During the first postnatal week in rodents, cholinergic retinal waves initiate in starburst amacrine cells (SACs), propagating to retinal ganglion cells (RGCs) and visual centers, essential for visual circuit refinement. By modulating exocytosis in SACs, dynamic changes in the protein kinase A (PKA) activity can regulate the spatiotemporal patterns of cholinergic waves. Previously, cysteine string protein-α (CSPα) is found to interact with the core exocytotic machinery by PKA-mediated phosphorylation at serine 10 (S10). However, whether PKA-mediated CSPα phosphorylation may regulate cholinergic waves via SACs remains unknown. Here, we examined how CSPα phosphorylation in SACs regulates cholinergic waves. First, we identified that CSPα1 is the major isoform in developing rat SACs and the inner plexiform layer during the first postnatal week. Using SAC-specific expression, we found that the CSPα1-PKA-phosphodeficient mutant (CSP-S10A) decreased wave frequency, but did not alter the wave spatial correlation compared to control, wild-type CSPα1 (CSP-WT), or two PKA-phosphomimetic mutants (CSP-S10D and CSP-S10E). These suggest that CSPα-S10 phosphodeficiency in SACs dampens the frequency of cholinergic waves. Moreover, the level of phospho-PKA substrates was significantly reduced in SACs overexpressing CSP-S10A compared to control or CSP-WT, suggesting that the dampened wave frequency is correlated with the decreased PKA activity. Further, compared to control or CSP-WT, CSP-S10A in SACs reduced the periodicity of wave-associated postsynaptic currents (PSCs) in neighboring RGCs, suggesting that these RGCs received the weakened synaptic inputs from SACs overexpressing CSP-S10A. Finally, CSP-S10A in SACs decreased the PSC amplitude and the slope to peak PSC compared to control or CSP-WT, suggesting that CSPα-S10 phosphodeficiency may dampen the speed of the SAC-RGC transmission. Thus, via PKA-mediated phosphorylation, CSPα in SACs may facilitate the SAC-RGC transmission, contributing to the robust frequency of cholinergic waves.

The dorsal lateral geniculate nucleus (dLGN) of the thalamus is the principal conduit for visual information from retina to visual cortex. Viewed initially as a simple relay, recent studies in the mouse reveal far greater complexity in the way input from the retina is combined, transmitted, and processed in dLGN. Here we consider the structural and functional organization of the mouse retinogeniculate pathway by examining the patterns of retinal projections to dLGN and how they converge onto thalamocortical neurons to shape the flow of visual information to visual cortex.

Often mislabeled as a simple relay of sensory information, the thalamus is a complicated structure with diverse functions. This diversity is exemplified by roles visual thalamus plays in processing and transmitting light-derived stimuli. Such light-derived signals are transmitted to the thalamus by retinal ganglion cells (RGCs), the sole projection neurons of the retina. Axons from RGCs innervate more than ten distinct nuclei within thalamus, including those of the lateral geniculate complex. Nuclei within the lateral geniculate complex of nocturnal rodents, which include the dorsal lateral geniculate nucleus (dLGN), ventral lateral geniculate nucleus (vLGN), and intergeniculate leaflet (IGL), are each densely innervated by retinal projections, yet, exhibit distinct cytoarchitecture and connectivity. These features suggest that each nucleus within this complex plays a unique role in processing and transmitting light-derived signals. Here, we review the diverse cytoarchitecture and connectivity of these nuclei in nocturnal rodents, in an effort to highlight roles for dLGN in vision and for vLGN and IGL in visuomotor, vestibular, ocular, and circadian function.

Early in development, before the onset of vision, the retina establishes direction-selective responses. During this time period, the retina spontaneously generates bursts of action potentials that propagate across its extent. The precise spatial and temporal properties of these “retinal waves” have been implicated in the formation of retinal projections to the brain. However, their role in the development of direction selective circuits within the retina has not yet been determined. We addressed this issue by combining multielectrode array and cell-attached recordings to examine mice that lack the CaV3.2 subunit of T-type Ca2+ channels (CaV3.2 KO) because these mice exhibit disrupted waves during the period that direction selective circuits are established. We found that the spontaneous activity of these mice displays wave-associated bursts of action potentials that are altered from that of control mice: the frequency of these bursts is significantly decreased and the firing rate within each burst is reduced. Moreover, the projection patterns of the retina demonstrate decreased eye-specific segregation in the dorsal lateral geniculate nucleus (dLGN). However, after eye-opening, the direction selective responses of CaV3.2 KO direction selective ganglion cells (DSGCs) are indistinguishable from those of wild-type DSGCs. Our data indicate that although the temporal properties of the action potential bursts associated with retinal waves are important for activity-dependent refining of retinal projections to central targets, they are not critical for establishing direction selectivity in the retina.

Retinal ganglion cell (RGC) dendritic atrophy is an early feature of many forms of retinal degeneration, providing a challenge to RGC classification. The characterization of these changes is complicated by the possibility that selective labeling of any particular class can confound the estimation of dendritic remodeling. To address this issue we have developed a novel, robust, and quantitative RGC classification based on proximal dendritic features which are resistant to early degeneration. RGCs were labeled through the ballistic delivery of DiO and DiI coated tungsten particles to whole retinal explants of 20 adult Brown Norway rats. RGCs were grouped according to the Sun classification system. A comprehensive set of primary and secondary dendrite features were quantified and a new classification model derived using principal component (PCA) and discriminant analyses, to estimate the likelihood that a cell belonged to any given class. One-hundred and thirty one imaged RGCs were analyzed; according to the Sun classification, 24% (n = 31) were RGCA, 29% (n = 38) RGCB, 32% (n = 42) RGCC, and 15% (n = 20) RGCD. PCA gave a 3 component solution, separating RGCs based on descriptors of soma size and primary dendrite thickness, proximal dendritic field size and dendritic tree asymmetry. The new variables correctly classified 73.3% (n = 74) of RGCs from a training sample and 63.3% (n = 19) from a hold out sample indicating an effective model. Soma and proximal dendritic tree morphological features provide a useful surrogate measurement for the classification of RGCs in disease. While a definitive classification is not possible in every case, the technique provides a useful safeguard against sample bias where the normal criteria for cell classification may not be reliable.

We have investigated the distribution of ON- and OFF-center responses in the lateral geniculate nucleus of ferrets with normal and abnormal retinal projections. Electrophysiological recordings in normal pigmented animals confirm previous studies on mustelids showing that ON-center responses are found in the anterior, inner parts of laminae A and AI and OFF-center responses in posterior, outer leaflets. In albino animals, lamina A displays normal patterns of ON/OFF segregation but in lamina AI, which receives an abnormal crossed retinal projection, no consistent patterns of segregation are found. Following monocular enucleation on the day of birth, the uncrossed projection in pigmented ferrets remains expanded across the LGN. Anatomically and physiologically, this projection is segregated into two leaflets: an anterior, inner ON-center leaflet and a posterior outer OFF-center leaflet. We conclude that the persistence of ON/OFF segregation, independent of geniculate location, suggests that self-sorting of retinal input is an important factor in generating the segregation.

The relative timing of a number of events during the development of the visual system has recently been suggested to be consistent across a number of mammalian species (Dreher & Robinson, 1988). Some conflicting reports, however, had suggested that the precocial guinea pig might represent an exception to the generalized scheme. A quantitative study was thus carried out on the development of the optic nerve and retina of the guinea pig. Consistent with the prediction of a stable relative time course of mammalian visual development, axons and growth cones were found in the optic stalk from the 24th postconceptional day (40% of the period from conception to eye opening —the cecal period), the peak number of axons was observed on the 32nd postconceptional day (56% of the cecal period), and the phase of rapid axonal loss extended to the 39th and 42nd postconceptional days (68–74% of the cecal period). The number of axons in the adult optic nerve (117,000) represented about 37% of the peak number of axons. Additional observations indicated that during development of the optic nerve the mean axonal diameter increased approximately threefold from 0.31 μm to 1.06 μm. As in other mammals studied so far, myelination was first noted after the period of rapid axonal loss and continued until in the adult 97% of axons were found to be myelinated. In the retina, the presence of pyknotic profiles in the ganglion cell layers extends throughout the periods of loss of the optic nerve axons. Finally, the presence of pyknotic profiles in the amacrine sublayer suggest that in the guinea pig, as in other mammalian species, there is a loss of displaced amacrine cells as well as ganglion cells from the ganglion cell layer.

The results of previous studies suggest that class-specific interactions contribute to the development of the different classes of retinal ganglion cells. We tested this hypothesis by examining the morphologies and distributions of alpha (α) cells in regions of mature cat retina selectively depleted of beta (β) cells as a result of visual cortex lesions at birth. We find that α cells in regions of central retina depleted of β cells are abnormally large while α cells in regions of peripheral retina depleted of β cells are abnormally small. The normal central-to-peripheral α cell soma-size gradient is absent in hemiretinae depleted of β cells. The dendritic fields of α cells in the border of β-cell-depleted hemiretina extend preferentially into the β-cell-poor hemiretina. In spite of this, α cell bodies retain their normal retinal distribution and remain distributed in a nonrandom mosaic-like pattern. Thus, it appears that the development of α retinal ganglion cells is influenced by interactions both with other α cells (class-specific interactions) and with surrounding β cells (nonclass-specific interactions).

A specific morphological class of ganglion cell, the alpha cell, was first defined in cat retina. Alpha cells have since been found in a wide range of mammalian retinae, including several orders of placental and marsupial mammals. Characteristically, they have the largest somata and a large dendritic field with a typical branching pattern. They occur as inner and outer stratifying subpopulations, presumably corresponding to ON-center and OFF-center receptive fields. In all species, alpha cells account for less than 10% of the ganglion cells, their somata are regularly spaced, and their dendritic fields evenly and economically cover the retina in a mosaic-like fashion. The morphology of alpha cells and many features, both of single cells and of the population, are conserved across species with different habitats and life-styles. This suggests that alpha cells are a consistent obligatory ganglion cell type in every mammalian retina and probably subserve some fundamental task(s) in visual performance.

Some general rules about the construction principles of ganglion cell classes are inferred from the alpha cells, stressing the importance of population parameters for the definition of a class. The principle, that a functionally and morphologically homogeneous population should have a regular arrangement and a complete and even coverage of the retina to perform its part in image processing at each retinal location, is especially evident across species and across ganglion cell types.

A new form of receptive field map, the response-component map, was developed to identify points within a receptive field that produce similar response patterns. The fields were probed with discretely flashed small spots of light. The magnitudes of the responses to stimulus onset and to stimulus offset elicited at each point were represented on the map by a vector radiating from the position representing the location of that point. Thus, response-component maps preserve the spatial distributions of responsivity and temporal nonlinearities. Points with similar response patterns were identified from a scatterplot in which the response at each spatial position was located in a plane representing the angles of the response-component vectors. Points with similar response characteristics that were also spatially contiguous were considered as a distinct response subregion within the receptive field.

Barely 10% of the receptive fields of goldfish ganglion cells mapped with this technique proved as simple as the traditional concentric field described for retinal cells. In at least 17% of the cases, the field showed three concentric rings, with a very small “inner center” within the center of the field. In at least 50% of the cases, response subregions of different type lay side by side, rather than in a concentric configuration. Some subregions could be differentiated by the relative strengths of the responses to onset and offset of the stimulus spot, supporting the hypothesis that a push-pull system generates ganglion cell responses. Subregions were evident in successive mappings of the same cell, demonstrating they are not due to the vagaries of individual responses. They probably represent the spatial domains (or their intersections) of individual interneurons distal to the retinal ganglion cells. It is possible that position within the receptive field may be coded by the temporal pattern of the responses.

γ-aminobutyric acid (GABA) evokes large whole-cell currents in solitary mammalian retinal ganglion cells studied by the patch-clamp method. This evidence suggests that GABA acts directly on the retinal ganglion cells as an inhibitory transmitter as it does elsewhere in the mammalian central nervous system. Here, single-channel recordings of the currents underlying the GABA-induced responses were studied in outside-out patches of cell membrane. In some other preparations, single GABAA channels recorded in the excised patch configuration have been shown to have altered properties in comparison to responses elicited during whole-cell recording. For example, in cortical neurons single GABA-activated channels in excised patches display accelerated desensitization kinetics as well as rapid rundown of the response. Therefore, in retinal ganglion cells, responses generated by GABA in cell-free patches were compared to whole-cell responses. After determining that the responses to GABA in acutely isolated outside-out patches were indeed similar to those of the whole-cell currents in retinal ganglion cells, the unitary conductances were studied. It was determined that these single-channel events resemble those reported in other nervous tissues with 4 elementary conductances of ~10 pS, 19–22 pS, 30–33 pS, and 45–50 pS at 33–35°C.

Intracellular injections of Neurobiotin were used to determine whether the major ganglion cell classes of the macaque monkey retina, the magnocellular-projecting parasol, and the parvocellular-projecting midget cells showed evidence of cellular coupling similar to that recently described for cat retinal ganglion cells. Ganglion cells were labeled with the fluorescent dye acridine orange in an in vitro, isolated retina preparation and were selectively targeted for intracellular injection under direct microscopic control. The macaque midget cells, like the beta cells of the cat's retina, showed no evidence of tracer coupling when injected with Neurobiotin. By contrast, Neurobiotin-filled parasol cells, like cat alpha cells, showed a distinct pattern of tracer coupling to each other (homotypic coupling) and to amacrine cells (heterotypic coupling).

In instances of homotypic coupling, the injected parasol cell was surrounded by a regular array of 3–6 neighboring parasol cells. The somata and proximal dendrites of these tracer-coupled cells were lightly labeled and appeared to costratify with the injected cell. Analysis of the nearest-neighbor distances for the parasol cell clusters showed that dendritic-field overlap remained constant as dendritic-field size increased from 100–400 μm in diameter.

At least two amacrine cell types showed tracer coupling to parasol cells. One amacrine type had a small soma and thin, sparsely branching dendrites that extended for 1–2 mm in the inner plexiform layer. A second amacrine type had a relatively large soma, thick main dendrites, and distinct, axon-like processes that extended for at least 2–3 mm in the inner plexiform layer. The main dendrites of the large amacrine cells were closely apposed to the dendrites of parasol cells and may be the site of Neurobiotin transfer between the two cell types. We suggest that the tracer coupling between neighboring parasol cells takes place indirectly via the dendrites of the large amacrine cells and provides a mechanism, absent in midget cells, for increasing parasol cell receptive-field size and luminance contrast sensitivity.

We have examined the retinal decussation patterns in pigmented ferrets that were either wild-type sable or heterozygous with one albino gene. Unilateral injections of horseradish peroxidase were made into the optic tract and labeled ganglion cells visualized in retinal wholemounts. In both wild-type and heterozygous ferrets, those ganglion cells in the temporal retina with the largest cell bodies projected only to the contralateral side of the brain. The total number of ipsilaterally projecting ganglion cells did not differ with the genotype of the animal. The numbers ranged from 5471–6759 cells. Unlike the cat, there is no difference in retinal decussation patterns in wild-type sable ferrets and heterozygous ferrets carrying one albino gene.

We have examined the time course of branch formation in neurites of retinal ganglion cells isolated from adult goldfish (Carassius auratus). These neurites elongate at approximately 13 μm/h, and usually branch by bifurcation of growth cones at their tips. The times elapsed between branchings in different neurites of single cells can be described by a Poisson distribution with a mean interval of approximately 2 h. As predicted by this distribution, a relatively large number of branchings occur simultaneously in different neurites of individual cells. Simultaneous branchings of neurites elongating at a common rate generate branch points that lay equidistant from their soma. Since similar branching patterns can be seen in dendrites of retinal ganglion and amacrine cells in situ, these results are consistent with the possibility that dendrites of individual neurons branch synchronously and grow at common rates during development.

The center of the receptive field of some retinal ganglion cells exhibits an interesting fine structure: the relative amplitudes of responses to onset and responses to offset of a small spot of light varies systematically as the spot is positioned at various places within the center. Although this pattern may appear complicated, a simple model can account for it in detail. The model postulates that the ganglion cell receives input from separate ON and OFF processes within the center of its receptive field. These processes have the form of Gaussian functions and are laterally displaced from each other. These central ON and OFF input processes are not associated with the additional antagonistic surround of the receptive field.

The model is examined for various parameters of the input processes. The observed systematic variation in the ratio of offset to onset responses is predicted when the two processes are of nearly equal width (standard deviation of the Gaussians). Receptive fields made of more than two Gaussians produce various patterns, depending on the relative standard deviations of the Gaussians. Oblong fields, reminiscent of those found in visual cortex, may be generated from a relatively small number of circularly symmetric Gaussian processes.

Action potentials were recorded from isolated goldfish retinae maintained in a superfusate of Ringer's solution. Responses to flashes of light and maintained discharges were obtained from 84 cells. The properties of these cells were compared to those in two other goldfish preparations: the isolated retina maintained in a flow of moist oxygen and the self-respiring fish. Maintained discharges of cells in the superfused retinae tended to have lower mean firing rates, higher variability, and weaker high-pass properties than had been observed in the previous preparations. These properties seemed insensitive to the particular formulae used to superfuse the retinae.

Cobalt, which disables synapses, dramatically reduced maintained firing and eliminated photic responses. Cells that did fire in the presence of cobalt generally had low variabilities before cobalt was added; their firing in cobalt was considerably more variable than the baseline. Nevertheless, cobalt did not seem to change the temporal dependency (high-pass properties) of the maintained discharges. The cholinergic agonist carbachol had an excitatory effect upon 71% of the cells tested. Bursty or oscillatory firing in cobalt was rendered more regular by the addition of carbachol. With the exception of the mean firing rate, none of the statistical properties of the maintained discharge differed in cobalt plus carbachol from those in normal Ringer's solution.

There was a tendency for the statistical properties of the maintained discharge after the treatment to approach those of the previously reported preparations; the treatment was at least partially responsible for the drift in properties. The results are discussed in terms of the possible sources of variability in the ganglion cell's discharge, with particular reference to the high-pass filter that appears to act upon it.

Neighboring ganglion cells in the retinae of vertebrates show cross-correlation between their impulse trains. Cross-correlation is found both in maintained discharges and during responses to visual stimulation. There has been speculation about the function of this statistical dependence, but little is known about its genesis. This study examines the statistics of the interimpulse intervals preceding and those following impulses that coincide with an impulse in the other train. Short intervals are rarer than expected preceding a coincidence, regardless of the form of the cross-correlation. Short intervals are more common than expected following a coincidence when the cross-correlation is positive, but rarer than expected following coincidences during negative cross-correlation. These results contradict the extant models for cross-correlation, but may be explained by the multiplicative combination of a variable common input and the variability within each cell. In addition, the lag (relative timing of coincident impulses in the two cells) is found to be related to the maintained discharge rates of the cells, implying that the lags may be explained without invoking specific delay circuits.

Acute hypoxemia (low PaO2) leads to changes in oxygen consumption and electrical responses of the outer retina of cats, but inner retinal ERG components and ganglion cell responses have been shown to be quite resistant to hypoxemia. The purpose of this study was to determine whether the resistance of the inner retina depends on (1) the stimulus conditions, specifically the degree of light adaptation; and (2) the ability of the photoreceptors to increase glycolysis during hypoxemia. To address these issues, recordings of single ganglion cell action potentials and of the b-wave and scotopic threshold response (STR) of the electroretinogram (ERG) were made from the eyes of anesthetized cats during hypoxemia alone and hypoxemia plus hypoglycemia. Ganglion cells appeared to be equally resistant to hypoxemia at high and low backgrounds (3.3 to 9.7 log equivalent quanta(555 nm)-deg-2-s-1), and the STR, recorded with dim stimuli during dark adaptation, when photoreceptor oxygen consumption is most susceptible to hypoxemia, was unchanged until PaO2 was below 30 mm Hg. The amplitude of the b-wave was similarly resistant to hypoxemia when the animal was normoglycemic. During hypoglycemia, however, both the b-wave and the STR became more sensitive to hypoxemia, beginning to change at PaO2s as high as 50 mm Hg when blood glucose was 40–50 mg/dl. It is argued that hypoglycemia limits or prevents the increased glycolytic ATP production that would ordinarily occur when the photoreceptor oxygen supply decreases, and that increased photoreceptor glycolysis is essential in the protection of the retina against mild hypoxemia.

The present study provides evidence that the adult mammalian retina is highly sensitive to the excitotoxic action of NMDA. In particular, we have investigated the effects of a single intravitreal injection of different doses of N-methyl-D-aspartate (NMDA) (2–200 nmoles) on the adult rat retina. Morphological evaluation of transverse sections of retinae demonstrated a dose-dependent loss of cells in the ganglion cell layer (GCL) and a reduction in the thickness of the inner plexiform layer. No obvious alterations were noted in the more distal retinal layers. Quantitative analyses of Nissl-stained whole-mounted retinae revealed that administration of 20 nmoles of NMDA resulted in a 70% loss of cells with a soma diameter greater than 8 μm (presumed retinal ganglion cells); a 20% loss of cells with a soma diameter smaller than 8 μm (presumed displaced amacrine cells) was also observed. In addition, NMDA produced a dose-dependent decrease of retinal choline acetyltransferase (ChAT) activity, suggesting that NMDA affects cholinergic amacrine cells as well. MK-801, a non-competitive NMDA antagonist, completely prevented the NMDA-induced loss of cells in the GCL and blocked, in a dose-dependent manner, the NMDA-induced decrease of ChAT activity. The excitotoxic action of NMDA observed in these experiments is thus likely mediated through the NMDA receptor subtype. This ”in vivo” model may be utilized to identify potential drugs that antagonize or limit the deleterious effects consequent to NMDA receptor overstimulation in the central nervous system.