from previous voltage and current recordings. PMID:17488978
HIRANO, ARLENE A.; BRANDSTÄTTER, JOHANN HELMUT; VILA, ALEJANDRO; BRECHA, NICHOLAS C.
Horizontal cells mediate inhibitory feed-forward and feedback communication in the outer retina; however, mechanisms that underlie transmitter release from mammalian horizontal cells are poorly understood. Toward determining whether the molecular machinery for exocytosis is present in horizontal cells, we investigated the localization of syntaxin-4, a SNARE protein involved in targeting vesicles to the plasma membrane, in mouse, rat, and rabbit retinae using immunocytochemistry. We report robust expression of syntaxin-4 in the outer plexiform layer of all three species. Syntaxin-4 occurred in processes and tips of horizontal cells, with regularly spaced, thicker sandwich-like structures along the processes. Double labeling with syntaxin-4 and calbindin antibodies, a horizontal cell marker, demonstrated syntaxin-4 localization to horizontal cell processes; whereas, double labeling with PKC antibodies, a rod bipolar cell (RBC) marker, showed a lack of co-localization, with syntaxin-4 immunolabeling occurring just distal to RBC dendritic tips. Syntaxin-4 immunolabeling occurred within VGLUT-1-immunoreactive photoreceptor terminals and underneath synaptic ribbons, labeled by CtBP2/RIBEYE antibodies, consistent with localization in invaginating horizontal cell tips at photoreceptor triad synapses. Vertical sections of retina immunostained for syntaxin-4 and peanut agglutinin (PNA) established that the prominent patches of syntaxin-4 immunoreactivity were adjacent to the base of cone pedicles. Horizontal sections through the OPL indicate a one-to-one co-localization of syntaxin-4 densities at likely all cone pedicles, with syntaxin-4 immunoreactivity interdigitating with PNA labeling. Pre-embedding immuno-electron microscopy confirmed the subcellular localization of syntaxin-4 labeling to lateral elements at both rod and cone triad synapses. Finally, co-localization with SNAP-25, a possible binding partner of syntaxin-4, indicated co-expression of these SNARE proteins in
Arman, A Cyrus; Sampath, Alapakkam P
The nervous system frequently integrates parallel streams of information to encode a broad range of stimulus strengths. In mammalian retina it is generally believed that signals generated by rod and cone photoreceptors converge onto cone bipolar cells prior to reaching the retinal output, the ganglion cells. Near absolute visual threshold a specialized mammalian retinal circuit, the rod bipolar pathway, pools signals from many rods and converges on depolarizing (AII) amacrine cells. However, whether subsequent signal flow to OFF ganglion cells requires OFF cone bipolar cells near visual threshold remains unclear. Glycinergic synapses between AII amacrine cells and OFF cone bipolar cells are believed to relay subsequently rod-driven signals to OFF ganglion cells. However, AII amacrine cells also make glycinergic synapses directly with OFF ganglion cells. To determine the route for signal flow near visual threshold, we measured the effect of the glycine receptor antagonist strychnine on response threshold in fully dark-adapted retinal cells. As shown previously, we found that response threshold for OFF ganglion cells was elevated by strychnine. Surprisingly, strychnine did not elevate response threshold in any subclass of OFF cone bipolar cell. Instead, in every OFF cone bipolar subclass strychnine suppressed tonic glycinergic inhibition without altering response threshold. Consistent with this lack of influence of strychnine, we found that the dominant input to OFF cone bipolar cells in darkness was excitatory and the response threshold of the excitatory input varied by subclass. Thus, in the dark-adapted mouse retina, the high absolute sensitivity of OFF ganglion cells cannot be explained by signal transmission through OFF cone bipolar cells.
Educational technology units must continually monitor their strategic plans to ensure that they are aligned with the realities of their institutions. Strategic dissonance occurs when previously successful strategies are no longer achieving the same positive outcomes. The Virtual Retina CD-ROM project is used in this paper as an example of…
Hoon, Mrinalini; Okawa, Haruhisa; Della Santina, Luca; Wong, Rachel O L
Structure and function are highly correlated in the vertebrate retina, a sensory tissue that is organized into cell layers with microcircuits working in parallel and together to encode visual information. All vertebrate retinas share a fundamental plan, comprising five major neuronal cell classes with cell body distributions and connectivity arranged in stereotypic patterns. Conserved features in retinal design have enabled detailed analysis and comparisons of structure, connectivity and function across species. Each species, however, can adopt structural and/or functional retinal specializations, implementing variations to the basic design in order to satisfy unique requirements in visual function. Recent advances in molecular tools, imaging and electrophysiological approaches have greatly facilitated identification of the cellular and molecular mechanisms that establish the fundamental organization of the retina and the specializations of its microcircuits during development. Here, we review advances in our understanding of how these mechanisms act to shape structure and function at the single cell level, to coordinate the assembly of cell populations, and to define their specific circuitry. We also highlight how structure is rearranged and function is disrupted in disease, and discuss current approaches to re-establish the intricate functional architecture of the retina.
Funke, Sebastian; Perumal, Natarajan; Beck, Sabine; Gabel-Scheurich, Silke; Schmelter, Carsten; Teister, Julia; Gerbig, Claudia; Gramlich, Oliver W.; Pfeiffer, Norbert; Grus, Franz H.
Glaucoma related proteomic changes have been documented in cell and animal models. However, proteomic studies investigating on human retina samples are still rare. In the present work, retina samples of glaucoma and non-glaucoma control donors have been examined by a state-of-the-art mass spectrometry (MS) workflow to uncover glaucoma related proteomic changes. More than 600 proteins could be identified with high confidence (FDR < 1%) in human retina samples. Distinct proteomic changes have been observed in 10% of proteins encircling mitochondrial and nucleus species. Numerous proteins showed a significant glaucoma related level change (p < 0.05) or distinct tendency of alteration (p < 0.1). Candidates were documented to be involved in cellular development, stress and cell death. Increase of stress related proteins and decrease of new glaucoma related candidates, ADP/ATP translocase 3 (ANT3), PC4 and SRFS1-interacting protein 1 (DFS70) and methyl-CpG-binding protein 2 (MeCp2) could be documented by MS. Moreover, candidates could be validated by Accurate Inclusion Mass Screening (AIMS) and immunostaining and supported for the retinal ganglion cell layer (GCL) by laser capture microdissection (LCM) in porcine and human eye cryosections. The workflow allowed a detailed view into the human retina proteome highlighting new molecular players ANT3, DFS70 and MeCp2 associated to glaucoma. PMID:27425789
Alabboud, Ied; Muyo, Gonzalo; Gorman, Alistair; Mordant, David; McNaught, Andrew; Petres, Clement; Petillot, Yvan R.; Harvey, Andrew R.
Hyperspectral imaging of the retina presents a unique opportunity for direct and quantitative mapping of retinal biochemistry - particularly of the vasculature where blood oximetry is enabled by the strong variation of absorption spectra with oxygenation. This is particularly pertinent both to research and to clinical investigation and diagnosis of retinal diseases such as diabetes, glaucoma and age-related macular degeneration. The optimal exploitation of hyperspectral imaging however, presents a set of challenging problems, including; the poorly characterised and controlled optical environment of structures within the retina to be imaged; the erratic motion of the eye ball; and the compounding effects of the optical sensitivity of the retina and the low numerical aperture of the eye. We have developed two spectral imaging techniques to address these issues. We describe first a system in which a liquid crystal tuneable filter is integrated into the illumination system of a conventional fundus camera to enable time-sequential, random access recording of narrow-band spectral images. Image processing techniques are described to eradicate the artefacts that may be introduced by time-sequential imaging. In addition we describe a unique snapshot spectral imaging technique dubbed IRIS that employs polarising interferometry and Wollaston prism beam splitters to simultaneously replicate and spectrally filter images of the retina into multiple spectral bands onto a single detector array. Results of early clinical trials acquired with these two techniques together with a physical model which enables oximetry map are reported.
Recently, we have proposed that the fluorescence spectra of sheep retina can be well correlated to the presence or absence of scrapie. Scrapie is the most widespread TSE (transmissible spongiform encephalopathy) affecting sheep and goats worldwide. Mice eyes have been previously reported as a model ...
Yazulla, S; Studholme, K M
The zebrafish retina is rapidly becoming a major preparation for the study of molecular genetic mechanisms underlying neural development and visual behavior. Studies utilizing retinal mutants would benefit by the availability of a data base on the distribution of neurotransmitter systems in the wild-type fish. To this end, the neurochemical anatomy of the zebrafish retina was surveyed by light microscopic immunocytochemistry. An extensive series of 60 separate antibodies were used to describe the distribution of major transmitter systems and a variety of neuron-associated membrane channels and proteins. These include markers (i.e., antibodies against enzymes, receptors, transporters) for transmitters: GABA, glycine, glutamate, biogenic amines, acetylcholine, cannabinoids and neuropeptides; as well as a sample of voltage-gated channels and synapse associated membrane proteins. Discussion of the comparative localization of these antibodies is restricted to other teleost fishes, particularly goldfish. Overall, there was great similarity in the distribution of the various markers, as might be expected. However, there were some notable differences, including several antibodies that did not label zebrafish at all, even though goldfish retinas that were processed in parallel, labeled beautifully. This survey is extensive, but not exhaustive, and hopefully will serve as a valuable resource for future studies of the zebrafish retina.
To understand mechanisms of neurotoxicity in susceptible populations, we examined age-related changes in constitutive gene expression in the retinas of young (4mos), middle-aged (11 mos) and aged (23 mos) male Long Evans rats. Derived from a pouch of the forebrain during develop...
Cang, Jianhua; Rentería, René C; Kaneko, Megumi; Liu, Xiaorong; Copenhagen, David R; Stryker, Michael P
The visual cortex is organized into retinotopic maps that preserve an orderly representation of the visual world, achieved by topographically precise inputs from the lateral geniculate nucleus. We show here that geniculocortical mapping is imprecise when the waves of spontaneous activity in the retina during the first postnatal week are disrupted genetically. This anatomical mapping defect is present by postnatal day 8 and has functional consequences, as revealed by optical imaging and microelectrode recording in adults. Pharmacological disruption of these retinal waves during the first week phenocopies the mapping defect, confirming both the site and the timing of the disruption in neural activity responsible for the defect. Analysis shows that the geniculocortical miswiring is not a trivial or necessary consequence of the retinogeniculate defect. Our findings demonstrate that disrupting early spontaneous activity in the eye alters thalamic connections to the cortex.
The pigment epithelial cell of the retina actively participates in two aspects of lipid metabolism: (a) the fatty acid esterification of vitamin A and its storage and transport to the photoreceptors, and (b) the phagocytosis and degradation of the lipoprotein membrane disks shed from the photoreceptor cells. Study of the pigment epithelial cells of adult albino and pigmented rodents has revealed the abundance of an organelle, microperoxisomes, not previously known to exist in this cell type. The metabolism, transport, and storage of lipids are major functions of other cell types which possess large numbers of microperoxisomes associated with a highly developed smooth endoplasmic reticulum. Microperoxisomes were encountered, but relatively rarely, in Muller cells and vascular endothelial cells. A tubular system in photoreceptor terminals is reactive in the cytochemical procedure used to visualize microperoxisomes. PMID:1168648
Zhang, Jian; Wu, Samuel M
In the tiger salamander retina, visual signals are transmitted to the inner retina via six morphologically distinct types of photoreceptors: large/small rods, large/small single cones, and double cones composed of principal and accessory members. The objective of this study was to determine the morphology of these photoreceptors and their synaptic interconnection with bipolar cells and horizontal cells in the outer plexiform layer (OPL). Here we showed that glutamate antibodies labeled all photoreceptors and recovering antibodies strongly labeled all cones and weakly labeled all rods. Antibodies against calbindin selectively stained accessory members of double cones. Antibodies against S-cone opsin stained small rods, a subpopulation of small single cones, and the outer segments of accessory double cones and a subtype of unidentified single cones. On average, large rods and small S-cone opsin positive rods accounted for 98.6% and 1.4% of all rods, respectively. Large/small cones, principle/accessory double cones, S-cone opsin positive small single cones, and S-cone opsin positive unidentified single cones accounted for about 66.9%, 23%, 4.5%, and 5.6% of the total cones, respectively. Moreover, the differential connection between rods/cones and bipolar/horizontal cells and the wide distribution of AMPA receptor subunits GluR2/3 and GluR4 at the rod/cone synapses were observed. These results provide anatomical evidence for the physiological findings that bipolar/horizontal cells in the salamander retina are driven by rod/cone inputs of different weights, and that AMPA receptors play an important role in glutamatergic neurotransmission at the first visual synapses. The different photoreceptors selectively contacting bipolar and horizontal cells support the idea that visual signals may be conveyed to the inner retina by different functional pathways in the outer retina.
Schaefer, Kellie A.; Toral, Marcus A.; Velez, Gabriel; Cox, Allison J.; Baker, Sheila A.; Borcherding, Nicholas C.; Colgan, Diana F.; Bondada, Vimala; Mashburn, Charles B.; Yu, Chen-Guang; Geddes, James W.; Tsang, Stephen H.; Bassuk, Alexander G.; Mahajan, Vinit B.
Purpose We characterize calpain-5 (CAPN5) expression in retinal and neuronal subcellular compartments. Methods CAPN5 gene variants were classified using the exome variant server, and RNA-sequencing was used to compare expression of CAPN5 mRNA in the mouse and human retina and in retinoblastoma cells. Expression of CAPN5 protein was ascertained in humans and mice in silico, in mouse retina by immunohistochemistry, and in neuronal cancer cell lines and fractionated central nervous system tissue extracts by Western analysis with eight antibodies targeting different CAPN5 regions. Results Most CAPN5 genetic variation occurs outside its protease core; and searches of cancer and epilepsy/autism genetic databases found no variants similar to hyperactivating retinal disease alleles. The mouse retina expressed one transcript for CAPN5 plus those of nine other calpains, similar to the human retina. In Y79 retinoblastoma cells, the level of CAPN5 transcript was very low. Immunohistochemistry detected CAPN5 expression in the inner and outer nuclear layers and at synapses in the outer plexiform layer. Western analysis of fractionated retinal extracts confirmed CAPN5 synapse localization. Western blots of fractionated brain neuronal extracts revealed distinct subcellular patterns and the potential presence of autoproteolytic CAPN5 domains. Conclusions CAPN5 is moderately expressed in the retina and, despite higher expression in other tissues, hyperactive disease mutants of CAPN5 only manifest as eye disease. At the cellular level, CAPN5 is expressed in several different functional compartments. CAPN5 localization at the photoreceptor synapse and with mitochondria explains the neural circuitry phenotype in human CAPN5 disease alleles. PMID:27152965
Poon, Annie Wing Hoi; Ma, Emilie Xiao Hang; Vadivel, Arul; Jung, Suna; Khoja, Zehra; Stephens, Laurel; Thébaud, Bernard; Wintermark, Pia
ABSTRACT Many premature newborns develop bronchopulmonary dysplasia (BPD), a chronic lung disease resulting from prolonged mechanical ventilation and hyperoxia. BPD survivors typically suffer long-term injuries not only to the lungs, but also to the brain and retina. However, currently it is not clear whether the brain and retinal injuries in these newborns are related only to their prematurity, or also to BPD. We investigated whether the hyperoxia known to cause histologic changes in the lungs similar to BPD in an animal model also causes brain and retinal injuries. Sprague Dawley rat pups were exposed to hyperoxia (95% O2, ‘BPD’ group) or room air (21% O2, ‘control’ group) from postnatal day 4–14 (P4–14); the rat pups were housed in room air between P14 and P28. At P28, they were sacrificed, and their lungs, brain, and eyes were extracted. Hematoxylin and eosin staining was performed on lung and brain sections; retinas were stained with Toluidine Blue. Hyperoxia exposure resulted in an increased mean linear intercept in the lungs (P<0.0001). This increase was associated with a decrease in some brain structures [especially the whole-brain surface (P=0.02)], as well as a decrease in the thickness of the retinal layers [especially the total retina (P=0.0008)], compared to the room air control group. In addition, a significant negative relationship was observed between the lung structures and the brain (r=−0.49, P=0.02) and retina (r=−0.70, P=0.0008) structures. In conclusion, hyperoxia exposure impaired lung, brain, and retina structures. More severe lung injuries correlated with more severe brain and retinal injuries. This result suggests that the same animal model of chronic neonatal hyperoxia can be used to simultaneously study lung, brain and retinal injuries related to hyperoxia. PMID:26988760
Poon, Annie Wing Hoi; Ma, Emilie Xiao Hang; Vadivel, Arul; Jung, Suna; Khoja, Zehra; Stephens, Laurel; Thébaud, Bernard; Wintermark, Pia
Many premature newborns develop bronchopulmonary dysplasia (BPD), a chronic lung disease resulting from prolonged mechanical ventilation and hyperoxia. BPD survivors typically suffer long-term injuries not only to the lungs, but also to the brain and retina. However, currently it is not clear whether the brain and retinal injuries in these newborns are related only to their prematurity, or also to BPD. We investigated whether the hyperoxia known to cause histologic changes in the lungs similar to BPD in an animal model also causes brain and retinal injuries. Sprague Dawley rat pups were exposed to hyperoxia (95% O2, 'BPD' group) or room air (21% O2, 'control' group) from postnatal day 4-14 (P4-14); the rat pups were housed in room air between P14 and P28. At P28, they were sacrificed, and their lungs, brain, and eyes were extracted. Hematoxylin and eosin staining was performed on lung and brain sections; retinas were stained with Toluidine Blue. Hyperoxia exposure resulted in an increased mean linear intercept in the lungs (P<0.0001). This increase was associated with a decrease in some brain structures [especially the whole-brain surface (P=0.02)], as well as a decrease in the thickness of the retinal layers [especially the total retina (P=0.0008)], compared to the room air control group. In addition, a significant negative relationship was observed between the lung structures and the brain (r=-0.49,P=0.02) and retina (r=-0.70,P=0.0008) structures. In conclusion, hyperoxia exposure impaired lung, brain, and retina structures. More severe lung injuries correlated with more severe brain and retinal injuries. This result suggests that the same animal model of chronic neonatal hyperoxia can be used to simultaneously study lung, brain and retinal injuries related to hyperoxia.
Molnar, Alyosha; Hsueh, Hain-Ann; Roska, Botond; Werblin, Frank S
In the mammalian retina, complementary ON and OFF visual streams are formed at the bipolar cell dendrites, then carried to amacrine and ganglion cells via nonlinear excitatory synapses from bipolar cells. Bipolar, amacrine and ganglion cells also receive a nonlinear inhibitory input from amacrine cells. The most common form of such inhibition crosses over from the opposite visual stream: Amacrine cells carry ON inhibition to the OFF cells and carry OFF inhibition to the ON cells ("crossover inhibition"). Although these synapses are predominantly nonlinear, linear signal processing is required for computing many properties of the visual world such as average intensity across a receptive field. Linear signaling is also necessary for maintaining the distinction between brightness and contrast. It has long been known that a subset of retinal outputs provide exactly this sort of linear representation of the world; we show here that rectifying (nonlinear) synaptic currents, when combined thorough crossover inhibition can generate this linear signaling. Using simple mathematical models we show that for a large set of cases, repeated rounds of synaptic rectification without crossover inhibition can destroy information carried by those synapses. A similar circuit motif is employed in the electronics industry to compensate for transistor nonlinearities in analog circuits.
Kuse, Yoshiki; Tsuruma, Kazuhiro; Sugitani, Sou; Izawa, Hiroshi; Ohno, Yuta; Shimazawa, Masamitsu; Hara, Hideaki
Progranulin (PGRN) is a secreted growth factor associated with embryo development, tissue repair, and inflammation. In a previous study, we showed that adipose-derived stem cell-conditioned medium (ASC-CM) is rich in PGRN. In the present study, we investigated whether PGRN is associated with retinal regeneration in the mammalian retina. We evaluated the effect of ASC-CM using the N-methyl-N-nitrosourea-induced retinal damage model in mice. ASC-CM promoted the differentiation of photoreceptor cells following retinal damage. PGRN increased the number of BrdU+ cells in the outer nuclear layer following retinal damage some of which were Rx (retinal precursor cell marker) positive. PGRN also increased the number of rhodopsin+ photoreceptor cells in primary retinal cell cultures. SU11274, a hepatocyte growth factor (HGF) receptor inhibitor, attenuated the increase. These findings suggest that PGRN may affect the differentiation of retinal precursor cells to photoreceptor cells through the HGF receptor signaling pathway. PMID:27030285
Grimes, William N; Li, Wei; Chávez, Andrés E; Diamond, Jeffrey S
In the mammalian retina, A17 amacrine cells provide reciprocal inhibitory feedback to rod bipolar cells, thereby shaping the time course of visual signaling in vivo. Previous results have indicated that A17 feedback can be triggered by Ca(2+) influx through Ca(2+)-permeable AMPA receptors and can occur independently of voltage-gated Ca(2+) (Ca(v)) channels, whose presence and functional role in A17 dendrites have not yet been explored. We combined electrophysiology, calcium imaging and immunohistochemistry and found that L-type Ca(v) channels in rat A17 amacrine cells were located at the sites of reciprocal synaptic feedback and that their contribution to GABA release was diminished by large-conductance Ca(2+)-activated potassium (BK) channels, which suppress postsynaptic depolarization in A17s and limit Ca(v) channel activation. We also found that BK channels, by limiting GABA release from A17s, regulate the flow of excitatory synaptic transmission through the rod pathway.
Lüke, Matthias; Januschowski, Kai; Beutel, Julia; Warga, Max; Grisanti, Salvatore; Peters, Swaantje; Schneider, Toni; Lüke, Christoph; Bartz-Schmidt, Karl Ulrich; Szurman, Peter
A good clinical experience of intravitreal triamcinolone acetonide (TA) has been reported in several studies, but there are growing indications that epiretinal crystals of TA exhibit retinal toxicity. To investigate the effects of TA on retinal function we used a model of an electrophysiological in vitro technique for testing retinal toxicity. Isolated bovine retinas were perfused with an oxygen saturated nutrient solution. The electroretinogram (ERG) was recorded as a transretinal potential using Ag/AgCl electrodes. After reaching stable ERG-amplitudes TA at the maximum solubility equilibrium (36 microg/ml) was either applied to the nutrient solution for 45 min or TA was administered epiretinally at concentrations (1 mg/ml, 4 mg/ml, 8 mg/ml, 20 mg/ml and 40 mg/ml) above the maximum solubility equilibrium to assure direct contact of the TA crystals with the isolated perfused retinas. After that the retinas were reperfused for 75 min with the standard nutrient solution. The percentage of a- and b-wave reduction directly after the application and at the washout was calculated. To assess the effects of TA at the level of the ganglion cell layer a Viability/Cytotoxicity Kit for mammalian cells was used. No changes of the ERG-amplitudes were detected during the exposure to 36 microg/ml TA for 45 min (b-wave: 9.6 microV+/-2.1 vs. 8 microV+/-2.1 (p=0.135); a-wave: -11 microV+/-2.7 vs. -10.6 microV+/-2.3 (p=0.889)) and at the washout (b-wave: 8 microV+/-2.1 vs. 8.3 microV+/-2.4 (p=0.18); a-wave: -10.6 microV+/-2.3 vs. -12 microV+/-2.6 (p=0.225)). At concentrations higher than 1mg/ml TA induced a decrease of the a- and b-wave in a concentration dependent manner. These changes were reversible for concentrations of TA up to 20mg/ml (b-wave: 9 microV+/-2.4 vs. 6.6 microV+/-2.5 (p=0.08); a-wave: -11.4 microV+/-2.0 vs. -11.2 microV+/-2.2 (p=0.37)), but irreversible at 40 mg/ml even at the end of the washout (b-wave: 9.8 microV+/-1.9 vs. 3 microV+/-1.7 (p=0.009); a-wave: -9
Heiduschka, P; Romann, I; Stieglitz, T; Thanos, S
Adult mammalian optic nerve axons are able to regenerate, when provided with the permissive environment of an autologous peripheral nerve graft, which is usually the sciatic nerve. This study demonstrates the ability of adult rat optic nerve axons to regenerate through the preformed perforations of a polyimide electrode carrier implanted at the interface between the proximal stump of the cut optic nerve and the stump of the peripheral nerve piece used for grafting. Evidence that retinal ganglion cells regenerated their axons through the perforated electrode carrier was obtained by retrograde labeling with a fluorescent dye deposited into the sciatic nerve graft beyond the nerve-carrier-nerve junction. The number of regenerating cells could be enhanced by injecting neuroprotective drugs like aurintricarboxylic acid and cortisol intravitreally. A second line of evidence was obtained by immunohistochemical staining with antibodies to neurofilament. Third, electrical activity of the regenerating nerves was recorded after stimulating the retina with a flash of light. The results suggest that a regenerating central nerve tract may serve as an experimental model to implant artificial microdevices to monitor the physiological and topographical properties of neurites passing through the device or to stimulate them, thus interfering with their potential to grow. This study reports for the first time that the optic nerve has unique properties, which aids in the realization of these goals.
Rapaport, D H
The development of synaptic ribbons in rod and cone photoreceptor terminals of the cat retina was studied using quantitative electron microscopy. At the region of the area centralis, synaptic ribbon profiles are initially recognized at PCD (postconception day) 59. Synaptic ribbon density increases rapidly, reaching a peak of 0.55 ribbons/micron 3 at PCD 68 (postnatal day 3) and maintains approximately that value for an additional 8 d. Following PCD 76, ribbon density begins to decrease, to 0.37 ribbons/microns 3 at PCD 82 and 0.25 ribbons/microns 3 at PCD 102. Although ribbon density drops by approximately 50% during this 39-d period, the outer plexiform layer (OPL) volume at the area centralis increases by about 20%. Ribbon density continues to decrease gradually over a protracted period to reach a final adult value of 0.11-0.14 ribbons/microns 3. During the period of high ribbon density, rod spherules with two, or even three ribbon profiles, were routinely observed. In contrast, in the adult, spherules with more than one ribbon profile are only rarely encountered. During development, the length of synaptic ribbon profiles increases from a mean of 0.22 microns at PCD 62 to the 0.47 microns mean length found in the adult.
Ferramosca, Alessandra; Zara, Vincenzo
After ejaculation, the mammalian male gamete must undergo the capacitation process, which is a prerequisite for egg fertilization. The bioenergetics of sperm capacitation is poorly understood despite its fundamental role in sustaining the biochemical and molecular events occurring during gamete activation. Glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) are the two major metabolic pathways producing ATP which is the primary source of energy for spermatozoa. Since recent data suggest that spermatozoa have the ability to use different metabolic substrates, the main aim of this work is to present a broad overview of the current knowledge on the energy-producing metabolic pathways operating inside sperm mitochondria during capacitation in different mammalian species. Metabolism of glucose and of other energetic substrates, such as pyruvate, lactate, and citrate, is critically analyzed. Such knowledge, besides its obvious importance for basic science, could eventually translate into the development of novel strategies for treatment of male infertility, artificial reproduction, and sperm selection methods. PMID:24791005
McCain, Christy M
Elevational gradients hold enormous potential for understanding general properties of biodiversity. Like latitudinal gradients, the hypotheses for diversity patterns can be grouped into historical explanations, climatic drivers, and spatial hypotheses. The spatial hypotheses include the species-area effect and spatial constraint (mid-domain effect null models). I test these two spatial hypotheses using regional diversity patterns for mammals (non-volant small mammals and bats) along 34 elevational gradients spanning 24.4 degrees S-40.4 degrees N latitude. There was high variability in the fit to the species-area hypothesis and the mid-domain effect. Both hypotheses can be eliminated as primary drivers of elevational diversity. Area and spatial constraint both represent sources of error rather than mechanisms underlying these mammalian diversity patterns. Similar results are expected for other vertebrate taxa, plants, and invertebrates since they show comparable distributions of elevational diversity patterns to mammalian patterns.
Tong, Nianting; Zhang, Zhenzhen; Gong, Yuanyuan; Yin, Lili
Abstract Objective Diosmin, a natural flavone glycoside, possesses antioxidant activity and has been used to alleviate ischemia/reperfusion (I/R) injury. The aim of this study was to clarify whether the administration of diosmin has a protective effect against I/R injury induced using the high intraocular pressure (IOP) model in rat retina, and to determine the possible antioxidant mechanisms involved. Methods Retinal I/R injury was induced in the rats by elevating the IOP to 110 mmHg for 60 min. Diosmin (100 mg/kg) or vehicle solution was administered intragastrically 30 min before the onset of ischemia and then daily after I/R injury until the animals were sacrificed. The levels of malondialdehyde (MDA) and the activities of total-superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) in the retinal tissues were determined 24 h after I/R injury. At 7 days post-I/R injury, electroretinograms (ERGs) were recorded, and the density of surviving retinal ganglion cells (RGCs) was estimated by counting retrograde tracer-labeled cells in whole-mounted retinas. Retinal histological changes were also examined and quantified using light microscopy. Results Diosmin significantly decreased the MDA levels and increased the activities of T-SOD, GSH-Px, and CAT in the retina of rats compared with the ischemia group (P<0.05), and suppressed the I/R-induced reduction in the a- and b-wave amplitudes of the ERG (P<0.05). The thickness of the entire retina, inner nuclear layer, inner plexiform layer, and outer retinal layer and the number of cells in the ganglion cell layer were significantly less after I/R injury (P<0.05), and diosmin remarkably ameliorated these changes on retinal morphology. Diosmin also attenuated the I/R-induced loss of RGCs of the rat retina (P<0.05). Conclusion Diosmin protected the retina from I/R injury, possibly via a mechanism involving the regulation of oxidative parameters. PMID:22509733
Evaluation of the repeated-dose liver micronucleus assay using N-nitrosomorpholine in young adult rats: report on collaborative study by the Collaborative Study Group for the Micronucleus Test (CSGMT)/Japanese Environmental Mutagen Society (JEMS)-Mammalian Mutagenicity Study (MMS) Group.
Hayashi, Aya; Kosaka, Mizuki; Kimura, Aoi; Wako, Yumi; Kawasako, Kazufumi; Hamada, Shuichi
The present study was conducted to evaluate the suitability of a repeated-dose liver micronucleus (LMN) assay in young adult rats as a collaborative study by the Mammalian mutagenicity study (MMS) group. All procedures were performed in accordance with the standard protocols of the MMS Group. Six-week-old male Crl:CD(SD) rats (5 animals/group) received oral doses of the hepatocarcinogen N-nitrosomorpholine (NMOR) at 0 (control), 5, 10, and 30mg/kg/day (10mL/kg) for 14 days. Control animals received vehicle (water). Hepatocytes were collected from the liver 24h after the last dose, and the number of micronucleated hepatocytes (MNHEPs) was determined by microscopy. The number of micronucleated immature erythrocytes (MNIMEs) in the femoral bone marrow was also determined. The liver was examined using histopathologic methods after formalin fixation. The results showed statistically significant and dose-dependent increases in the number of MNHEPs in the liver at doses of 10mg/kg and greater when compared with the vehicle control. However, no significant increase was noted in the number of MNIMEs in the bone marrow at doses of up to 30mg/kg. Histopathology of the liver revealed hypertrophy and single cell necrosis of hepatocytes at doses of 5mg/kg and above. These results showed that the induction of micronuclei by NMOR was detected by the repeated-dose LMN assay, but not by the repeated-dose bone marrow micronucleus assay.
Pegg, Anthony E.
Summary Polyamines are ubiquitous small basic molecules that play multiple essential roles in mammalian physiology. Their cellular content is highly regulated and there is convincing evidence that altered metabolism is involvement in many disease states. Drugs altering polyamine levels may therefore have a variety of important targets. This review will summarize the current state of understanding of polyamine metabolism and function, the regulation of polyamine content, and heritable pathological conditions that may be derived from altered polyamine metabolism. PMID:19603518
Ben-Ari, Matan; Inbar, Moshe
The direct trophic links between mammalian herbivores and plant-dwelling insects have been practically ignored. Insects are ubiquitous on plants consumed by mammalian herbivores and are thus likely to face the danger of being incidentally ingested by a grazing mammal. A few studies have shown that some herbivorous hemipterans are able to avoid this peril by dropping to the ground upon detecting the heat and humidity on the mammal's breath. We hypothesized that if this risk affects the entire plant-dwelling insect community, other insects that share this habitat are expected to develop similar escape mechanisms. We assessed the ability of three species (adults and larvae) of coccinellid beetles, important aphid predators, to avoid incidental ingestion. Both larvae and adults were able to avoid incidental ingestion effectively by goats by dropping to the ground, demonstrating the importance of this behavior in grazed habitats. Remarkably, all adult beetles escaped by dropping off the plant and none used their functional wings to fly away. In controlled laboratory experiments, we found that human breath caused 60-80% of the beetles to drop. The most important component of mammalian herbivore breath in inducing adult beetles and larvae to drop was the combination of heat and humidity. The fact that the mechanism of dropping in response to mammalian breath developed in distinct insect orders and disparate life stages accentuates the importance of the direct influence of mammalian herbivores on plant-dwelling insects. This direct interaction should be given its due place when discussing trophic interactions.
Ben-Ari, Matan; Inbar, Moshe
The direct trophic links between mammalian herbivores and plant-dwelling insects have been practically ignored. Insects are ubiquitous on plants consumed by mammalian herbivores and are thus likely to face the danger of being incidentally ingested by a grazing mammal. A few studies have shown that some herbivorous hemipterans are able to avoid this peril by dropping to the ground upon detecting the heat and humidity on the mammal's breath. We hypothesized that if this risk affects the entire plant-dwelling insect community, other insects that share this habitat are expected to develop similar escape mechanisms. We assessed the ability of three species (adults and larvae) of coccinellid beetles, important aphid predators, to avoid incidental ingestion. Both larvae and adults were able to avoid incidental ingestion effectively by goats by dropping to the ground, demonstrating the importance of this behavior in grazed habitats. Remarkably, all adult beetles escaped by dropping off the plant and none used their functional wings to fly away. In controlled laboratory experiments, we found that human breath caused 60–80% of the beetles to drop. The most important component of mammalian herbivore breath in inducing adult beetles and larvae to drop was the combination of heat and humidity. The fact that the mechanism of dropping in response to mammalian breath developed in distinct insect orders and disparate life stages accentuates the importance of the direct influence of mammalian herbivores on plant-dwelling insects. This direct interaction should be given its due place when discussing trophic interactions. PMID:23424674
Fujieda, Hiroki; Sasaki, Hiroshi
Brain-derived neurotrophic factor (BDNF) regulates many aspects of neuronal development, including survival, axonal and dendritic growth and synapse formation. Despite recent advances in our understanding of the functional significance of BDNF in retinal development, the retinal cell types expressing BDNF remains poorly defined. The goal of the present study was to determine the localization of BDNF in the mammalian retina, with special focus on the subtypes of amacrine cells, and to characterize, at the cellular level, the effects of constant light exposure during early postnatal period on retinal expression of BDNF. Retinas from 3-week-old rats reared in a normal light cycle or constant light were subjected to double immunofluorescence staining using antibodies to BDNF and retinal cell markers. BDNF immunoreactivity was localized to ganglion cells, cholinergic amacrine cells and dopaminergic amacrine cells, but not to AII amacrine cells regardless of rearing conditions. Approximately 75% of BDNF-positive cells in the inner nuclear layer were cholinergic amacrine cells in animals reared in a normal lighting condition. While BDNF immunoreactivity in ganglion cells and cholinergic amacrine cells was significantly increased by constant light rearing, which in dopaminergic amacrine cells was apparently unaltered. The overall structure of the retina and the density of ganglion cells, cholinergic amacrine cells and AII amacrine cells were unaffected by rearing conditions, whereas the density of dopaminergic amacrine cells was significantly increased by constant light rearing. The present results indicate that cholinergic amacrine cells are the primary source of BDNF in the inner nuclear layer of the rat retina and provide the first evidence that cholinergic amacrine cells may be involved in the visual activity-dependent regulation of retinal development through the production of BDNF. The present data also suggest that the production or survival of dopaminergic amacrine
Bucci, Diego; Rodriguez-Gil, Juan Enrique; Vallorani, Claudia; Spinaci, Marcella; Galeati, Giovanna; Tamanini, Carlo
Mammalian cells use glucides as a substrate that can be catabolized through glycolitic pathways or oxidative phosphorylation, used as a source of reducing potential, or used for anabolic aims. An important role in supplying cells with energy is played by different membrane proteins that can actively (sodium-dependent glucose transporters) or passively (glucose transporters; GLUT) transport hexoses through the lipidic bilayer. In particular, GLUTs are a family of 13 proteins that facilitate the transport of sugars and have a peculiar distribution in different tissues as well as a particular affinity for substrates. These proteins are also present in mature sperm cells, which, in fact, need carriers for uptake energetic sources that are important for maintaining cell basic activity as well as specific functions, such as motility and fertilization ability. Likewise, several GLUTs have been studied in various mammalian species (man, bull, rat, mouse, boar, dog, stallion, and donkey) to point out both their actual presence or absence and their localization on plasma membrane. The aim of this work is to give an overall picture of the studies available on GLUTs in mammalian spermatozoa at this moment, pointing out the species peculiarity, the possible role of these proteins, and the potential future research on this item.
Hughes, Steven; Welsh, Laura; Katti, Christiana; González-Menéndez, Irene; Turton, Michael; Halford, Stephanie; Sekaran, Sumathi; Peirson, Stuart N.; Hankins, Mark W.; Foster, Russell G.
Photosensitive retinal ganglion cells (pRGCs) respond to light from birth and represent the earliest known light detection system to develop in the mouse retina. A number of morphologically and functionally distinct subtypes of pRGCs have been described in the adult retina, and have been linked to different physiological roles. We have previously identified two distinct isoforms of mouse melanopsin, Opn4L and Opn4S, which are generated by alternate splicing of the Opn4 locus. These isoforms are differentially expressed in pRGC subtypes of the adult mouse retina, with both Opn4L and Opn4S detected in M1 type pRGCs, and only Opn4L detected in M2 type pRGCs. Here we investigate the developmental expression of Opn4L and Opn4S and show a differential profile of expression during postnatal development. Opn4S mRNA is detected at relatively constant levels throughout postnatal development, with levels of Opn4S protein showing a marked increase between P0 and P3, and then increasing progressively over time until adult levels are reached by P10. By contrast, levels of Opn4L mRNA and protein are low at birth and show a marked increase at P14 and P30 compared to earlier time points. We suggest that these differing profiles of expression are associated with the functional maturation of M1 and M2 subtypes of pRGCs. Based upon our data, Opn4S expressing M1 type pRGCs mature first and are the dominant pRGC subtype in the neonate retina, whereas increased expression of Opn4L and the maturation of M2 type pRGCs occurs later, between P10 and P14, at a similar time to the maturation of rod and cone photoreceptors. We suggest that the distinct functions associated with these cell types will develop at different times during postnatal development. PMID:22496826
Hughes, Steven; Welsh, Laura; Katti, Christiana; González-Menéndez, Irene; Turton, Michael; Halford, Stephanie; Sekaran, Sumathi; Peirson, Stuart N; Hankins, Mark W; Foster, Russell G
Photosensitive retinal ganglion cells (pRGCs) respond to light from birth and represent the earliest known light detection system to develop in the mouse retina. A number of morphologically and functionally distinct subtypes of pRGCs have been described in the adult retina, and have been linked to different physiological roles. We have previously identified two distinct isoforms of mouse melanopsin, Opn4L and Opn4S, which are generated by alternate splicing of the Opn4 locus. These isoforms are differentially expressed in pRGC subtypes of the adult mouse retina, with both Opn4L and Opn4S detected in M1 type pRGCs, and only Opn4L detected in M2 type pRGCs. Here we investigate the developmental expression of Opn4L and Opn4S and show a differential profile of expression during postnatal development. Opn4S mRNA is detected at relatively constant levels throughout postnatal development, with levels of Opn4S protein showing a marked increase between P0 and P3, and then increasing progressively over time until adult levels are reached by P10. By contrast, levels of Opn4L mRNA and protein are low at birth and show a marked increase at P14 and P30 compared to earlier time points. We suggest that these differing profiles of expression are associated with the functional maturation of M1 and M2 subtypes of pRGCs. Based upon our data, Opn4S expressing M1 type pRGCs mature first and are the dominant pRGC subtype in the neonate retina, whereas increased expression of Opn4L and the maturation of M2 type pRGCs occurs later, between P10 and P14, at a similar time to the maturation of rod and cone photoreceptors. We suggest that the distinct functions associated with these cell types will develop at different times during postnatal development.
Clark, Damon A; Demb, Jonathan B
Sensory systems use receptors to extract information from the environment and neural circuits to perform subsequent computations. These computations may be described as algorithms composed of sequential mathematical operations. Comparing these operations across taxa reveals how different neural circuits have evolved to solve the same problem, even when using different mechanisms to implement the underlying math. In this review, we compare how insect and mammalian neural circuits have solved the problem of motion estimation, focusing on the fruit fly Drosophila and the mouse retina. Although the two systems implement computations with grossly different anatomy and molecular mechanisms, the underlying circuits transform light into motion signals with strikingly similar processing steps. These similarities run from photoreceptor gain control and spatiotemporal tuning to ON and OFF pathway structures, motion detection, and computed motion signals. The parallels between the two systems suggest that a limited set of algorithms for estimating motion satisfies both the needs of sighted creatures and the constraints imposed on them by metabolism, anatomy, and the structure and regularities of the visual world.
Evans, J W; Zawadzki, R J; Liu, R; Chan, J; Lane, S; Werner, J S
Imaging the structure and correlating it with the biochemical content of the retina holds promise for fundamental research and for clinical applications. Optical coherence tomography (OCT) is commonly used to image the 3D structure of the retina and while the added functionality of biochemical analysis afforded by Raman scattering could provide critical molecular signatures for clinicians and researchers, there are many technical challenges to combining these imaging modalities. We present an ex vivo OCT microscope combined with Raman spectroscopy capable of collecting morphological and molecular information about a sample simultaneously. The combined instrument will be used to investigate remaining technical challenges to combine these imaging modalities, such as the laser power levels needed to achieve a Raman signal above the noise level without damaging the sample.
Maple, Bruce R; Zhang, Jian; Pang, Ji-Jie; Gao, Fan; Wu, Samuel M
In immunocytochemical studies of the tiger salamander retina, 17% of neurons in the outer nuclear layer did not label for recoverin, a photoreceptor marker. Lucifer yellow injection showed a population of cells in the ONL to be displaced bipolar cells, with axon terminals that stratified exclusively in the OFF sublamina of the inner plexiform layer (IPL), and predominately within the cone-dominated region of the OFF sublamina. Glutamate generated a dendritic cationic conductance increase in all displaced bipolar cells tested, and typical cone-dominated bipolar cell light responses were observed among displaced cells that stratified in the central IPL. We conclude that displaced bipolar cells in the tiger salamander retina are entirely OFF-center cells, and predominately cone-dominated cells.
Koschmieder, Ingo; Müller, Lothar
Imaging of the retina of the human eye is an essential aid for medical diagnosis. The technical realization of photos of the ocular fundus is not trivial because of the optical properties of the eye. Established devices to obtain images are so called fundus cameras with digital documentation capabilities. New procedures do not need the use of pupils enlarging measures at the patient and work with infrared illumination. The quality of the diagnostic findings depends on the one hand fundamentally on the lay-out of the optical design of the fundus camera. On the other hand there are limitations caused by the eye itself which is always a part of the beam path. Both impacts define the attainable results. Special applications deal with the stereoscopic imaging of the retina or with spectral reflection characteristics.
Rajala, Raju V. S.
The phosphoinositide (PI) cycle, discovered over 50 years ago by Mabel and Lowell Hokin, describes a series of biochemical reactions that occur on the inner leaflet of the plasma membrane of cells in response to receptor activation by extracellular stimuli. Studies from our laboratory have shown that the retina and rod outer segments (ROSs) have active PI metabolism. Biochemical studies revealed that the ROSs contain the enzymes necessary for phosphorylation of phosphoinositides. We showed that light stimulates various components of the PI cycle in the vertebrate ROS, including diacylglycerol kinase, PI synthetase, phosphatidylinositol phosphate kinase, phospholipase C, and phosphoinositide 3-kinase (PI3K). This article describes recent studies on the PI3K-generated PI lipid second messengers in the control and regulation of PI-binding proteins in the vertebrate retina. PMID:19638643
The goal of the retinal prosthesis project is the development of an implantable microelectrode array that can be used to supply visually-driven electrical input to cells in the retina, bypassing nonfunctional rod and cone cells, thereby restoring vision to blind individuals. This goal will be achieved through the study of the fundamentals of electrical engineering, vision research, and biomedical engineering with the aim of acquiring the knowledge needed to engineer a high-density microelectrode-tissue hybrid sensor that will restore vision to millions of blind persons. The modeling and simulation task within this project is intended to address the question how best to stimulate, and communicate with, cells in the retina using implanted microelectrodes.
Bull, Natalie D; Martin, Keith R
Retinal degenerative diseases are the leading cause of incurable blindness worldwide. Furthermore, existing pharmacological and surgical interventions are only partially effective in halting disease progression, thus adjunctive neuroprotective strategies are desperately needed to preserve vision. Stem cells appear to possess inherent neuroprotective abilities, at least in part by providing neurotrophic support to injured neurons. Advances in stem cell biology offer the hope of new therapies for a broad range of neurodegenerative conditions, including those of the retina. Experimental cell-mediated therapies also hint at the tantalizing possibility of achieving retinal neuronal replacement and regeneration, once cells are lost to the disease process. This article summarizes the latest advances in cell therapies for neuroprotection and regeneration in neurodegenerative pathologies of both the inner and outer retina.
Ölveczky, Bence P.; Baccus, Stephen A.; Meister, Markus
An important task in vision is to detect objects moving within a stationary scene. During normal viewing this is complicated by the presence of eye movements that continually scan the image across the retina, even during fixation. To detect moving objects, the brain must distinguish local motion within the scene from the global retinal image drift due to fixational eye movements. We have found that this process begins in the retina: a subset of retinal ganglion cells responds to motion in the receptive field centre, but only if the wider surround moves with a different trajectory. This selectivity for differential motion is independent of direction, and can be explained by a model of retinal circuitry that invokes pooling over nonlinear interneurons. The suppression by global image motion is probably mediated by polyaxonal, wide-field amacrine cells with transient responses. We show how a population of ganglion cells selective for differential motion can rapidly flag moving objects, and even segregate multiple moving objects.
Zanello, Susana B.; Theriot, Corey; Westby, Christian; Boyle, Richard
Several stress environmental factors are combined in a unique fashion during spaceflight, affecting living beings widely across their physiological systems. Recently, attention has been placed on vision changes in astronauts returning from long duration missions. Alterations include hyperoptic shift, globe flattening, choroidal folds and optic disc edema, which are probably associated with increased intracranial pressure. These observations justify a better characterization of the ocular health risks associated with spaceflight. This study investigates the impact of spaceflight on the biology of the mouse retina. Within a successful tissue sharing effort, eyes from albino Balb/cJ mice aboard STS-133 were collected for histological analysis and gene expression profiling of the retina at 1 and 7 days after landing. Both vivarium and AEM (Animal Enclosure Module) mice were used as ground controls. Oxidative stress-induced DNA damage was higher in the flight samples compared to controls on R+1, and decreased on R+7. A trend toward higher oxidative and cellular stress response gene expression was also observed on R+1 compared to AEM controls, and these levels decreased on R+7. Several genes coding for key antioxidant enzymes, namely, heme-oxygenase-1, peroxiredoxin, and catalase, were among those upregulated after flight. Likewise, NF B and TGFbeta1, were upregulated in one flight specimen that overall showed the most elevated oxidative stress markers on R+1. In addition, retinas from vivarium control mice evidenced higher oxidative stress markers, NF B and TGFbeta1, likely due to the more intense illumination in vivarium cages versus the AEM. These preliminary data suggest that spaceflight represents a source of environmental stress that translates into oxidative and cellular stress in the retina, which is partially reversible upon return to Earth. Further work is needed to dissect the contribution of the various spaceflight factors (microgravity, radiation) and to
Qin, Zhenxia; Yang, Jing; Klassen, Henry J.; Aswad, Dana W.
Purpose. To determine the propensity of retinal proteins for spontaneous damage via formation of isoaspartyl sites, a common type of protein damage that could contribute to retinal disease. Methods. Tissue extracts were obtained from retinas and brains of control mice and from mice in which the gene for protein L-isoaspartate O-methyltransferase (PIMT; an enzyme that repairs isoaspartyl protein damage) was knocked out. PIMT expression in these extracts was measured by Western blot, and its specific activity was assayed by monitoring the rate of [3H]methyl transfer from S-adenosyl-[methyl-3H]L-methionine to γ-globulin. Isoaspartate levels in extracts were measured by their capacity to accept [3H]methyl groups via the PIMT-catalyzed methylation reaction. To compare molecular weight distributions of isoaspartyl-rich proteins in retina versus brain, proteins from PIMT knockout (KO) and control mice were separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF). Isoaspartyl proteins were 3H-labeled on-blot using a PIMT overlay and imaged by autoradiography. Results. When normalized to the β-actin content of each tissue, retina was found to be nearly identical to brain with regard to expression and activity of PIMT and its propensity to accumulate isoaspartyl sites when PIMT is absent. The two tissues show distinct differences in the molecular weight distribution of isoaspartyl proteins. Conclusions. The retina is rich in PIMT activity and contains a wide range of proteins that are highly susceptible to this type of protein damage. Recoverin may be one such protein. Isoaspartate formation, along with oxidation, should be considered as a potential source of protein dysfunction and autoimmunity in retinal disease. PMID:24550364
Weltzien, Felix; Percival, Kumiko A; Martin, Paul R; Grünert, Ulrike
About 15 parallel ganglion cell pathways transmit visual signals to the brain, but the interneuron (bipolar and amacrine) populations providing input to ganglion cells remain poorly understood in primate retina. We carried out a quantitative analysis of the inner nuclear layer in the retina of the marmoset (Callithrix jacchus). Vertical Vibratome sections along the horizontal meridian were processed with immunohistochemical markers. Image stacks were taken with a confocal microscope, and densities of cell populations were determined. The density of flat midget bipolar cells fell from 15,746 cells/mm(2) at 1 mm (8 deg) to 7,827 cells/mm(2) at 3 mm (25 deg). The rod bipolar cell density fell from 8,640 cells/mm(2) at 1 mm to 4,278 cells/mm(2) at 3 mm, but the ratio of the two bipolar cell types did not change with eccentricity. The amacrine cell density ranged from 30,000 cells/mm(2) at 8 deg to less than 15,000 cells/mm(2) at 25 deg, but throughout the retina, the ratio of glycinergic to γ-aminobutyric acid (GABA)ergic to amacrine cells remained relatively constant. The fractions of rod bipolar, cone bipolar, amacrine, Müller, and horizontal cells of all cells in the inner nuclear layer were comparable in central and peripheral retina. Marmosets had lower proportions of midget bipolar and rod bipolar in comparison with macaque. These differences were correlated with differences in rod and cone densities between the two species and did not reflect fundamental differences in the wiring between the two species.
Sargoy, Allison; Barnes, Steven; Brecha, Nicholas C; Pérez De Sevilla Müller, Luis
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.
Besharse, Joseph C; McMahon, Douglas G
Ocular clocks, first identified in the retina, are also found in the retinal pigment epithelium (RPE), cornea, and ciliary body. The retina is a complex tissue of many cell types and considerable effort has gone into determining which cell types exhibit clock properties. Current data suggest that photoreceptors as well as inner retinal neurons exhibit clock properties with photoreceptors dominating in nonmammalian vertebrates and inner retinal neurons dominating in mice. However, these differences may in part reflect the choice of circadian output, and it is likely that clock properties are widely dispersed among many retinal cell types. The phase of the retinal clock can be set directly by light. In nonmammalian vertebrates, direct light sensitivity is commonplace among body clocks, but in mice only the retina and cornea retain direct light-dependent phase regulation. This distinguishes the retina and possibly other ocular clocks from peripheral oscillators whose phase depends on the pace-making properties of the hypothalamic central brain clock, the suprachiasmatic nuclei (SCN). However, in mice, retinal circadian oscillations dampen quickly in isolation due to weak coupling of its individual cell-autonomous oscillators, and there is no evidence that retinal clocks are directly controlled through input from other oscillators. Retinal circadian regulation in both mammals and nonmammalian vertebrates uses melatonin and dopamine as dark- and light-adaptive neuromodulators, respectively, and light can regulate circadian phase indirectly through dopamine signaling. The melatonin/dopamine system appears to have evolved among nonmammalian vertebrates and retained with modification in mammals. Circadian clocks in the eye are critical for optimum visual function where they play a role fine tuning visual sensitivity, and their disruption can affect diseases such as glaucoma or retinal degeneration syndromes.
Buschini, Elisa; Piras, Antonio; Nuzzi, Raffaele; Vercelli, Alessandro
Inflammation protects from dangerous stimuli, restoring normal tissue homeostasis. Inflammatory response in the nervous system ("neuroinflammation") has distinct features, which are shared in several diseases. The retina is an immune-privileged site, and the tight balance of immune reaction can be disrupted and lead to age-related macular disease (AMD) and to its peculiar sign, the druse. Excessive activation of inflammatory and immunological cascade with subsequent induction of damage, persistent activation of resident immune cells, accumulation of byproducts that exceeds the normal capacity of clearance giving origin to a chronic local inflammation, alterations in the activation of the complement system, infiltration of macrophages, T-lymphocytes and mast-cells from the bloodstream, participate in the mechanisms which originate the drusen. In addition, aging of the retina and AMD involve also para-inflammation, by which immune cells react to persistent stressful stimuli generating low-grade inflammation, aimed at restoring function and maintaining tissue homeostasis by varying the set point in relation to the new altered conditions. This mechanism is also seen in the normal aging retina, but, in the presence of noxious stimuli as in AMD, it can become chronic and have an adverse outcome. Finally, autophagy may provide new insights to understand AMD pathology, due to its contribution in the removal of defective proteins. Therefore, the AMD retina can represent a valuable model to study neuroinflammation, its mechanisms and therapy in a restricted and controllable environment. Targeting these pathways could represent a new way to treat and prevent both exudative and dry forms of AMD.
Peichl, L; Künzle, H; Vogel, P
The retinae of insectivores have been rarely studied, and their photoreceptor arrangements and expression patterns of visual pigments are largely unknown. We have determined the presence and distribution of cones in three species of shrews (common shrew Sorex araneus, greater white-toothed shrew Crocidura russula, dark forest shrew Crocidura poensis; Soricidae) and in the lesser hedgehog tenrec Echinops telfairi (Tenrecidae). Special cone types were identified and quantified in flattened whole retinae by antisera/antibodies recognizing the middle-to-long-wavelength-sensitive (M/L-)cone opsin and the short-wavelength-sensitive (S-)cone opsin, respectively. A combination of immunocytochemistry with conventional histology was used to assess rod densities and cone/rod ratios. In all four species the rods dominate at densities of about 230,000-260,000/mm2. M/L- and S-cones are present, comprising between 2% of the photoreceptors in the nocturnal Echinops telfairi and 13% in Sorex araneus that has equal diurnal and nocturnal activity phases. This suggests dichromatic color vision like in many other mammals. A striking feature in all four species are dramatically higher S-cone proportions in ventral than in dorsal retina (0.5% vs. 2.5-12% in Sorex, 5-15% vs. 30-45% in Crocidura poensis, 3-12% vs. 20-50% in Crocidura russula, 10-30% vs. 40-70% in Echinops). The functional and comparative aspects of these structural findings are discussed.
Kim, Andy Jeesu; Chang, Janet Ya-An; Shi, Liheng; Chang, Richard Cheng-An; Ko, Michael Lee; Ko, Gladys Yi-Ping
Purpose The purpose of this study was to determine the effects of metformin on dysfunctional retinas in obesity-induced type 2 diabetic mice. Methods A high-fat diet (HFD)-induced diabetic mouse model (C57BL/6J) was used in this study. After 2 months of the HFD regimen, HFD mice were given daily metformin through oral gavage. Body weights, glucose tolerance, and retinal light responses were monitored regularly. Fluorescein angiography (FA) was used to assess changes in retinal vasculature. Ocular tissues (retina, vitreous, and lens) were harvested and analyzed for molecular changes as determined by immunofluorescent staining, Western blot analysis, and cytokine profiling. Results Starting 1 month after the diet regimen, mice fed the HFD had mildly compromised retinal light responses as measured by electroretinography (ERG), which worsened over time compared to that in the control. In HFD mice treated with metformin, systemic glucose levels reverted back to normal, and their weight gain slowed. Metformin reversed HFD-induced changes in phosphorylated protein kinase B (pAKT), extracellular signal-regulated kinase (pERK), and 5′AMP-activated protein kinase (pAMPK) in the retina. However, metformin treatments for 3 months did not restore the retinal light responses nor lessen the HFD-induced retinal neovascularization, even though it did reduce intraocular inflammation. Conclusions Although metformin was able to reverse systemic changes induced by HFD, it was not able to restore HFD-caused retinal light responses or deter neovascularization. PMID:28114566
de Juan, Joaquín; Ferrone, Claudia; Giannini, Daniela; Huang, David; Koch, Giorgio; Russo, Valentina; Tan, Ou; Bruni, Carlo
Optical coherence tomography (OCT) has recently become one of the primary methods for noninvasive probing of the human retina. The pseudoimage formed by OCT (the so-called B-scan) varies probabilistically across pixels due to complexities in the measurement technique. Hence, sensitive automatic procedures of diagnosis using OCT may exploit statistical analysis of the spatial distribution of reflectance. In this paper, we perform a statistical study of retinal OCT data. We find that the stretched exponential probability density function can model well the distribution of intensities in OCT pseudoimages. Moreover, we show a small, but significant correlation between neighbor pixels when measuring OCT intensities with pixels of about 5 µm. We then develop a simple joint probability model for the OCT data consistent with known retinal features. This model fits well the stretched exponential distribution of intensities and their spatial correlation. In normal retinas, fit parameters of this model are relatively constant along retinal layers, but varies across layers. However, in retinas with diabetic retinopathy, large spikes of parameter modulation interrupt the constancy within layers, exactly where pathologies are visible. We argue that these results give hope for improvement in statistical pathology-detection methods even when the disease is in its early stages. PMID:20304733
Du, Jianhai; Rountree, Austin; Cleghorn, Whitney M.; Contreras, Laura; Lindsay, Ken J.; Sadilek, Martin; Gu, Haiwei; Djukovic, Danijel; Raftery, Dan; Satrústegui, Jorgina; Kanow, Mark; Chan, Lawrence; Tsang, Stephen H.; Sweet, Ian R.; Hurley, James B.
Production of energy in a cell must keep pace with demand. Photoreceptors use ATP to maintain ion gradients in darkness, whereas in light they use it to support phototransduction. Matching production with consumption can be accomplished by coupling production directly to consumption. Alternatively, production can be set by a signal that anticipates demand. In this report we investigate the hypothesis that signaling through phototransduction controls production of energy in mouse retinas. We found that respiration in mouse retinas is not coupled tightly to ATP consumption. By analyzing metabolic flux in mouse retinas, we also found that phototransduction slows metabolic flux through glycolysis and through intermediates of the citric acid cycle. We also evaluated the relative contributions of regulation of the activities of α-ketoglutarate dehydrogenase and the aspartate-glutamate carrier 1. In addition, a comprehensive analysis of the retinal metabolome showed that phototransduction also influences steady-state concentrations of 5′-GMP, ribose-5-phosphate, ketone bodies, and purines. PMID:26677218
Pohl-Guimarães, Fernanda; Calaza, Karin da Costa; Yamasaki, Edna Nanami; Kubrusly, Regina Célia Cussa; Reis, Ricardo Augusto de Melo
Several mechanisms underlying ethanol action in GABAergic synapses have been proposed, one of these mechanisms is on GABA release. Here, we report that in ovo exposure to ethanol induces an increase on GABA release in the embryonic chick retina. Eleven-day-old chick embryos (E11) received an injection of either phosphate buffer saline (PBS) or ethanol (10%, v/v, diluted in PBS), and were allowed to develop until E16. A single glutamate stimulus (2 mM) showed approximately a 40% increase on GABA release in E16 retinas when compared to controls. The effect was dependent on NMDA receptors and GAD65 mRNA levels, which were increased following the ethanol treatment. However, the numbers of GABA-, GAD-, and NR1-immunoreactive cells, and the expression levels of these proteins, were not affected. We conclude that ethanol treatment at a time point when synapses are being formed during development selectively increases GABA release in the retina via a NMDA receptor-dependent process.
Schiller, Peter H.
In the retina, several parallel channels originate that extract different attributes from the visual scene. This review describes how these channels arise and what their functions are. Following the introduction four sections deal with these channels. The first discusses the “ON” and “OFF” channels that have arisen for the purpose of rapidly processing images in the visual scene that become visible by virtue of either light increment or light decrement; the ON channel processes images that become visible by virtue of light increment and the OFF channel processes images that become visible by virtue of light decrement. The second section examines the midget and parasol channels. The midget channel processes fine detail, wavelength information, and stereoscopic depth cues; the parasol channel plays a central role in processing motion and flicker as well as motion parallax cues for depth perception. Both these channels have ON and OFF subdivisions. The third section describes the accessory optic system that receives input from the retinal ganglion cells of Dogiel; these cells play a central role, in concert with the vestibular system, in stabilizing images on the retina to prevent the blurring of images that would otherwise occur when an organism is in motion. The last section provides a brief overview of several additional channels that originate in the retina. PMID:20876118
Contín, M A; Benedetto, M M; Quinteros-Quintana, M L; Guido, M E
Light is the visible part of the electromagnetic radiation within a range of 380–780 nm; (400–700 on primates retina). In vertebrates, the retina is adapted to capturing light photons and transmitting this information to other structures in the central nervous system. In mammals, light acts directly on the retina to fulfill two important roles: (1) the visual function through rod and cone photoreceptor cells and (2) non-image forming tasks, such as the synchronization of circadian rhythms to a 24 h solar cycle, pineal melatonin suppression and pupil light reflexes. However, the excess of illumination may cause retinal degeneration or accelerate genetic retinal diseases. In the last century human society has increased its exposure to artificial illumination, producing changes in the Light/Dark cycle, as well as in light wavelengths and intensities. Although, the consequences of unnatural illumination or light pollution have been underestimated by modern society in its way of life, light pollution may have a strong impact on people's health. The effects of artificial light sources could have direct consequences on retinal health. Constant exposure to different wavelengths and intensities of light promoted by light pollution may produce retinal degeneration as a consequence of photoreceptor or retinal pigment epithelium cells death. In this review we summarize the different mechanisms of retinal damage related to the light exposure, which generates light pollution. PMID:26541085
Contín, M A; Benedetto, M M; Quinteros-Quintana, M L; Guido, M E
Light is the visible part of the electromagnetic radiation within a range of 380-780 nm; (400-700 on primates retina). In vertebrates, the retina is adapted to capturing light photons and transmitting this information to other structures in the central nervous system. In mammals, light acts directly on the retina to fulfill two important roles: (1) the visual function through rod and cone photoreceptor cells and (2) non-image forming tasks, such as the synchronization of circadian rhythms to a 24 h solar cycle, pineal melatonin suppression and pupil light reflexes. However, the excess of illumination may cause retinal degeneration or accelerate genetic retinal diseases. In the last century human society has increased its exposure to artificial illumination, producing changes in the Light/Dark cycle, as well as in light wavelengths and intensities. Although, the consequences of unnatural illumination or light pollution have been underestimated by modern society in its way of life, light pollution may have a strong impact on people's health. The effects of artificial light sources could have direct consequences on retinal health. Constant exposure to different wavelengths and intensities of light promoted by light pollution may produce retinal degeneration as a consequence of photoreceptor or retinal pigment epithelium cells death. In this review we summarize the different mechanisms of retinal damage related to the light exposure, which generates light pollution.
Burns, S A; Elsner, A E
We investigated the low-frequency temporal response of the retina by measuring the corneal electroretinogram elicited by flickering lights. A sum of two temporal sine-wave modulations was used to generate difference frequencies between a 36-Hz standard stimulus and a series of low-frequency stimuli. The response of the retina at the difference frequency did not change as the low-frequency component of the stimulus was varied from 0.5 to 4 Hz. We also replicated an earlier study, stimulating the retina with a sum of two sine waves that were varied in average frequency but keeping the difference frequency constant. These data showed no change in the amplitude of the difference frequency as the average stimulus frequency was varied from 8 to almost 40 Hz. Taken together, the two sets of data support the notion that the in vivo early retinal response is low pass and extends without attenuation to frequencies greater than 30 Hz, in contrast to the sensitivity of the visual system measured by psychophysical techniques.
Wang, Benquan; Yao, Xincheng
Intrinsic optical signal (IOS) imaging is a promising noninvasive method for advanced study and diagnosis of eye diseases. Before pursuing clinical applications, more IOS studies employing animal models are necessary to establish the relationship between IOS distortions and eye diseases. Ample mouse models are available for investigating the relationship between IOS distortions and eye diseases. However, in vivo IOS imaging of mouse retinas is challenging due to the small ocular lens (compared to frog eyes) and inevitable eye movements. We report here in vivo IOS imaging of mouse retinas using a custom-designed functional OCT. The OCT system provided high resolution (3 μm) and high speed (up to 500 frames/s) imaging of mouse retinas. An animal holder equipped with a custom designed ear bar and bite bar was used to minimize eye movement due to breathing and heartbeats. Residual eye movement in OCT images was further compensated by accurate image registration. Dynamic OCT imaging revealed rapid IOSs from photoreceptor outer segments immediately (<10 ms) after the stimulation delivery, and unambiguous IOS changes were also observed from inner retinal layers with delayed time courses compared to that of photoreceptor IOSs.
Bastawrous, Andrew; Giardini, Mario Ettore; Bolster, Nigel M; Peto, Tunde; Shah, Nisha; Livingstone, Iain AT; Weiss, Helen A.; Hu, Sen; Rono, Hillary; Kuper, Hannah; Burton, Matthew
Importance Visualization and interpretation of the optic nerve and retina is an essential part of most physical examinations. Objectives To design and validate a smartphone-based retinal adapter enabling image capture and remote grading of the retina Design, setting and participants Validation study comparing the grading of optic nerves from smartphones images with those of a Digital Fundus Camera. Both image sets were independently graded at Moorfields Eye Hospital Reading Centre. Nested within the six-year follow-up of the Nakuru Eye Disease Cohort in Kenya: 1,460adults (2,920eyes) aged 55years and above were recruited consecutively from the Study. A sub-set of 100 optic disc images from both methods were further used to validate a grading app for the optic nerves. Main outcome(s) and measure(s) Vertical cup-to-disc-ratio (VCDR) for each test was compared, in terms of agreement (Bland-Altman & weighted Kappa) and test-retest variability (TRV). Results 2,152 optic nerve images were available from both methods (additionally 371 from reference but not Peek, 170 from Peek but not the reference and 227 from neither the reference camera or Peek). Bland-Altman analysis demonstrated a difference of the average of 0.02 with 95% limits of agreement between -0.21 and 0.17 and a weighted Kappa coefficient of 0.69 (excellent agreement). An experienced retinal photographer was compared to a lay photographer (no health care experience prior to the study) with no observable difference in image acquisition quality between them. Conclusions and relevance Non-clinical photographers using the low-cost Peek Retina adapter and smartphone were able to acquire optic nerve images at a standard that enabled comparable independent remote grading of the images to those acquired using a desktop retinal camera operated by an ophthalmic assistant. The potential for task-shifting and the detection of avoidable causes of blindness in the most at risk communities makes this an attractive public
Katyal, Sachin; Glubrecht, Darryl D.; Li, Lei; Gao, Zhihua; Godbout, Roseline
Background The Reelin-Dab1 signaling pathway plays a critical role in the positioning of migrating neurons, dendrite formation and lamination in the developing central nervous system. We have previously identified two alternatively spliced forms of Dab1 in the developing chick retina: an early form, Dab1-E, expressed in retinal progenitor cells, and a late form, Dab1 or Dab1-L, expressed in amacrine and ganglion cells. Compared to Dab1-L, Dab1-E lacks two exons that encode two Src family kinase (SFK) phosphorylation sites. Principal Findings Both Dab1-L and Dab1-E-like transcripts were identified in human fetal retina. Expression of human Dab1-L in primary chick retinal cultures resulted in Reelin-mediated induction of SFK phosphorylation and formation of neurite-like processes. In contrast, human Dab1-E-expressing cells retained an undifferentiated morphology. The human Dab1 gene is located within a common fragile site, and it has been postulated that it may function as a tumor suppressor. Analysis of Dab1 splice forms in retinoblastoma and neuroblastoma tumor cells revealed relative enrichment of Dab1-L-like (includes exons 7 and 8) and Dab1-E-like (excludes exons 7 and 8) transcripts in retinoblastoma and neuroblastoma, respectively. Treatment of retinoblastoma cell line RB522A with Reelin resulted in increased tyrosine phosphorylation of Dab1. As Nova2 has previously been implicated in the exclusion of exons 9B and 9C in Dab1, we examined the expression of this splicing factor in neuroblastoma and retinoblastoma cell lines. Nova2 was only detected in neuroblastoma cells, suggesting a correlation between Nova2 expression and increased levels of Dab1-E-like splice forms in neuroblastoma. Conclusions These results indicate that alternative splicing of Dab1 is conserved in avian and mammalian species, with Dab1-L driving SFK phosphorylation in both species. Dab1-E- and Dab-L-like isoforms are also expressed in childhood neural tumors, with preferential enrichment
Stella, Salvatore L.; Li, Stefanie; Sabatini, Andrea; Vila, Alejandro; Brecha, Nicholas C.
Glutamate is the major excitatory neurotransmitter in the retina, and most glutamatergic neurons express one of the three known vesicular glutamate transporters (VGLUT1, 2, or 3). However, the expression profiles of these transporters vary greatly in the retina. VGLUT1 is expressed by photoreceptor and bipolar cell terminals, and VGLUT2 appears to be predominately expressed by ganglion cells, and perhaps Müller cells, cone photoreceptor terminals, and horizontal cells in some species. The discovery of a third vesicular glutamate transporter, VGLUT3, has brought about speculation concerning its role and function based on its expression in amacrine cells. To address this we studied the postnatal development of VGLUT3 from day 0 through adult in the rat retina, and compared this with the expression patterns of VGLUT1 and VGLUT2. VGLUT3 expression was restricted to a population of amacrine cells. Expression of VGLUT3 was first observed at postnatal day 10 (P10) in the soma and some processes, which extensively arborized in both the ON and OFF sublamina of the IPL by P15. In contrast, VGLUT1 and VGLUT2 expression appeared earlier than VGLUT3; with VGLUT1 initially detected at P5 in photoreceptor terminals and P6 in bipolar terminals, and VGLUT2 immunoreactivity initially detected at P0 in ganglion cell bodies, and remained prominent throughout all stages of development. Interestingly, VGLUT3 has extensive somatic expression throughout development, which could be involved in non-synaptic modulation by glutamate in developing retina, and could influence trophic and extra-synaptic neuronal signaling by glutamate in the inner retina. PMID:18482716
Dong, Cecilia Q.; Panagis, Lampros; Kamthan, Gautam; Ren, Lizhen; Rozenboym, Anna; Perera, Tarique D.; Coplan, Jeremy D.; Danias, John
Purpose To determine whether short-term pressure elevation affects complement gene expression in the retina in vitro and in vivo. Methods Muller cell (TR-MUL5) cultures and organotypic retinal cultures from adult mice and monkeys were subjected to either 24-h or 72-h of pressure at 0, 15, 30, and 45 mmHg above ambient. C57BL/6 mice were subjected to microbead-induced intraocular pressure (IOP) elevation for 7 days. RNA and protein were extracted and used for analysis of expression levels of complement component genes and complement component 1, q subcomponent (C1q) and complement factor H (CFH) immunoblotting. Results mRNA levels of complement genes and C1q protein levels in Muller cell cultures remained the same for all pressure levels after exposure for either 24 or 72 h. In primate and murine organotypic cultures, pressure elevation did not produce changes in complement gene expression or C1q and CFH protein levels at either the 24-h or 72-h time points. Pressure-related glial fibrillary acidic protein (GFAP) mRNA expression changes were detected in primate retinal organotypic cultures (analysis of variance [ANOVA]; p<0.05). mRNA expression of several other genes changed as a result of time in culture. Eyes subjected to microbead-induced IOP elevation had no differences in mRNA expression of complement genes and C1q protein levels (ANOVA; p>0.05 for both) with contralateral control and naïve control eyes. Conclusions Short-term elevation of pressure in vitro as well as short-term (1 week) IOP elevation in vivo does not seem to dramatically alter complement system gene expression in the retina. Prolonged expression to elevated pressure may be necessary to affect the complement system expression. PMID:24505213
Lužnik, Zala; Parekh, Mohit; Bertolin, Marina; Griffoni, Carlo; Ponzin, Diego
Summary Retinal degenerative diseases are one of the main clinical causes of incurable and severe visional impairment. Thus, extensive research effort is put into the development of new causal therapeutic options. Promisingly, a number of studies showed regenerative capacity in specific retinal regions (the ciliary epithelium, retinal pigmented epithelium, iris, and Müller glia cells). However, most recent research studies are based on animal models or in vitro cultured cells, probably because of the limited availability of human posterior eye tissues (vitreous, retina, and choroid). To address this, we showed in our previous reports that eye banks with large numbers of globes collected yearly could set up biorepositories/biobanks where these precious tissues are isolated, quality controlled, and finally stored for scientists and clinicians wanting to access human tissues and test their own hypotheses. These precious human posterior eye tissues could be used for further research purposes, epidemiological studies, and target validation of newly developed drugs. In addition, this could be a promising and challenging option to retrieve potential retinal stem and progenitor cells from different parts of the retina and could be a breakthrough in the future delivery of ex vivo prepared customized (histocompatible) retinal tissue on scaffolds for transplantation purposes. In this Perspective, we will consider how the biorepositories could influence the future strategies for retinal stem cell therapies. Significance Retinal degenerative diseases are one of the main causes of severe vision impairment and regenerative medicine is attracting much attention as a potential therapy. Although highly desirable, the reactivation and proliferation of endogenous stem cells in vivo is not sufficient to generate enough cells to restore visual function after retinal injury. Thus, the replacement of exogenously derived normal donor cells is a promising solution. The challenge is to
Wang, Li-Ting; Wang, Shih-Jong; Hsu, Shih-Hsien
Wntless (GPR177) protein is a newly identified regulator of Wnt signals in Drosophila, but its cellular function in mammals is still unclear. In this study, we explored the expression pattern and potential cellular function of Wntless in mammalian cells. Wntless mRNA was expressed in many mouse tissues, including the spleen, lung, kidney, thymus, and stomach, and lower levels of expression were detected in the mouse brain and testis. Expression of Wntless protein analyzed by Western blot and immunohistochemical staining was only detected in the submucosa, muscle, ganglia, and nerve cells of murine large intestines. Both immunofluorescence staining and subcellular fraction extraction analysis revealed that endogenous Wntless protein was expressed predominantly in the cytoplasmic organelles with a morphologically dot-shaped distribution. Furthermore, overexpression of Wntless could be corrected by and may activate the nuclear factor-κB (NF-κB) signaling pathway in cancer (HeLa) cells. These results suggest that Wntless plays a role in signaling regulation during the formation of cancer in addition to its role as a retromer protein in mammalian systems.
Manley, Geoffrey A
Evolution of the cochlea and high-frequency hearing (>20 kHz; ultrasonic to humans) in mammals has been a subject of research for many years. Recent advances in paleontological techniques, especially the use of micro-CT scans, now provide important new insights that are here reviewed. True mammals arose more than 200 million years (Ma) ago. Of these, three lineages survived into recent geological times. These animals uniquely developed three middle ear ossicles, but these ossicles were not initially freely suspended as in modern mammals. The earliest mammalian cochleae were only about 2 mm long and contained a lagena macula. In the multituberculate and monotreme mammalian lineages, the cochlea remained relatively short and did not coil, even in modern representatives. In the lineage leading to modern therians (placental and marsupial mammals), cochlear coiling did develop, but only after a period of at least 60 Ma. Even Late Jurassic mammals show only a 270 ° cochlear coil and a cochlear canal length of merely 3 mm. Comparisons of modern organisms, mammalian ancestors, and the state of the middle ear strongly suggest that high-frequency hearing (>20 kHz) was not realized until the early Cretaceous (~125 Ma). At that time, therian mammals arose and possessed a fully coiled cochlea. The evolution of modern features of the middle ear and cochlea in the many later lineages of therians was, however, a mosaic and different features arose at different times. In parallel with cochlear structural evolution, prestins in therian mammals evolved into effective components of a new motor system. Ultrasonic hearing developed quite late-the earliest bat cochleae (~60 Ma) did not show features characteristic of those of modern bats that are sensitive to high ultrasonic frequencies.
Hirata, Akimasa; Takano, Yukinori; Fujiwara, Osamu; Dovan, Thanh; Kavet, Robert
For magnetic field exposures at extremely low frequencies, the electrostimulatory response with the lowest threshold is the magnetophosphene, a response that corresponds to an adult exposed to a 20 Hz magnetic field of nominally 8.14 mT. In the IEEE standard C95.6 (2002), the corresponding in situ field in the retinal locus of an adult-sized ellipsoidal was calculated to be 53 mV m-1. However, the associated dose in the retina and brain at a high level of resolution in anatomically correct human models is incompletely characterized. Furthermore, the dose maxima in tissue computed with voxel human models are prone to staircasing errors, particularly for the low-frequency dosimetry. In the analyses presented in this paper, analytical and quasi-static finite-difference time-domain (FDTD) solutions were first compared for a three-layer sphere exposed to a uniform 50 Hz magnetic field. Staircasing errors in the FDTD results were observed at the tissue interface, and were greatest at the skin-air boundary. The 99th percentile value was within 3% of the analytic maximum, depending on model resolution, and thus may be considered a close approximation of the analytic maximum. For the adult anatomical model, TARO, exposed to a uniform magnetic field, the differences in the 99th percentile value of in situ electric fields for 2 mm and 1 mm voxel models were at most several per cent. For various human models exposed at the magnetophosphene threshold at three orthogonal field orientations, the in situ electric field in the brain was between 10% and 70% greater than the analytical IEEE threshold of 53 mV m-1, and in the retina was lower by roughly 50% for two horizontal orientations (anterior-posterior and lateral), and greater by about 15% for a vertically oriented field. Considering a reduction factor or safety factors of several folds applied to electrostimulatory thresholds, the 99th percentile dose to a tissue calculated with voxel human models may be used as an estimate of
Hirata, Akimasa; Takano, Yukinori; Fujiwara, Osamu; Dovan, Thanh; Kavet, Robert
For magnetic field exposures at extremely low frequencies, the electrostimulatory response with the lowest threshold is the magnetophosphene, a response that corresponds to an adult exposed to a 20 Hz magnetic field of nominally 8.14 mT. In the IEEE standard C95.6 (2002), the corresponding in situ field in the retinal locus of an adult-sized ellipsoidal was calculated to be 53 mV m(-1). However, the associated dose in the retina and brain at a high level of resolution in anatomically correct human models is incompletely characterized. Furthermore, the dose maxima in tissue computed with voxel human models are prone to staircasing errors, particularly for the low-frequency dosimetry. In the analyses presented in this paper, analytical and quasi-static finite-difference time-domain (FDTD) solutions were first compared for a three-layer sphere exposed to a uniform 50 Hz magnetic field. Staircasing errors in the FDTD results were observed at the tissue interface, and were greatest at the skin-air boundary. The 99th percentile value was within 3% of the analytic maximum, depending on model resolution, and thus may be considered a close approximation of the analytic maximum. For the adult anatomical model, TARO, exposed to a uniform magnetic field, the differences in the 99th percentile value of in situ electric fields for 2 mm and 1 mm voxel models were at most several per cent. For various human models exposed at the magnetophosphene threshold at three orthogonal field orientations, the in situ electric field in the brain was between 10% and 70% greater than the analytical IEEE threshold of 53 mV m(-1), and in the retina was lower by roughly 50% for two horizontal orientations (anterior-posterior and lateral), and greater by about 15% for a vertically oriented field. Considering a reduction factor or safety factors of several folds applied to electrostimulatory thresholds, the 99th percentile dose to a tissue calculated with voxel human models may be used as an
Xu, Min; Nagati, Jason S.; Xie, Jian; Li, Jiwen; Walters, Holly; Moon, Young-Ah; Gerard, Robert D.; Huang, Chou-Long; Comerford, Sarah A.; Hammer, Robert E.; Horton, Jay D.; Chen, Rui; Garcia, Joseph A.
Endocrine erythropoietin (Epo), which is synthesized in the kidney or liver of adult mammals, controls erythrocyte production and is regulated by the stress-responsive transcription factor Hypoxia Inducible Factor 2 (HIF-2). We previously reported that the lysine acetyltransferase Cbp is required for HIF-2α acetylation and efficient HIF-2 dependent Epo induction during hypoxia. We now show these processes require acetate-dependent acetyl CoA synthetase 2 (Acss2). In Hep3B hepatoma cells and in Epo-generating organs of hypoxic or acutely anemic mice, acetate levels increase and Acss2 is required for HIF-2α acetylation, Cbp/HIF-2α complex formation and recruitment to the Epo enhancer, and efficient Epo induction. In acutely anemic mice, acetate supplementation augments stress erythropoiesis in an Acss2-dependent manner. In acquired and genetic chronic anemia mouse models, acetate supplementation also increases Epo expression and resting hematocrits. Thus, a mammalian stress-responsive acetate switch controls HIF-2 signaling and Epo induction during pathophysiological states marked by tissue hypoxia. PMID:25108527
Ceramide, the backbone structure of all sphingolipids, as well as a minor component of cellular membranes, has a unique role in the skin, by forming the epidermal permeability barrier at the extracellular domains of the outermost layer of the skin, the stratum corneum, which is required for terrestrial mammalian survival. In contrast to the role of ceramide in forming the permeability barrier, the signaling roles of ceramide and its metabolites have not yet been recognized. Ceramide and/or its metabolites regulate proliferation, differentiation, and apoptosis in epidermal keratinocytes. Recent studies have further demonstrated that a ceramide metabolite, sphingosine-1-phosphate, modulates innate immune function. Ceramide has already been applied to therapeutic approaches for treatment of eczema associated with attenuated epidermal permeability barrier function. Pharmacological modulation of ceramide and its metabolites' signaling can also be applied to cutaneous disease prevention and therapy. The author here describes the signaling roles of ceramide and its metabolites in mammalian cells and tissues, including the epidermis. This article is part of a Special Issue entitled The Important Role of Lipids in the Epidermis and their Role in the Formation and Maintenance of the Cutaneous Barrier. Guest Editors: Kenneth R. Feingold and Peter Elias.
Peyman, G A; Conway, M D; House, B J
We evaluated the effects of argon-green (514.5 nm) and CW neodymium YAG (1060 nm) wavelengths on experimentally detached retinas of primates. Neither laser produced damage to the sensory retina of the fovea. The argon green wavelength, which was absorbed by haemoglobin in the vessel or by extravasated red blood cells, created vasospasm and nerve fiber layer damage. The beam of the CW YAG was not absorbed by haemoglobin; therefore, no vasospasm could be produced on experimentally detached retinas.
Rapp, L.M.; Jose, J.G.; Pitts, D.G.
Quantitative autoradiography was used to study the incorporation of /sup 3/H-thymidine into the retina of albino rats following in vivo exposure to 300-nm radiation. Relative to background labeling in unexposed eyes, there was 8-20 times as much label per unit area in the outer nuclear layer, inner nuclear layer, and ganglion cells of 300-nm exposed retinas. The photoreceptor inner segments also showed thymidine labeling in both control and exposed retinas.
Evaluation of the repeated-dose liver and gastrointestinal tract micronucleus assays with 22 chemicals using young adult rats: summary of the collaborative study by the Collaborative Study Group for the Micronucleus Test (CSGMT)/The Japanese Environmental Mutagen Society (JEMS) - Mammalian Mutagenicity Study Group (MMS).
Hamada, Shuichi; Ohyama, Wakako; Takashima, Rie; Shimada, Keisuke; Matsumoto, Kazumi; Kawakami, Satoru; Uno, Fuyumi; Sui, Hajime; Shimada, Yasushi; Imamura, Tadashi; Matsumura, Shoji; Sanada, Hisakazu; Inoue, Kenji; Muto, Shigeharu; Ogawa, Izumi; Hayashi, Aya; Takayanagi, Tomomi; Ogiwara, Yosuke; Maeda, Akihisa; Okada, Emiko; Terashima, Yukari; Takasawa, Hironao; Narumi, Kazunori; Wako, Yumi; Kawasako, Kazufumi; Sano, Masaki; Ohashi, Nobuyuki; Morita, Takeshi; Kojima, Hajime; Honma, Masamitsu; Hayashi, Makoto
The repeated-dose liver micronucleus (RDLMN) assay using young adult rats has the potential to detect hepatocarcinogens. We conducted a collaborative study to assess the performance of this assay and to evaluate the possibility of integrating it into general toxicological studies. Twenty-four testing laboratories belonging to the Mammalian Mutagenicity Study Group, a subgroup of the Japanese Environmental Mutagen Society, participated in this trial. Twenty-two model chemicals, including some hepatocarcinogens, were tested in 14- and/or 28-day RDLMN assays. As a result, 14 out of the 16 hepatocarcinogens were positive, including 9 genotoxic hepatocarcinogens, which were reported negative in the bone marrow/peripheral blood micronucleus (MN) assay by a single treatment. These outcomes show the high sensitivity of the RDLMN assay to hepatocarcinogens. Regarding the specificity, 4 out of the 6 non-liver targeted genotoxic carcinogens gave negative responses. This shows the high organ specificity of the RDLMN assay. In addition to the RDLMN assay, we simultaneously conducted gastrointestinal tract MN assays using 6 of the above carcinogens as an optional trial of the collaborative study. The MN assay using the glandular stomach, which is the first contact site of the test chemical when administered by oral gavage, was able to detect chromosomal aberrations with 3 test chemicals including a stomach-targeted carcinogen. The treatment regime was the 14- and/or 28-day repeated-dose, and the regime is sufficiently promising to incorporate these methods into repeated-dose toxicological studies. The outcomes of our collaborative study indicated that the new techniques to detect chromosomal aberrations in vivo in several tissues worked successfully.
Cao, Nan; Cao, Fengmei; Lin, Yabin; Bai, Tingzhu; Song, Shengyu
For a new kind of retina-like senor camera and a traditional rectangular sensor camera, dual cameras acquisition and display system need to be built. We introduce the principle and the development of retina-like senor. Image coordinates transformation and interpolation based on sub-pixel interpolation need to be realized for our retina-like sensor's special pixels distribution. The hardware platform is composed of retina-like senor camera, rectangular sensor camera, image grabber and PC. Combined the MIL and OpenCV library, the software program is composed in VC++ on VS 2010. Experience results show that the system can realizes two cameras' acquisition and display.
Background Irradiation with light wavelengths from the far red (FR) to the near infrared (NIR) spectrum (600 nm -1000 nm) has been shown to have beneficial effects in several disease models. In this study, we aim to examine whether 670 nm red light pretreatment can provide protection against hyperoxia-induced damage in the C57BL/6J mouse retina. Adult mice (90–110 days) were pretreated with 9 J/cm2 of 670 nm light once daily for 5 consecutive days prior to being placed in hyperoxic environment (75% oxygen). Control groups were exposed to hyperoxia, but received no 670 nm light pretreatment. Retinas were collected after 0, 3, 7, 10 or 14 days of hyperoxia exposure (n = 12/group) and prepared either for histological analysis, or RNA extraction and quantitative polymerase chain reaction (qPCR). Photoreceptor damage and loss were quantified by counting photoreceptors undergoing cell death and measuring photoreceptor layer thickness. Localization of acrolein, and cytochrome c oxidase subunit Va (Cox Va) were identified through immunohistochemistry. Expression of heme oxygenase-1 (Hmox-1), complement component 3 (C3) and fibroblast growth factor 2 (Fgf-2) genes were quantified using qPCR. Results The hyperoxia-induced photoreceptor loss was accompanied by reduction of metabolic marker, Cox Va, and increased expression of oxidative stress indicator, acrolein and Hmox-1. Pretreatment with 670 nm red light reduced expression of markers of oxidative stress and C3, and slowed, but did not prevent, photoreceptor loss over the time course of hyperoxia exposure. Conclusion The damaging effects of hyperoxia on photoreceptors were ameliorated following pretreatment with 670 nm light in hyperoxic mouse retinas. These results suggest that pretreatment with 670 nm light may provide stability to photoreceptors in conditions of oxidative stress. PMID:24134095
Kokona, Despina; Charalampopoulos, Ioannis; Pediaditakis, Iosif; Gravanis, Achille; Thermos, Kyriaki
The aim of the present study was to investigate the neuroprotective properties of the endogenous neurosteroid dehydroepiandrosterone (DHEA) in an in vivo model of retinal excitotoxicity, and the involvement of Nerve Growth Factor (NGF) in its actions. Adult Sprague-Dawley rats (250-300 g) received intravitreally (RS)-alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid hydrobromide (AMPA; 42 nmol/eye) alone or in combination with DHEA (10(-8), 10(-7), 10(-6) M), or PBS (50 mM, control group). To examine the involvement of NGF and its TrkA receptor in the pharmacological effects of DHEA, animals received AMPA and NGF (60 pg/eye) in the absence or presence of a TrkA receptor inhibitor (Calbiochem 648450, 10(-6) M) or AMPA, DHEA (10(-6) M) and TrkA receptor inhibitor (10(-6), 10(-5) M). Immunohistochemistry studies [choline acetyltransferase (ChAT), brain nitric oxide synthetase (bNOS), calbindin, and TUNEL] and fluorescence-activated cell sorting (FACS) were used to examine retinal cell loss and protection. TrkA receptor immunoreactivity (-IR) and colocalization studies with relevant markers were also performed. AMPA (42 nmol) treatment resulted in a loss of bNOS, ChAT and calbindin immunoreactivities 24 h after its administration. DHEA, administered intravitreally, protected the retina from excitotoxicity in a dose-dependent manner. This effect was mimicked by NGF, and reversed by the NGF TrkA receptor inhibitor. The TrkA receptor is expressed in ganglion cells of rat retina. TUNEL staining and FACS analysis substantiated the neuroprotective actions of DHEA. These results demonstrate for the first time that the neurosteroid DHEA, administered intravitreally, protects the retina from AMPA excitotoxicity. An NGF TrkA receptor mechanism appears to be involved in this neuroprotection.
Rocha, F A F; Saito, C A; Silveira, L C L; de Souza, J M; Ventura, D F
The turtle retina has been extensively used for the study of chromatic processing mechanisms. Color opponency has been previously investigated with trichromatic paradigms, but behavioral studies show that the turtle has an ultraviolet (UV) channel and a tetrachromatic visual system. Our laboratory has been working in the characterization of neuronal responses in the retina of vertebrates using stimuli in the UV-visible range of the electromagnetic spectrum. In the present investigation, we recorded color-opponent responses from turtle amacrine and ganglion cells to UV and visible stimuli and extended our previous results that UV color-opponency is present at the level of the inner nuclear layer. We recorded from 181 neurons, 36 of which were spectrally opponent. Among these, there were 10 amacrine (5%), and 26 ganglion cells (15%). Morphological identification of color-opponent neurons was possible for two ganglion cell classes (G17 and G22) and two amacrine cell classes (A22 and A23b). There was a variety of cell response types and a potential for complex processing of chromatic stimuli, with intensity- and wavelength-dependent response components. Ten types of color opponency were found in ganglion cells and by adding previous results from our laboratory, 12 types of opponent responses have been found. The majority of the ganglion cells were R+UVBG- and RG+UVB-color-opponents but there were other less frequent types of chromatic opponency. This study confirms the participation of a UV channel in the processing of color opponency in the turtle inner retina and shows that the turtle visual system has the retinal mechanisms to allow many possible chromatic combinations.
Figueiredo, Isabel N; Moura, Susana; Neves, Júlio S; Pinto, Luís; Kumar, Sunil; Oliveira, Carlos M; Ramos, João D
In this work we propose a novel method for identifying individuals based on retinal fundus image matching. The method is based on the image registration of retina blood vessels, since it is known that the retina vasculature of an individual is a signature, i.e., a distinctive pattern of the individual. The proposed image registration consists of a multiscale affine registration followed by a multiscale elastic registration. The major advantage of this particular two-step image registration procedure is that it is able to account for both rigid and non-rigid deformations either inherent to the retina tissues or as a result of the imaging process itself. Afterwards a decision identification measure, relying on a suitable normalized function, is defined to decide whether or not the pair of images belongs to the same individual. The method is tested on a data set of 21721 real pairs generated from a total of 946 retinal fundus images of 339 different individuals, consisting of patients followed in the context of different retinal diseases and also healthy patients. The evaluation of its performance reveals that it achieves a very low false rejection rate (FRR) at zero FAR (the false acceptance rate), equal to 0.084, as well as a low equal error rate (EER), equal to 0.053. Moreover, the tests performed by using only the multiscale affine registration, and discarding the multiscale elastic registration, clearly show the advantage of the proposed approach. The outcome of this study also indicates that the proposed method is reliable and competitive with other existing retinal identification methods, and forecasts its future appropriateness and applicability in real-life applications.
Berkowitz, Bruce A.; Bredell, Bryce X.; Davis, Christopher; Samardzija, Marijana; Grimm, Christian; Roberts, Robin
Purpose Excessive and continuously produced free radicals in the outer retina are implicated in retinal aging and the pathogenesis of sight-threatening retinopathies, yet measuring outer retinal oxidative stress in vivo remains a challenge. Here, we test the hypothesis that continuously produced paramagnetic free radicals from the outer retina can be measured in vivo using high-resolution (22-μm axial resolution) 1/T1magnetic resonance imaging (MRI) without and with a confirmatory quench (quench-assisted MRI). Methods Low-dose sodium iodate–treated and diabetic C57Bl6/J mice (and their controls), and rod-dominated (129S6) or cone-only R91W;Nrl−/− mice were studied. In dark-adapted groups, 1/T1 was mapped transretinally in vivo without or with (1) the antioxidant combination of methylene blue (MB) and α-lipoic acid (LPA), or (2) light exposure; in subgroups, retinal superoxide production was measured ex vivo (lucigenin). Results In the sodium iodate model, retinal superoxide production and outer retina-specific 1/T1 values were both significantly greater than normal and corrected to baseline with MB+LPA therapy. Nondiabetic mice at two ages and 1.2-month diabetic mice (before the appearance of oxidative stress) had similar transretinal 1/T1 profiles. By 2.3 months of diabetes, only outer retinal 1/T1 values were significantly greater than normal and were corrected to baseline with MB+LPA therapy. In mice with healthy photoreceptors, a light quench caused 1/T1 of rods, but not cones, to significantly decrease from their values in the dark. Conclusions Quench-assisted MRI is a feasible method for noninvasively measuring normal and pathologic production of free radicals in photoreceptors/RPE in vivo. PMID:26670830
Headington, Kenneth; Choi, Stacey S.; Nickla, Debora; Doble, Nathan
Purpose The chick eye is extensively used as a model in the study of myopia and its progression; however, analysis of the photoreceptor mosaic has required the use of excised retina due to the uncorrected optical aberrations in the lens and cornea. This study implemented high resolution adaptive optics (AO) retinal imaging to visualize the chick cone mosaic in vivo. Methods The New England College of Optometry (NECO) AO fundus camera was modified to allow high resolution in vivo imaging on 2 six-week-old White Leghorn chicks (Gallus gallus domesticus) – labeled chick A and chick B. Multiple, adjacent images, each with a 2.5° field of view, were taken and subsequently montaged together. This process was repeated at varying retinal locations measured from the tip of the pecten. Automated software was used to determine the cone spacing and density at each location. Voronoi analysis was applied to determine the packing arrangement of the cones. Results In both chicks, cone photoreceptors were clearly visible at all retinal locations imaged. Cone densities measured at 36° nasal-12° superior retina from the pecten tip for chick A and 40° nasal-12° superior retina for chick B were 21,714±543 and 26,105±653 cones/mm2 respectively. For chick B, a further 11 locations immediately surrounding the pecten were imaged, with cone densities ranging from 20,980±524 to 25,148±629 cones/mm2. Conclusion In vivo analysis of the cone density and its packing characteristics are now possible in the chick eye through AO imaging, which has important implications for future studies of myopia and ocular disease research. PMID:21950701
Bauer, Patrik Maximilian; Zalis, Marina Castro; Abdshill, Hodan; Deierborg, Tomas; Johansson, Fredrik; Englund-Johansson, Ulrica
Background Disease progression in retinal neurodegeneration is strongly correlated to immune cell activation, which may have either a neuroprotective or neurotoxic effect. Increased knowledge about the immune response profile and retinal neurodegeneration may lead to candidate targets for treatments. Therefore, we have used the explanted retina as a model to explore the immune response and expression of the immune modulator galectin-3 (Gal-3), induced by the cultivation per se and after additional immune stimulation with lipopolysaccharide (LPS), and how this correlates with retinal neurotoxicity. Methods Post-natal mouse retinas were cultured in a defined medium. One group was stimulated with LPS (100 ng/ml, 24 h). Retinal architecture, apoptotic cell death, and micro- and macroglial activity were studied at the time of cultivation (0 days in vitro (DIV)) and at 3, 4 and 7 DIV using morphological staining, biochemical- and immunohistochemical techniques. Results Our results show that sustained activation of macro- and microglia, characterized by no detectable cytokine release and limited expression of Gal-3, is not further inducing apoptosis additional to the axotomy-induced apoptosis in innermost nuclear layer. An elevated immune response was detected after LPS stimulation, as demonstrated primarily by release of immune mediators (i.e. interleukin 2 (IL-2), IL-6, KC/GRO (also known as CLCX1) and tumour necrosis factor-α (TNF-α)), increased numbers of microglia displaying morphologies of late activation stages as well as Gal-3 expression. This was accompanied with increased apoptosis in the two additional nuclear layers, and damage to retinal gross architecture. Conclusion We demonstrate that an immune response characterized by sustained and increased release of cytokines, along with an increase in Gal-3 expression, is accompanied by significant increased neurotoxicity in the explanted retina. Further investigations using the current setting may lead to
Eberhart, Anja; Feodorova, Yana; Song, Congdi; Wanner, Gerhard; Kiseleva, Elena; Furukawa, Takahisa; Kimura, Hiroshi; Schotta, Gunnar; Leonhardt, Heinrich; Joffe, Boris; Solovei, Irina
To improve light propagation through the retina, the rod nuclei of nocturnal mammals are uniquely changed compared to the nuclei of other cells. In particular, the main classes of chromatin are segregated in them and form regular concentric shells in order; inverted in comparison to conventional nuclei. A broad study of the epigenetic landscape of the inverted and conventional mouse retinal nuclei indicated several differences between them and several features of general interest for the organization of the mammalian nuclei. In difference to nuclei with conventional architecture, the packing density of pericentromeric satellites and LINE-rich chromatin is similar in inverted rod nuclei; euchromatin has a lower packing density in both cases. A high global chromatin condensation in rod nuclei minimizes the structural difference between active and inactive X chromosome homologues. DNA methylation is observed primarily in the chromocenter, Dnmt1 is primarily associated with the euchromatic shell. Heterochromatin proteins HP1-alpha and HP1-beta localize in heterochromatic shells, whereas HP1-gamma is associated with euchromatin. For most of the 25 studied histone modifications, we observed predominant colocalization with a certain main chromatin class. Both inversions in rod nuclei and maintenance of peripheral heterochromatin in conventional nuclei are not affected by a loss or depletion of the major silencing core histone modifications in respective knock-out mice, but for different reasons. Maintenance of peripheral heterochromatin appears to be ensured by redundancy both at the level of enzymes setting the epigenetic code (writers) and the code itself, whereas inversion in rods rely on the absence of the peripheral heterochromatin tethers (absence of code readers).
Cenci, Riccardo; Bedeschi, Franco; Marino, Pietro; Morello, Michael J.; Ninci, Daniele; Piucci, Alessio; Punzi, Giovanni; Ristori, Luciano; Spinella, Franco; Stracka, Simone; Tonelli, Diego; Walsh, John
We report on the performance of a specialized processor capable of reconstructing charged particle tracks in a realistic LHC silicon tracker detector, at the same speed of the readout and with sub-microsecond latency. The processor is based on an innovative pattern-recognition algorithm, called "artificial retina algorithm", inspired from the vision system of mammals. A prototype of the processor has been designed, simulated, and implemented on Tel62 boards equipped with high-bandwidth Altera Stratix III FPGA devices. The prototype is the first step towards a real-time track reconstruction device aimed at processing complex events of high-luminosity LHC experiments at 40 MHz crossing rate.
Masland, Richard H.; Mills, John W.
Photoreceptor cells of the rabbit retina accumulate choline from the extracellular environment by an overall process that has a high affinity for choline. These cells do not synthesize acetylcholine; instead, the choline taken up is incorporated into phosphorylcholine and eventually phospholipid. A mechanism for efficient choline accumulation is presumably concomitant to the photoreceptor cell's synthesis of large amounts of membrane for outer segment membrane renewal. Its existence in the photoreceptor cell supports previous evidence that high-affinity choline uptake is not confined to neurons that release acetylcholine, but may be present wherever large amounts of choline are required.
Liets, Lauren C; Eliasieh, Kasra; van der List, Deborah A; Chalupa, Leo M
The aging nervous system is known to manifest a variety of degenerative and regressive events. Here we report the unexpected growth of dendrites in the retinas of normal old mice. The dendrites of many rod bipolar cells in aging mice were observed to extend well beyond their normal strata within the outer plexiform layer to innervate the outer nuclear layer where they appeared to form contacts with the spherules of rod photoreceptors. Such dendritic sprouting increased with age and was evident at all retinal eccentricities. These results provide evidence of retinal plasticity associated with normal aging.
This dissertation develops laser Raman and fluorescence based spectroscopy for the noninvasive detection of medically important pigments in the human retina. Large-scale epidemiological studies have recently shown that the pigments lutein and zeaxanthin, located in the ˜1 mm diameter macular area of the retina, protect the eye from phototoxic blue light and/or oxidative damage. Resonance Raman spectroscopy (RRS) can detect and monitor macular pigments in intact human eyes quantitatively by recording the Raman scattered light originating from the highly specific stretching vibrations of the pigment molecules' conjugated carbon backbone. This dissertation develops RRS from a spatially averaged measuring approach to spatially resolved imaging. For this purpose, a filter-based Raman imaging setup with speckle-free illumination was constructed that permits detection of macular pigments at physiological concentrations with eye-safe laser excitation levels. Subsequently, RRS images would be obtained from the living human retina. The images demonstrate quantitative as well as micron-scale, spatially resolved RRS detection of the whole macular pigment distribution. The RRS images reveal important physiological details of a subject's macular pigment distribution such as the peaked pigment concentration in the center of the macula, and the rapidly dropping pigment concentration towards the periphery of the macula. As an alternative to direct RRS imaging of macular pigments, this dissertation explores an indirect imaging approach of macular pigments, based on excitation spectroscopy of lipofuscin. A dual-wavelength laser apparatus was constructed that excites the lipofuscin fluorescence at wavelengths inside and outside the spectral range of macular pigment absorption, and that allows one to image the fluorescence intensities in a large section of the retina centered on the macula. Measuring the lipofuscin fluorescence intensities inside and outside the macula area at the two
Gonda, Steve R.; Helmstetter, Charles E.; Thornton, Maureen
A method and bioreactor for the continuous production of synchronous (same age) population of mammalian cells have been invented. The invention involves the attachment and growth of cells on an adhesive-coated porous membrane immersed in a perfused liquid culture medium in a microgravity analog bioreactor. When cells attach to the surface divide, newborn cells are released into the flowing culture medium. The released cells, consisting of a uniform population of synchronous cells are then collected from the effluent culture medium. This invention could be of interest to researchers investigating the effects of the geneotoxic effects of the space environment (microgravity, radiation, chemicals, gases) and to pharmaceutical and biotechnology companies involved in research on aging and cancer, and in new drug development and testing.
Damuth, John; MacFadden, Bruce J.
This valuable collection of essays presents and evaluates techniques of body-mass estimation and reviews current and potential applications of body-size estimates in paleobiology. Papers discuss explicitly the errors and biases of various regression techniques and predictor variables, and the identification of functionally similar groups of species for improving the accuracy of estimates. At the same time other chapters review and discuss the physiological, ecological, and behavioral correlates of body size in extant mammals; the significance of body-mass distributions in mammalian faunas; and the ecology and evolution of body size in particular paleofaunas. Coverage is particularly detailed for carnivores, primates, and ungulates, but information is also presented on marsupials, rodents, and proboscideans.
Maderson, P F A
A 1972 model for the evolutionary origin of hair suggested a primary mechanoreceptor role improving behavioral thermoregulation contributed to the success of late Paleozoic mammal-like reptiles. An insulatory role appeared secondarily subsequent to protohair multiplication. That model is updated in light of new data on (a) palaeoecology of mammalian ancestors; (b) involvement of HRPs in keratinization; (c) lipogenic lamellar bodies that form the barrier to cutaneous water loss; and (d) growth factors involved in hair follicle embryogenesis and turnover. It is now proposed that multiplication of sensory protohairs caused by mutations in patterning genes initially protected the delicate barrier tissues and eventually produced the minimal morphology necessary for an insulatory pelage. The latter permitted Mesozoic mammals to occupy the nocturnal niche 'in the shadow of dinosaurs'. When the giant reptiles became extinct, mammals underwent rapid radiation and reemerged as the dominant terrestrial vertebrates.
Márquez, Javier; Cardona, Carolina; Campos-Sandoval, José A; Peñalver, Ana; Tosina, Marta; Matés, José M; Martín-Rufián, Mercedes
Glutamine/glutamate homeostasis must be exquisitely regulated in mammalian brain and glutaminase (GA, E.C. 184.108.40.206) is one of the main enzymes involved. The products of GA reaction, glutamate and ammonia, are essential metabolites for energy and biosynthetic purposes but they are also hazardous compounds at concentrations beyond their normal physiological thresholds. The classical pattern of GA expression in mammals has been recently challenged by the discovery of novel transcript variants and protein isoforms. Furthermore, the interactome of brain GA is also starting to be uncovered adding a new level of regulatory complexity. GA may traffic in brain and unexpected locations, like cytosol and nucleus, have been found for GA isoforms. Finally, the expression of GA in glial cells has been reported and its potential implications in ammonia homeostasis are discussed.
Smith, Richard S; Peterlin, Zita; Araneda, Ricardo C
Mammalian species have evolved a large and diverse number of odorant receptors (ORs). These proteins comprise the largest family of G-protein-coupled receptors (GPCRs) known, amounting to ~1,000-different receptors in the rodent. From the perspective of olfactory coding, the availability of such a vast number of chemosensory receptors poses several fascinating questions; in addition, such a large repertoire provides an attractive biological model to study ligand-receptor interactions. The limited functional expression of these receptors in heterologous systems, however, has greatly hampered attempts to deorphanize them. We have employed a successful approach that combines electrophysiological and imaging techniques to analyze the response profiles of single sensory neurons. Our approach has enabled us to characterize the "odor space" of a population of native aldehyde receptors and the molecular range of a genetically engineered receptor, OR-I7.
Nakada, K; Inoue, K; Hayashi, J
We generated mice with deletion mutant mtDNA by its introduction from somatic cells into mouse zygotes. Expressions of disease phenotypes are limited to tissues expressing mitochondrial dysfunction. Considering that all these mice share the same nuclear background, these observations suggest that accumulation of the mutant mtDNA and resultant expressions of mitochondrial dysfunction are responsible for expression of disease phenotypes. On the other hand, mitochondrial dysfunction and expression of clinical abnormalities were not observed until the mutant mtDNA accumulated predominantly. This protection is due to the presence of extensive and continuous interaction between exogenous mitochondria from cybrids and recipient mitochondria from embryos. Thus, we would like to propose a new hypothesis on mitochondrial biogenesis, interaction theory of mitochondria: mammalian mitochondria exchange genetic contents, and thus lost the individuality and function as a single dynamic cellular unit.
Farley, Katherine I.; Surovtseva, Yulia; Merkel, Janie; Baserga, Susan J.
The nucleolus is responsible for the production of ribosomes, essential machines which synthesize all proteins needed by the cell. The structure of human nucleoli is highly dynamic and is directly related to its functions in ribosome biogenesis. Despite the importance of this organelle, the intricate relationship between nucleolar structure and function remains largely unexplored. How do cells control nucleolar formation and function? What are the minimal requirements for making a functional nucleolus? Here we review what is currently known regarding mammalian nucleolar formation at nucleolar organizer regions (NORs), which can be studied by observing the dissolution and reformation of the nucleolus during each cell division. Additionally, the nucleolus can be examined by analyzing how alterations in nucleolar function manifest in differences in nucleolar architecture. Furthermore, changes in nucleolar structure and function are correlated with cancer, highlighting the importance of studying the determinants of nucleolar formation. PMID:25670395
Sekirnjak, Chris; Hottowy, Pawel; Sher, Alexander; Dabrowski, Wladyslaw; Litke, Alan M.; Chichilnisky, E. J.
Current epiretinal implants contain a small number of electrodes with diameters of a few hundred microns. Smaller electrodes are desirable to increase the spatial resolution of artificial sight. To lay the foundation for the next generation of retinal prostheses, we assessed the stimulation efficacy of micro-fabricated arrays of 61 platinum disk electrodes with diameters 8-12 μm, spaced 60 μm apart. Isolated pieces of rat, guinea pig, and monkey retina were placed on the multi-electrode array ganglion cell side down and stimulated through individual electrodes with biphasic, charge-balanced current pulses. Spike responses from retinal ganglion cells were recorded either from the same or a neighboring electrode. Most pulses evoked only 1-2 spikes with short latencies (0.3-10 ms), and rarely was more than one recorded ganglion cell stimulated. Threshold charge densities for eliciting spikes in ganglion cells were typically below 0.15 mC/cm2 for pulse durations between 50 and 200 μs, corresponding to charge thresholds of ˜ 100 pC. Stimulation remained effective after several hours and at frequencies up to 100 Hz. Application of cadmium chloride did not abolish evoked spikes, implying direct activation. Thus, electrical stimulation of mammalian retina with small-diameter electrodes is achievable, providing high temporal and spatial precision with low charge densities.
Puck, T T; Krystosek, A; Chan, D C
A theory is presented proposing that genetic regulation in mammalian cells is at least a two-tiered effect; that one level of regulation involves the transition between gene exposure and sequestration; that normal differentiation requires a different spectrum of genes to be exposed in each separate state of differentiation; that the fiber systems of the cell cytoskeleton and the nuclear matrix together control the degree of gene exposure; that specific phosphorylation of these elements causes them to assume a different organizational network and to impose a different pattern of sequestration and exposure on the elements of the genome; that the varied gene phosphorylation mechanisms in the cell are integrated in this function; that attachment of this network system to specific parts of the chromosomes brings about sequestration or exposure of the genes in their neighborhood in a fashion similar to that observed when microtubule elements attach through the kinetochore to the centromeric DNA; that one function of repetitive sequences is to serve as elements for the final attachment of this fibrous network to the specific chromosomal loci; and that at least an important part of the calcium manifestation as a metabolic trigger of different differentiation states involves its acting as a binding agent to centers of electronegativity, in particular proteins and especially phosphorylated groups, so as to change the conformation of the fiber network that ultimately controls gene exposure in the mammalian cell. It would appear essential to determine what abnormal gene exposures and sequestrations are characteristic of each type of cancer; which agonists, if any, will bring about reverse transformation; and whether these considerations can be used in therapy.
Manley, Geoffrey A
The hearing organs of amniote vertebrates show large differences in their size and structure between the species' groups. In spite of this, their performance in terms of hearing sensitivity and the frequency selectivity of auditory-nerve units shows unexpectedly small differences. The only substantial difference is that therian, defined as live-bearing, mammalian groups are able to hear ultrasonic frequencies (above 15-20 kHz), whereas in contrast monotreme (egg laying) mammals and all non-mammalian amniotes cannot. This review compares the structure and physiology of the cochleae of the main groups and asks the question as to why the many structural differences seen in therian mammals arose, yet did not result in greater differences in physiology. The likely answers to this question are found in the history of the mammals during the Cretaceous period that ended 65 million years ago. During that period, the therian cochlea lost its lagenar macula, leading to a fall in endolymph calcium levels. This likely resulted in a small revolution and an auditory crisis that was compensated for by a subsequent series of structural and physiological adaptations. The end result was a system of equivalent performance to that independently evolved in other amniotes but with the additional - and of course "unforeseen" - advantage that ultrasonic-frequency responses became an available option. That option was not always availed of, but in most groups of therian mammals it did evolve and is used for communication and orientation based on improved sound localization, with micro-bats and toothed whales relying on it for prey capture.
Zile, M. R.; Richardson, K.; Cowles, M. K.; Buckley, J. M.; Koide, M.; Cowles, B. A.; Gharpuray, V.; Cooper, G. 4th
BACKGROUND: The purpose of this study was to determine whether changes in the constitutive properties of the cardiac muscle cell play a causative role in the development of diastolic dysfunction. METHODS AND RESULTS: Cardiocytes from normal and pressure-hypertrophied cats were embedded in an agarose gel, placed on a stretching device, and subjected to a change in stress (sigma), and resultant changes in cell strain (epsilon) were measured. These measurements were used to examine the passive elastic spring, viscous damping, and myofilament activation. The passive elastic spring was assessed in protocol A by increasing the sigma on the agarose gel at a constant rate to define the cardiocyte sigma-versus-epsilon relationship. Viscous damping was assessed in protocol B from the loop area between the cardiocyte sigma-versus-epsilon relationship during an increase and then a decrease in sigma. In both protocols, myofilament activation was minimized by a reduction in [Ca2+]i. Myofilament activation effects were assessed in protocol C by defining cardiocyte sigma versus epsilon during an increase in sigma with physiological [Ca2+]i. In protocol A, the cardiocyte sigma-versus-epsilon relationship was similar in normal and hypertrophied cells. In protocol B, the loop area was greater in hypertrophied than normal cardiocytes. In protocol C, the sigma-versus-epsilon relation in hypertrophied cardiocytes was shifted to the left compared with normal cells. CONCLUSIONS: Changes in viscous damping and myofilament activation in combination may cause pressure-hypertrophied cardiocytes to resist changes in shape during diastole and contribute to diastolic dysfunction.
Vinson, Gavin P.
The standard model of adrenocortical zonation holds that the three main zones, glomerulosa, fasciculata, and reticularis each have a distinct function, producing mineralocorticoids (in fact just aldosterone), glucocorticoids, and androgens respectively. Moreover, each zone has its specific mechanism of regulation, though ACTH has actions throughout. Finally, the cells of the cortex originate from a stem cell population in the outer cortex or capsule, and migrate centripetally, changing their phenotype as they progress through the zones. Recent progress in understanding the development of the gland and the distribution of steroidogenic enzymes, trophic hormone receptors, and other factors suggests that this model needs refinement. Firstly, proliferation can take place throughout the gland, and although the stem cells are certainly located in the periphery, zonal replenishment can take place within zones. Perhaps more importantly, neither the distribution of enzymes nor receptors suggest that the individual zones are necessarily autonomous in their production of steroid. This is particularly true of the glomerulosa, which does not seem to have the full suite of enzymes required for aldosterone biosynthesis. Nor, in the rat anyway, does it express MC2R to account for the response of aldosterone to ACTH. It is known that in development, recruitment of stem cells is stimulated by signals from within the glomerulosa. Furthermore, throughout the cortex local regulatory factors, including cytokines, catecholamines and the tissue renin-angiotensin system, modify and refine the effects of the systemic trophic factors. In these and other ways it more and more appears that the functions of the gland should be viewed as an integrated whole, greater than the sum of its component parts. PMID:27378832
Costa, Liliana; Faustino, Maria Amparo F; Neves, Maria Graça P M S; Cunha, Angela; Almeida, Adelaide
Photodynamic inactivation (PDI) has been used to inactivate microorganisms through the use of photosensitizers. The inactivation of mammalian viruses and bacteriophages by photosensitization has been applied with success since the first decades of the last century. Due to the fact that mammalian viruses are known to pose a threat to public health and that bacteriophages are frequently used as models of mammalian viruses, it is important to know and understand the mechanisms and photodynamic procedures involved in their photoinactivation. The aim of this review is to (i) summarize the main approaches developed until now for the photodynamic inactivation of bacteriophages and mammalian viruses and, (ii) discuss and compare the present state of the art of mammalian viruses PDI with phage photoinactivation, with special focus on the most relevant mechanisms, molecular targets and factors affecting the viral inactivation process.
Bandaranayake, Ashok D.; Almo, Steven C.
Mammalian protein production platforms have had a profound impact in many areas of basic and applied research, and an increasing number of blockbuster drugs are recombinant mammalian proteins. With global sales of these drugs exceeding US$120 billion per year, both industry and academic research groups continue to develop cost effective methods for producing mammalian proteins to support preclinical and clinical evaluations of potential therapeutics. While a wide range of platforms have been successfully exploited for laboratory use, the bulk of recent biologics have been produced in mammalian cell lines due to the requirement for post translational modification and the biosynthetic complexity of the target proteins. In this review we highlight the range of mammalian expression platforms available for recombinant protein production, as well as advances in technologies for the rapid and efficient selection of highly productive clones. PMID:24316512
Costa, Liliana; Faustino, Maria Amparo F.; Neves, Maria Graça P. M. S.; Cunha, Ângela; Almeida, Adelaide
Photodynamic inactivation (PDI) has been used to inactivate microorganisms through the use of photosensitizers. The inactivation of mammalian viruses and bacteriophages by photosensitization has been applied with success since the first decades of the last century. Due to the fact that mammalian viruses are known to pose a threat to public health and that bacteriophages are frequently used as models of mammalian viruses, it is important to know and understand the mechanisms and photodynamic procedures involved in their photoinactivation. The aim of this review is to (i) summarize the main approaches developed until now for the photodynamic inactivation of bacteriophages and mammalian viruses and, (ii) discuss and compare the present state of the art of mammalian viruses PDI with phage photoinactivation, with special focus on the most relevant mechanisms, molecular targets and factors affecting the viral inactivation process. PMID:22852040
Lundh von Leithner, Peter; Ciurtin, Coziana
Purpose The vertebrate retina develops from the center to the periphery. In amphibians and fish the retinal margin continues to proliferate throughout life, resulting in retinal expansion. This does not happen in mammals. However, some mammalian peripheral retinal pigment epithelial (RPE) cells continue to divide, perhaps as a vestige of this mechanism. The RPE cells are adjacent to the ciliary margin, a known stem cell source. Here we test the hypothesis that peripheral RPE is fundamentally different from central RPE by challenging different regions with microscopic laser burns and charting differential responses in terms of levels of proliferation and the regions over which this proliferation occurs. Methods Microscopic RPE lesions were undertaken in rats at different eccentricities and the tissue stained for proliferative markers Ki67 and bromodeoxyuridine (BrdU) and the remodeling metalloproteinase marker 2 (MMP2). Results All lesions produced local RPE proliferation and tissue remodeling. Significantly more mitosis resulted from peripheral than central lesions. Unexpectedly, single lesions also resulted in RPE cells proliferating across the entire retina. Their number did not increase linearly with lesion number, indicating that they may be a specific population. All lesions repaired and formed apparently normal relations with the neural retina. Repaired RPE was albino. Conclusions These results highlight regional RPE differences, revealing an enhanced peripheral repair capacity. Further, all lesions have a marked impact on both local and distant RPE cells, demonstrating a pan retinal signaling mechanism triggering proliferation across the tissue plane. The RPE cells may represent a distinct population as their number did not increase with multiple lesions. The fact that repairing cells were hypopigmented is of interest because reduced pigment is associated with enhanced proliferative capacities in the developing neural retina. PMID:20360994
Holtmaat, Anthony; Svoboda, Karel
Synaptic plasticity in adult neural circuits may involve the strengthening or weakening of existing synapses as well as structural plasticity, including synapse formation and elimination. Indeed, long-term in vivo imaging studies are beginning to reveal the structural dynamics of neocortical neurons in the normal and injured adult brain. Although the overall cell-specific morphology of axons and dendrites, as well as of a subpopulation of small synaptic structures, are remarkably stable, there is increasing evidence that experience-dependent plasticity of specific circuits in the somatosensory and visual cortex involves cell type-specific structural plasticity: some boutons and dendritic spines appear and disappear, accompanied by synapse formation and elimination, respectively. This Review focuses on recent evidence for such structural forms of synaptic plasticity in the mammalian cortex and outlines open questions.
Claudepierre, Thomas; Manglapus, Mary K.; Marengi, Nathan; Radner, Stephanie; Champliaud, Marie-France; Tasanen, Kaisa; Bruckner-Tuderman, Leena; Hunter, Dale D.; Brunken, William J.
The ectoderm gives rise not only to the skin but also to the entire CNS. This common embryonic lineage suggests that some molecular isoforms might serve analogous functions in both tissues. Indeed, not only are laminins important components of dermal adhesion mechanisms, but they also regulate some aspects of synaptic development in both the CNS and the PNS. In the skin, laminins are part of a hemidesmosome complex essential for basal keratinocyte adhesion that includes collagen XVII (BP180) and BPAG1 (dystonin/BP230). Here, we show that CNS neurons also express collagen XVII and BPAG1 and that these molecules are expressed in the adult and developing retina. In the retina, isoforms of collagen XVII and BPAG1 are colocalized with laminins at photoreceptor synapses and around photoreceptor outer segments; both molecules are expressed by rods, whereas cones express collagen XVII but not BPAG1. Moreover, biochemical data demonstrate that collagen XVII complexes with retinal laminins. We propose that collagen XVII and BPAG1 isoforms may help to anchor elements of the rod photoreceptor cytomatrix to the extracellular matrix. PMID:15880472
Gilliam, Jared C; Wensel, Theodore G
In order to identify candidate cation channels important for retinal physiology, 28 TRP channel genes were surveyed for expression in the mouse retina. Transcripts for all TRP channels were detected by RT-PCR and sequencing. Northern blotting revealed that mRNAs for 12 TRP channel genes are enriched in the retina. The strongest signals were observed for TRPC1, TRPC3, TRPM1, TRPM3, and TRPML1, and clear signals were obtained for TRPC4, TRPM7, TRPP2, TRPV2, and TRPV4. In situ hybridization and immunofluorescence revealed widespread expression throughout multiple retinal layers for TRPC1, TRPC3, TRPC4, TRPML1, PKD1, and TRPP2. Striking localization of enhanced mRNA expression was observed for TRPC1 in the photoreceptor inner segment layer, for TRPM1 in the inner nuclear layer (INL), for TRPM3 in the INL, and for TRPML1 in the outer plexiform and nuclear layers. Strong immunofluorescence signal in cone outer segments was observed for TRPM7 and TRPP2. TRPC5 immunostaining was largely confined to INL cells immediately adjacent to the inner plexiform layer. TRPV2 antibodies stained photoreceptor axons in the outer plexiform layer. Expression of TRPM1 splice variants was strong in the ciliary body, whereas TRPM3 was strongly expressed in the retinal pigmented epithelium.
The cephalopod retina contains two kinds of photopigments, rhodopsin and retinochrome. For many years retinochrome has been thought to be localized in the inner segments of the visual cells, whereas rhodopsin is in the outer segments. However, it is now clear that retinochrome can be extracted also from fragments of outer segments. In the dark- adapted retina of Loligo pealei retinochrome is distributed half-and- half in the inner and outer segments. Todarodes pacificus contains much more retinochrome than Loligo, and it is more abundant in the outer than in the inner segments. The outer segments of Loligo contain retinochrome and metarhodopsin in addition to rhodopsin, whether squids are kept in the dark or in the light. But there is extremely little metarhodopsin (about 3% of rhodopsin) even in light-adapted eyes. The inner segments contain only retinochrome, and much less in the light than in the dark. On the other hand, retinochrome in the outer segments increases markedly during light adaptation. These facts suggest the possibility that some retinochrome moves forward from the inner to the outer segments during light adaptation and there reacts with metarhodopsin to promote regeneration of rhodopsin. PMID:6620
Zhai, Yi-Ran; Zhao, Yong; Zhong, Jie; Li, Ke; Lu, Cui-Xin; Zhang, Bing
A new sphere-mapping algorithm called sector mapping is introduced to map sector images to the sphere of an eyeball. The proposed sector-mapping algorithm is evaluated and compared with the plane-mapping algorithm adopted in previous work. A simulation that maps an image of concentric circles to the sphere of the eyeball and an analysis of the difference in distance between neighboring points in a plane and sector were used to compare the two mapping algorithms. A three-dimensional model of a whole retina with clear retinal detachment was generated using the Visualization Toolkit software. A comparison of the mapping results shows that the central part of the retina near the optic disc is stretched and its edges are compressed when the plane-mapping algorithm is used. A better mapping result is obtained by the sector-mapping algorithm than by the plane-mapping algorithm in both the simulation results and real clinical retinal detachment three-dimensional reconstruction.
Wong, D.; Lois, N.
AIM—To describe a new surgical technique for foveal relocation, and to report the outcome in nine patients treated with this procedure. METHODS—Nine consecutive patients with subfoveal choroidal neovascular membranes (CNVMs) secondary to age related macular degeneration underwent foveal relocation surgery by redistribution of the neurosensory retina (RNR). The technique involved induction of a retinal detachment via a single retinotomy, relocation of the fovea by "sweeping" the retinal tissue with a retinal brush, and stabilisation of the retina in its new location using perfluorocarbon liquid peroperatively and silicone oil postoperatively. RESULTS—In eight of nine eyes successful relocation of the fovea was achieved; in one eye the CNVM remained in a subfoveal location postoperatively. Visual acuity improved in two eyes, remained unchanged in three, and decreased in four eyes after a median follow up of 4 months (range 2.5-6 months). Complications included rupture of a foveal cyst with the development of a macular hole in one eye and epimacular membrane formation in another eye. In two eyes, macular retinal vessel closure occurred at the time of laser photocoagulation; one of these eyes later developed cystoid macular oedema and the other an epiretinal membrane. Recurrence of the CNVM was observed in one eye, but was controlled with further laser treatment. CONCLUSIONS—Foveal relocation by RNR appears to be feasible, obviating the need for extensive retinotomies or scleral shortening. PMID:10729290
de Wit, G C
A retinal scanning display is a new kind of display that directly uses the retina as a projection screen. This differs from, for example, a TV which first creates an image on a screen outside the eye. Although a retinal scanning display needs no screen, the principle of creating an image is similar to that of a TV. Where the TV uses an electron beam to create (scan) a raster pattern on a screen, a retinal scanning display uses a beam of light to scan a raster pattern on the retina. The way to get from an equally illuminated raster pattern to an image is to modulate the intensity of the beam as it scans. Since the eye does not look at a physical screen, people often wonder what the image of a retinal scanning display will look like. Upon seeing the image, the general response is: 'It looks just like a normal display'. In fact it is a normal display, but one that has many advantages compared to other kind of displays, such as: possibility of high brightness, large color gamut, possibility of high-resolution and good image quality.
Biswas, Sonia; Haselier, Christine; Mataruga, Anja; Thumann, Gabriele; Walter, Peter; Müller, Frank
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
Wu, S M; Maple, B R
Physiological and pharmacological mechanisms of glutamatergic, GABAergic and glycinergic synapses in the tiger salamander retina were studied. We used immunocytochemical and autoradiographic methods to study localizations of these neurotransmitters and their uptake transporters; and electrophysiological methods (intracellular, extracellular and whole cell patch electrode recordings) to study the light responses, miniature postsynaptic currents and neurotransmitter-induced postsynaptic currents in various retinal neurons. Our results are consistent with the following scheme: Glutamate is used by the photoreceptor and bipolar cell output synapses and the release of glutamate is largely mediated by calcium-dependent vesicular processes. The postsynaptic glutamate receptors in DBCs are L-AP4 receptors, in HBCs, HCs and ganglion cells are the kainate/AMPA and NMDA receptors. Subpopulations of HCs make GABAergic synapses on cones and gate chloride condunctance through GABAA receptors. GABAergic HCs do not make feedforward synapses on bipolar cell dendrites and the neurotransmitter identity of the HCs making feedforward synapses is unknown. Subpopulations of amacrine cells make GABAergic synapses on bipolar cell synaptic terminals, other amacrine cells and ganglion cells and GABA gates chloride conductances in theses cells. Glycinergic amacrine cells make synapses on bipolar cell synaptic terminals, other amacrine cells and ganglion cells and glycine opens postsynaptic chloride channels. Glycinergic interplexiform cells make synapses on bipolar cells in the outer retina and glycine released from these cells open chloride channels in bipolar cell dendrites.
Gellermann, Werner; Ermakov, Igor V.; McClane, Robert W.; Bernstein, Paul S.
We have used resonance Raman scattering as a novel, non- invasive, in-vivo optical technique to measure the concentration of carotenoid pigment in the human retina. Using argon laser excitation we are able to measure two strong carotenoid resonance Raman signals at 1159 and 1525 wave numbers, respectively. The required laser power levels are within the limits given by safety standards for ocular exposure. Of the approximately ten carotenoid pigment found in normal human serum, the species lutein and zeaxanthin are concentrated in high amounts in the cells of the human macula, which is an approximately 5 mm diameter area of the retina in which the visual acuity is highest. These carotenoids give the macula a characteristic yellow coloration, and it is speculated that these molecules function as filter to attenuate photochemical damage and/or image degradation under bright UV/blue light exposures. In addition, they are thought to act as free-radical scavenging antioxidants. Studies have shown that there may be a link between macular degenerative diseases, the leading cause of blindness in the elderly in the US, and the presence or absence of the carotenoids. We describe an instrument capable of measuring the macular carotenoids in human subjects in a non-invasive, rapid and quantitative way.
Zhao, Lin; Feng, Zhihui; Zou, Xuan; Cao, Ke; Xu, Jie; Liu, Jiankang
Retina is particularly susceptible to aging as oxidative damage accumulates within retina, leading to age-related retinal dysfunction or even visual loss. However, the underlying mechanisms still remain obscure and effective therapeutic strategy is urgently in need. Here, we quested for the answer particularly focusing on mitochondrial homeostasis and O-GlcNAcylation in rat retina. By comparing expression of electron transfer chain complexes and key factors in mitochondrial biogenesis and dynamics in retinas of aged and young Sprague-Dawley rats, we found that mitochondrial Complex I, II, IV and V were increased in aged retina with decreased mtTFA and Mfn2. Also, we noticed that p38 and JNK of MAPK signaling were substantially more activated in aged retina, suggesting stress induction. In addition, we found that pan-O-GlcNAcylation was remarkably stronger with lower OGA expression in aged retina. To further elucidate the roles of Mfn2 and O-GlcNAcylation, we employed ARPE-19 cells and found that ATP production, oxygen consumption, and mitochondrial membrane potential were reduced and ROS level was increased by Mfn2 knockdown, while treating with PUGNAc or UDP-GlcNAc heightened oxygen consumption and reduced ROS. Our results suggest disrupted mitochondrial homeostasis may increase oxidative stress; yet enhanced O-GlcNAcylation might defend against oxidative stress and promote mitochondrial respiration in aged retina.
Volland, Stefanie; Esteve-Rudd, Julian; Hoo, Juyea; Yee, Claudine; Williams, David S
Mouse models have greatly assisted our understanding of retinal degenerations. However, the mouse retina does not have a macula, leading to the question of whether the mouse is a relevant model for macular degeneration. In the present study, a quantitative comparison between the organization of the central mouse retina and the human macula was made, focusing on some structural characteristics that have been suggested to be important in predisposing the macula to stresses leading to degeneration: photoreceptor density, phagocytic load on the RPE, and the relative thinness of Bruch's membrane. Light and electron microscopy measurements from retinas of two strains of mice, together with published data on human retinas, were used for calculations and subsequent comparisons. As in the human retina, the central region of the mouse retina possesses a higher photoreceptor cell density and a thinner Bruch's membrane than in the periphery; however, the magnitudes of these periphery to center gradients are larger in the human. Of potentially greater relevance is the actual photoreceptor cell density, which is much greater in the mouse central retina than in the human macula, underlying a higher phagocytic load for the mouse RPE. Moreover, at eccentricities that correspond to the peripheral half of the human macula, the rod to cone ratio is similar between mouse and human. Hence, with respect to photoreceptor density and phagocytic load of the RPE, the central mouse retina models at least the more peripheral part of the macula, where macular degeneration is often first evident.
Qian, Xiuqing; Zhang, Kunya; Liu, Zhicheng
A method is proposed to determine the mechanical properties of retina based on in vivo experiments and numerical simulations. First, saline water was injected into the anterior chamber of the right eye of a cat to cause acute high intraocular pressure. After the eye was scanned using optical coherence tomography under different acute high intraocular pressures, the images of the retina in vivo were obtained and the thickness of the retina was calculated. Then, the three-dimensional structure of the optic nerve head including the retina and the choroid were reconstructed using image processing technology. Three different material models for the retina and the choroid were taken and the finite element models of the optic nerve head were constructed. Finally, an inverse method was proposed to determine the parameters of a constitutive model of the retina and of the choroid simultaneously. The results showed that the deformation of the retina can be properly simulated taking into consideration the nonlinear elastic properties of the retina and of the choroid.
Hoo, Juyea; Yee, Claudine; Williams, David S.
Mouse models have greatly assisted our understanding of retinal degenerations. However, the mouse retina does not have a macula, leading to the question of whether the mouse is a relevant model for macular degeneration. In the present study, a quantitative comparison between the organization of the central mouse retina and the human macula was made, focusing on some structural characteristics that have been suggested to be important in predisposing the macula to stresses leading to degeneration: photoreceptor density, phagocytic load on the RPE, and the relative thinness of Bruch’s membrane. Light and electron microscopy measurements from retinas of two strains of mice, together with published data on human retinas, were used for calculations and subsequent comparisons. As in the human retina, the central region of the mouse retina possesses a higher photoreceptor cell density and a thinner Bruch’s membrane than in the periphery; however, the magnitudes of these periphery to center gradients are larger in the human. Of potentially greater relevance is the actual photoreceptor cell density, which is much greater in the mouse central retina than in the human macula, underlying a higher phagocytic load for the mouse RPE. Moreover, at eccentricities that correspond to the peripheral half of the human macula, the rod to cone ratio is similar between mouse and human. Hence, with respect to photoreceptor density and phagocytic load of the RPE, the central mouse retina models at least the more peripheral part of the macula, where macular degeneration is often first evident. PMID:25923208
Teeters, Jeffrey L.; Werblin, Frank S.
The retina computes to let us see, but can we see the retina compute? Until now, the answer has been no, because the unconscious nature of the processing hides it from our view. Here the authors describe a method of seeing computations performed throughout the retina. This is achieved by using neurophysiological data to construct a model of the retina, and using a special-purpose image processing computer (PIPE) to implement the model in real time. Processing in the model is organized into stages corresponding to computations performed by each retinal cell type. The final stage is the transient (change detecting) ganglion cell. A CCD camera forms the input image, and the activity of a selected retinal cell type is the output which is displayed on a TV monitor. By changing the retina cell driving the monitor, the progressive transformations of the image by the retina can be observed. These simulations demonstrate the ubiquitous presence of temporal and spatial variations in the patterns of activity generated by the retina which are fed into the brain. The dynamical aspects make these patterns very different from those generated by the common DOG (Difference of Gaussian) model of receptive field. Because the retina is so successful in biological vision systems, the processing described here may be useful in machine vision.
Olivier, S S; Jones, S M; Chen, D C; Zawadzki, R J; Choi, S S; Laut, S P; Werner, J S
Optical coherence tomography (OCT) sees the human retina sharply with adaptive optics. In vivo cellular visualization of the human retina at micrometer-scale resolution is possible by enhancing Fourier-domain optical-coherence tomography with adaptive optics, which compensate for the eye's optical aberrations.
PHOTOSENSITIVITY(BIOLOGICAL), *CANCER, * AMINO ACIDS , RETINA, PHOTOCHEMICAL REACTIONS, RETINA, PATHOLOGY, RESPIRATION, GLYCOLYSIS, FISHES, ELECTROPHYSIOLOGY, FORMALDEHYDE, CATALYSTS, LIGHT, STIMULATION(PHYSIOLOGY), CHEMORECEPTORS, OXYGEN, ULTRAVIOLET RADIATION, MICE, ITALY.
This review summarizes the current knowledge about the ontogenetic development of the circadian system in mammals. The developmental changes of overt rhythms are discussed, although the main focus of the review is the underlying neuronal and molecular mechanisms. In addition, the review describes ontogenetic development, not only as a process of morpho-functional maturation. The need of repeated adaptations and readaptations due to changing developmental stage and environmental conditions is also considered. The review analyzes mainly rodent data, obtained from the literature and from the author's own studies. Results from other species, including humans, are presented to demonstrate common features and species-dependent differences. The review first describes the development of the suprachiasmatic nuclei as the central pacemaker system and shows that intrinsic circadian rhythms are already generated in the mammalian fetus. As in adult organisms, the period length is different from 24 h and needs continuous correction by environmental periodicities, or zeitgebers. The investigation of the ontogenetic development of the mechanisms of entrainment reveals that, at prenatal and early postnatal stages, non-photic cues deriving from the mother are effective. Light-dark entrainment develops later. At a certain age, both photic and non-photic zeitgebers may act in parallel, even though the respective time information is 12 h out of phase. That leads to a temporary internal desynchronization. Because rhythmic information needs to be transferred to effector organs, the corresponding neural and humoral signalling pathways are also briefly described. Finally, to be able to transform a rhythmic signal into an overt rhythm, the corresponding effector organs must be functionally mature. As many of these organs are able to generate their own intrinsic rhythms, another aspect of the review is dedicated to the development of peripheral oscillators and mechanisms of their entrainment
Cringle, Stephen J; Yu, Dao-Yi
A disrupted oxygen environment in the retina of severely premature neonates is thought to be a key factor in the development of retinopathy of prematurity (ROP). This review describes our understanding of intraretinal oxygen distribution and consumption in a range of animal models, including species with naturally avascular retinas and models of induced occlusion of the retinal vasculature. The influence of graded systemic hyperoxia on retinal oxygenation is also discussed along with modulation of retinal oxygen metabolism. The differences in retinal oxygenation between developing and mature retinas are also described. Comparisons are made with studies in the monkey retina in order to assess possible similarities in behaviour between rat and human retinas. Pathogenesis mechanism and possible intervention strategies during the diseased processes in ROP are proposed based on our current knowledge.
Boshuo Wang; Weiland, James D
Electrical impedance of the retina is a critical factor in retinal prostheses, determining the intraretinal current flow and potential distribution of electrical stimulation. Previous resistivity measurements in retina were limited to healthy retina, and didn't include mouse models, a common and important animal model in retinal research. This experimental study measured the resistivity profiles of wild-type, rd1, and rd10 mice, providing basis for computational simulations and predictive modeling studies. The peak resistance frequency method has been utilized to measure the resistivity profiles of the retina cross section, and the results show agreement with previous studies in retina of normal rats and embryonic chicks. Retinal degeneration affects the width of the profile, which is in agreement with histological measurements. Degeneration also results in lower peak resistivity. The results indicate that, on the mesoscopic scale, resistivity is dominated by spatial factors, while influence of remodeling on the cellular level is not apparent under such scale.
Lam, Dominic M. K.
Goldfish retinas incubated with L-glutamate-14C (UL) were found to synthesize γ-aminobutyric acid-14C (GABA-14C) The accumulation of newly synthesized GABA was enhanced by physiological stimulation of the retina with flashing light; and this increase was directly proportional to the logarithm of the light intensity. The total GABA content was also higher in light-stimulated than in dark-adapted retinas, although the glutamate content remained unchanged No differences were found in the cell-free activities of glutamate decarboxylase (EC 4 1.1 15) and GABA-glutamate transaminase (EC 220.127.116.11) extracted from light-stimulated and dark-adapted retinas. These findings, together with other physiological and morphologcal evidence, suggest that GABA plays a functional role in synaptic transmission in the goldfish retina PMID:4339278
Gonzalez‐Cordero, Anai; West, Emma L.; Han, Ya‐Ting; Welby, Emily; Naeem, Arifa; Blackford, Samuel J. I.; Bainbridge, James W. B.; Pearson, Rachael A.; Ali, Robin R.
Abstract Loss of photoreceptors due to retinal degeneration is a major cause of untreatable blindness. Cell replacement therapy, using pluripotent stem cell‐derived photoreceptor cells, may be a feasible future treatment. Achieving safe and effective cell replacement is critically dependent on the stringent selection and purification of optimal cells for transplantation. Previously, we demonstrated effective transplantation of post‐mitotic photoreceptor precursor cells labelled by fluorescent reporter genes. As genetically labelled cells are not desirable for therapy, here we developed a surface biomarker cell selection strategy for application to complex pluripotent stem cell differentiation cultures. We show that a five cell surface biomarker panel CD73(+)CD24(+)CD133(+)CD47(+)CD15(−) facilitates the isolation of photoreceptor precursors from three‐dimensional self‐forming retina differentiated from mouse embryonic stem cells. Importantly, stem cell‐derived cells isolated using the biomarker panel successfully integrate and mature into new rod photoreceptors in the adult mouse retinae after subretinal transplantation. Conversely, unsorted or negatively selected cells do not give rise to newly integrated rods after transplantation. The biomarker panel also removes detrimental proliferating cells prior to transplantation. Notably, we demonstrate how expression of the biomarker panel is conserved in the human retina and propose that a similar selection strategy will facilitate isolation of human transplantation‐competent cells for therapeutic application. Stem Cells 2015;33:2469—2482 PMID:25982268
Lakowski, Jorn; Gonzalez-Cordero, Anai; West, Emma L; Han, Ya-Ting; Welby, Emily; Naeem, Arifa; Blackford, Samuel J I; Bainbridge, James W B; Pearson, Rachael A; Ali, Robin R; Sowden, Jane C
Loss of photoreceptors due to retinal degeneration is a major cause of untreatable blindness. Cell replacement therapy, using pluripotent stem cell-derived photoreceptor cells, may be a feasible future treatment. Achieving safe and effective cell replacement is critically dependent on the stringent selection and purification of optimal cells for transplantation. Previously, we demonstrated effective transplantation of post-mitotic photoreceptor precursor cells labelled by fluorescent reporter genes. As genetically labelled cells are not desirable for therapy, here we developed a surface biomarker cell selection strategy for application to complex pluripotent stem cell differentiation cultures. We show that a five cell surface biomarker panel CD73(+)CD24(+)CD133(+)CD47(+)CD15(-) facilitates the isolation of photoreceptor precursors from three-dimensional self-forming retina differentiated from mouse embryonic stem cells. Importantly, stem cell-derived cells isolated using the biomarker panel successfully integrate and mature into new rod photoreceptors in the adult mouse retinae after subretinal transplantation. Conversely, unsorted or negatively selected cells do not give rise to newly integrated rods after transplantation. The biomarker panel also removes detrimental proliferating cells prior to transplantation. Notably, we demonstrate how expression of the biomarker panel is conserved in the human retina and propose that a similar selection strategy will facilitate isolation of human transplantation-competent cells for therapeutic application.
Mangiamele, Lisa A; Gomez, Julia R; Curtis, Nancy J; Thompson, Richmond R
Estradiol rapidly (within 30 minutes) influences a variety of sociosexual behaviors in both mammalian and nonmammalian vertebrates, including goldfish, in which it rapidly stimulates approach responses to the visual cues of females. Such rapid neuromodulatory effects are likely mediated via membrane-associated estrogen receptors; however, the localization and distribution of such receptors within the nervous system is not well described. To begin to address this gap, we identified GPER/GPR30, a G-protein-coupled estrogen receptor, in goldfish (Carassius auratus) neural tissue and used reverse-transcription polymerase chain reaction (RT-PCR) and in situ hybridization to test if GPR30 is expressed in the brain regions that might mediate visually guided social behaviors in males. We then used immunohistochemistry to determine whether GPR30 colocalizes with isotocin-producing cells in the preoptic area, a critical node in the highly conserved vertebrate social behavior network. We used quantitative (q)PCR to test whether GPR30 mRNA levels differ in males in breeding vs. nonbreeding condition and in males that were socially interacting with a female vs. a rival male. Our results show that GPR30 is expressed in the retina and in many brain regions that receive input from the retina and/or optic tectum, as well as in a few nodes in the social behavior network, including cell populations that produce isotocin. J. Comp. Neurol. 525:252-270, 2017. © 2016 Wiley Periodicals, Inc.
Davenport, Christopher M.; Detwiler, Peter B.; Dacey, Dennis M.
Negative feedback from horizontal cells to cone photoreceptors is regarded as the critical pathway for the formation of the antagonistic surround of retinal neurons, yet the mechanism by which horizontal cells accomplish negative feedback has been difficult to determine. Recent evidence suggests that feedback uses a novel, non-GABAergic pathway that directly modulates the calcium current in cones. In non-mammalian vertebrates, enrichment of retinal pH buffering capacity attenuates horizontal cell feedback, supporting one model in which feedback occurs by horizontal cell modulation of the extracellular pH in the cone synaptic cleft. Here we test the effect of exogenous pH buffering on the response dynamics of H1 horizontal cells and the center-surround receptive field structure of parasol ganglion cells in the macaque monkey retina. Enrichment of the extracellular buffering capacity with HEPES selectively attenuates surround antagonism in parasol ganglion cells. The H1 horizontal cell light response includes a slow, depolarizing component that is attributed to negative feedback to cones. This part of the response is attenuated by HEPES and other pH buffers in a dose-dependent manner that is correlated with predicted buffering capacity. The selective effects of pH buffering on the parasol cell surround and H1 cell light response suggests that, in primate retina, horizontal cell feedback to cones is mediated via a pH-dependent mechanism and is a major determinant of the ganglion cell receptive field surround. PMID:18184788
Uludag, H; De Vos, P; Tresco, P A
Entrapment of mammalian cells in physical membranes has been practiced since the early 1950s when it was originally introduced as a basic research tool. The method has since been developed based on the promise of its therapeutic usefulness in tissue transplantation. Encapsulation physically isolates a cell mass from an outside environment and aims to maintain normal cellular physiology within a desired permeability barrier. Numerous encapsulation techniques have been developed over the years. These techniques are generally classified as microencapsulation (involving small spherical vehicles and conformally coated tissues) and macroencapsulation (involving larger flat-sheet and hollow-fiber membranes). This review is intended to summarize techniques of cell encapsulation as well as methods for evaluating the performance of encapsulated cells. The techniques reviewed include microencapsulation with polyelectrolyte complexation emphasizing alginate-polylysine capsules, thermoreversible gelation with agarose as a prototype system, interfacial precipitation and interfacial polymerization, as well as the technology of flat sheet and hollow fiber-based macroencapsulation. Four aspects of encapsulated cells that are critical for the success of the technology, namely the capsule permeability, mechanical properties, immune protection and biocompatibility, have been singled out and methods to evaluate these properties were summarized. Finally, speculations regarding future directions of cell encapsulation research and device development are included from the authors' perspective.
Many mammalian species use chemosignals to coordinate reproduction by altering the physiology and behavior of both sexes. Chemosignals prime reproductive physiology so that individuals become sexually mature and active at times when mating is most probable and suppress it when it is not. Once in reproductive condition, odors produced and deposited by both males and females are used to find and select individuals for mating. The production, dissemination and appropriate responses to these cues are modulated heavily by organizational and activational effects of gonadal sex steroids and thereby intrinsically link chemical communication to the broader reproductive context. Many compounds have been identified as "pheromones" but very few have met the expectations of that term: a unitary, species-typical substance that is both necessary and sufficient for an experience-independent behavioral or physiological response. In contrast, most responses to chemosignals are dependent or heavily modulated by experience, either in adulthood or during development. Mechanistically, chemosignals are perceived by both main and accessory (vomeronasal) olfactory systems with the importance of each system tied strongly to the nature of the stimulus rather than to the response. In the central nervous system, the vast majority of responses to chemosignals are mediated by cortical and medial amygdala connections with hypothalamic and other forebrain structures. Despite the importance of chemosignals in mammals, many details of chemical communication differ even among closely related species and defy clear categorization. Although generating much research and public interest, strong evidence for the existence of a robust chemical communication among humans is lacking.
Burman, Chloe; Ktistakis, Nicholas T
Autophagy is a fundamental intracellular trafficking pathway conserved from yeast to mammals. It is generally thought to play a pro-survival role, and it can be up regulated in response to both external and intracellular factors, including amino acid starvation, growth factor withdrawal, low cellular energy levels, endoplasmic reticulum (ER) stress, hypoxia, oxidative stress, pathogen infection, and organelle damage. During autophagy initiation a portion of the cytosol is surrounded by a flat membrane sheet known as the isolation membrane or phagophore. The isolation membrane then elongates and seals itself to form an autophagosome. The autophagosome fuses with normal endocytic traffic to mature into a late autophagosome, before fusing with lysosomes. The molecular machinery that enables formation of an autophagosome in response to the various autophagy stimuli is almost completely identified in yeast and-thanks to the observed conservation-is also being rapidly elucidated in higher eukaryotes including mammals. What are less clear and currently under intense investigation are the mechanism by which these various autophagy components co-ordinate in order to generate autophagosomes. In this review, we will discuss briefly the fundamental importance of autophagy in various pathophysiological states and we will then review in detail the various players in early autophagy. Our main thesis will be that a conserved group of heteromeric protein complexes and a relatively simple signalling lipid are responsible for the formation of autophagosomes in mammalian cells.
Ris, H; Witt, P L
The structure of the mammalian trilaminar kinetochore was investigated using stereo electron microscopy of chromosomes in hypotonic solutions which unraveled the chromosome but maintained microtubules. Mouse and Chinese hamster ovary cells were arrested in Colcemid and allowed to reform microtubules after Colcemid was removed. Recovered cells were then swelled, lysed or spread in hypotonic solutions which contained D2O to preserve microtubules. The chromosomes were observed in thin and thick sections and as whole mounts using high voltage electron microscopy. Bundles of microtubules were seen directly attached to chromatin, indicating that the kinetochore outer layer represents a differential arrangement of chromatin, continuous with the body of the chromosome. In cells fixed wihout pretreatment, the outer layer could be seen to be composed of hairpin loops of chromatin stacked together to form a solid layer. The hypotonically-induced unraveling of the outer layer was found to be reversible, and the typical 300 nm thick disk reformed when cells were returned to isotonic solutions. Short microtubules, newly nucleated after Colcemid removal, were found not to be attached to the kinetochore out layer, but were situated in the fibrous corona on the external surface of the outer layer. This was verified by observation of thick sections in stereo which made it possible to identify microtubules ends within the section. Thus, kinetochore microtubules are nucleated within the fibrous corona, and subsequently become attached to the outer layer.
Gmünder, Felix K.; Suter, Robert N.; Kiess, M.; Urfer, R.; Nordau, C.-G.; Cogoli, A.
Equipment used in space for the cultivation of mammalian cells does not meet the usual standard of earth bound bioreactors. Thus, the development of a space worthy bioreactor is mandatory for two reasons: First, to investigate the effect on single cells of the space environment in general and microgravity conditions in particular, and second, to provide researchers on long term missions and the Space Station with cell material. However, expertise for this venture is not at hand. A small and simple device for animal cell culture experiments aboard Spacelab (Dynamic Cell Culture System; DCCS) was developed. It provides 2 cell culture chambers, one is operated as a batch system, the other one as a perfusion system. The cell chambers have a volume of 200 μl. Medium exchange is achieved with an automatic osmotic pump. The system is neither mechanically stirred nor equipped with sensors. Oxygen for cell growth is provided by a gas chamber that is adjacent to the cell chambers. The oxygen gradient produced by the growing cells serves to maintain the oxygen influx by diffusion. Hamster kidney cells growing on microcarriers were used to test the biological performance of the DCCS. On ground tests suggest that this system is feasible.
Simoens, P; Ghoshal, N G
The arterial supply of the optic nerve and the retina of four fetal and ten adult sheep was studied by various injection techniques and verified by histological examination in one neonatal lamb. The optic nerve was supplied by the rostral cerebral artery, including the rostral hypophysial artery, the internal ophthalmic artery and its chiasmatic rete, and recurrent branches from the choroidoretinal arteries. The retinal arterioles were supplied by the choroidoretinal arteries, which coursed rostrally in the external sheath of the optic nerve and perforated the area cribrosa of the sclera. The dorsal choroidoretinal artery arose variably from the external ophthalmic artery and its terminal branches. It supplied the rostral portion of the optic nerve, the choroid and, except in one sheep, all retinal arterioles. The ventral choroidoretinal artery arose variably from the anastomotic branch between the external and internal ophthalmic arteries. It supplied the rostral aspect of the optic nerve and the choroid and continued in the fetal lambs as the hyaloid artery. In one specimen it also supplied the lateral retinal arterioles. There was no evidence for a central retinal artery and a continuous arterial circle of the optic nerve. Images Fig. 6 Fig. 7 Fig. 8 PMID:7333956
Park, Silvia J.H.; Borghuis, Bart G.; Rahmani, Pouyan; Zeng, Qiang
Visual processing in the retina depends on coordinated signaling by interneurons. Photoreceptor signals are relayed to ∼20 ganglion cell types through a dozen excitatory bipolar interneurons, each responsive to light increments (ON) or decrements (OFF). ON and OFF bipolar cell pathways become tuned through specific connections with inhibitory interneurons: horizontal and amacrine cells. A major obstacle for understanding retinal circuitry is the unknown function of most of the ∼30–40 amacrine cell types, each of which synapses onto a subset of bipolar cell terminals, ganglion cell dendrites, and other amacrine cells. Here, we used a transgenic mouse line in which vasoactive intestinal polypeptide-expressing (VIP+) GABAergic interneurons express Cre recombinase. Targeted whole-cell recordings of fluorescently labeled VIP+ cells revealed three predominant types: wide-field bistratified and narrow-field monostratified cells with somas in the inner nuclear layer (INL) and medium-field monostratified cells with somas in the ganglion cell layer (GCL). Bistratified INL cells integrated excitation and inhibition driven by both ON and OFF pathways with little spatial tuning. Narrow-field INL cells integrated excitation driven by the ON pathway and inhibition driven by both pathways, with pronounced hyperpolarizations at light offset. Monostratified GCL cells integrated excitation and inhibition driven by the ON pathway and showed center-surround spatial tuning. Optogenetic experiments showed that, collectively, VIP+ cells made strong connections with OFF δ, ON-OFF direction-selective, and W3 ganglion cells but weak, inconsistent connections with ON and OFF α cells. Revealing VIP+ cell morphologies, receptive fields and synaptic connections advances our understanding of their role in visual processing. SIGNIFICANCE STATEMENT The retina is a model system for understanding nervous system function. At the first stage, rod and cone photoreceptors encode light and
Guttmann, T; Vítek, A; Pivec, L
High resolution melting profiles of different mammalian DNAs are presented. Melting curves of various mammalian DNAs were compared with respect to the degree of asymmetry, first moment, transition breath and Tmi of individual subtransitions. Quantitative comparison of the shape of all melting curves was made. Correlation between phylogenetical relations among mammals and shape of the melting profiles of their DNAs was demonstrated. The difference between multi-component heterogeneity of mammalian DNAs found by optical melting analysis and sedimentation in CsCl-netropsin density gradient is also discussed. PMID:840642
Kaiya, Hiroyuki; Kangawa, Kenji; Miyazato, Mikiya
The growth hormone secretagogue-receptor (GHS-R) was discovered in humans and pigs in 1996. The endogenous ligand, ghrelin, was discovered 3 years later, in 1999, and our understanding of the physiological significance of the ghrelin system in vertebrates has grown steadily since then. Although the ghrelin system in non-mammalian vertebrates is a subject of great interest, protein sequence data for the receptor in non-mammalian vertebrates has been limited until recently, and related biological information has not been well organized. In this review, we summarize current information related to the ghrelin receptor in non-mammalian vertebrates. PMID:23882259
Taylor, Scott M; Loew, Ellis R; Grace, Michael S
The Atlantic tarpon, Megalops atlanticus, is a large piscivorous fish that supports economically important recreational fisheries in the Gulf of Mexico, Caribbean, and Florida Atlantic coast. Megalops atlanticus undergoes ontogenetic shifts in morphology, hatching in the open ocean as larvae (less than 1 cm in length), moving into hypoxic turbid mangrove marshes as juveniles (around 10 cm in length), and then moving into coastal oceanic waters as adults (over 100 cm in length). In this study, photoreceptor distributions, opsin distributions, and photoreceptor absorbance characteristics were studied with light microscopy, transmission electron microscopy, antiopsin immunofluorescence, and microspectrophotometry, respectively, at four ecologically distinct life-history stages--premetamorphic larva, settlement stage, juvenile, and adult. The purposes of this study were 1) to determine the extent to which the retina of M. atlanticus changes over the course of development and 2) to relate these retinal changes with ecological shifts between developmental stages. The new data presented here indicate that the M. atlanticus retina changes substantially in rod and cone distributions and absorbance characteristics over the course of development and that these changes correlate closely with those in habitat and behavior. We show that M. atlanticus has a rod-dominated retina at the larval stage (which is unusual for teleost larvae) and that the scotopic visual system becomes far better developed with maturity, adding a substantial tapetum and high densities of small, bundled, and stacked rod cells. We also show that there are shifts in cone and rod spectral sensitivities and an increase in the diversity of spectrally distinct cone classes, including the addition of ultraviolet cones as fish mature into adults.
Gellermann, Werner; Ermakov, Igor V.; Ermakova, Maia R.; McClane, Robert W.; Zhao, Da-You; Bernstein, Paul S.
We have used resonant Raman scattering spectroscopy as a novel, noninvasive, in vivo optical technique to measure the concentration of the macular carotenoid pigments lutein and zeaxanthin in the living human retina of young and elderly adults. Using a backscattering geometry and resonant molecular excitation in the visible wavelength range, we measure the Raman signals originating from the single- and double-bond stretch vibrations of the π-conjugated molecule's carbon backbone. The Raman signals scale linearly with carotenoid content, and the required laser excitation is well below safety limits for macular exposure. Furthermore, the signals decline significantly with increasing age in normal eyes. The Raman technique is objective and quantitative and may lead to a new method for rapid screening of carotenoid pigment levels in large populations at risk for vision loss from age-related macular degeneration, the leading cause of blindness in the elderly in the United States.
Rodríguez-Sánchez, Iràm Pablo; Garza-Rodríguez, Maria Lourdes; Mohamed-Noriega, Karim; Voruganti, Venkata Saroja; Tejero, Maria Elizabeth; Delgado-Enciso, Ivan; Ibave, Diana Cristina Perez; Schlabritz-Loutsevitch, Natalia E.; Mohamed-Noriega, Jibran; Martinez-Fierro, Margarita L; Reséndez-Pérez, Diana; Cole, Shelley A; Cavazos-Adame, Humberto; Comuzzie, Anthony G.; Mohamed-Hamsho, Jesús; Barrera-Saldaña, Hugo Alberto
Background Olfactomedin-like is a polyfunctional polymeric glycoprotein. This family has at least four members. One member of this family is OLFML3, which is preferentially expressed in placenta but is also detected in other adult tissues including the liver and heart. However, the orthologous rat gene is expressed in the iris, sclera, trabecular meshwork, retina, and optic nerve. Methods OLFML3 amplification was performed by RT-PCR from human and baboon ocular tissues. The products were cloned and sequenced. Results We report OFML3 expression in human and baboon eye. The full CDS has 1221 bp, from which a OFR of 406 amino acid was obtained. The baboon OLFML3 gene nucleotidic sequence has 98%, and amino acidic 99% similarity with humans. Conclusions OLFML3 expression in human and baboon ocular tissues and its high similarity make the baboon a powerful model to deduce the physiological and/or metabolic function of this protein in the eye. PMID:23398349
Abdallah, Samer S; Ramahi, Omar; Bizheva, Kostadinka
Finite Difference Time Domain (FDTD) method was developed to model changes in the light scattering properties of retinal photoreceptors resulting from the functional response of living retina to external light stimulation. Physiological processes such as membrane hyper-polarization and conformation changes of rhodopsin in the photoreceptors outer segment (OS) were simulated by varying the optical properties of the cell organelles. The FDTD code was validated by comparing the results from a 2D simulation of light scattering from an infinite cylinder to the Mie analytical solution for the same geometry. Results from the FDTD simulations show that hyper-polarization of the outer cell membrane is the least likely cause for the observed increase in light scattering in photoreceptors. Other computational data suggests that the experimentally observed changes in reflectivity are most likely related to cell dynamics and to cell volume changes.
Barranco, Francisco; Díaz, Javier; Ros, Eduardo; del Pino, Begoña
We present a bioinspired model for detecting spatiotemporal features based on artificial retina response models. Event-driven processing is implemented using four kinds of cells encoding image contrast and temporal information. We have evaluated how the accuracy of motion processing depends on local contrast by using a multiscale and rank-order coding scheme to select the most important cues from retinal inputs. We have also developed some alternatives by integrating temporal feature results and obtained a new improved bioinspired matching algorithm with high stability, low error and low cost. Finally, we define a dynamic and versatile multimodal attention operator with which the system is driven to focus on different target features such as motion, colors, and textures.
Finlay, Barbara L
Evolutionary and other functional accounts of the retina and its normal development highlight different aspects of control of its growth and form than genomic and mechanistic accounts. Discussing examples from opsin expression, developmental regulation of the eye's size and optical quality, regulation of eye size with respect to brain and body size, and the development of the fovea, these different aspects of control are contrasted. Contributions of mouse models, particularly with regard to relative timing of events in different species are reviewed, introducing a Web-based utility for exploration of timing issues (www.translatingtime.net). Variation at the individual level, in early experience, and also across species is an essential source of information to understand normal development and its pathologies.
Conway, Bevil R; Chatterjee, Soumya; Field, Greg D; Horwitz, Gregory D; Johnson, Elizabeth N; Koida, Kowa; Mancuso, Katherine
Color has become a premier model system for understanding how information is processed by neural circuits, and for investigating the relationships among genes, neural circuits, and perception. Both the physical stimulus for color and the perceptual output experienced as color are quite well characterized, but the neural mechanisms that underlie the transformation from stimulus to perception are incompletely understood. The past several years have seen important scientific and technical advances that are changing our understanding of these mechanisms. Here, and in the accompanying minisymposium, we review the latest findings and hypotheses regarding color computations in the retina, primary visual cortex, and higher-order visual areas, focusing on non-human primates, a model of human color vision.
Allaman-Pillet, Nathalie; Oberson, Anne; Bustamante, Mauro; Tasinato, Andrea; Hummler, Edith; Schorderet, Daniel F
BIGH3 is a secreted protein, part of the extracellular matrix where it interacts with collagen and integrins on the cell surface. BIGH3 can play opposing roles in cancer, acting as either tumor suppressor or promoter, and its mutations lead to different forms of corneal dystrophy. Although many studies have been carried out, little is known about the physiological role of BIGH3. Using the cre-loxP system, we generated a mouse model with disruption of the Bigh3 genomic locus. Bigh3 silencing did not result in any apparent phenotype modifications, the mice remained viable and fertile. We were able to determine the presence of BIGH3 in the retinal pigment epithelium (RPE). In the absence of BIGH3, a transient decrease in the apoptotic process involved in retina maturation was observed, leading to a transient increase in the INL thickness at P15. This phenomenon was accompanied by an increased activity of the pro-survival ERK pathway.
Mutter, Marion; Swietek, Natalia; Münch, Thomas A
Blindness is one of the most devastating conditions affecting the quality of life. Hereditary degenerative diseases, such as retinitis pigmentosa, are characterized by the progressive loss of photoreceptors, leading to complete blindness. No treatment is known, the current state-of-the-art of restoring vision are implanted electrode arrays. As a recently discovered alternative, optical neuromodulators, such as channelrhodopsin, allow new strategies for treating these diseases by imparting light-sensitivity onto the remaining retinal neurons after photoreceptor cell death. Retinal degeneration is a heterogeneous set of diseases with diverse secondary effects on the retinal circuitry. Successful treatment strategies have to take into account this diversity, as only the existing retinal hardware can serve as substrate for optogenetic intervention. The goal is to salvage the retinal ruins and to revert the leftover tissue into a functional visual sensor that operates as optimally as possible. Here, we discuss three different successful approaches that have been applied to degenerated mouse retina.
Willis, Gregory L.; Moore, Cleo; Armstrong, Stuart Maxwell
Critical analysis of recent research suggesting that light pollution causes Parkinson's disease (PD) reveals that such a hypothesis is unsustainable in the context of therapeutic use of light in treating various neuropsychiatric conditions. Reinterpretation of their findings suggests that retinal damage caused by prolonged light exposure may have contributed to the observed enhancement of experimental PD. To test this hypothesis further, forty-two Sprague Dawley rats received microinjections of 6-hydroxydopamine (6-OHDA), 1-methyl-4-phenyl-2, 4, 6-tetrahydropyridine (MPTP), paraquat or rotenone into the vitreal mass in doses so minute that the effects could not be attributed to diffusion into brain. Significant changes in five motor parameters consistent with symptoms of experimental PD were observed. These findings support the interpretation that the retina is involved in the control of motor function and in the aetiology of PD. PMID:24473093
Baehr, Wolfgang; Frederick, Jeanne M.
Naturally occurring and laboratory generated animal models serve as powerful tools with which to investigate the etiology of human retinal degenerations, especially retinitis pigmentosa (RP), Leber congenital amaurosis (LCA), cone dystrophies (CD) and macular degeneration (MD). Much progress has been made in elucidating gene defects underlying disease, in understanding mechanisms leading to disease, and in designing molecules for translational research and gene-based therapy to interfere with the progression of disease. Key to this progress has been study of naturally occurring murine and canine retinal degeneration mutants. This article will review the history, phenotypes and gene defects of select animal models with outer retina (photoreceptor and retinal pigment epithelium) degeneration phenotypes. PMID:19375447
Malicki, J; Pooranachandran, N; Nikolaev, A; Fang, X; Avanesov, A
The vertebrate retina is remarkably conserved in evolution. Its relative simplicity and well-defined architecture make it particularly suitable for developmental and functional analysis of neuronal networks in the vertebrate central nervous system. The zebrafish model is at the forefront of these studies. It makes it possible to apply a wide variety of parallel embryological, genetic, and imaging tools to study the eye. Here we discuss experimental approaches that range from cell lineage analysis to the imaging of synaptic calcium currents and atomic force microscopy. These methods are currently used in zebrafish to model morphogenetic events during early development of the eye primordium, cell fate decisions during retinal neurogenesis, and the differentiation and function of the many fine structural features that underlie the detection and processing of light stimuli in the eye.
Gellermann, Werner; Emakov, Igor V.; McClane, Robert W.
We have generated high spatial resolution images showing the distribution of carotenoid macular pigments in the human retina using Raman spectroscopy. A low level of macular pigments is associated with an increased risk of developing age-related macular degeneration, a leading cause of irreversible blindness. Using excised human eyecups and resonant excitation of the pigment molecules with narrow bandwidth blue light from a mercury arc lamp, we record Raman images originating from the carbon-carbon double bond stretch vibrations of lutein and zeaxanthin, the carotenoids comprising human macular pigments. Our Raman images reveal significant differences among subjects, both in regard to absolute levels as well as spatial distribution within the macula. Since the light levels used to obtain these images are well below established safety limits, this technique holds promise for developing a rapid screening diagnostic in large populations at risk for vision loss from age-related macular degeneration.
Rodmell, Paul I; Crowe, John A; Gorman, Alastair; Harvey, Andrew R; Muyo, Gonzalo; Mordant, David J; McNaught, Andy I; Morgan, Stephen P
A Monte Carlo simulation of light propagation through the retina has been developed to understand the path-length distributions within the retinal vessel. For full-field illumination, the path-length distribution within the vessel comprises directly backscattered light and light that has passed once or twice through the vessel. The origins of these light path-length distributions can be better understood by investigating different combinations of single-point illumination and detection positions. Perhaps the most significant observation is that illumination at the edges of the vessel, rather than over the whole field of view, and detection directly above the vessel capture only the light that has taken a single pass through the vessel. This path-length distribution is tightly constrained around the diameter of the vessel and can potentially provide enhancements for oxygen saturation imaging. The method could be practically implemented using an offset-pinhole confocal imaging system or structured light illumination.
Cuadros, M A; García-Martín, M; Martin, C; Ríos, A
The existence of specialised phagocytic cells is described in regions of the retinal neuroepithelium undergoing intense cell death during early differentiation of the avian embryo retina (2.5-5 days of incubation). These results were obtained using routine techniques for light microscopy, acid phosphatase histochemistry and immunocytochemical staining with antibodies MB-1 and QH-1, both specific for quail endothelial cells and all blood cells except mature erythrocytes. Specialised phagocytes were distinguishable from neuroepithelial cells on the basis of morphological criteria: in the former, the nucleus was not oval in shape and was not oriented perpendicular to basement membrane neuroepithelium. The cytoplasm of the specialised phagocytes was often filled with dead cell fragments. In contrast to neuroepithelial cells, the specialised phagocytes showed acid phosphatase activity and were labelled with both MB-1 and QH-1 antibodies in normal quail embryos and chick----quail yolk sac chimeras. Moreover, some acid phosphatase positive and MB-1/QH-1 positive cells also appeared in the presumptive vitreous body, at the edges of the optic cup and in the surrounding mesenchyme. As the vitreal cells and the specialised phagocytes of the neural retina were immunolabelled in chick----quail yolk sac chimeras, we conclude that they are derived from haemopoietic cells in the yolk sac. Some images suggest that these cells enter the vitreous body from the surrounding mesenchyme and traverse the basement membrane of the neuroepithelium in the optic disc region to give rise to the specialised phagocytes of the retinal neuroepithelium. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Fig. 12 Fig. 13 Fig. 14 Fig. 15 Fig. 16 Fig. 17 Fig. 18 Fig. 19 Fig. 20 PMID:1769889
Fan, Jie; Wu, Bill X.; Crosson, Craig E.
Purpose Acid sphingomyelinase (ASMase) catalyzes the hydrolysis of sphingomyelin to ceramide and mediates multiple responses involved in inflammatory and apoptotic signaling. However, the role ASMase plays in ischemic retinal injury has not been investigated. The purpose of this study was to investigate how reduced ASMase expression impacts retinal ischemic injury. Methods Changes in ceramide levels and ASMase activity were determined by high performance liquid chromatography-tandem mass spectrometry analysis and ASMase activity. Retinal function and morphology were assessed by electroretinography (ERG) and morphometric analyses. Levels of TNF-α were determined by ELISA. Activation of p38 MAP kinase was assessed by Western blot analysis. Results In wild-type mice, ischemia produced a significant increase in retinal ASMase activity and ceramide levels. These increases were associated with functional deficits as measured by ERG analysis and significant structural degeneration in most retinal layers. In ASMase+/− mice, retinal ischemia did not significantly alter ASMase activity, and the rise in ceramide levels were significantly reduced compared to levels in retinas from wild-type mice. In ASMase+/− mice, functional and morphometric analyses of ischemic eyes revealed significantly less retinal degeneration than in injured retinas from wild-type mice. The ischemia-induced increase in retinal TNF-α levels was suppressed by the administration of the ASMase inhibitor desipramine, or by reducing ASMase expression. Conclusions Our results demonstrate that reducing ASMase expression provides partial protection from ischemic injury. Hence, the production of ceramide and subsequent mediators plays a role in the development of ischemic retinal injury. Modulating ASMase may present new opportunities for adjunctive therapies when treating retinal ischemic disorders. PMID:27571014
Garanto, Alejandro; Mandal, Nawajes A.; Egido-Gabás, Meritxell; Marfany, Gemma; Fabriàs, Gemma; Anderson, Robert E.; Casas, Josefina; Gonzàlez-Duarte, Roser
Sphingolipids (SPLs) are finely tuned structural compounds and bioactive molecules involved in membrane fluidity and cellular homeostasis. The core sphingolipid, ceramide (CER), and its derivatives, regulate several crucial processes in neuronal cells, among them cell differentiation, cell–cell interactions, membrane conductance, synaptic transmission, and apoptosis. Mutations in Ceramide Kinase-Like (CERKL) cause autosomal recessive Retinitis Pigmentosa and Cone Rod Dystrophy. The presence of a conserved lipid kinase domain and the overall similarity with CERK suggested that CERKL might play a role in the SPL metabolism as a CER kinase. Unfortunately, CERKL functi