Glycine receptors of A-type ganglion cells of the mouse retina

2007 ◽  
Vol 24 (4) ◽  
pp. 471-487 ◽  
Author(s):  
SRIPARNA MAJUMDAR ◽  
LIANE HEINZE ◽  
SILKE HAVERKAMP ◽  
ELENA IVANOVA ◽  
HEINZ WÄSSLE
Keyword(s):  
Fluorescent Protein ◽  
Ganglion Cells ◽  
Glycine Receptor ◽  
Mouse Retina ◽  
Inhibitory Input ◽  
Visual Channel ◽  
Fast Kinetics ◽  
Transgenic Mouse Line ◽  
Green Fluorescent ◽  
Α Subunits

A-type ganglion cells of the mouse retina represent the visual channel that transfers temporal changes of the outside world very fast and with high fidelity. In this study we combined anatomical and physiological methods in order to study the glycinergic, inhibitory input of A-type ganglion cells. Immunocytochemical studies were performed in a transgenic mouse line whose ganglion cells express green fluorescent protein (GFP). The cells were double labeled for GFP and the four α subunits of the glycine receptor (GlyR). It was found that most of the glycinergic input of A-type cells is through fast, α1-expressing synapses. Whole-cell currents were recorded from A-type ganglion cells in retinal whole mounts. The response to exogenous application of glycine and spontaneous inhibitory postsynaptic currents (sIPSCs) were measured. By comparing glycinergic currents recorded in wildtype mice and in mice with specific deletions of GlyRα subunits (Glra1spd-ot,Glra2−/−,Glra3−/−), the subunit composition of GlyRs of A-type ganglion cells could be further defined. Glycinergic sIPSCs of A-type ganglion cells have fast kinetics (decay time constant τ = 3.9 ± 2.5 ms, mean ± SD). Glycinergic sIPSCs recorded inGlra2−/−andGlra3−/−mice did not differ from those of wildtype mice. However, the number of glycinergic sIPSCs was significantly reduced inGlra1spd-otmice and the remaining sIPSCs had slower kinetics than in wildtype mice. The results show that A-type ganglion cells receive preferentially kinetically fast glycinergic inputs, mediated by GlyRs composed of α1 and β subunits.

scholarly journals A high-speed, bright, red fluorescent voltage sensor to detect neural activity

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Connor Beck ◽  
Yiyang Gong
Keyword(s):  
Blue Light ◽  
Neural Activity ◽  
High Speed ◽  
Fluorescent Protein ◽  
Resonance Energy ◽  
Fluorescent Sensors ◽  
Fast Kinetics ◽  
Spectral Separation ◽  
Green Fluorescent

Abstract Genetically encoded voltage indicators (GEVIs) have emerged as a technology to optically record neural activity with genetic specificity and millisecond-scale temporal resolution using fluorescence microscopy. GEVIs have demonstrated ultra-fast kinetics and high spike detection fidelity in vivo, but existing red-fluorescent voltage indicators fall short of the response and brightness achieved by green fluorescent protein-based sensors. Furthermore, red-fluorescent GEVIs suffer from incomplete spectral separation from green sensors and blue-light-activated optogenetic actuators. We have developed Ace-mScarlet, a red fluorescent GEVI that fuses Ace2N, a voltage-sensitive inhibitory rhodopsin, with mScarlet, a bright red fluorescent protein (FP). Through fluorescence resonance energy transfer (FRET), our sensor detects changes in membrane voltage with high sensitivity and brightness and has kinetics comparable to the fastest green fluorescent sensors. Ace-mScarlet’s red-shifted absorption and emission spectra facilitate virtually complete spectral separation when used in combination with green-fluorescent sensors or with blue-light-sensitive sensors and rhodopsins. This spectral separation enables both simultaneous imaging in two separate wavelength channels and high-fidelity voltage recordings during simultaneous optogenetic perturbation.

The anemia of the newborn induces erythropoietin expression in the developing mouse retina

2010 ◽  
Vol 299 (1) ◽  
pp. R111-R118 ◽  
Author(s):  
N. Scheerer ◽  
N. Dünker ◽  
S. Imagawa ◽  
M. Yamamoto ◽  
N. Suzuki ◽  
...  
Keyword(s):  
Transforming Growth Factor ◽  
Fluorescent Protein ◽  
Ganglion Cells ◽  
Retinal Development ◽  
Transforming Growth Factor Β ◽  
Neuroprotective Activity ◽  
Mouse Retina ◽  
Mrna Levels ◽  
Neuroprotective Effects

The hematopoietic hormone erythropoietin (Epo), regularly produced by the kidneys and the liver, is also expressed in neuronal tissue, where it has been found to mediate paracrine neuroprotective effects. In most studies exploring the rescue effects of Epo, apoptosis was exogenously induced by different cell death stimuli. Herein, we set out to study the expression and function of Epo in physiologically occurring apoptosis in a model of retinal development. We made use of an organotypic retinal wholemount culture system that resembles the physiological in vivo situation with cell connections still retained. Epo mRNA expression in the retina, liver, and kidney showed a significant increase during early development, coinciding with the anemia of the newborn. In the retina of Epo-green fluorescent protein transgenic mice, Epo-expressing cells were identified and found to be distributed in the retinal ganglion cell layer. Treatment of retinal wholemount cultures with recombinant Epo resulted in a significant decrease of apoptotic ganglion cells as well as photoreceptor cells throughout retinal development. Moreover, transforming growth factor-β-induced apoptosis was completely antagonized by Epo when both factors were simultaneously applied. Investigations on the signaling pathway revealed a decrease in Bax mRNA levels in Epo-treated retinal cells. We conclude that Epo exerts wide and prolonged neuroprotective activity in physiologically occurring apoptosis and thus contributes to proper retinal development.

scholarly journals Intrinsic and Extrinsic Light Responses in Melanopsin-Expressing Ganglion Cells During Mouse Development

2008 ◽  
Vol 100 (1) ◽  
pp. 371-384 ◽  
Author(s):  
Tiffany M. Schmidt ◽  
Kenichiro Taniguchi ◽  
Paulo Kofuji
Keyword(s):  
Fluorescent Protein ◽  
Ganglion Cells ◽  
Plexiform Layer ◽  
Artificial Chromosome ◽  
Inner Plexiform Layer ◽  
Mouse Development ◽  
Light Responses ◽  
Functional Changes ◽  
Electrophysiological Recordings ◽  
Green Fluorescent

Melanopsin (Opn4) is a photopigment found in a subset of retinal ganglion cells (RGCs) that project to various brain areas. These neurons are intrinsically photosensitive (ipRGCs) and are implicated in nonimage-forming responses to environmental light such as the pupillary light reflex and circadian entrainment. Recent evidence indicates that ipRGCs respond to light at birth, but questions remain as to whether and when they undergo significant functional changes. We used bacterial artificial chromosome transgenesis to engineer a mouse line in which enhanced green fluorescent protein (EGFP) is expressed under the control of the melanopsin promoter. Double immunolabeling for EGFP and melanopsin demonstrates their colocalization in ganglion cells of mutant mouse retinas. Electrophysiological recordings of ipRGCs in neonatal mice (postnatal day 0 [P0] to P7) demonstrated that these cells responded to light with small and sluggish depolarization. However, starting at P11 we observed ipRGCs that responded to light with a larger and faster onset (<1 s) and offset (<1 s) depolarization. These faster, larger depolarizations were observed in most ipRGCs by early adult ages. However, on application of a cocktail of synaptic blockers, we found that all cells responded to light with slow onset (>2.5 s) and offset (>10 s) depolarization, revealing the intrinsic, melanopsin-mediated light responses. The extrinsic, cone/rod influence on ipRGCs correlates with their extensive dendritic stratification in the inner plexiform layer. Collectively, these results demonstrate that ipRGCs make use of melanopsin for phototransduction before eye opening and that these cells further integrate signals derived from the outer retina as the retina matures.

Functional integrity of green fluorescent protein conjugated glycine receptor channels

1999 ◽  
Vol 38 (6) ◽  
pp. 785-792 ◽  
Author(s):  
Brigitte David-Watine ◽  
Spencer L. Shorte ◽  
Sergio Fucile ◽  
Didier de Saint Jan ◽  
Henri Korn ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Glycine Receptor ◽  
Functional Integrity ◽  
Green Fluorescent

Thioacylation is required for targeting G-protein subunit Go1α to detergent-insoluble caveolin-containing membrane domains

2001 ◽  
Vol 355 (2) ◽  
pp. 323-331 ◽  
Author(s):  
Francesca GUZZI ◽  
Deborah ZANCHETTA ◽  
Bice CHINI ◽  
Marco PARENTI
Keyword(s):  
Plasma Membrane ◽  
Fluorescent Protein ◽  
Transmembrane Helix ◽  
Membrane Domains ◽  
Protein Subunit ◽  
Caveolin 1 ◽  
V2 Receptor ◽  
Green Fluorescent ◽  
Membrane Compartmentalization ◽  
Α Subunits

α-Subunits of heterotrimeric Gi-like proteins (αi, αo and αz) associate with the cytoplasmic leaflet of the plasma membrane by means of N-terminally linked myristic acid and palmitic acid. An additional role for palmitate has been recently suggested by the observation that fusion with the palmitoylated N-terminus of αi1 relocalizes cytosolic green-fluorescent-protein reporter to low buoyancy, Triton-insoluble membrane domains (TIFF; Triton-insoluble floating fraction), enriched with caveolin-1 [Galbiati, Volonté, Meani, Milligan, Lublin, Lisanti and Parenti (1999) J. Biol. Chem 274, 5843-5850]. Here we show that, upon transient expression in transfected COS-7 cells, myristoylated and palmitoylated αo (αowt, where wt is wild-type) is exclusively found in TIFF, from where non-palmitoylated αowt and αoC3S (Cys3 → Ser) mutant are excluded. Moreover, αo fused to N-terminally truncated human vasopressin V2 receptor (V2TR-αo), lacking myristate and palmitate, still localizes at the plasma membrane by means of first transmembrane helix of V2R, but is excluded from TIFF. Likewise, αoC3S does not partition into TIFF, even when its membrane avidity is enhanced by co-expression of βγ-subunits. Thus membrane association, in the absence of added palmitate, is not sufficient to confer partitioning of αo within TIFF, suggesting that palmitoylation is a signal for membrane compartmentalization of dually acylated α-subunits

scholarly journals Asymmetric Distributions of Achromatic Bipolar Cells in the Mouse Retina

2022 ◽  
Vol 15 ◽  
Author(s):  
Zachary J. Sharpe ◽  
Angela Shehu ◽  
Tomomi Ichinose
Keyword(s):  
Visual Processing ◽  
Fluorescent Protein ◽  
Bipolar Cells ◽  
Mouse Retina ◽  
Temporal Region ◽  
Retinal Tissue ◽  
Evolutionary Changes ◽  
Asymmetric Distributions ◽  
Nasal Region ◽  
Green Fluorescent

In the retina, evolutionary changes can be traced in the topography of photoreceptors. The shape of the visual streak depends on the height of the animal and its habitat, namely, woods, prairies, or mountains. Also, the distribution of distinct wavelength-sensitive cones is unique to each animal. For example, UV and green cones reside in the ventral and dorsal regions in the mouse retina, respectively, whereas in the rat retina these cones are homogeneously distributed. In contrast with the abundant investigation on the distribution of photoreceptors and the third-order neurons, the distribution of bipolar cells has not been well understood. We utilized two enhanced green fluorescent protein (EGFP) mouse lines, Lhx4-EGFP (Lhx4) and 6030405A18Rik-EGFP (Rik), to examine the topographic distributions of bipolar cells in the retina. First, we characterized their GFP-expressing cells using type-specific markers. We found that GFP was expressed by type 2, type 3a, and type 6 bipolar cells in the Rik mice and by type 3b, type 4, and type 5 bipolar cells in the Lhx4 mice. All these types are achromatic. Then, we examined the distributions of bipolar cells in the four cardinal directions and three different eccentricities of the retinal tissue. In the Rik mice, GFP-expressing bipolar cells were more highly observed in the nasal region than those in the temporal retina. The number of GFP cells was not different along with the ventral-dorsal axis. In contrast, in the Lhx4 mice, GFP-expressing cells occurred at a higher density in the ventral region than in the dorsal retina. However, no difference was observed along the nasal-temporal axis. Furthermore, we examined which type of bipolar cells contributed to the asymmetric distributions in the Rik mice. We found that type 3a bipolar cells occurred at a higher density in the temporal region, whereas type 6 bipolar cells were denser in the nasal region. The asymmetricity of these bipolar cells shaped the uneven distribution of the GFP cells in the Rik mice. In conclusion, we found that a subset of achromatic bipolar cells is asymmetrically distributed in the mouse retina, suggesting their unique roles in achromatic visual processing.

scholarly journals Biomolecular Contrast Agents for Optical Coherence Tomography

2019 ◽  
Author(s):  
George J. Lu ◽  
Li-dek Chou ◽  
Dina Malounda ◽  
Amit K. Patel ◽  
Derek S. Welsbie ◽  
...  
Keyword(s):  
Optical Coherence Tomography ◽  
Contrast Agents ◽  
Fluorescent Protein ◽  
Mouse Retina ◽  
Optical Coherence ◽  
Gas Vesicles ◽  
Cellular Functions ◽  
Green Fluorescent

ABSTRACTOptical coherence tomography (OCT) has gained wide adoption in biological and medical imaging due to its exceptional tissue penetration, 3D imaging speed and rich contrast. However, OCT plays a relatively small role in molecular and cellular imaging due to the lack of suitable biomolecular contrast agents. In particular, while the green fluorescent protein has provided revolutionary capabilities to fluorescence microscopy by connecting it to cellular functions such as gene expression, no equivalent reporter gene is currently available for OCT. Here we introduce gas vesicles, a unique class of naturally evolved gas-filled protein nanostructures, as the first genetically encodable OCT contrast agents. The differential refractive index of their gas compartments relative to surrounding aqueous tissue and their nanoscale motion enables gas vesicles to be detected by static and dynamic OCT at picomolar concentrations. Furthermore, the OCT contrast of gas vesicles can be selectively erasedin situwith ultrasound, allowing unambiguous assignment of their location. In addition, gas vesicle clustering modulates their temporal signal, enabling the design of dynamic biosensors. We demonstrate the use of gas vesicles as reporter genes in bacterial colonies and as purified contrast agentsin vivoin the mouse retina. Our results expand the utility of OCT as a unique photonic modality to image a wider variety of cellular and molecular processes.

scholarly journals Genetic access to functionally distinct ganglion cell types in the Somatostatin-IRES-Cre mouse retina

2019 ◽  
Author(s):  
James W. Fransen ◽  
Bart G. Borghuis
Keyword(s):  
Ganglion Cells ◽  
Visual Image ◽  
Cell Types ◽  
Luminance Contrast ◽  
Mouse Retina ◽  
Mouse Line ◽  
Visual Responses ◽  
Transgenic Mouse Line ◽  
Inhibitory Circuits ◽  
Distinct Cell

AbstractRetinal ganglion cells (GCs) are a functionally diverse neuron population that encodes and transmits distinct representations of the visual image on the retina to target nuclei in the brain. Independent studies of visually-evoked responses, cell morphology, and gene expression each suggest that GCs in mouse may comprise as many as forty distinct cell types. To date, only a subset of these types have been characterized in detail, and for most genetic access is still lacking. Thus, the majority of identified GC types remains inaccessible for targeted electrophysiology and functional imaging, precluding efficient studies of their response properties, and the cell-intrinsic mechanisms and presynaptic circuits that generate these properties. Here we show that an existing mouse line that is commonly used for studies of cortical inhibitory circuits – Somatostatin-IRES-Cre (Sst-Cre), consistently labels an understudied subset of four GC types with distinct visual responses. We characterized these types both anatomically and functionally using Cre-dependent reporter mouse lines and confocal fluorescence imaging, calcium imaging, and whole-cell electrophysiology. We show that one of the labeled GC types is suppressed by luminance contrast, while another matches a recently described orientation-selective GC type. Our results give new information about these two identified GC types, and establish the utility of the Sst-Cre transgenic mouse line for studies of recently identified GC circuits in the mouse retina.

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