Research Article
Co-expression of murine opsins facilitates identifying the site of cone adaptation
- BJÖRN EKESTEN, PETER GOURAS, JANOS HARGITAI
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- 30 October 2002, pp. 389-393
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Murine cones contain two opsins in the same cone, one ultraviolet (UV) and the other middle-wavelength sensitive (M). A long-wavelength flash only affecting M-opsin suppresses the cone electroretinogram (ERG) produced by light absorption of UV-cone opsin raising the hypothesis that activation of M-cone opsin suppresses UV-cone opsin responses in the same cone. Here we show that pharmacologic blockade of synaptic transmission in the superfused murine retina, which eliminates interaction from second-order neurons, fails to prevent suppression of the UV-opsin driven pathway by long-wavelength stimuli. This proves that the antagonism must be occurring in the same cone, co-expressing both opsins. Our results show that UV-opsin suppression successively ceases in presence of the M-opsin activating background light, which implies that cone light adaptation is controlled at the opsin stage, before activation of transducin. It also reveals the time course of a transient desensitization of cones due to post-opsin factors in the transduction cascade.
Distribution of protein kinase C isoforms in the cat retina
- BOZENA FYK-KOLODZIEJ, WENHUI CAI, ROBERTA G. POURCHO
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- 12 November 2002, pp. 549-562
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Immunocytochemical localization was carried out for five isoforms of protein kinase C (PKC) in the cat retina. In common with other mammalian species, PKCα was found in rod bipolar cells. Staining was also seen in a small population of cone bipolar cells with axon terminals ramifying near the middle of the inner plexiform layer (IPL). PKCβI was localized to rod bipolar cells, one class of cone bipolar cell, and numerous amacrine and displaced amacrine cells. Staining for PKCβII was seen in three types of cone bipolar cells as well as in amacrine and ganglion cells. Immunoreactivity for both PKCε and PKCζ was found in rod bipolar cells; PKCε was also seen in a population of cone bipolar cells and a few amacrine and ganglion cells whereas PKCζ was found in all ganglion cells. Double-label immunofluorescence studies showed that dendrites of the two PKCβII-positive OFF-cone bipolar cells exhibit immmunoreactivity for the kainate-selective glutamate receptor GluR5. The third PKCβII cone bipolar is an ON-type cell and did not stain for GluR5. The retinal distribution of these isoforms of PKC is consistent with a role in modulation of various aspects of neurotransmission including synaptic vesicle release and regulation of receptor molecules.
Development of glutamatergic synapses in the rat retina: The postnatal expression of ionotropic glutamate receptor subunits
- IRIS HACK, PETER KOULEN, LEO PEICHL, JOHANN HELMUT BRANDSTÄTTER
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- 28 June 2002, pp. 1-13
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We examined the distribution of the AMPA glutamate receptor subunits GluR1 to GluR4, of the kainate receptor subunits GluR6/7 and KA2, and of the glutamate receptor subunits δ1/2, during postnatal development of the rat retina by immunocytochemistry and light microscopy using receptor subunit specific antisera. The various ionotropic glutamate receptor subunits were expressed early in postnatal rat retina, and most of the subunits, with the exception of δ1/2, were found in both synaptic layers of rat retina. The glutamate receptor subunits studied showed differences in their time of appearance, their spatial distribution patterns, and in their expression levels in the developing rat retina. Interestingly, most of the AMPA receptor subunits were expressed earlier than the kainate receptor subunits in the two synaptic layers of the retina, indicating that AMPA glutamate receptors play an important role in early postnatal glutamatergic synaptic transmission. We also studied the ultrastructural localization of the AMPA glutamate receptor subunits GluR1 to GluR4 by immunocytochemistry and electron microscopy in the inner plexiform layer of the mature rat retina. Most of the subunits were found postsynaptic to the ribbon synapses of OFF-cone, ON-cone, and rod bipolar cells. The results of this study suggest an involvement of ionotropic glutamate receptors in processes of synaptic maturation and the formation of synaptic circuitries in the developing plexiform layers of the retina. Furthermore, AMPA and kainate receptors play a role in synaptic processing and in the development of both the scotopic and photopic pathways in the rat retina.
Coupling pattern of S1 and S2 amacrine cells in the rabbit retina
- WEI LI, JIAN ZHANG, STEPHEN C. MASSEY
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- 02 July 2002, pp. 119-131
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Previous studies have shown that indoleamine-accumulating cells (IACs) in the rabbit retina consist of two main cell types: S1 and S2 amacrine cells (Vaney, 1986; Sandell & Masland, 1986). Both cell types are wide-field GABA amacrine cells that make reciprocal synaptic contacts with rod bipolar cell terminals (Ehinger & Holmgren, 1979; Strettoi et al., 1990). We have examined the coupling pattern of S1 and S2 amacrine cells after the intracellular injection of Neurobiotin. Our results may be summarized as follows: (1) S1 amacrine cells were extensively coupled and their dendrites formed a network similar to but less dense than the matrix stained with an antibody to serotonin. (2) Morphological observations and cluster analysis, based on a scattergram, showed that the vast majority of coupled cells were S1 amacrine cells, accounting for approximately half of the total IACs. The rest of the uncoupled IACs were S2 amacrine cells. (3) Sometimes, two adjacent varicosities, one from an injected S1 and one from a coupled S1, contacted a single rod bipolar terminal. (4) S2 amacrine cells were also coupled but much less than the S1s. (5) Rarely, crossover coupling between S1 and S2 amacrine cells was observed. These results suggest that the extensive coupling between S1 amacrine cells, combined with a larger dendritic field, may contribute a wide-field component to the inhibitory surround of the rod pathway. By comparison, the smaller, weakly coupled S2 amacrine cells may provide a local component.
D2-like dopamine receptors promote interactions between calcium and chloride channels that diminish rod synaptic transfer in the salamander retina
- WALLACE B. THORESON, SALVATORE L. STELLA, ERIC J. BRYSON, JOHN CLEMENTS, PAUL WITKOVSKY
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- 05 September 2002, pp. 235-247
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Activation of D2-like dopamine receptors in rods with quinpirole stimulates L-type calcium currents (ICa). This result appears inconsistent with studies showing that D2-like dopamine receptor activation diminishes rod signals in second-order retinal neurons. Since small reductions in [Cl−]i can inhibit photoreceptor ICa, we tested the hypothesis that enhancement of ICa with the D2/D4 receptor agonist, quinpirole, increases calcium-activated chloride currents (ICl(Ca)) causing an efflux of Cl− from rods that would provide a negative feedback inhibition of ICa. In agreement with studies from Xenopus, quinpirole reduced rod input to second-order neurons of tiger salamander retina without significantly altering rod voltage responses. Quinpirole also diminished the amplitude of depolarization-evoked increases in [Ca2+]i measured with Fura-2 in rods, a finding consistent with inhibition of synaptic transmission from rods. Electrophysiological and Cl−-imaging experiments indicated ECl in rods is ∼ −20 mV. Quinpirole enhanced ICl(Ca) and elicited an efflux of Cl− at the resting potential. A similar Cl− efflux was produced by extracellular replacement of 24 mM Cl− with CH3SO4− and this low Cl− solution inhibited Ca2+responses to a similar degree as quinpirole did. When ICl(Ca) was inhibited with niflumic acid, quinpirole enhanced both ICa and depolarization-evoked increases in [Ca2+]i. Furthermore, with niflumic acid, quinpirole no longer inhibited rod inputs into horizontal and bipolar cells. These results suggest an initial enhancement of ICa by quinpirole is followed by a stimulation of Cl− currents, including ICl(Ca). The net result is a Cl− efflux that inhibits depolarization-evoked increases in [Ca2+]i and synaptic transmission from rods.
Noninvasive recording and response characteristics of the rat dc-electroretinogram
- NEAL S. PEACHEY, J. BRETT STANTON, ALAN D. MARMORSTEIN
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- 30 January 2003, pp. 693-701
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In response to light, the retinal pigment epithelium (RPE) generates a series of potentials that can be recorded using the dc-electroretinogram (dc-ERG). As these potentials can be related to specific cellular events, they provide information about RPE function and how that may be altered by disease or experimental manipulation. The purposes of the present study were to define a noninvasive means for recording the rat dc-ERG, to use this to define the stimulus–response properties of the major components, and to relate these results to measures of the rat electrooculogram (EOG). Parallel studies were conducted in two strains of rats (Long-Evans, LE; Sprague-Dawley, SD) that are commonly used in vision research. Rats were sedated with ketamine/xylazine and placed on a heating pad. Ag/AgCl wire electrodes were bridged with capillary tubes filled with Hanks balanced salt solution. The active electrode was placed in contact with the corneal surface and referenced to a second electrode placed within the orbit. The dc-ERG signal was amplified (dc-100 Hz), digitized, and stored offline. The duration of full-field flash stimuli was controlled using a mechanical shutter and flash luminance was controlled with neutral density filters. EOGs were recorded using subdermal platinum needle electrodes placed near the eye. In response to a 5-min light exposure, the dc-ERG of LE and SD rats included a distinct b-wave, after potential, c-wave, fast oscillation, and a slow potential of positive polarity the characteristics of which are consistent with a light peak.
Modeling receptive-field structure of koniocellular, magnocellular, and parvocellular LGN cells in the owl monkey (Aotus trivigatus)
- XIANGMIN XU, A.B. BONDS, VIVIEN A. CASAGRANDE
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- 30 January 2003, pp. 703-711
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Most cells in the retina and lateral geniculate nucleus (LGN) of primates have a concentric center/surround receptive-field organization. Details of the relationship between center and surround often can be used to predict how cells respond to visual stimuli. Models of the receptive-field organization and center/surround relationships also are useful when comparing cell classes. In the present study, we used the difference-of-Gaussians (DOG) model to quantitatively examine and compare the receptive-field center/surround organization of koniocellular (K), magnocellular (M), and parvocellular (P) LGN cells of owl monkeys. We obtained estimates of receptive-field center size (rc) and center sensitivity (Kc), and surround size (rs), and surround sensitivity (Ks) from 62 K, M, and P LGN cells by fitting their spatial-frequency responses with a DOG function (Rodieck, 1965; Croner & Kaplan, 1995). The DOG function not only accounted for the responses of P and M cells, but also provided a good description of K-cell responses. We found that at matched eccentricities of less than 15 deg, K cells had the largest rc and rs among the three cell classes. K cells also had the lowest Kc and Ks.
Contextual influences on the directional responses of tectal cells in pigeons
- HONG-JIN SUN, JIAN ZHAO, TRACY L. SOUTHALL, BIN XU
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- 02 July 2002, pp. 133-144
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Contrary to the traditional view that receptive fields are limited in spatial extent, recent studies have indicated that the response of neurons to a local stimulus within the receptive field can be modulated by stimulation of the surrounding region. Here we quantified the nature of these contextual effects on visual motion responses of neurons in the pigeon's optic tectum using standard extracellular recording techniques. All of the cells tested responded well to a test spot moving across their receptive fields. When a background pattern was moved in the same or in a similar direction as that of the test spot, the responses of most deep tectal neurons to the test spot were maximally inhibited. Movement of the background in the opposite or near opposite direction produced minimal inhibition or even facilitation. For some deep tectal neurons, this directionally selective modulation by the moving background was maintained when the background motion was paired with different movement directions of the test spot (including both the preferred and least preferred directions). Thus, this selectivity for opposing motion was independent of the absolute direction of either the test spot or the background, a finding which is consistent with the results reported by Frost and Nakayama (1983), although they did not include all test spot directions. For some other neurons, identified here for the first time, the background movement selectively modulated the response only when the test spot moved in the neuron's preferred directions. These neurons lost selectivity for opposing motion when the test spot moved in nonpreferred directions. The significance of these contextual effects on the motion response of tectal neurons may be related to how the brain distinguishes self-induced motion from object motion and segregates figure from ground.
AMPA glutamate receptor subunit 2 in normal and visually deprived macaque visual cortex
- MARGARET T.T. WONG-RILEY, PAULETTE JACOBS
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- 12 November 2002, pp. 563-573
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Glutamate and its various receptors are known to play an important role in excitatory synaptic transmission throughout the CNS, including the primary visual cortex. Among subunits of the AMPA receptors (α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid), subunit 2 (GluR2) is of special significance because it controls their Ca2+ permeability. In the past, this subunit has been studied mostly in conjunction with other AMPA subunits. The present study sought to determine if GluR2 alone has a distinct laminar distribution in the normal macaque visual cortex, and if its pattern correlated with that of cytochrome oxidase (CO) under normal and monocularly deprived conditions. In the normal adult cortex, GluR2 immunoreactivity (ir) had a patchy distribution in layers II/III, in register with CO-rich puffs. GluR2-ir highlighted the upper border of layer II, the lower border of layer IV (previously termed IVCβdark) and, most prominently, layer VI. Labeled neurons were primarily of the pyramidal type present in the upper border and lower half of layer VI, layers II/III, and scattered in layers V and upper IVB. Labeled nonpyramidal cells were large in layer IVB and small in IVCβdark. Notably, the bulk of CO-rich layers IVC and IVA had very low levels of GluR2-ir. At fetal day 13, however, GluR2 labeling showed a honeycomb-like pattern in layer IVA not found in the adult. A fragment of GluR2 cDNA was generated from a human cDNA library, and in situ hybridization revealed an expression pattern similar to that of GluR2 proteins. After 1–4 weeks of monocular impulse blockade with tetrodotoxin (TTX), alternating rows of strong and weak GluR2-ir in layers VI and II/III appeared in register with CO-labeled dark and light ocular dominance columns in layer IVC and puffs in II/III, respectively. Our results indicate that various cortical layers are differentially influenced by glutamate. The bulk of the major geniculate-recipient layers IVC and IVA have low levels of GluR2, presumably favoring synaptic transmission via Ca2+-permeable glutamate receptors. GluR2 plays a more important role in supragranular and infragranular layers, where the initial geniculate signals are further modified and are transmitted to other cortical and subcortical centers. The maintenance of GluR2 in these output layers is governed by visual input and neuronal activity, as monocular impulse blockade induced a down-regulation of this subunit in deprived ocular dominance columns.
NMDA-evoked [Ca2+]i increase in salamander retinal ganglion cells: Modulation by PKA and adrenergic receptors
- YI HAN, SAMUEL M. WU
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- 05 September 2002, pp. 249-256
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Application of NMDA induces a depolarization and increase of intracellular calcium concentration ([Ca2+]i) in retinal ganglion cells, which cause ganglion cell death in models of glaucoma. In the present study, we investigated the pharmacological mechanism of how NMDA-evoked increase in calcium could be modulated in dissociated retinal ganglion cells from tiger salamander. In these neurons, protein kinase A (PKA) up-regulated the NMDA-evoked [Ca2+]i increase. In the presence of 8-bromo-cAMP or forskolin to stimulate PKA, the elevation level of [Ca2+]i induced by NMDA became even higher; In the presence of H-89, a PKA inhibitor, the NMDA-evoked [Ca2+]i increase was attenuated. In addition, applications of adrenergic compounds were also found to influence the NMDA-evoked [Ca2+]i increase. UK-14,304, a selective α2 agonist, reduced the elevation level of [Ca2+]i caused by NMDA. In contrast, isoproterenol, a β agonist, augmented the NMDA-evoked [Ca2+]i increase. These adrenergic regulations were due to direct activation of adrenoceptors, since modulations of both UK-14,304 and isoproterenol on the NMDA-evoked [Ca2+]i increase were abolished by their respective antagonists. Furthermore, adrenergic regulations were mediated through a PKA-related pathway since PKA inhibitor blocked adrenergic regulations. The possible modulatory site(s) by PKA was also discussed.
The pupillary and ciliary components of the cat Edinger-Westphal nucleus: A transsynaptic transport investigation
- JONATHAN T. ERICHSEN, PAUL J. MAY
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- 28 June 2002, pp. 15-29
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The distribution of preganglionic motoneurons supplying the ciliary ganglion in the cat was defined both qualitatively and quantitatively. These cells were retrogradely labeled directly, following injections of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) into the ciliary ganglion, or were transsynaptically labeled following injections of WGA into the vitreous chamber. Almost half of the cells are distributed rostral to the oculomotor nucleus, both in and lateral to the anteromedian nucleus. Of the remaining preganglionic motoneurons, roughly 20% of the total are located dorsal to the oculomotor nucleus. Strikingly few of these neurons are actually found within the Edinger-Westphal nucleus proper. Instead, the majority are found in the adjacent supraoculomotor area or along the midline between the two somatic nuclei. An additional population, roughly 30% of the total, is located ventral to the oculomotor nucleus. This study also provides evidence for a functional subdivision of this preganglionic population. Pupil-related preganglionic motoneurons were transsynaptically labeled by injecting WGA into the anterior chamber, while lens-related preganglionic motoneurons were transsynaptically labeled by injecting WGA into the ciliary muscle. The results suggest that the pupil-related preganglionic motoneurons, that is, those controlling the iris sphincter pupillae muscle, are located rostrally, in and lateral to the anteromedian nucleus. In contrast, lens-related preganglionic motoneurons, that is, those controlling the ciliary muscle are particularly prevalent caudally, both dorsal and ventral to the oculomotor nucleus. Thus, the cat intraocular muscle preganglionic innervation is spatially organized with respect to function, despite the dispersed nature of its distribution.
Photoreceptor inner segments in monkey and human retina: Mitochondrial density, optics, and regional variation
- Q.V. HOANG, R.A. LINSENMEIER, C.K. CHUNG, C.A. CURCIO
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- 30 October 2002, pp. 395-407
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The present work quantifies aspects of photoreceptor structure related to mitochondria, inner segment dimensions, and optical properties, as a basis for furthering our understanding of rod and cone function. Electron-microscopic analyses were performed on the retina of one stumptail macaque (Macaca arctoides) to obtain stereological measurements of ellipsoid mitochondrial density, and sizes and shapes of outer and inner segments. In addition, the distribution of mitochondria and the optical properties of human foveal cones were examined with electron microscopy and Nomarski differential interference contrast (NDIC) imaging. Mitochondria comprised 74–85% of cone ellipsoids and 54–66% of rod ellipsoids in macaque. Ellipsoid volume increased with eccentricity by 2.4-fold for rods and more than 6-fold for cones over eccentricities to 12.75 mm, while the volume of the outer segment supported by the ellipsoid was essentially constant for both rods and cones. Per unit volume of outer segment, cones contained ten times as much mitochondria as rods. In human fovea, as in the rest of the retina, most cone mitochondria were located in the distal inner segment. In the foveal center, however, there are also mitochondria in the myoid, as well as in the outer fiber, proximal to the external limiting membrane (ELM). Analyses of the optical aperture of human foveal cones, the point at which their refractive index clearly differs from the extrareceptoral space, showed that it correlated well with the location of mitochondria, except in the foveal center, where the aperture appeared proximal to the ELM. While mitochondria have an important metabolic function, we suggest that the striking differences between rods and cones in mitochondrial content are unlikely to be determined by metabolic demand alone. The numerous cone mitochondria may enhance the waveguide properties of cones, particularly in the periphery.
Responses in extrastriate cortex to optic flow during simulated turns
- HELEN SHERK, JONG-NAM KIM
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- 30 October 2002, pp. 409-419
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An observer locomoting along a straight path sees a pattern of optic flow in which images move approximately radially outward from his heading point. If the observer turns, the optic flow field changes markedly because each object's image now has a component of horizontal motion added to its “optic flow” motion. We tested the responses of 326 cells in the cat's extrastriate area LS (lateral suprasylvian visual area) to movies simulating the optic flow seen during locomotion in a straight line, and during various simulated turns to the left and right. About 60% of 326 cells tested responded to optic flow simulating turns. Of most interest was a subset of cells, 15% of the total, that had “turn-selective” responses. They responded significantly better to turns in a particular direction (usually to the contralateral side) than to turns in the opposite direction or to optic flow simulating straight-ahead locomotion. For each cell, we generated a display of fronto-parallel motion with a direction and speed that matched the image motion in the preferred turn movie, as seen at the receptive-field center. Most turn-selective cells responded significantly better to their preferred turn movie than to this fronto-parallel stimulus. We examined the role of cells' selectivity for stimulus direction, speed, and acceleration in determining cell preference for particular turns. Direction preference played some role for most cells, but about a third of the cells preferred turn movies that did not reflect their direction selectivity. Other factors, including the presence of opposing motion, only rarely appeared to determine cell preferences for particular turn movies.
Calcium-induced calcium release and calcium buffering in retinal horizontal cells
- EDUARDO SOLESSIO, ERIC M. LASATER
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- 30 January 2003, pp. 713-725
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Calcium plays an integral role in intracellular signaling and process control in neurons. In the outer retina, it is a key component to the phototransduction cycle and neurotransmitter release in photoreceptor and bipolar cell terminals. It also contributes to the responses of horizontal and bipolar cells. In the dark, horizontal cells are depolarized and calcium enters via calcium permeant AMPA receptors and voltage-activated calcium channels. As a result, horizontal cells must be capable of handling high calcium loads without sustaining damage. The aim of this study was to examine the components determining the intracellular calcium levels in H2 horizontal cells in the retina of white bass. Calcium responses were evoked in isolated cells by depolarizing voltage steps and monitored by conventional imaging techniques. The responses consisted of two components: calcium entry through voltage-gated calcium channels and subsequent release from intracellular stores by calcium-induced calcium release (CICR). Under control conditions, changes in calcium levels reached 541 nM on average from a basal level of 60 nM. When release from CICR stores was blocked with ryanodine or dantrolene, calcium levels barely reached 180 nM. The threshold level needed to trigger CICR was dependent on the duration of the applied depolarization and increased in response to shorter pulses.
Residual eye-movements in macaque and their effects on visual responses of neurons
- JASON FORTE, JONATHAN W. PEIRCE, JAMES M. KRAFT, JOHN KRAUSKOPF, PETER LENNIE
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- 28 June 2002, pp. 31-38
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We recorded continuously, with high precision, the positions of the eyes in anesthetized macaque monkeys prepared for physiological recording. Most recordings were made after the infusion of muscle relaxant to immobilize the eyes; in some cases we also were able to record eye position for periods before the eyes were immobilized. In all monkeys, the eyes moved continuously by as much as 0.5 deg over a 10-min sampling period. The average distance moved was proportional to the square root of the sampling period, as would be expected from a random walk. The movements had three distinct components: slow drifts, and two rhythms driven by the pulse and respiration. The rhythmic movements occurred only under paralysis: they were not discernible in measurements made before the infusion of muscle relaxant. The movements of the eye in the paralyzed animal can have substantial effects on the measured physiological characteristics of neurons. For excursions in the midrange of those we observed, a neuron's sensitivity to a spatial frequency of 10 cycle/deg might be underestimated by as much as a factor of three, depending on the method by which responses were averaged. We show how the effects of eye-movements can be mitigated by appropriate data analysis.
A structural basis for omnidirectional connections between starburst amacrine cells and directionally selective ganglion cells in rabbit retina, with associated bipolar cells
- E.V. FAMIGLIETTI
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- 02 July 2002, pp. 145-162
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Directionally selective (DS) ganglion cells of rabbit retina are of two principal types. ON DS ganglion cells prefer low velocity in one of three directions of movement and project axons to the accessory optic system (AOS), whereas ON–OFF DS ganglion cells prefer higher velocity in one of four directions and project to tectum and thalamus. Each has a distinct, recognizable dendritic morphology, based upon the correlation of form, physiology, and central projections. In previous Golgi studies, ON and ON–OFF DS cells were found to be partly co-stratified, and ON–OFF DS cells were found to co-stratify with starburst amacrine (SA) cells, the cholinergic amacrine cells of the retina, which also contain elevated levels of GABA. SA cells are radially symmetrical, have synaptic boutons in a distal annular zone of its dendritic tree, are presynaptic primarily to ganglion cell dendrites, co-stratify with ON–OFF DS ganglion cells, and contain the neurotransmitters shown pharmacologically to be involved in DS responses. For these reasons, SA cells are thought to play a role in the DS mechanism. Several models of this mechanism have utilized SA cell dendritic geometry in a centrifugal, radial format to impose directional inputs on DS ganglion cells.
Transgenic expression of a GFP-rhodopsin COOH-terminal fusion protein in zebrafish rod photoreceptors
- BRIAN D. PERKINS, PAMELA M. KAINZ, DONALD M. O'MALLEY, JOHN E. DOWLING
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- 05 September 2002, pp. 257-264
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To facilitate the identification and characterization of mutations affecting the retina and photoreceptors in the zebrafish, a transgene expressing green fluorescent protein (GFP) fused to the C-terminal 44 amino acids of Xenopus rhodopsin (Tam et al., 2000) under the control of the 1.3-kb proximal Xenopus opsin promoter was inserted into the zebrafish genome. GFP expression was easily observed in a ventral patch of retinal cells at 4 days postfertilization (dpf). Between 45–50% of the progeny from the F1, F2, and F3 generations expressed the transgene, consistent with a single integration event following microinjection. Immunohistochemical analysis demonstrated that GFP is expressed exclusively in rod photoreceptors and not in the UV, blue, or red/green double cones. Furthermore, GFP is localized to the rod outer segments with little to no fluorescence in the rod inner segments, rod cell bodies, or rod synapse regions, indicating proper targeting and transport of the GFP fusion protein. Application of exogenous retinoic acid (RA) increased the number of GFP-expressing cells throughout the retina, and possibly the level of expressed rhodopsin. When bred to a zebrafish rod degeneration mutant, fewer GFP-expressing rods were seen in living mutants as compared to wild-type siblings. This transgenic line will facilitate the search for recessive and dominant mutations affecting rod photoreceptor development and survival as well as proper rhodopsin expression, targeting, and transport.
Retinal ganglion cells projecting to the optic tectum and visual thalamus of lizards
- ALINO MARTINEZ-MARCOS, ENRIQUE LANUZA, FERNANDO MARTINEZ-GARCIA
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- 12 November 2002, pp. 575-581
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Retinal ganglion cells projecting to the optic tectum and visual thalamus have been investigated in the lizard, Podarcis hispanica. Injections of biotinylated dextran-amine in the optic tectum reveal seven morphological cell varieties including one displaced ganglion cell type. Injections in the visual thalamus yield similar ganglion cell classes plus four giant ganglion cells, including two displaced ganglion cell types. The present study constitutes the first comparison of tectal versus thalamic ganglion cell types in reptiles. The situation found in lizards is similar to that reported in mammals and birds where some cell types projecting to the thalamus are larger than those projecting to the mesencephalic roof. The presence of giant retino-thalamic ganglion cells with specific dendritic arborizations in sublaminae A and B of the inner plexiform layer suggests that parts of the visual thalamus of lizards could be implicated in movement detection, a role that might be played by the ventral lateral geniculate nucleus, which is involved in our tracer injections.
Melatonin receptor mRNA localization and rhythmicity in the retina of the domestic chick, Gallus domesticus
- ARJUN K. NATESAN, VINCENT M. CASSONE
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- 05 September 2002, pp. 265-274
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The indoleamine hormone melatonin is synthesized and released by photoreceptors during the night within the chick retina, and confers timing information to modulate retinal physiology. Three subtypes of melatonin receptor with nearly identical pharmacological profiles have been described in chickens and are present in the retina. In this study, the spatial localization and temporal pattern of the mRNA for each of these receptors within the retina are described. The localization and rhythmicity of receptor mRNA were analyzed using in situ hybridization and RNase protection assay, respectively, with probes against specific nucleotide sequences encoding these receptors. Mel1A and Mel1C receptor mRNA have similar patterns of expression, primarily in the inner segments of photoreceptors, vitread portion of the inner nuclear layer, and in the retinal ganglion cell layer. Mel1B receptor mRNA is expressed at higher levels in the retina, with expression in photoreceptors, throughout the inner nuclear layer, and in the ganglion cell layer. Mel1A receptor mRNA is rhythmic in both light:dark (LD) cycles and in constant darkness (DD); Mel1A peaks during midday and mid-subjective day, respectively. Mel1C receptor mRNA is also rhythmically expressed in LD, but with a lower amplitude, such that transcript is high during the day and low during the night. In DD, Mel1C rhythms become 180 deg out of phase with a slight increase at night. Mel1B mRNA expression was highly variable and arrhythmic.
Differential expression of K+ currents in mammalian retinal bipolar cells
- HUI-JUAN HU, ZHUO-HUA PAN
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- 02 July 2002, pp. 163-173
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Whole-cell voltage-clamp recordings were performed to investigate voltage-dependent K+ currents in acutely isolated retinal cone bipolar cells (CBCs) from the rat. The physiological and pharmacological properties of the currents were compared with those in rod bipolar cells (RBCs). The K+ currents were found to be much larger in CBC than in RBCs. In addition, the currents in CBCs were activated and inactivated at more negative potentials. Based on the apparent inactivation property of the currents, CBCs were found to fall into two groups of cells that differed in the inactivation kinetics of IK(V) but did not correlate to the ON- and OFF-type. The IK(V) for the group of CBCs showing faster inactivation, as well as for all RBCs, contained two components with decay time constants around 0.1 and 1 s. The IK(V) for the group of CBCs showing slower inactivation only contained the slower component. Furthermore, three components of IK(V) were observed based on tetraethylammonium (TEA) sensitivity: high-sensitive, low-sensitive, and resistant component. The IK(V) for a portion of CBCs showing faster inactivation, as well as for all RBCs, contained all three components. The IK(V) for the remaining CBCs, including all of those CBCs showing slower inactivation, only contained the latter two components. This study reveals a differential expression of K+ currents in rat retinal bipolar cells, suggesting that K+ channels may play an important role in bipolar cell processing in mammalian retinas.